ArticleLiterature Review

A Requirement for Copper in Angiogenesis

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Abstract

Although two decades have passed since copper was shown to stimulate blood vessel formation in the avascular cornea of rabbits, only recently have clinical trials established that Cu privation by diet or by Cu chelators diminishes a tumor's ability to mount an angiogenic response. These data have shed new light on the functional role of Cu in microvessel development and, of equal importance, stimulated new nutritional models of cancer therapeutic intervention.

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... Cancer cells and cells in atherosclerosis have higher Cu levels and more predominant nuclear localization of Cu in some cells than normal tissues (9,51). Cu chelation prevents tumor growth and neointimal thickening after vascular injury (27,58,65,119). Furthermore, nuclear accumulation of Cu is strongly associated with proliferation of hepatocytes in ATP7B Ϫ/Ϫ mice, an animal model of Wilson disease, before the appearance of histopathological alterations (72). Because dysregulation of cell proliferation and migration is frequently associated with a number of diseases, an understanding of the molecular mechanism by which Cu stimulates cell proliferation/migration will potentially provide the targets for new therapeutic strategies. ...
... Cu plays an important role in physiological and pathological angiogenesis (27,28,46,58,65,193). Cu or Cu complexes directly stimulate angiogenesis in a number of model systems including the rabbit corneal system (159) and various source of endothelial cells (6,129). ...
... Cu plays an important role in physiological and pathological angiogenesis (27,28,46,58,65,193). Cu or Cu complexes directly stimulate angiogenesis in a number of model systems including the rabbit corneal system (159) and various source of endothelial cells (6,129). Furthermore, Cu chelators inhibit tumor growth and angiogenic responses (27,28,58,65) in numerous animal and xenograft models (58). Cu metabolism appears to be altered in tumors (8,75), while angiogenic lesions in cancer have higher Cu levels in cell nuclei than normal tissues and serum Cu levels themselves appear to be elevated in a number of tumor types (36) including breast cancer (61). ...
Article
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Copper (Cu) is an essential micronutrient but excess Cu is potentially toxic. Its important propensity to cycle between two oxidation states accounts for its frequent presence as a co-factor in many physiological processes through Cu-containing enzymes, including mitochondrial energy production (via cytochrome c oxidase), protection against oxidative stress (via superoxide dismutase), and extracellular matrix stability (via lysyl oxidase). Since free Cu is potentially toxic, the bioavailability of intracellular Cu is tightly controlled by Cu transporters and Cu chaperones. Recent evidence reveals that these Cu transport systems play an essential role in the physiological responses of cardiovascular cells, including cell growth, migration, angiogenesis and wound repair. In response to growth factors, cytokines and hypoxia, their expression, subcellular localization and function are tightly regulated. Cu transport systems and their regulators have also been linked to various cardiovascular pathophysiologies such as hypertension, inflammation, atherosclerosis, diabetes, cardiac hypertrophy and cardiomyopathy. A greater appreciation of the central importance of Cu transporters and Cu chaperones in cell signaling and gene expression in cardiovascular biology offers the possibility of identifying new therapeutic targets for cardiovascular disease.
... Copper (Cu) has important roles in angiogenesis through unknown mechanisms [20][21][22][23] . Cellular Cu entry occurs mainly through the Cu transporter CTR1 (ref. ...
... Furthermore, mCtr1-KO ECs showed almost complete inhibition of VEGF-induced signalling events (Fig. 2e). As Cu entry is required for activating the Cu-dependent enzyme LOX, which is involved in angiogenesis [20][21][22]33,41,42 , and for Cu binding to MEK1/2, which increases p-ERK1/2 (ref. 32 ), we examined the role of Cu in VEGF-induced signalling. ...
... How the newly identified CTR1-VEGFR2 complex interacts with VEGFR2-binding proteins to facilitate internalization is worthy of future investigation. Cu entry is involved in direct activation of MEK by Cu binding, stimulating phosphorylation of ERK 32 , growth factor signalling by FGF and insulin, and activation of the Cu enzyme LOX, which is involved in angiogenesis [20][21][22]33,41,42 . We found that VEGF-induced CTR1-Cys 189 OH formation promotes the phosphorylation of MEK1/2 and ERK1/2 in a Cu-transport-independent manner in ECs. ...
Article
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Vascular endothelial growth factor receptor type 2 (VEGFR2, also known as KDR and FLK1) signalling in endothelial cells (ECs) is essential for developmental and reparative angiogenesis. Reactive oxygen species and copper (Cu) are also involved in these processes. However, their inter-relationship is poorly understood. Evidence of the role of the endothelial Cu importer CTR1 (also known as SLC31A1) in VEGFR2 signalling and angiogenesis in vivo is lacking. Here, we show that CTR1 functions as a redox sensor to promote angiogenesis in ECs. CTR1-depleted ECs showed reduced VEGF-induced VEGFR2 signalling and angiogenic responses. Mechanistically, CTR1 was rapidly sulfenylated at Cys189 at its cytosolic C terminus after stimulation with VEGF, which induced CTR1–VEGFR2 disulfide bond formation and their co-internalization to early endosomes, driving sustained VEGFR2 signalling. In vivo, EC-specific Ctr1-deficient mice or CRISPR–Cas9-generated redox-dead Ctr1(C187A)-knockin mutant mice had impaired developmental and reparative angiogenesis. Thus, oxidation of CTR1 at Cys189 promotes VEGFR2 internalization and signalling to enhance angiogenesis. Our study uncovers an important mechanism for sensing reactive oxygen species through CTR1 to drive neovascularization.
... In order to achieve the appropriate mechanical properties values and to address the problems of mechanical properties instability and strain rate sensitivity simultaneously, suitable alloying strategy is one of the solutions. Copper (Cu) is an essential trace element in the human body, and Cu ions have the functions of promoting endothelialization and antibacterial [19,29]. Besides, Cu has a medium solid solubility in Zn alloys (~2.75 wt% at 425 • C), and its solubility decreases with decreasing the temperature [30], so it is feasible Zn-Cu alloy can achieve solid solution strengthening and precipitation strengthening. ...
... mg/L that inhibited cell viability (endothelial and smooth muscle cells) was in line with previous studies [32,46,47], and tolerance of endothelial cells to Zn 2+ content is better than that of smooth muscle cells. The trace Cu 2+ release has a positive effect on endothelial cell viability [29,32], but accelerated Zn 2+ releases in 0Mn and 0.4Mn alloys could weaken or offset this effect, as shown in Fig. 12a and c. Although in vitro cytotoxicity deteriorated in 100% concentration extracts, suitable diluted extracts were recommended for in vitro cytotoxicity evaluation of pure Mg [48], and a similar method should be applicable to biodegradable Zn-based alloys. ...
Article
Recently, zinc (Zn) alloy has been considered as a promising biodegradable material due to its excellent physiological degradable behavior and acceptable biocompatibility. However, poor mechanical performance limits its application as vascular stents. In this study, novel biodegradable Zn-2.2Cu-xMn (x = 0.4, 0.7, and 1.0 wt%) alloys with suitable mechanical performance were investigated. The effects of Mn addition on microstructure, mechanical properties, and in vitro degradation of Zn-2.2Cu-xMn alloys were systematically investigated. After adding Mn, dynamic recrystallization (DRX) during hot extrusion was promoted, resulting in slightly finer grain size, higher DRXed regions ratio, and weaker texture. And volume fraction and number density of second phase precipitates (micron, submicron, and nano-sized ε and MnZn13 phase) and the concentration of (Cu, Mn) in the matrix were increased. Therefore, Zn-2.2Cu-xMn alloys exhibited suitable mechanical performances (strength >310 MPa, elongation >30%) mainly due to the combination effects of grain refinement, solid solution strengthening, second phase precipitation hardening, and texture weakening. Moreover, the alloys maintained good stability of mechanical properties within 18 months and good elongation over 15% even at a high strain rate of 0.1 s⁻¹. In addition, the alloys presented appropriate in vitro degradation rates in a basically uniform degradation mode and acceptable in vitro cytocompatibility. The above results indicated that the newly designed biodegradable Zn-2.2Cu-0.4Mn alloy with suitable comprehensive mechanical properties, appropriate degradation behavior, and acceptable cytocompatibility is a promising candidate for vascular stents.
... Numerous studies have shown that an elevated level of copper is directly correlated with cancer progression. Additionally, copper concentration correlated with the age of patients and the stage of cancer [20][21][22][23][24]. Excess of copper ions was found in the serum and tissues of patients with, among others: breast [24,25], prostate [26,27], colorectal [28], lung [29] and brain [30] cancers, compared to healthy people. ...
... Previously it was mentioned that cancer cells exhibit an increased concentration of copper ions [20][21][22][23][24]. Moreover, it is broadly known that some quinoline derivatives possess metal ions complexing properties [18] what is essential for their anti-proliferative properties [51]. ...
Article
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Small molecule nitrogen heterocycles are very important structures, widely used in the design of potential pharmaceuticals. Particularly, derivatives of 8-hydroxyquinoline (8-HQ) are successfully used to design promising anti-cancer agents. Conjugating 8-HQ derivatives with sugar derivatives, molecules with better bioavailability, selectivity, and solubility are obtained. In this study, 8-HQ derivatives were functionalized at the 8-OH position and connected with sugar derivatives (D-glucose or D-galactose) substituted with different groups at the anomeric position, using copper(I)-catalyzed 1,3-dipolar azide-alkyne cycloaddition (CuAAC). Glycoconjugates were tested for inhibition of the proliferation of cancer cell lines (HCT 116 and MCF-7) and inhibition of β-1,4-galactosyltransferase activity, which overexpression is associated with cancer progression. All glycoconjugates in protected form have a cytotoxic effect on cancer cells in the tested concentration range. The presence of additional amide groups in the linker structure improves the activity of glycoconjugates, probably due to the ability to chelate metal ions present in many types of cancers. The study of metal complexing properties confirmed that the obtained glycoconjugates are capable of chelating copper ions, which increases their anti-cancer potential.
... Lysyl oxidase is an enzyme that catalyses the formation of aldehyde-based crosslinks in collagen [33]. Copper has also been found to be an essential element for angiogenesis, leading to hypotheses regarding its value in promoting the vascularisation of bone [629][630][631][632][633]. It has been shown that copper-containing scaffolds in vivo are able to promote blood vessel formation due to superior collagen formation. ...
Thesis
One of the greatest requirements of modern medicine is the ability to treat patients suffering from osteoarthritis (OA) and bone fractures. Currently, there is no long-term therapy for OA; symptoms can be managed with anti-inflammatories and analgesics until they worsen to the extent that the damage becomes debilitating, and joint arthroplasty, is necessitated. However, these replacements are not perfect; firstly, there is the need for surgery and secondly, if the patient is young, the prosthetic can deteriorate, engendering further surgery. Bone fractures are regularly seen in orthopaedic clinics and are commonly repaired using fixation techniques or biomaterials. After any intervention, the fracture site can remain compromised, potentially engendering re-fracture and/or further surgical involvement. Regenerative strategies for both OA and bone fracture aim to alleviate pain, whilst maintaining or restoring damaged tissues to healthy states. Mesenchymal stem/stromal cells (MSC) are thought to facilitate tissue repair via either progenitor or secreaome functions. BM-MSC have, in previous work, been investigated as a therapy for OA via either their direct application or through their secreted Extracellular Vesicles (EV). In this study, MSC have been successfully isolated from bone marrow, and from these isolated cells, EV have been captured and characterised. The isolated EV have been shown to be readily internalised by chondrocytes and, in order to determine the method of EV internalisation by chondrocytes, in vitro drug inhibition studies were performed on labelled EV. Via inhibition of the caveolin dependent endocytosis pathway, EV uptake was prevented, thus indicating that this method of endocytosis is the method of EV internalisation. In regenerative medicine for knee OA, it is likely that MSC and EV would be injected into the knee. In order to determine if the MSC and EV would reside in the joint, both were labelled with gold nanostars and Supra Magnetic Iron Oxide Nanoparticles (SPION). These labelled cells and EV were then injected into a sheep stifle 1 week post creation of an OA model (meniscal transection model). These labelled cells and EV could then be seen within the knee for up to 4 weeks post injection, as ascertained via Magnetic Resonance Imaging (MRI) and MultiSpectral Optoacoustic Tomography (MSOT). Upon evaluating the regenerative effects of the MSC and EV, no difference in cartilage damage could be seen. During bone fracture, MSC and osteoblasts are recruited to the site of injury. Bioglasses have been used previously as a material to improve bone repair through the release of ions and conditioning the local environment. Our work has shown that conditioned media from bioglasses can influence both MSC and osteoblasts to augment the bone repair process. Through screening bioglasses on MSC and osteoblasts, the potential for bioglasses to alter MSC derived EV to promote osteogenesis has been shown. As a conclusion, this study has shown that the BM-MSC are a source of EV, and that both the MSC and EV can potentially be used in a musculoskeletal scenario of regenerative medicine.
... CP usually has antioxidant properties, but in conditions such as diabetes mellitus and hyperhomocysteinemia, it promotes vasculopathic effects that include lipid oxidation, the negation of nitric oxide bioactivity, and endothelial cell apoptosis via its copper content [29]. Moreover, copper stimulates blood vessel formation by promoting endothelial cell migration and angiogenesis [30,31]. These findings suggest that CP is involved in the development of inflammation, apoptosis, and angiogenesis. ...
Article
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Purpose To examine the difference in the vitreal protein profiles of patients with proliferative diabetic retinopathy (PDR) with and without preoperative intravitreal conbercept (IVC) treatment. Methods Liquid chromatography-tandem mass spectrometry- (LC-MS/MS-) based proteomic methods were used to determine the protein profiles of the vitreous humor in patients with PDR treated with (IVC group; n = 9) and without (PDR group; n = 8) preoperative IVC. Gene ontology (GO) annotation and REACTOME pathway analysis were obtained to overview differentially expressed proteins between each group. Intravitreal levels of apolipoprotein A-II (APOA2) and ceruloplasmin (CP) were measured using enzyme-linked immunosorbent assays. Results 307 proteins were expressed differentially between PDR and IVC groups, including 218 proteins downregulated in response to IVC. The most notable GO annotations in level 3 and REACTOME pathways describing the differentially expressed proteins were “innate immune response” and “platelet degranulation.” The intravitreal levels of APOA2 and CP were lower in the IVC group than in the PDR group (p < 0.01). Conclusions In addition to decreasing the intravitreal vascular endothelial growth factor level, IVC may alter the vitreal protein profile in patients with PDR, with the differentially regulated proteins involved in the immune response, platelet degranulation, complement activation, and inflammation.
... Angiogenesis plays a crucial role in efficient wound healing and tissue regeneration, especially in cardiovascular patch implanting position. Copper has previously been suggested to be proangiogenic by multiple mechanisms that include releasing various angiogenic factors and stimulating endothelial cell proliferation [50,51]. In this study, we found that Cu@TP-dBPs had an improved proangiogenic effect both in vitro and in vivo. ...
Article
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Decellularization is a promising technique to produce natural scaffolds for tissue engineering applications. However, non-crosslinked natural scaffolds disfavor application in cardiovascular surgery due to poor biomechanics and rapid degradation. Herein, we proposed a green strategy to crosslink and functionalize acellular scaffolds via the self-assembly of copper@tea polyphenol nanoparticles (Cu@TP NPs), and the resultant nanocomposite acellular scaffolds were named as Cu@TP-dBPs. The crosslinking degree, biomechanics, denaturation temperature and resistance to enzymatic degradation of Cu@TP-dBPs were comparable to those of glutaraldehyde crosslinked dBPs (Glut-dBPs). Furthermore, Cu@TP-dBPs were biocompatible and had abilities to inhibit bacterial growth and promote the formation of capillary-like networks. Subcutaneous implantation models demonstrated that Cu@TP-dBPs were free of calcification and allowed for host cell infiltration at day 21. Cardiac patch graft models confirmed that Cu@TP-dBP patches showed improved ingrowth of functional blood vessels and remodeling of extracellular matrix at day 60. These results suggested that Cu@TP-dBPs not only had comparable biomechainics and biostability to Glut-dBPs, but also had several advantages over Glut-dBPs in terms of anticalcification, remodeling and integration capabilities. Particularly, they were functional patches possessing antibacterial and proangiogenic activities. These material properties and biological functions made Cu@TP-dBPs a promising functional acellular patch for cardiovascular applications.
... It is elevated in conditions of acute inflammation. Cp contains seven copper atoms per molecule, participates in copper transport and metabolism, and has ferroxidase activity [9,10]. Furthermore, Cp is involved in the modulation of coagulation and angiogenesis and the inactivation of biogenic amines [11,12]. ...
Article
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We investigated whether the additional determination of ceruloplasmin (Cp) levels could improve the prognostic value of N-terminal pro-B-type natriuretic peptide (NT-proBNP) in heart failure (HF) patients in a 1-year follow-up. Cp and NT-proBNP levels and clinical and laboratory parameters were assessed simultaneously at baseline in 741 HF patients considered as possible heart transplant recipients. The primary endpoint (EP) was a composite of all-cause death (non-transplant patients) or heart transplantation during one year of follow-up. Using a cut-off value of 35.9 mg/dL for Cp and 3155 pg/mL for NT-proBNP (top interquartile range), a univariate Cox regression analysis showed that Cp (hazard ratio (HR) = 2.086; 95% confidence interval (95% CI, 1.462–2.975)), NT-proBNP (HR = 3.221; 95% CI (2.277–4.556)), and the top quartile of both Cp and NT-proBNP (HR = 4.253; 95% CI (2.795–6.471)) were all risk factors of the primary EP. The prognostic value of these biomarkers was demonstrated in a multivariate Cox regression model using the top Cp and NT-proBNP concentration quartiles combined (HR = 2.120; 95% CI (1.233–3.646)). Lower left ventricular ejection fraction, VO2 max, lack of angiotensin-converting enzyme inhibitor or angiotensin receptor blocker therapy, and nonimplantation of an implantable cardioverter-defibrillator were also independent risk factors of a poor outcome. The combined evaluation of Cp and NT-proBNP had advantages over separate NT-proBNP and Cp assessment in selecting a group with a high 1-year risk. Thus multi-biomarker assessment can improve risk stratification in HF patients.
... Sufficient evidence shows that Cu stimulates and is required for VEGF expression [55,57]. However, this regulating activity is not the only action of Cu in angiogenesis; Cu also affects the expression and activation of other factors involved in blood vessel formation and maturation [58]. Therefore, Cu promoting blood vessel formation has a significant advantage over the effect of VEGF alone because Cu promotes the formation and maturation of new blood vessels. ...
Preprint
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Preeclampsia (PE) is characterized by a series of clinical features such as hypertension and proteinuria associated with endothelial dysfunction and the impairment of placenta vascular endothelial integrity. This study aimed to investigate the effect of serum copper (Cu) level on some angiogenesis-related factors including vascular endothelial growth factor-A (VEGF-A), soluble Fms-like tyrosine kinase-1 (sVEGF-R1), soluble endoglin (sEng) and cerruloplasmin (Cp) in Iraqi women with preeclampsia (PE) and control pregnant women. Therefore, 60 women with PE in addition to 30 healthy pregnant women were enrolled in the study. Serum concentration of sEng, VEGF-A, sVEGF-R1, and Cu in PE group significantly increased (p<0.05) in the PE group compared with that in the control group. Increased production of antiangiogenic factors, soluble VEGF-A and sEng contribute to the pathophysiology of PE, indicating the involvement of these parameters in the angiogenic balance in patients with PE. Tests for between-subject effects showed that the circulating angiogenesis factors and Cu were significantly associated with the presence of PE. Serum Cu level was significantly correlated with VEGF- A and VEGF-R1 levels but not with sEng. Multiple regression analysis revealed that only Cp and BP can significantly predict the complications in women with PE. In conclusion, serum Cu has a role in the angiogenesis in women with PE and may be a new drug target in the prevention or treatment of PE.
... It's well known that copper (Cu) is one of the essential trace elements for the survival of living organisms and mainly exists in the form of bivalent copper (Cu 2+ ) [9]. Cu has beneficial effects on osteogenesis, angiogenesis, and antimicrobial [10][11][12]. Early studies on mice diet supplemented with Cu demonstrated that Cu exerted effect on improving bone mineralization and increasing weight of femurs [13], and Cu depletion could cause osteoporosis that was shown radiologically and histologically [14,15]. ...
Article
Development of multifunctional bone grafting biomaterials with both osteogenesis and angiogenesis properties have earned increasing interest in the field of regenerative medicine. In the present investigation, copper-doped β-tricalcium phosphate (Cu-TCP) powders were successfully synthesized. And Cu-containing calcium phosphate cement (Cu-CPC) was acquired through uniformly mixing CPC and Cu-TCP powders, with Cu-TCP serving as the donor of Cu2+. Cu-CPC exhibited suitable setting time, and the incorporation of Cu-TCP aggregating into CPC exhibited positive effect on the compressive strength while Cu2+ was in lower concentration. Investigation results showed that Cu-CPC had relatively low releasing amount of Cu2+, which was attributed to the re-bonding of Cu2+ into the newly formed HA crystals on surface. In vitro osteogenesis and angiogenesis properties of Cu-CPC were systematically evaluated through co-culture with mouse bone marrow stromal cells (mBMSCs) and human umbilical vein endothelial cells (HUVECs) respectively. The results indicated dose-dependent biological functions of Cu2+ in Cu-CPCs. The mBMSCs and HUVECs showed well activity and attachment morphology on TCP/CPC, 0.05 Cu-TCP/CPC, 0.1 Cu-TCP/CPC. The upregulated osteogenic-related genes expression and angiogenic-related genes expression were detected with lower Cu2+ content. Taken together, Cu-containing CPC is of great potential for the regeneration of vascularized new bone.
... These mechanisms may contribute to the repair and maintenance of the functional integrity of blood vessels, providing insight into central roles for IQGAP1 and ATP7A in vascular homeostasis. Of note, Cu levels are significantly increased in cancer and atherosclerotic lesions (12,43), and Cu chelators prevent neointima thickening in response to PDGF promotes IQGAP1 translocation from the cytoplasm to the leading edge, thereby stimulating lamellipodia formation via recruiting ATP7A and Rac1, which in turn promotes directional VSMC migration involved in neointimal formation. ATP7A may transports copper to the secretory copper enzymes at the leading edge, which may promote extracellular matrix (ECM) remodeling and VSMC migration. ...
Article
Vascular smooth muscle cell (VSMC) migration contributes to neointimal formation after vascular injury. We previously demonstrated that copper (Cu) transporter ATP7A is involved in PDGF-induced VSMC migration in Cu- and Rac1-dependent manner. Underlying mechanism is still unknown. Here we show that ATP7A interacts with IQGAP1, a Rac1- and receptor tyrosine kinase-binding scaffold proteins, which mediates PDGF-induced VSMC migration and vascular remodeling. In cultured rat aortic SMCs, PDGF stimulation rapidly promoted ATP7A association with IQGAP1 and Rac1, and their translocation to lipid rafts and leading edge. Co-transfection assay revealed that ATP7A directly bound to N-terminal domain of IQGAP1. Functionally, either ATP7A or IQGAP1 depletion using siRNA significantly inhibited PDGF-induced VSMC migration without additive effects, suggesting that IQGAP1 and ATP7A are in the same axis to promote migration. Furthermore, IQGAP1 siRNA blocked PDGF-induced ATP7A association with Rac1 as well as their translocation to leading edge while PDGF-induced IQGAP1 translocation was not affected by ATP7A siRNA or Cu chelator. Overexpression of mutant IQGAP1 lacking Rac1 binding site prevented PDGF-induced translocation of Rac1, but not ATP7A, to the leading edge, thereby inhibiting lamellipodia formation and VSMC migration. In vivo, ATP7A colocalized with IQGAP1 at neointimal VSMC in mice wire injury model, while neointimal formation and extracellular matrix deposition induced by vascular injury were inhibited in ATP7A mutant mice with reduced Cu transporter function. In summary, IQGAP1 functions as ATP7A and Rac1 binding scaffold protein to organize PDGF-dependent ATP7A translocation to the lamellipodial leading edge, thereby promoting VSMC migration and vascular remodeling.
... Copper (Cu) is one of the well-known antibacterial metals [16] and can offer a high antibacterial activity for other metals with proper Cu addition [17][18][19]. Cu is also one of the essential elements in the human body and plays an important role in the immune system [20,21]. A small amount of copper content in alloys or bioactive glass has been reported to improve the cytocompatibility and cell viability [22,23]. ...
... Copper (Cu) is an essential trace element necessary for development of connective tissues, nerve coverings and bone growth [91]. Cu promotes the proliferation of vascular endothelial cells, thus accelerating the revascularization process [92,93]. Cu deficiency causes normocytic anemia, neuropenia and abnormal glucose and cholesterol metabolism [94]. ...
Article
Statement of significance: Numerous studies on magnesium and iron materials have been reported to date, in an effort to formulate bioabsorbable stents with tailorable mechanical characteristics and corrosion behavior. Crucial concerns regarding poor ductility and remarkably rapid corrosion of magnesium, and very slow degradation of iron, seem to be still not desirably fulfilled. Zinc was introduced as a potential implant material in 2013 due to its promising biodegradability and biocompatibility. Since then, extensive investigations have been made toward development of zinc alloys that meet clinical benchmarks for vascular scaffolding. This review critically surveys the zinc alloys developed since 2013 from metallurgical and biodegradation points of view. Microstructural features, mechanical, corrosion and in vivo performances of these new alloys are thoroughly reviewed and evaluated.
... This finding supports previous research into this area which links copper and angiogenesis. Once the proportion of metal in the alloys has increased, it acted as a pleiotrophic agent assisted in angiogenesis, which eased the release of mediators of angiogenesis [73,78,79]. The contribution of Cu to the stimulation of hypoxia-inducible factor-1 (HIF-1) as the transcription factor and its effect on regulating the expression of VEGF was already approved, a platform on which vascular systems in the skeletal muscles can be developed [80]. ...
Article
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There have been several attempts to design innovative biomaterials as surface coatings to enhance the biological performance of biomedical implants. The objective of this study was to design multifunctional Cu/a-C:H thin coating depositing on the Ti-6Al-4V alloy (TC4) via magnetron sputtering in the presence of Ar and CH4 for applications in bone implants. Moreover, the impact of Cu amount and sp²/sp³ ratio on the interior stress, corrosion behavior, mechanical properties, and tribological performance and biocompatibility of the resulting biomaterial was discussed. X-ray photoelectron spectroscopy (XPS) revealed that the sp²/sp³ portion of the coating was enhanced for samples having higher Cu contents. The intensity of the interior stress of the Cu/a-C:H thin bio-films decreased by increase of Cu content as well as the sp²/sp³ ratio. By contrast, the values of Young's modulus, the H³/E² ratio, and hardness exhibited no significant difference with enhancing Cu content and sp²/sp³ ratio. However, there was an optimum Cu content (36.8 wt.%) and sp²/sp³ ratio (4.7) that it is feasible to get Cu/a-C:H coating with higher hardness and tribological properties. Electrochemical impedance spectroscopy test results depicted significant improvement of Ti-6Al-4V alloy corrosion resistance by deposition of Cu/a-C:H thin coating at an optimum Ar/CH4 ratio. Furthermore, Cu/a-C:H thin coating with higher Cu contents showed better antibacterial properties and higher angiogenesis and osteogenesis activities. The coated samples inhibited the growth of bacteria as compared to the uncoated sample (p < 0.05). In addition, such coating composition can stimulate angiogenesis, osteogenesis and control host response, thereby increasing the success rate of implants. Moreover, Cu/a-C:H thin films encouraged development of blood vessels on the surface of titanium alloy when the density of grown blood vessels was increased with enhancing the Cu amount of the films. It is speculated that such coating can be a promising candidate for enhancing the osseointegration features.
... Cu, an essential micronutrient and catalytic cofactor, plays an important role in physiological repair processes such as wound healing and angiogenesis as well as several pathophysiologies including tumor growth and atherosclerosis (Harris 2004 Das et al. 2016). A previous study detected concentrated Cu in tumor tissues and angiogenic lesions and showed that it promoted neovascularization, while Cu chelators such as BCS inhibited tumor growth and angiogenic responses (Daniel et al. 2004). ...
Article
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Superoxide dismutase (SOD) 3, a copper (Cu)-containing anti-oxidative enzyme, plays a key role in extracellular redox homeostasis. Cu chaperone antioxidant-1 (Atox-1) not only delivers Cu ions to SOD3 at the trans-Golgi network, it also functions as a transcription factor of SOD3; however, the role of Atox-1 in the regulation of SOD3 during the monocytic differentiation of THP-1 cells has not yet been elucidated. A treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA) induced the expression of the Cu transport protein ATP7A in THP-1 cells. On the other hand, the nuclear translocation of Atox-1 was detected in TPA-treated THP-1 cells, and was suppressed in the presence of the Cu chelator, bathocuproinedisulfonic acid. Furthermore, Atox-1 bound to the SOD3 promoter region in TPA-treated THP-1 cells. The overexpression of Atox-1 in THP-1 cells significantly enhanced TPA-elicited SOD3 expression, whereas its knockdown suppressed this induction. The present results demonstrate that Atox-1 functions as a key molecule in TPA-elicited SOD3 expression.
... Consequently, 300% higher collagen solubility was found to lead to copper-deficiency and brittle bone [359]. Recently, being discovered as an essential element in angiogenesis [360] and to initiate endothelia cells towards angiogenesis [361,362], the application of copper ions as an alternative therapeutic agent in promoting vascularization has attracted increasing attentions [362e365]. Since vascularization plays a critical role in bone healing [37], it is reasonable to conduct relevant research. ...
Article
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Bone grafts have been predominated used to treat bone defects, delayed union or non-union, and spinal fusion in orthopaedic clinically for a period of time, despite the emergency of synthetic bone graft substitutes. Nevertheless, the integration of allogeneic grafts and synthetic substitutes with host bone was found jeopardized in long-term follow-up studies. Hence, the enhancement of osteointegration of these grafts and substitutes with host bone is considerably important. To address this problem, addition of various growth factors, such as bone morphogenetic proteins (BMPs), parathyroid hormone (PTH) and platelet rich plasma (PRP), into structural allografts and synthetic substitutes have been considered. Although clinical applications of these factors have exhibited good bone formation, their further application was limited due to high cost and potential adverse side effects. Alternatively, bioinorganic ions such as magnesium, strontium and zinc are considered as alternative of osteogenic biological factors. Hence, this paper aims to review the currently available bone grafts and bone substitutes as well as the biological and bio-inorganic factors for the treatments of bone defect.
... 49,50 Copper enhances angiogenesis by upregulating the activity of vascular endothelial growth factor (VEGF). 51 Given their role in bone metabolism, ECM trace elements have been incorporated into bone regenerating bioceramics to improve their performance. For example, proliferation of MC3T3-E1 cells is upregulated when cultured on composite ceramics enriched in zinc compared to cells cultured on non-zinc containing ceramics, and more bone formation has been detected around those implants in rabbit femora. ...
Article
Bone regeneration is a major challenge in cranio-facial and orthopedic surgery. Several biomaterials have been developed to overcome the limitations of autogenic bone graft, the gold standard for bone regeneration. Among these materials, decellularized extracellular matrix (ECM) has offered promising outcomes in bone regeneration. The ECMs include bioactive molecules that modulate cell proliferation and differentiation, thus, providing an excellent microenvironment for cell recruitment. ECM can be derived from tissues such as bone, cartilage, skin, and developing tooth germs, Alternatively, it can also be derived in vitro from cells such as osteoblasts, chondrocytes, adipose stem cells (ASCs), mesenchymal cells (MSCs) and embryonic stem cells (ESCs). Even though cell-derived ECM has shown promising results in enhancing bone regeneration, the cost involved in preparation process may hinder its routinely use. ECM immunogenicity is eliminated by decellularization processes that involve the use of physico-chemical methods, detergents, enzymatic treatments, or combination of these methods. Interestingly, ECM small molecules such as glycosaminoglycans and self-assembling peptides reveal good impact upon enhancing bone regeneration. This review addresses the state-of-the-art on the use of ECM for bone regeneration.
... It's well known that copper (Cu) is one of the essential trace elements for the survival of living organisms and mainly exists in the form of bivalent copper (Cu 2+ ) [9]. Cu has beneficial effects on osteogenesis, angiogenesis, and antimicrobial [10][11][12]. Early studies on mice diet supplemented with Cu demonstrated that Cu exerted effect on improving bone mineralization and increasing weight of femurs [13], and Cu depletion could cause osteoporosis that was shown radiologically and histologically [14,15]. ...
Article
Mesoporous bioactive glass (MBG) is a type of material with high biological activity and excellent biocompatibility. Because of its high specific surface area and adjustable surface morphology, MBG is usable for loading and delivering molecules. In our previous report, MBG particles were used as gene vectors and showed good transfection rate. In this paper, MBG, prepared through a sacrificial liquid template method in sol–gel process, was covered with polyglycerol (PG) and the resulting MBG-PG was further functionalized with octaarginine (Arg 8. More specifically, MBG-PG-Arg 8 particles were synthesized by PG functionalization of MBG through ring-opening polymerization of glycidol on the MBG surface, followed by multistep organic transformations (–OH→ –OTs (tosylate)→ –N 3 in the PG layer and click conjugation of the Arg 8 terminated with propargyl glycine. MBG-PG-Arg8 was successfully taken up by cells more efficiently due to the cellpenetrating property of Arg 8, and thus showed higher plasmid DNA loading and cell transfection efficiency than MBG modified with amino groups. This novel arginine-functionalized MBG may be a good candidate as a vector for gene delivery with higher efficiency.
... Altered redox status, such as increased levels of copper and ROS, has long been observed in cancer cells compared with normal cells [6][7][8]. Moreover, enhanced levels of copper appear to be an essential co-factor for angiogenesis and correlate with cancer stage and/or progression [6,9,10]. For these reasons, targeting copper transport and metabolism as anticancer strategies has gained tremendous interest [4,5,[11][12][13]. ...
Article
A main biochemical property of cancer cells, compared with normal cells, is altered redox status including increased levels of copper to maintain their malignant phenotypes. Thus, increasing copper accumulation, by using ionophores, to disrupt abnormal redox homeostasis of cancer cells may be an important anticancer strategy. Naturally occurring molecules with extraordinarily diverse chemical scaffolds are an important source of inspiration for developing copper ionophores. Dietary flavonoids are well-characterized copper chelators and show cancer chemopreventive potential, but their ionophoric role for redox-active copper and the related biological implications have remained unknown. This study reports, for the first time, the structural basis, chemical driving forces and biological implications of flavones (a widely distributed subgroup of flavonoids) as Cu(II) ionophores, and also provides new insights into cancer chemopreventive mechanism of flavones bearing 3(or 5)-hydroxy-4-keto group. 3-Hydroxyflavone surfaced as a potent Cu(II) ionophore to induce the mitochondria-dependent apoptosis of cancer cells in a redox intervention fashion via sequential proton-loss Cu(II) chelation, GSH-driving releasing of copper and protonation-dependent efflux of the neutral ligand.
... It is usually used for treating Wilson's disease. Recently, its potential in treating cancer has been unveiled; growing evidence demonstrates that TETA plays a role against cancer by inhibiting telomerase [31], in antiangiogenesis [32], and in suppressing cancer cell proliferation by modulating metabolism [33], as well as in induction of cell withered death [34]. However, none of these studies focused on the effect of TETA's chemical properties on cancer, specifically the two amino groups, and their role as a potential catalyst for Asc autoxidation. ...
Article
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Breast cancer is characterized by overexpression of superoxide dismutase (SOD) and downregulation of catalase and more resistance to hydrogen peroxide (H 2 O 2 ) than normal cells. Thus, relatively high H 2 O 2 promotes breast cancer cell growth and proliferation. However, excessive intracellular H 2 O 2 leads to death of breast cancer cells. In cancer cells, high level ascorbic acid (Asc) is able to be autoxidized and thus provides an electron to oxygen to generate H 2 O 2 . In the present study, we demonstrated that triethylenetetramine (TETA) enhances Asc autoxidation and thus elevates H 2 O 2 production in MCF-7 cells. Furthermore, Asc/TETA combination significantly impaired cancer cell viability, while having much milder effects on normal cells, indicating Asc/TETA could be a promising therapy for breast cancer. Moreover, SOD1 and N-acetyl-L-cysteine failed to improve MCF-7 cells viability in the presence of Asc/TETA, while catalase significantly inhibited the cytotoxicity of Asc/TETA to breast cancer cells, strongly suggesting that the selective cytotoxicity of Asc/TETA to cancer cells is H 2 O 2 -dependent. In addition, Asc/TETA induces RAS/ERK downregulation in breast cancer cells. Animal studies confirmed that Asc/TETA effectively suppressed tumor growth in vivo. In conclusion, TETA synergizes pharmacologic Asc autoxidation and H 2 O 2 overproduction in breast cancer cells, which suppresses RAS/ERK pathway and results in apoptosis.
... 53 This pro-angiogenic effect of copper was primarily through the regulation of a series of adhesive and growth-promoting factors involved in angiogenesis, such as VEGF. 54 Therefore, it was reasonable to suggest that copper is involved in the regulation of transcription factors for these pro-angiogenic genes. ...
Article
Impact statement: Copper promotes angiogenesis, but the mechanistic insights have not been fully elucidated until recently. In addition, the significance of copper promotion of angiogenesis in myocardial regeneration was increasingly revealed. Copper critically participates in the regulation of hypoxia-inducible factor 1 (HIF-1) of angiogenic gene expression. Interestingly, myocardial ischemia causes copper efflux from the heart, leading to suppression of angiogenesis, although HIF-1α, the critical subunit of HIF-1, remains accumulated in the ischemic myocardium. Strategies targeting copper specific delivery to the ischemic myocardium lead to selective activation of HIF-1-regulated angiogenic gene expression. Vascularization of the ischemic myocardium re-establishes the tissue injury microenvironment, and rebuilds the conduit for communication between the tissue injury signals and the remote regenerative responses including stem cells. This process promotes myocardial regeneration. Thus, a simple and effective copper supplementation to the ischemic myocardium would become a novel therapeutic approach to the treatment of patients with ischemic heart diseases.
... Copper, an essential micronutrient, plays an important role in physiological processes including wound repair and angiogenesis. (27,28) Since copper may exist in oxidized and reduced states in the body, this metal have functions as co-factor in redox enzymes. (29) We previously reported that the loss of superoxide dismutase 3, a copper-containing secretory antioxidant enzyme, may result in the accumulation of intracellular ROS in tumor cells, and ultimately exacerbate tumor progression. ...
Article
Copper is one of the essential micronutrients, and copper-containing enzymes contribute to crucial functions in the body. Lysyl oxidase is a copper-containing enzyme that remodels the extracellular matrix by cross-linking collagen and elastin. The overexpression of lysyl oxidase was recently shown to promote tumor metastasis. M2-like macrophages were also found to significantly accumulate in the tumor microenvironment, and correlated with a poor patient’s outcome. We speculate that M2-like macrophages promote tumor progression via lysyl oxidase expression. Epigenetics, a mitotically heritable change in gene expression without any change in DNA sequencing, is also associated with tumor progression. However, the relationship between lysyl oxidase expression in M2-like macrophages and epigenetics remains unclear. Lysyl oxidase expression was significantly induced in human leukemic THP-1 cell-derived M2-like macrophages. Furthermore, the level of histone H3 tri-methylation at lysine 27 was decreased, and a pre-treatment with a H3K27 demethylase inhibitor notably suppressed lysyl oxidase expression in M2-like macrophages. Lysyl oxidase derived from M2-like macrophages also enhanced breast cancer cell migration, and this was suppressed by a H3K27 demethylase inhibitor. The present results suggest the mechanism of lysyl oxidase expression in M2-like macrophages as an aspect of epigenetics, particularly histone methylation.
... W. , anti-infection (X. Y. Wang, Dong, et al., 2019), bone healing (Liu et al., 2018), endothelialization (Harris, 2014), and vascularization (Niu et al., 2016; J. Y. properties. Also, Cu was added to polymer materials by methods, including physical blending (Tabesh, Salimijazi, Kharaziha, & Hejazi, 2018) and chemical bonding (Li et al., 2018). ...
Article
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Vascular implant interventional medical catheter will contact with blood firstly after implantation. The anticoagulation and antibacterial functions of this device will determine the success or failure. Copper (Cu) has been verified to possess multi‐biofunctions, but it was challenging to add the Cu metal to most materials. Take advantage of its functionality; Cu has been grafted on the material surface to improve the anticoagulation function and accelerate endothelialization. In this study, a Cu‐bearing chitosan coating was prepared on the catheter to endow the anticoagulation and anti‐infection functions. Besides, properties characterization and functional evaluation of the coated medical catheter were investigated. Dynamic blood clotting and platelet adhesion tests were carried out to evaluate the anticoagulation property. Besides this, the antibacterial test was used to estimate the anti‐infection function. The surface energy and Cu ions release from the coating were detected and calculated by contact angles and immersion tests, respectively. The results of attenuated total reflectance Fourier transform infrared spectroscopy (ATR‐FTIR) and X‐ray photoelectron spectroscopy (XPS) revealed that Cu ions were grafted in the chitosan coating. Thermogravimetric analysis (TA) result showed the concentration of Cu ions in the coating. The results of dynamic blood clotting, platelet adhesion, and antibacterial tests revealed that Cu grafted in chitosan would improve the blood compatibility and anti‐infection property. The surface properties and Cu ions release behavior of Cu‐bearing coating revealed the reasons for multi‐biofunctions. This study indicated that the Cu‐bearing chitosan coating could endow the vascular implant interventional device anticoagulation and anti‐infection functions, which has excellent potential for clinical application.
... The addition of these alloying element, particularly, Cu and Mn can improve the mechanical properties, not only strength, but also elongation. Cu is an essential trace element required for bone growth and connectivity, and enhances the proliferation of vascular endothelial cells, and revascularization [184,185]. In addition, Cu deficiency leads to normocytic cholesterol metabolism and neutropenia [186]. ...
Article
Biodegradable metals (BMs) gradually degrade in vivo by releasing corrosion products once exposed to the physiological environment in the body. Complete dissolution of biodegradable implants assists tissue healing, with no implant residues in the surrounding tissues. In recent years, three classes of BMs have been extensively investigated, including magnesium (Mg)-based, iron (Fe)-based, and zinc (Zn)-based BMs. Among these three BMs, Mg-based materials have undergone the most clinical trials. However, Mg-based BMs generally exhibit faster degradation rates, which may not match the healing periods for bone tissue, whereas Fe-based BMs exhibit slower and less complete in vivo degradation. Zn-based BMs are now considered a new class of BMs due to their intermediate degradation rates, which fall between those of Mg-based BMs and Fe-based BMs, thus requiring extensive research to validate their suitability for biomedical applications. In the present study, recent research and development on Zn-based BMs are reviewed in conjunction with discussion of their advantages and limitations in relation to existing BMs. The underlying roles of alloy composition, microstructure, and processing technique on the mechanical and corrosion properties of Zn-based BMs are also discussed.
... Cu, a key microelement essential to organisms and a cofactor of many enzymes, is involved in angiogenesis and lymphangiogenesis, such as a stimulator of angiogenesis and endothelial cell migration in the vascular system and a local regulator for the growth or regression of new blood vessels [14][15][16][17][18]. Cu depletion has been shown to inhibit angiogenesis in a wide variety of cancer cells and xenograft systems [19][20][21]. ...
Article
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Molecular transport and cell circulation between tissues and organs through blood and lymphatic vessels are essential for physiological homeostasis in vertebrates. Despite the report of its association with vessel formation in solid tumors, the biological effects of Copper (Cu) accumulation on angiogenesis and lymphangiogenesis during embryogenesis are still unknown. In this study, we unveiled that intersegmental blood circulation was partially blocked in Cu²⁺-stressed zebrafish embryos and cell migration and tube formation were impaired in Cu²⁺-stressed mammalian HUVECs. Specifically, Cu²⁺-stressed embryos showed down-regulation in the expression of amotl2 and its downstream pERK1/2-foxm1-MMP2/9 regulatory axis, and knockdown/knockout of foxm1 in zebrafish embryos phenocopied angiogenesis defects, while FOXM1 knockdown HUVECs phenocopied cell migration and tube formation defects, indicating that excessive Cu²⁺-induced angiogenesis defects and blocked cell migration via down-regulating amotl2-pERK1/2-foxm1-MMP2/9 regulatory axis in both embryos and mammalian cells. Additionally, thoracic duct was revealed to be partially absent in Cu²⁺-stressed zebrafish embryos. Specifically, Cu²⁺-stressed embryos showed down-regulation in the expression of ccbe1 (a gene with pivotal function in lymphangiogenesis) due to the hypermethylation of the E2F7/8 binding sites on ccbe1 promoter to reduce their binding enrichment on the promoter, contributing to the potential mechanisms for down-regulation of ccbe1 and the formation of lymphangiogenesis defects in Cu²⁺-stressed embryos and mammalian cells. These integrated data demonstrate that Cu²⁺ stress impairs angiogenesis and lymphangiogenesis via down-regulation of pERK1/2-foxm1-MMP2/9 axis and epigenetic regulation of E2F7/8 transcriptional activity on ccbe1 expression, respectively.
... New blood vessels are formed in a multistep coordinated sequence, essentially comprising the breakdown of the vessel's basement membrane, the migration and proliferation of the endothelial cells within the ECM, and the tube's formation. Copper activates many components involved in angiogenesis, like VEGF, angiogenin, ceruloplasmin, SPARC, NFkB; conversely, decreased copper availability impairs their function [127]. Several studies on in vitro tumor models and xenografts address copper availability in blood vessel growth [76]. ...
Article
Copper homeostasis is strictly regulated by protein transporters and chaperones, to allow its correct distribution and avoid uncontrolled redox reactions. Several studies address copper as involved in cancer development and spreading (epithelial to mesenchymal transition, angiogenesis). However, being endogenous and displaying a tremendous potential to generate free radicals, copper is a perfect candidate, once opportunely complexed, to be used as a drug in cancer therapy with low adverse effects. Copper ions can be modulated by the organic counterpart, after complexed to their metalcore, either in redox potential or geometry and consequently reactivity. During the last four decades, many copper complexes were studied regarding their reactivity toward cancer cells, and many of them could be a drug choice for phase II and III in cancer therapy. Also, there is promising evidence of using ⁶⁴Cu in nanoparticles as radiopharmaceuticals for both positron emission tomography (PET) imaging and treatment of hypoxic tumors. However, few compounds have gone beyond testing in animal models, and none of them got the status of a drug for cancer chemotherapy. The main challenge is their solubility in physiological buffers and their different and non-predictable mechanism of action. Moreover, it is difficult to rationalize a structure-based activity for drug design and delivery. In this review, we describe the role of copper in cancer, the effects of copper-complexes on tumor cell death mechanisms, and point to the new copper complexes applicable as drugs, suggesting that they may represent at least one component of a multi-action combination in cancer therapy.
... However, the ductility significantly declined with respect to pure Zn, while Zn-Cu alloys have elongation values above 30%, which successfully accomplishes the criteria for cardiovascular stents applications [27]. Moreover, Cu is well-known for its antibacterial properties [1,[28][29][30] and it is reported to stimulate endothelial cell migration and angiogenesis [31][32][33]. Alloying with peritectic-forming elements such as Cu or eutectic-forming elements such as Mg, refines the microstructure of pure Zn and enhances its mechanical properties [34,35]. The importance of controlling topography, chemistry, and mechanical properties at micro and nanoscale is fundamental for obtaining the desired cellular response [28]. ...
Article
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In the recent decades, zinc (Zn) and its alloys have been drawing attention as promising candidates for bio-resorbable cardiovascular stents due to its degradation rate more suitable than magnesium (Mg) and iron (Fe) alloys. However, its mechanical properties need to be improved in order to meet the criteria for vascular stents. This work investigates the mechanical properties, biodegradability and biocompatibility of Zn-Mg and Zn-Cu alloys in order to determine a proper alloy composition for optimal stent performance. Nanoindentation measurements are performed to characterize the mechanical properties at the nanoscale as a function of the Zn microstructure variations induced by alloying. The biodegradation mechanisms are discussed and correlated to microstructure, mechanical performance and bacterial/cell response. Addition of Mg or Cu alloying elements refined the microstructure of Zn and enhanced yield strength (YS) and ultimate tensile strength (UTS) proportional to the volume fraction of secondary phases. Zn-1Mg showed the higher YS and UTS and better performance in terms of degradation stability in Hanks' solution. Zn-Cu alloys presented an antibacterial effect for S. aureus controlled by diffusion mechanisms and by contact. Biocompatibility was dependent on the degradation rate and the nature of the corrosion products.
... However, unbound or "free" Cu ions are not generally encountered in cells in either valence state (for a detailed discussion of the chemical speciation of Cu in cells, see Fahrni [1]). Cu is essential for many physiological processes including iron homeostasis [2], angiogenesis [3], neurotransmitter biosynthesis [4], immune function (including neutrophil activation and macrophage function) [5] and energy metabolism [6]. ...
Article
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Essential metals such as copper (Cu) and zinc (Zn) are important cofactors in diverse cellular processes, while metal imbalance may impact or be altered by disease state. Cu is essential for aerobic life with significant functions in oxidation-reduction catalysis. This redox reactivity requires precise intracellular handling and molecular-to-organismal levels of homeostatic control. As the central organ of Cu homeostasis in vertebrates, the liver has long been associated with Cu storage disorders including Wilson Disease (WD) (heritable human Cu toxicosis), Idiopathic Copper Toxicosis and Endemic Tyrolean Infantile Cirrhosis. Cu imbalance is also associated with chronic liver diseases that arise from hepatitis viral infection or other liver injury. The labile redox characteristic of Cu is often discussed as a primary mechanism of Cu toxicity. However, work emerging largely from the study of WD models suggests that Cu toxicity may have specific biochemical consequences that are not directly attributable to redox activity. This work reviews Cu toxicity with a focus on the liver and proposes that Cu accumulation specifically impacts Zn-dependent processes. The prospect that Cu toxicity has specific biochemical impacts that are not entirely attributable to redox may promote further inquiry into Cu toxicity in WD and other Cu-associated disorders.
... Early in 1980, McAuslan and Gole showed that Cu can stimulate blood vessel formation by introducing CuSO 4 into anterior chamber implants at micromolar levels [34]. Later studies further suggested that Cu could not only stimulate the proliferation and migration of endothelial cells to promote the formation of new blood vessels, but also inhibit the proliferation of vascular smooth muscle cells and thrombosis [22,35]. These results prompted more intense research on Cu in vessels. ...
Article
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Copper is an indispensable trace metal element in the human body, which is mainly absorbed in the stomach and small intestine and excreted into the bile. Copper is an important component and catalytic agent of many enzymes and proteins in the body, so it can influence human health through multiple mechanisms. Based on the biological functions and benefits of copper, an increasing number of researchers in the field of biomaterials have focused on developing novel copper-containing biomaterials, which exhibit unique properties in protecting the cardiovascular system, promoting bone fracture healing, and exerting antibacterial effects. Copper can also be used in promoting incisional wounds healing, killing cancer cells, Positron Emission Tomography (PET) imaging, radioimmunological tracing and radiotherapy of cancer. In the present review, the biological functions of copper in the human body are presented, along with an overview of recent progress in our understanding of the biological applications and development of copper-containing materials. Furthermore, this review also provides the prospective on the challenges of those novel biomaterials for future clinical applications.
... mg day − 1 for adults [20]. Besides, Cu plays a vital part in the immune system [15][16][17], and has beneficial effects on endothelial cells proliferation [18,19]. Moreover, copper has been reported to enhance antibacterial property [21,22], which can minimize the risk of infection in clinical applications. ...
Article
Zn-3Cu-xMg (x = 0, 0.1, 0.5 and 1.0 wt.%) alloys were developed as potential biodegradable metallic materials in this study. The mechanical properties, corrosion behavior and in vitro cytocompatibility of Zn-3Cu-xMg alloys were studied systematically to evaluate the feasibility as biodegradable implant materials. The secondary phase in as-cast and as-extruded Zn-3Cu alloy was CuZn5 phase. Mg2Zn11 phase newly formed and precipitated by Mg addition. The volume fraction of Mg2Zn11 phase increased gradually with increasing Mg concentration. As a result, yield strength was improved from 213.7 to 426.7 MPa and increased by 99.7% while elongation decreased from 47.1% to 0.9%. Besides, biocompatibility was improved apparently and in vitro corrosion rates increased from 11.4 to 43.2 μm year− 1, which is more suitable for clinic application. The present research indicated that the newly developed alloys could be promising candidates for biomedical use due to the proper mechanical properties, degradation rate and acceptable biocompatibility.
... [30][31][32][33][34][35] Alternatively, copper ions (Cu 2+ ) are the bioactive components with the specific capability for promoting angiogenesis, which is originated from their performance on stabilizing the expression of hypoxia-inducible factor (HIF-1 ) and secretion of vascular endothelial growth factor (VEGF), further enhancing the recruitment and differentiation of cells during the blood vessel-producing procedure. [36][37][38][39][40][41][42] It has been fully revealed that Cu ions can facilitate the cell migration, angiogenesis, and collagen deposition for the specific biomedical implementation on accelerating wound healing. [43][44][45][46] As the necessary component for maintaining human health, the adults typically require the highest safe intake amount of Cu of 10 mg per day. ...
Article
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As an essential trace element in the human body, transitional metal copper (Cu) ions are the bioactive components within the body featuring dedicated biological effects such as promoting angiogenesis and influencing lipid/glucose metabolism. The recent substantial advances of nanotechnology and nanomedicine promote the emerging of distinctive Cu‐involved biomaterial nanoplatforms with intriguing theranostic performances in biomedicine, which are originated from the biological effects of Cu species and the physiochemical attributes of Cu‐composed nanoparticles. Based on the very‐recent significant progresses of Cu‐involved nanotheranostics, this work highlights and discusses the principles, progresses, and prospects on the elaborate design and rational construction of Cu‐composed functional nanoplatforms for a diverse array of biomedical applications, including photonic nanomedicine, catalytic nanotherapeutics, antibacteria, accelerated tissue regeneration, and bioimaging. The engineering of Cu‐based nanocomposites for synergistic nanotherapeutics is also exemplified, followed by revealing their intrinsic biological effects and biosafety for revolutionizing their clinical translation. Finally, the underlying critical concerns, unresolved hurdles, and future prospects on their clinical uses are analyzed and an outlook is provided. By entering the “Copper Age,” these Cu‐involved nanotherapeutic modalities are expected to find more broad biomedical applications in preclinical and clinical phases, despite the current research and developments still being in infancy.
... Cu 2+ is an essential element for living organisms [22,23]. It is a vital trace element that plays a key role in immunity, increasing the average rate of bone resorption in bone metabolism and improving collagen fiber precipitation [20,24,25]. It is also important in the areas of bone engineering and regeneration [20]. ...
Article
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The objectives of this study were to reduce the corrosion rate and increase the cytocompatibility of AZ31 Mg alloy. Two coatings were considered. One coating contained MgO (MAO/AZ31). The other coating contained Cu2+ (Cu/MAO/AZ31), and it was produced on the AZ31 Mg alloy via microarc oxidation (MAO). Coating characterization was conducted using a set of methods, including scanning electron microscopy, energy-dispersive spectrometry, X-ray photoelectron spectroscopy, and X-ray diffraction. Corrosion properties were investigated through an electrochemical test, and a H2 evolution measurement. The AZ31 Mg alloy with the Cu2+-containing coating showed an improved and more stable corrosion resistance compared with the MgO-containing coating and AZ31 Mg alloy specimen. Cell morphology observation and cytotoxicity test via Cell Counting Kit-8 assay showed that the Cu2+-containing coating enhanced the proliferation of L-929 cells and did not induce a toxic effect, thus resulting in excellent cytocompatibility and biological activity. In summary, adding Cu ions to MAO coating improved the corrosion resistance and cytocompatibility of the coating.
... Zinc increases the activity of alkaline phosphatase and stimulates osteoblasts to produce collagen (Hall, Dimai, & Farley, 1999;Seo, Cho, Kim, Shin, & Kwun, 2010); hence its deficiency impairs bone growth (Hosea, Taylor, Wood, Mollard, & Weiler, 2004;Oner, Bhaumick, & Bala, 1984). Copper is essential for vascular growth (Harris, 2004;Sen et al., 2002) as well as collagen formation and maturation (O'Dell, 1981;Opsahl et al., 1982). Manganese is involved in the synthesis of chondroitin sulfates, the main galactosamine-polysaccharides in bone matrix (Leach Jr, 1971;Leach, Muenster, & Wien, 1969). ...
Article
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The bone regenerative capacity of synthetic calcium phosphates (CaPs) can be enhanced through the enrichment with selected metal trace ions. However, defining the optimal elemental composition required for bone formation is challenging due to a large number of possible concentrations and combinations of these elements. We hypothesized that the ideal elemental composition exists in the inorganic phase of the bone extracellular matrix (ECM). To study our hypothesis, we first obtained natural hydroxyapatite through the calcination of bovine bone, which was then investigated its reactivity with acidic phosphates to produce CaP cements. Bioceramic scaffolds fabricated using these cements were assessed for their composition, properties, and in vivo regenerative performance and compared to controls. We found that natural hydroxyapatite can react with phosphoric acid to produce CaP cements with biomimetic trace metals. These cements present significantly superior in vivo bone regenerative performance compared to cements prepared using synthetic apatite. In summary, this study opens new avenues for further advancements in the field of CaP bone biomaterials by introducing a simple approach to develop biomimetic CaPs. This work also sheds light on the role of the inorganic phase of bone and its composition in defining the regenerative properties of natural bone xenografts.
Article
Copper nanoparticles are explored significantly for their antimicrobial activity, especially for antibiotic-resistant strain infections. However, copper has severe toxic responses and mostly it is due to its generation capability of reactive oxygen species (ROS) molecules while interacting with in vitro or in vivo systems. In the current study, wire shaped copper nanostructures were synthesized via microwave irradiation with single step doping of carbon nanodots (CDs). The synthesized material (CuCs) was characterized by UV-Vis spectroscopy, fluorescence spectroscopy, FTIR, TEM, FESEM, XRD, DLS, and XPS. The fluorescence spectroscopy, microscopy and Raman spectroscopy results suggested CuCs to work well as a bi-modal imaging nanoprobe (fluorescence/SERS). The cell culture studies prove significant cytocompatibility and ROS scavenging property of the samples with respect to control. Further, CuCs-gelatin nanocomposite thin films were prepared and implanted into rodent deep wound model. The histological study has showed enhanced angiogenesis in the subcutaneous region. The results were validated by immuno-histochemistry. The ROS scavenging and enhanced angiogenesis were validated via gene expression studies and a HIF-α induced enhanced angiogenesis mechanism was also proposed for better wound healing.
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In this work, mechanical properties, degradation behavior and biocompatibility of as-extruded and solid solution treated Mg-Cu binary alloy with 0.22 and 0.45 wt.% Cu additions have been investigated, and the effects of main microstructural parameters (grain size, dislocation, second phase distribution and content) on these properties are analyzed. Mechanical tests revealed that as-solutionized Mg-Cu alloys (with larger grain size and less content of Mg2Cu phase) exhibited better mechanical properties than the as-extruded alloys. Strengthening effects of the microstructural parameters have been analyzed and quantified. The results suggested that the inverse elevation of the yield strength was mainly related to the over-solubility Cu atom in the matrix. Immersion tests showed that the solid solution treatment improved the corrosion resistance of Mg-0.2Cu alloy (with 0.22 wt.% Cu) but deteriorated it seriously for Mg-0.5Cu alloy (with 0.45 wt.% Cu). The enhanced corrosion resistance of Mg-0.2Cu alloy was owing to the apparent decrease of Mg2Cu phase content. In contrast, the deterioration of corrosion resistance for Mg-0.5Cu alloy was attributed to the distribution change of the Mg2Cu phase from diffuse to local, because large localized Mg2Cu particles can lead to severer corrosion of the matrix. Cytotoxicity experiments revealed that the as-solutionized Mg-0.2Cu alloy with the least Mg2Cu phase content exhibited the best biocompatibility due to the slowest degradation rate.
Article
Xanthine oxidase (XO) is a key enzyme catalyzing hypoxanthine to xanthine and then uric acid causing hyperuricemia. A Cu(II) complex of chrysin was synthesized and characterized by UV–vis absorption, Fourier transform infrared, nuclear magnetic resonance (¹H NMR) and mass spectroscopy studies. The interaction of Cu(II)-complex with XO was investigated by spectroscopic methods and molecular simulation. The Cu(II)-chrysin complex exhibited a better inhibitory ability (IC50 = 0.82 ± 0.034 μM) against XO than its corresponding ligands chrysin and Cu2 + in a mix-competitive manner. The binding affinity of Cu(II)-chrysin complex with XO was much higher than that of chrysin. The hydrogen bonds and van der Waals forces played main roles in the binding. Analysis of circular dichroism spectra indicated that the complex induced the conformational change of XO. The molecular simulation found that the Cu(II)-chrysin complex inserted into the active cavity of XO with Cu acting as a bridge, occupying the catalytic center of the enzyme to avoid entry of the substrate xanthine, leading to the inhibition of XO. This study may provide new insights into the inhibition mechanism of the Cu(II)-chrysin complex as a promising XO inhibitor and its potential application for the treatment of hyperuricemia.
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Intrauterine adhesions (IUAs) caused by mechanical damage or infection increase the risk of infertility in women. Although numerous physical barriers such as balloon or hydrogel are developed for the prevention of IUAs, the therapeutic efficacy is barely satisfactory due to limited endometrial healing, which may lead to recurrence. Herein, a second near‐infrared (NIR‐II) light‐responsive shape memory composite based on the combination of cuprorivaite (CaCuSi4O10) nanosheets (CUP NSs) as photothermal conversion agents and polymer poly(d,l‐lactide‐co‐trimethylene carbonate) (PT) as shape memory building blocks is developed. The as‐prepared CUP/PT composite possesses excellent shape memory performance under NIR‐II light, and the improved operational feasibility as an antiadhesion barrier for the treatment of IUAs. Moreover, the released ions (Cu, Si) can stimulate the endometrial regeneration due to the angiogenic bioactivity. This study provides a new strategy to prevent IUA and restore the injured endometrium relied on shape memory composite with enhanced tissues reconstruction ability. A second near‐infrared light‐responsive shape memory composite, which is composed of cuprorivaite (CaCuSi4O10) nanosheets as photothermal conversion agents and polymer poly(d,l‐lactide‐co‐trimethylene carbonate) as shape memory building blocks, is developed for intrauterine adhesions prevention and endometrial regeneration.
Article
Chronic bacterial infection, local inflammation, and insufficient angiogenesis contribute to poor healing of diabetic wounds. Here, Cu2O/Pt nanocubes (CPN) are successfully developed with good biocompatibility for treatment of diabetic wounds in rats. The synthesized CPN are characterized using SEM, XPS, and XRD. CPN exhibit triple-enzyme mimetic activity: oxidase-like, peroxidase-like, and catalase-like activities. Moreover, CPN show significant antibacterial activity against Gram-negative and Gram-positive bacteria when combined with low concentration of H2O2, via generation of highly reactive ROS. CPN also exhibit significantly accelerated wound healing in a full-layer deprivation rat model infected by Staphylococcus aureus, which is ascribed to the constant release of copper ions, subsequently activating the VEGF/AKT/ERK1/2 signaling pathway and promoting angiogenesis. CPN are able to catalyze H2O2 to generate O2 for local hypoxia alleviation. Furthermore, in vivo results indicate that treatment with CPN promotes the expression of transforming growth factor and matrix metalloproteinases, causing enhanced cell proliferation and collagen deposition, as well as extracellular matrix remodeling. In contrast, CPN decrease the expression of proinflammatory cytokines, such as TNF-ɑ and IL-1β, which are induced by bacterial infection and hyperglycemia. These results suggest a novel strategy for the treatment of diabetic wound healing.
Article
Caffeic acid phenethyl ester (CAPE) is an active polyphenol of propolis from honeybee hives, and exhibits antioxidant and interesting pharmacological activities. However, in this study, we found that in the presence of Cu(II), CAPE exhibited pro-oxidative rather than antioxidant effect: synergistic DNA damage was induced by the combination of CAPE and Cu(II) together as measured by strand breakage in plasmid DNA and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) formation, which is dependent on the molar ratio of CAPE:Cu(II). Production of Cu(I) and H2O2 from the redox reaction between CAPE and Cu(II), and subsequent •OH formation was found to be responsible for the synergistic DNA damage. DNA sequencing investigations provided more direct evidence that CAPE/Cu(II) caused preferential cleavage at guanine, thymine and cytosine residues. Interestingly, we found there are competitive binding between CAPE and DNA with Cu(II)/Cu(I), which changed the redox activity of Cu(II)/Cu(I), via complementary applications of different analytical methods. The observed DNA damage was mainly attributed to the formation of DNA-Cu(II)/Cu(I) complexes, which is still redox active and initiated the redox reaction near the binding site between copper and DNA. Based on these data, we proposed that the synergistic DNA damage induced by CAPE/Cu(II) might be due to the competitive binding between CAPE and DNA with Cu, and site-specific production of •OH near the binding site of copper with DNA. Our findings may have broad biological implications for future research on the pro-oxidative effects of phenolic compounds in the presence of transition metals.
Article
Endothelialization is an important process after stenting in coronary artery. Recovery of the injured site timely can reduce the neointima formation and platelet absorbance, leading to a lower risk of in-stent restenosis. Copper is known to be critical in vascular construction. Thus a combination of copper with stent materials is a meaningful attempt. A copper bearing L605-Cu cobalt alloy was prepared and its effect on human umbilical vein endothelial cells (HUVECs) was evaluated in vitro in this study. It was found that HUVECs attached and stretched better on the surface of L605-Cu compared with L605, and the apoptosis of cells was decreased simultaneously. The migration and tube formation of HUVECs were also enhanced by the extract of L605-Cu. Furthermore, L605-Cu increased the mRNA expression of VEGF in HUVECs significantly. However it had no effect on the secretion of NO or mRNA expression of eNOS. The result of blood clotting test indicated that L605-Cu had better blood compatibility. These results above have demonstrated that the L605-Cu alloy is promising to be a new stent material with function of accelerating endothelialization. This article is protected by copyright. All rights reserved.
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Near-infrared (NIR) light-responsive shape-memory composites have shown great potential in various biomedical applications due to their amazing remote actuation property. However, most of them are bioinert and restricted in NIR-I biological window. Here, we developed a new biodegradable NIR-II-responsive shape memory composite by the incorporation of CaSiCu4O10 nanosheets (CUP NSs) into shape memory polymer of poly (D, L-lactide-co-trimethylene carbonate) (PDLLA-co-TMC, denoted as PT). The as-prepared CUP/PT composites possessed outstanding shape memory performance upon low power NIR-II laser irradiation (1064 nm, 0.2 W·cm⁻²), offering the possibility of deep-tissue applications. Moreover, due to the sustained release of bioactive Cu²⁺ and SiO4⁴⁻ ions, the composites exhibited strong stimulation of angiogenesis both in vitro and in vivo. Our work demonstrated the promising potential of CUP/PT composites as NIR-II light-responsive biomaterials with enhanced tissue reconstruction ability, especially for wound healing.
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Alle Organismen müssen Metallhomöostase sicherstellen, indem sie fluktuierende Mengen von Spurenelementen in der Umwelt bewältigen. Genetische und biochemische Studien in Säuger und Fliegen zeigten eine zentrale Rolle für den metalloregulatorische Protein MTF-1 (metal-responsive transcription factor) in der Homöostase von Übergangsmetallen. MTF-1 ist ein essentielles Gen in Säugern, währenddem es in Drosophila nicht überlebenswichtig ist. In beiden Fällen aber, in der Fliege und in Säugern, ist es ein kritischer Faktor für die Toleranz gegenüber Metallen und in Drosophila, unerwarteterweise auch gegenüber Kupfermangel. Obwohl grosse Fortschritte in der Biologie der Übergangsmetallen gemacht worden sind, bleibt das Wissen über Signalwege und Mechanismen der Antwort auf Metallstress lückenhaft. Auch ist wenig bekannt über die Gene, die in der unmittelbaren Handhabung von verschiedenen Metallen involviert sind. In dieser Studie versuchten wir, neue Komponenten der Antwort auf Metallstress in Drosophila melanogaster mit Hilfe einer Microarray basierten Transkriptom Analyse und genomweiter RNA Interferenz zu identifizieren. Zuerst wurde das Transkriptom von MTF-1 mutanten und wildtyp Larven verglichen. Beide Genotypen wurden in normalem Futter und solchem mit Metallzusätzen aufgezogen. Dadurch wurden neue, mögliche MTF-1 Zielgene gefunden, wie z.B. der Kupferimporter Ctr1B, der Zinkexporter ZnT-D1, der ABC Transporter CG10505 und die Ferritin-Gene Fer1HCH und Fer2LCH. Weitere biochemische und genetische Analysen in der Fliege bestätigten diese Kandidaten als MTF-1 Zielgene und erklärten einige Aspekte des Drosophila MTF-1 knockout Phänotyps. Ausserdem entdeckten wir eine Anzahl von Genen, die unabhängig von MTF-1 auf Metallzusatz reagierten. Unter diesen sind die Gene des Glutathion basierten Detoxifikation Signalwegs und die Gene involviert in Hitzeschock und Immunantwort. In der zweiten Analyse wurde ein genomweiter RNAi Screen in Drosophila S2 Zellkulturen durchgeführt, in Zusammenarbeit mit dem RNAi Screening Center an der Harvard Medical School. Der Screen identifizierte eine grosse Anzahl von Kandidatengenen, welche den Promotor vom Metallothionein A (MtnA) regulieren. MtnA ist eines der am Besten charakterisierten Zielgenen von MTF-1. Ein zweiter Screen mit hausinternen RNAi Verfahren wurde durchgeführt, um die Daten zu bestätigen. Einige Kandidatengene scheinen in einen Signalweg involviert zu sein, der den MtnA promotor reguliert. Weitere Studien sind notwendig für eine detaillierte Charakterisierung dieser Gene, und um ihren Platz in der Hierarchie der Antwort auf Metallstress zu bestimmen. All organisms have to ensure metal homeostasis by coping with fluctuating amounts of trace elements in the environment. Genetic and biochemical studies in mammals and Drosophila have established a central role for metal-responsive transcription factor MTF-1 in transition metal homeostasis. MTF-1 is an essential gene in mammals. In Drosophila, MTF-1 is not required for viability. However, in the fly, as in mammals, it is a critical factor for metal tolerance, and, unexpectedly, also for the copper starvation response. Even though great progress in the biology of transition metals has been made, there is still a lack of knowledge about the pathways and mechanisms leading to metal response and the genes involved in the immediate handling of various metals. In this study we attempted to identify novel components of the metal response using microarray based transcriptome analysis and genome-wide RNA interference technique in Drosophila melanogaster. Firstly, the transcriptome of MTF-1 mutant and wild type larvae, raised in normal or metal-supplemented food, was compared. This revealed new candidate MTF-1 target genes, such as the copper importer Ctr1B, the zinc exporter ZnT-D1, the ABC transporter CG10505, and the ferritin genes Fer1HCH and Fer2LCH. Further biochemical and genetic analyses in the fly established these candidates as genuine MTF-1 regulated genes and explained various aspects of Drosophila MTF-1 knockout phenotype. Moreover, we have uncovered a variety of genes that respond to metal load, independent of MTF-1. Among these are the genes involved in the glutathione- mediated detoxification pathway, and the genes of heat shock and immune responses. In the second analysis a genome-wide RNAi screen was done in cultured Drosophila S2 cells in collaboration with the RNAi Screening Center at Harvard Medical School. The screen identified a large number of candidate genes acting upstream of the promoter of metallothionein A, one of the best characterized target genes of MTF-1. A secondary screen using in house RNAi procedures was performed to validate the data. Several candidate genes appear to be involved in a pathway that regulates the metallothionein promoter. Future studies are needed for a detailed characterization of these genes and for placing them within the hierarchy of the metal stress response pathway.
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Altered redox status including higher levels of copper in cancer cells than in normal cells inspired many researchers to develop copper ionophores targeting this status. We have recently found that flavon-3-ol (3-HF) works as a potent Cu(II) ionophore by virtue of its keto-enol moiety. To further emphasize the significance of this moiety for developing Cu(II) ionophores, we herein designed a β-diketo analog of piperlongumine, PL-I, characterized by the presence of high proportion of the keto-enol form in dimethylsulfoxide and chloroform, and identified its keto-enol structure by NMR and theoretical calculations. Benefiting from deprotonation of its enolic hydroxyl group, this molecule is capable of facilitating the transport of Cu(II) through cellular membranes to disrupt redox homeostasis of human hepatoma HepG2 cells and trigger their death.
Chapter
Bone grafts have been predominately applied for the treatments of bone defect, delayed union, non-union and spinal fusion for a period of time despite of the emergency of synthetic graft substitutes. However, the integration of allogeneic bone graft to host bone is unsatisfactory in long-term follow-up. To address this clinical issue, bone morphogenetic proteins (BMPs), parathyroid hormone (PTH) and platelet-rich plasma (PRP) have been considered to incorporate with the allografts for achieving superior bone integration. Although the clinical outcomes of these approaches have exhibited effective bone formation, high application cost and potential adverse effects are concerned. In fact, bioinorganic ions such as magnesium, strontium, and zinc are considered as the alternative of osteogenic factors. Hence, this chapter aims to review the currently available bone grafts, bone substitutes, biological factors and bio-inorganic ions for the treatments of bone defect.
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Copper is an essential element in cells; it can act as either a recipient or a donor of electrons, participating in various reactions. However, an excess of copper ions in cells is detrimental as these copper ions can generate free radicals and increase oxidative stress. In multicellular organisms, copper metabolism involves uptake, distribution, sequestration, and excretion, at both the cellular and systemic levels. Mammalian enterocytes take in bioavailable copper ions from the diet in a Ctr1-dependent manner. After incorporation, cuprous ions are delivered to ATP7A, which pumps Cu+ from enterocytes into the blood. Copper ions arrive at the liver through the portal vein and are incorporated into hepatocytes by Ctr1. Then, Cu+ can be secreted into the bile or the blood via the Atox1/ATP7B/ceruloplasmin route. In the bloodstream, this micronutrient can reach peripheral tissues and is again incorporated by Ctr1. In peripheral tissue cells, cuprous ions are either sequestrated by molecules such as metallothioneins or targeted to utilization pathways by chaperons such as Atox1, Cox17, and CCS. Copper metabolism must be tightly controlled in order to achieve homeostasis and avoid disorders. A hereditary or acquired copper unbalance, including deficiency, overload, or misdistribution, may cause or aggravate certain diseases such as Menkes disease, Wilson disease, neurodegenerative diseases, anemia, metabolic syndrome, cardiovascular diseases, and cancer. A full understanding of copper metabolism and its roles in diseases underlies the identification of novel effective therapies for such diseases.
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Ceruloplasmin (CP) is a blue serum protein found in human serum; it carries approximately 95% of the total circulating copper (Cu) in healthy individuals. The relationship of CP with OS, inflammation, and DNA damage is known. Oxidative stress (OS), inflammation, and DNA damage are the main causes underlying atherosclerotic heart disease. Several studies have indicated a close association between high serum CP and several types of heart disease. However, the CP levels are still unknown in many heart diseases. To gather the studies of CP in heart disease and to prepare the ground for new studies for researchers, we designed this review.
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In this study, multifunctional tantalum copper composite nanotubes (TaCu-NTs) were coated on titanium for enhanced bacteriostatic, angiogenic and osteogenic properties. Three coatings of Ta, TaCu1 (Ta: Cu = 4:1 at.%), and TaCu2 (Ta: Cu = 1:1 at.%) were deposited on titanium by magnetron sputtering. The bare titanium and the three coatings were subsequently anodized into four kinds of nanotubes (NT) of TNT, Ta-NT, TaCu1-NT, and TaCu2-NT, respectively. The released copper ions measured by inductively coupled plasma atomic emission spectroscopy (ICP/AES) presented that TaCu2-NT coating released the highest amount of copper ions, which led to the best bacteriostasis against Escherichia coli and Staphylococcus aureus. Potentiodynamic polarization tests clarified that Ta-NT showed the highest corrosion resistance, followed by TaCu1-NT and TaCu2-NT. TaCu2-NT showed not only the best angiogenic property in terms of cell migration, tube formation, and real-time quantitative polymerase chain reaction (RT-qPCR) of human umbilical vein endothelial cells (HUVECs), but also the best osteogenic property in terms of cell viability, alkaline phosphatase activity, and mineralization of MC3T3-E1 cells. Therefore, TaCu2-NT coating has a greater potential than the other coatings of TNT, Ta-NT and TaCu1-NT in promoting bacteriostasis, angiogenesis and osteointegration for titanium implants.
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Bone related-bacterial diseases including wound infections and osteomyelitis (OM) remain a serious problem accompanied with amputation in most severe cases. In this work, we report an exceptional effective antibacterial alginate aerogel, which consists of tigecycline (TGC) and octahedral Cu crystal as an organo-inorganic synergy platform for antibacterial and local infection therapy applications. The alginate aerogel could greatly prolong the release of copper ions and maintain effective antibacterial concentration over 18 days. The result of in-vitro experiments demonstrated that the alginate aerogel has an exceptional effective function on antibacterial activity. Cytotoxicity tests indicated that the alginate aerogel has low biological toxicity (average cell viability >75%). These remarkable results suggested that the alginate aerogel exhibits great potential for the treatment of OM, and has a prosperous future of application in bone tissue engineering.
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Traditional bone wax has lots of shortcomings such as the risk of infection and inflammation and the ability to hinder osteogenesis that limit its clinical applications. In this study, we designed a novel biodegradable bone wax with desirable angiogenic and antibacterial ability and low foreign body reaction by mixing calcium sulfate, poloxamer, and cupric ions. To evaluate its biocompatibility and angiogenetic effect in vitro, we cultured human umbilical vein endothelial cells (HUVECs) with the indicated bone wax to observe cell viability and vessel-like tubular formation. The bone wax was then implanted in a critical-sized bone defect rat model for 4 and 8 weeks to successfully stimulate angiogenesis in vivo. Finally, the bone wax extract was incubated with Gram-positive Staphylococcus aureus to confirm its antibacterial ability. The copper-loaded biodegradable bone wax overcomes the drawbacks of traditional bone wax and provides a new approach for the treatment of bone injuries.
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The effect of dietary copper deficiency on tumour growth, neovascularisation and microvascular integrity was studied in the rat cremaster muscle. Male, weanling Sprague-Dawley rats were fed purified diets which were copper deficient (< 0.5 micrograms g-1 of diet) or copper adequate (5 micrograms g-1 of diet). Seven days after initiation of diets, a chondrosarcoma was implanted in the cremaster muscle of each rat. Five, 10 or 20 days after tumour implantation, rats were anesthetised and their cremasters prepared for observation by intravital microscopy. Intraarterial injection of fluorescein isothiocyanate-conjugated albumin and subsequent observation of fluorescence in the perivascular space indicated no difference in microvascular albumin leakage between the tumour vasculature of copper deficient and copper adequate rats. Neither tumour growth (assessed by wet weight), vascular density (assessed by light microscopy), nor any ultrastructural characteristics of the tumour or its vasculature (assessed by electron microscopy) were affected by copper deficiency. In view of findings by others which indicate changes in tumour characteristics with copper deficiency, we conclude that the copper dependency of tumour growth and vascularisation is a function of the type of tumour, the host tissue, or the conditions of copper depletion.
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Although the angiogenic proteins acidic fibroblast growth factor (FGF-1) and basic fibroblast growth factor (FGF-2) both interact with the transition metal copper, itself a putative modulator of angiogenesis, a role for copper in FGF function has not been established. Using nonreducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis, we detect the complete conversion of recombinant forms of human FGF-1 monomer protein to FGF-1 homodimers after exposure to copper ions. In contrast, not all forms of bovine FGF-1 isolated from bovine brain or a recombinant preparation of human FGF-2 completely formed homodimers after exposure to copper ions under similar conditions. Since the copper-induced FGF-1 homodimers reverted to the monomer form in the presence of dithiothreitol, specific alkylation of cysteine residues by pyridylethylation prevented FGF-1 homodimer formation, and preformed FGF-1 homodimers could not be dissociated by the metal chelator EDTA, FGF-1 dimer formation appeared to result from the formation of intermolecular disulfide bonds by copper-induced oxidation of sulfhydryl residues. FGF-1 homodimers bound with similar apparent affinity as FGF-1 monomers to immobilized copper ions, both eluting at 60 mM imidazole. Both human FGF-1 monomer and dimer forms had a 6-fold higher apparent affinity for immobilized copper ions, as compared with human FGF-2, which eluted in the monomer form at 10 mM imidazole. Further, in contrast to FGF-1 monomers, which dissociate from immobilized heparin in 1.0 M NaCl, preformed FGF-1 homodimers had reduced apparent affinity for immobilized heparin and eluted at 0.4 M NaCl. In contrast, the apparent affinity of human FGF-2 for immobilized heparin was unaffected after exposure to copper ions. Heparin appeared to modulate the formation of copper-induced intermolecular disulfide bonds for FGF-1 but not FGF-2, since co-incubation of heparin and copper with FGF-1 monomers resulted in dimers and other oligomeric complexes. FGF-1 copper-induced homodimers failed to induce mitogenesis in [3H]thymidine incorporation assays, an effect which could be reversed by treatment with dithiothreitol, whereas FGF-2-induced mitogenic activity was relatively unaffected by pretreatment with copper. The differences between human FGF-1 and FGF-2 in protein-copper interactions may be due to differing free thiol content and arrangement between the two proteins. A recombinant human FGF-1 mutant containing the two cysteines conserved throughout the FGF family of proteins but lacking a cysteine residue (Cys 131) present in wild-type human FGF-1 but not human FGF-2 readily formed copper-induced dimers.(ABSTRACT TRUNCATED AT 400 WORDS)
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Microvascular proliferation, a hallmark of malignant brain tumors, represents an attractive target of anticancer research, especially because of the quiescent nonproliferative endothelium of the normal brain. Cerebral neoplasms sequester copper, a trace metal that modulates angiogenesis. Using a rabbit brain tumor model, normocupremic animals developed large vascularized VX2 carcinomas. By contrast, small, circumscribed, relatively avascular tumors were found in the brains of rabbits copper-depleted by diet and penicillamine treatment (CDPT). The CDPT rabbits showed a significant decrease in serum copper, copper staining of tumor cell nuclei, microvascular density, the tumor volume, endothelial cell turnover, and an increase in the vascular permeability (breakdown of the blood-brain barrier), as well as peritumoral brain edema. In non-tumor-bearing animals, CDPT did not alter the vascular permeability or the brain water content. CDPT also inhibited the intracerebral growth of the 9L gliosarcoma in F-344 rats, with a similar increase of the peritumoral vascular permeability and the brain water content. CDPT failed to inhibit tumor growth and the vascularization of the VX2 carcinoma in the thigh muscle or the metastases to the lung, findings that may reflect regional differences in the responsiveness of the endothelium, the distribution of copper, or the activity of cuproenzymes. Metabolic and pharmacologic withdrawal of copper suppresses intracerebral tumor angiogenesis; angiosuppression is a novel biologic response modifier for the in situ control of tumor growth in the brain.
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To investigate the effects of D-penicillamine (D-Pen) on angiogenesis, we have studied the effects of this drug on in vitro proliferation of human endothelial cells (EC) and in vivo corneal neovascularization. D-Pen, in the presence of copper sulfate, suppressed tritiated thymidine ([3H]TdR) incorporation into EC in a dose-dependent manner. Significant inhibition was observed with D-Pen concentrations attainable in the serum and tissues of treated patients. Neither D-Pen nor copper ion alone significantly affected [3H]TdR incorporation into EC. The inhibition by D-Pen and copper was blocked by catalase (CAT) or horseradish peroxidase but not by boiled CAT or SOD. When rabbits were daily injected intravenously with D-Pen at the per kilogram dosage administered to rheumatoid patients, neovascularization as quantitated by the proliferation of corneal new blood vessels was significantly inhibited. These results suggest that hydrogen peroxide generated by D-Pen and copper exerts a pronounced antiangiogenic effect through inhibition of EC proliferation. It is, therefore, considered that D-Pen may suppress rheumatoid synovitis by reducing the number of small blood vessels available for the emigration of chronic inflammatory cells, and the proliferation of the synovial tissue.
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Salts of CuI, II, NiII, SnII, InIII and a sub-fraction of the cupoprotein ceruloplasmin induced phagokinesis of cultured aortal endothelial cells. A variant aortal endothelial cell line was highly sensitive; cells travelled up to 1000 μm in 24 h in response to 2× 10−6 M SnCl2. Other metal ions tested (ZnII, CoII, MnII, CrII, FeIII, AlIII, SbIII and MoII) were not active. The motility response of endothelial cells to Cu ions in vitro is proposed as a model system for studying early events in neovascularization and as a sensitive assay for detecting angiogenic activity in fractions from cells and tissues.
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High affinity fibroblast growth factor (FGF) receptors contain a cluster of acidic amino acids in their extracellular domains that is reminiscent of the calcium binding domains of some cell adhesion molecules. Based on this observation, we used a calcium blotting technique to show that FGFR-1 binds calcium and that calcium binding is not observed in a mutagenized form of the receptor that lacks the acidic box region. The acidic box also binds other divalent cations, including copper. This latter interaction appears unique since the binding of copper to FGFR-1 mediates the binding of the receptor to immobilized heparin. While this observation may help explain the angiogenic properties of copper, divalent cation binding to FGF receptors may also mediate the interaction between FGF receptors, cell adhesion molecules and other proteoglycan components of the extracellular matrix.
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Preclinical and in vitro studies have determined that copper is an important cofactor for angiogenesis. Tetrathiomolybdate (TM) was developed as an effective anticopper therapy for the initial treatment of Wilson's disease, an autosomal recessive disorder that leads to abnormal copper accumulation. Given the potency and uniqueness of the anticopper action of TM and its lack of toxicity, we hypothesized that TM would be a suitable agent to achieve and maintain mild copper deficiency to impair neovascularization in metastatic solid tumors. Following preclinical work that showed efficacy for this anticopper approach in mouse tumor models, we carried out a Phase I clinical trial in 18 patients with metastatic cancer who were enrolled at three dose levels of oral TM (90, 105, and 120 mg/day) administered in six divided doses with and in-between meals. Serum ceruloplasmin (Cp) was used as a surrogate marker for total body copper. Because anemia is the first clinical sign of copper deficiency, the goal of the study was to reduce Cp to 20% of baseline value without reducing hematocrit below 80% of baseline. Cp is a reliable and sensitive measure of copper status, and TM was nontoxic when Cp was reduced to 15-20% of baseline. The level III dose of TM (120 mg/ day) was effective in reaching the target Cp without added toxicity. TM-induced mild copper deficiency achieved stable disease in five of six patients who were copper deficient at the target range for at least 90 days.
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Fibroblast growth factor (FGF) 1 is known to be released in response to stress conditions as a component of a multiprotein aggregate containing the p40 extravescicular domain of p65 synaptotagmin (Syt) 1 and S100A13. Since FGF1 is a Cu2+-binding protein and Cu2+ is known to induce its dimerization, we evaluated the capacity of recombinant FGF1, p40 Syt1, and S100A13 to interact in a cell-free system and the role of Cu2+ in this interaction. We report that FGF1, p40 Syt1, and S100A13 are able to bind Cu2+ with similar affinity and to interact in the presence of Cu2+ to form a multiprotein aggregate which is resistant to low concentrations of SDS and sensitive to reducing conditions and ultracentrifugation. The formation of this aggregate in the presence of Cu2+ is dependent on the presence of S100A13 and is mediated by cysteine-independent interactions between S100A13 and either FGF1 or p40 Syt1. Interestingly, S100A13 is also able to interact in the presence of Cu2+ with Cys-free FGF1 and this observation may account for the ability of S100A13 to export Cys-free FGF1 in response to stress. Lastly, tetrathiomolybdate, a Cu2+ chelator, significantly represses in a dose-dependent manner the heat shock-induced release of FGF1 and S100A13. These data suggest that S100A13 may be involved in the assembly of the multiprotein aggregate required for the release of FGF1 and that Cu2+ oxidation may be an essential post-translational intracellular modifier of this process.
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Angiogenesis plays a central role in wound healing. Among many known growth factors, vascular endothelial growth factor (VEGF) is believed to be the most prevalent, efficacious, and long-term signal that is known to stimulate angiogenesis in wounds. Whereas a direct role of copper to facilitate angiogenesis has been evident two decades ago, the specific targets of copper action remained unclear. This report presents first evidence showing that inducible VEGF expression is sensitive to copper and that the angiogenic potential of copper may be harnessed to accelerate dermal wound contraction and closure. At physiologically relevant concentrations, copper sulfate induced VEGF expression in primary as well as transformed human keratinocytes. Copper shared some of the pathways utilized by hypoxia to regulate VEGF expression. Topical copper sulfate accelerated closure of excisional murine dermal wound allowed to heal by secondary intention. Copper-sensitive pathways regulate key mediators of wound healing such as angiogenesis and extracellular matrix remodeling. Copper-based therapeutics represents a feasible approach to promote dermal wound healing.
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The release of signal peptideless proteins occurs through nonclassical export pathways and the release of fibroblast growth factor (FGF)1 in response to cellular stress is well documented. Although biochemical evidence suggests that the formation of a multiprotein complex containing S100A13 and Synaptotagmin (Syt)1 is important for the release of FGF1, it is unclear where this intracellular complex is assembled. As a result, we employed real-time analysis using confocal fluorescence microscopy to study the spatio-temporal aspects of this nonclassical export pathway and demonstrate that heat shock stimulates the redistribution of FGF1 from a diffuse cytosolic pattern to a locale near the inner surface of the plasma membrane where it colocalized with S100A13 and Syt1. In addition, coexpression of dominant-negative mutant forms of S100A13 and Syt1, which both repress the release of FGF1, failed to inhibit the stress-induced peripheral redistribution of intracellular FGF1. However, amlexanox, a compound that is known to attenuate actin stress fiber formation and FGF1 release, was able to repress this process. These data suggest that the assembly of the intracellular complex involved in the release of FGF1 occurs near the inner surface of the plasma membrane and is dependent on the F-actin cytoskeleton.
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Copper plays an essential role in promoting angiogenesis. Tumors that become angiogenic acquire the ability to enter a phase of rapid growth and exhibit increased metastatic potential, the major cause of morbidity in cancer patients. We report that copper deficiency induced by tetrathiomolybdate (TM) significantly impairs tumor growth and angiogenesis in two animal models of breast cancer: an inflammatory breast cancer xenograft in nude mice and Her2/neu cancer-prone transgenic mice. In vitro, TM decreases the production of five proangiogenic mediators: (a) vascular endothelial growth factor; (b) fibroblast growth factor 2/basic fibroblast growth factor; (c) interleukin (IL)-1alpha; (d) IL-6; and (e) IL-8. In addition, TM inhibits vessel network formation and suppresses nuclear factor (NF)kappaB levels and transcriptional activity. Our study suggests that a major mechanism of the antiangiogenic effect of copper deficiency induced by TM is suppression of NFkappaB, contributing to a global inhibition of NFkappaB-mediated transcription of proangiogenic factors.
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Tetrathiomolybdate (TM), a copper-lowering agent, has been shown in preclinical murine tumor models to be antiangiogenic. We evaluated the antitumor activity of TM in patients with advanced kidney cancer in a Phase II trial. Fifteen patients with advanced kidney cancer were eligible to participate in this trial. TM was initiated p.o. at 40 mg three times a day with meals and 60 mg at bedtime to deplete copper. A target serum ceruloplasmin (CP) level of 5-15 mg/dl was defined as copper depletion. Doses of TM were reduced for grade 3-4 toxicity and to maintain a CP level in the target range. Once copper depletion was attained, patients underwent baseline tumor measurements and then again every 12 weeks for response assessment. Patients not exhibiting progressive disease at 12 weeks after copper depletion continued on treatment. Serum levels of Interleukin (IL)-6, IL-8, vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) were assayed pretreatment and at various time points on treatment. Dynamic contrast enhanced-magnetic resonance imaging (DCE-MRI) was performed on selected patients in an attempt to assess changes in tumor vascularity. All of the patients rapidly became copper depleted. Thirteen patients were evaluable for response. No patient had a complete response or PR. Four patients (31%) had stable disease for at least 6 months during copper depletion (median, 34.5 weeks). TM was well tolerated, with dose reductions most commonly occurring for grade 3-4 granulocytopenia of short duration not associated with febrile episodes. Serum levels of IL-6, IL-8, VEGF, and bFGF did not correlate with clinical activity. Serial DCE-MRI was performed only in four patients, and a decrease in vascularity seemed to correlate with necrosis of a tumor mass associated with tumor growth. TM is well tolerated and consistently depletes copper as measured by the serum CP level. Clinical activity was limited to stable disease for a median of 34.5 weeks in this Phase II trial in patients with advanced kidney cancer. Serum levels of proangiogenic factors IL-6, IL-8, VEGF, and bFGF may correlate with copper depletion but not with disease stability in this small cohort. TM may have a role in the treatment of kidney cancer in combination with other antiangiogenic therapies.
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The induction of an acute inflammatory response followed by the release of polypeptide cytokines and growth factors from peripheral blood monocytes has been implicated in mediating the response to vascular injury. Because the Cu2+-binding proteins IL-1alpha and fibroblast growth factor 1 are exported into the extracellular compartment in a stress-dependent manner by using intracellular Cu2+ to facilitate the formation of S100A13 heterotetrameric complexes and these signal peptideless polypeptides have been implicated as regulators of vascular injury in vivo, we examined the ability of Cu2+ chelation to repress neointimal thickening in response to injury. We observed that the oral administration of the Cu2+ chelator tetrathiomolybdate was able to reduce neointimal thickening after balloon injury in the rat. Interestingly, although immunohistochemical analysis of control neointimal sections exhibited prominent staining for MAC1, IL-1alpha, S100A13, and the acidic phospholipid phosphatidylserine, similar sections obtained from tetrathiomolybdate-treated animals did not. Further, adenoviral gene transfer of the IL-1 receptor antagonist during vascular injury also significantly reduced the area of neointimal thickening. Our data suggest that intracellular copper may be involved in mediating the response to injury in vivo by its ability to regulate the stress-induced release of IL-1alpha by using the nonclassical export mechanism employed by human peripheral blood mononuclear cells in vitro.
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SPARC is a transiently expressed extracellular matrix-binding protein that alters cell shape and regulates endothelial cell proliferation in vitro. In this study, we show that SPARC mRNA and protein are synthesized by endothelial cells during angiogenesis in vivo. SPARC and peptides derived from a cationic region of the protein (amino acids 113-130) stimulated the formation of endothelial cords in vitro; moreover, these peptides stimulated angiogenesis in vivo. Mapping of the active domain demonstrated that the sequence KGHK was responsible for most of the angiogenic activity; substitution of the His residue decreased the effect. We found that proteolysis of SPARC provided a source of KGHK, GHK, and longer peptides that contained these sequences. Although the Cu(2+)-GHK complex had been identified as a mitogen/morphogen in normal human plasma, we found KGHK and longer peptides to be potent stimulators of angiogenesis. SPARC113-130 and KGHK were shown to bind Cu2+ with high affinity; however, previous incubation with Cu2+ was not required for the stimulatory activity. Since a peptide from a second cationic region of SPARC (SPARC54-73) also bound Cu2+ but had no effect on angiogenesis, the angiogenic activity appeared to be sequence specific and independent of bound Cu2+. Thus, specific degradation of SPARC, a matrix-associated protein expressed by endothelial cells during vascular remodeling, releases a bioactive peptide or peptides, containing the sequence (K)GHK, that could regulate angiogenesis in vivo.
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Objective: To determine whether tetrathiomolybdate (TM), a powerful chelator of copper, is capable of lowering the body stores of copper and suppressing the growth of head and neck squamous cell carcinoma in an orthotopic murine model. Study design: In vivo, murine model. Methods: Twelve 8-week-old male C3H/HeJ mice were assigned to either a TM treatment group (n = 7) or a control group (n = 5). Serum samples were obtained from a single mouse in each group to measure the level of ceruloplasmin as a surrogate marker of total body copper on days 0, 4, and 7. Mice in both groups received a floor-of-mouth injection of 1.5 x 105 SCC VII/SF cells. After 7 to 10 days of tumor growth the treatment group received fresh water daily, to which TM was added to achieve an oral intake of 50 mg per mouse. The control group received only fresh drinking water daily. Tumor volume measurements were obtained every other day. Microvessel density counts were assessed in the tumors by Factor VIII analysis. Results: Measurable tumor growth was achieved in 100% of the mice by the tenth day. Total body copper was reduced by 28% from baseline levels in mice in the treatment group. The difference in mean tumor volume in the control group was 4.7 times greater than the TM-treated group at the completion of treatment (3004 mm3 and 633mm3, respectively). This accounted for an overall suppression rate of 79% (P =.008; two-tailed Student t test). In addition, microvessel density was reduced by 50% in the TM-treated group. Conclusion: In this initial study, the first of its kind in head and neck squamous cell carcinoma, we have demonstrated the ability of TM to significantly suppress both the growth of squamous cell carcinoma and tumor vascularity in this orthotopic murine model, suggesting its potential for efficacy in the treatment of this disease in humans.
Article
Copper ions stimulate proliferation of human umbilical artery and vein endothelial cells but not human dermal fibroblasts or arterial smooth muscle cells. Incubation of human umbilical vein endothelial cells for 48 h with 500 μM CuSO4 in a serum-free medium in the absence of exogenous growth factors results in a twofold increase in cell number, similar to the cell number increase induced by 20 ng/ml of basic fibroblast growth factor under the same conditions. Copper-induced proliferation of endothelial cells is not inhibited by 10% fetal bovine serum or by the presence of antibodies against a variety of angiogenic, growth, and chemotactic factors including angiogenin, fibroblast growth factors, epidermal growth factor, platelet-derived growth factor, tumor necrosis factor-α, transforming growth factor-β, macrophage/monocyte chemotactic and activating factor, and macrophage inflammatory protein-1α. Moreover, despite the previous observations that copper increased total specific binding of 125I-angiogenin to endothelial cells, binding to the 170 kDa receptor is not changed; hence, the mitogenic activity of angiogenin is not altered by copper. Copper-induced proliferation, along with early reports that copper induces migration of endothelial cells, may suggest a possible mechanism for the involvement of copper in the process of angiogenesis. J. Cell. Biochem. 69:326–335, 1998. © 1998 Wiley-Liss, Inc.
Article
Copper, cadmium, and disulfiram (an ionophore for copper) modulate the synthesis of several polypeptides in two clonal lines of bovine aortal endothelial cells. After treatment of Type 1 endothelial cells with 10(-3) M CuSO4 or 10(-5) M CdCl2 four cell-associated polypeptides (Mr = 28,000, 32,000, 73,000, and 83,000 daltons) were induced. In contrast, in Type 2 endothelial cells, which have cultural characteristics distinct from Type 1, only one new cell-associated protein (Mr = 32,000 daltons) was induced by CuSO4 or CdCl2. The same four polypeptides, described above, were induced by disulfiram (10(-7) M) in Type 1 endothelial cells. In contrast when Type 2 endothelial cells were treated with disulfiram the synthesis of only two new cell-associated proteins (Mr = 32,000 and 40,000 daltons) was induced. Other differences are revealed by analyses of proteins secreted into the growth medium. In particular low levels of only CuSO4 (10(-6) M) enhanced the synthesis in Type 2 cells of a protein (Mr = 220,000 daltons) identified as fibronectin. Since only copper ions induced fibronectin, we propose that the mechanism of induction of fibronectin synthesis, in contrast to the induction of cell-associated polypeptides, does not involve a sulphydryl-containing receptor molecule. It is suggested that the specific enhancement of fibronectin synthesis by copper ions may be a controlling event in the stimulation by copper ions of endothelial cell migration and angiogenesis.
Article
Low concentrations of copper sulphate, Dispirin, or Walker carcinoma extract elicit intraocular vascularization when tested by anterior chamber implants in rats. The response is markedly depressed by pre-treatment of animals with methylprednisolone acetate, suggesting that such induced vascularization is mediated by leucocytes. Since many agents inducing vascularization also induce migration of cultured endothelial cells, it raises the problem of how to isolate and study the action of possible leucocyte-derived angiogenic factors. Regardless of the identity of the natural angiogenic factor or factors, it is proposed that during blood vessel formation specialized endothelial cells migrate in response to an angiogenic signal and deposit fibronectin on which other cells can track and subsequently adhere, forming an endothelium.
Article
The involvement of leukocytes in corneal neovascularization has been known for a long time. Recent observations suggest that collagenase from leukocytes may be a common mediator for prostaglandin E1 (PGE1)- and copper-induced corneal neovascularization. This study was designed to investigate the effect of copper ion on collagenase activity from leukocytes and other sources and leukocyte infiltration in the corneal angiogenic process induced by PGE1. These results demonstrated that collagenase production from leukocytes was stimulated in a dose-dependent manner by copper ion but not by PGE1. Copper chloride 0.2 mM produced the highest stimulation. Copper ion had no effect on collagenase release from corneal fibroblasts and capillary endothelium. There were more polymorphonuclear leukocytes (PMN) in the prostaglandin E1 treated corneas than in the control. The time-course study showed that the appearance of PMN reached a peak on day 2 and new vessel growth could not be identified until day 4. These results supported an earlier suggestion that leukocytes play a role in corneal neovascularization and further suggested that copper in corneal neovascularization can stimulate the release of collagenase from leukocytes.
Article
The principal objective of our work is to sufficiently understand the mechanism of angiogenesis in the adult organism to allow interference with the process on a rational basis. It is apparent that several "factors" can trigger angiogenesis. To test these, we used the rabbit cornea mostly because it is avascular (i.e., the background is zero) and transparent (i.e., the newly formed capillaries that invade the cornea are easily visible in the unanaesthetized animal). Under these conditions, it was found that the cornea ready to be colonized by capillaries under the action of an angiogenesis effector becomes rich in copper ions and sialic acid. Motility of bovine capillary endothelium was utilized to analyze the angiogenesis process on the ground that mobilization of capillary endothelium is the first morphological event observed during angiogenesis in vivo and the methods to measure cell motility are reasonably accurate. With this approach it was found that heparin, fibronectin, and gangliosides are involved in the mobilization of capillary endothelium. The precise interaction among these three components is not yet clear.
Article
The purpose of this study was to determine the copper deposition and localization during the evolution of two murine mammary adenocarcinomas. In the normal tissue, the copper was located within the cytoplasm, whereas it was intra- and perinuclear in the tumors. The more angiogenic and metastatic tumor showed the higher percentage of copper-positive cells. In the tumor, copper deposits correlated well with its angiogenic and metastatic ability, but additional factors would be required for the process to be induced.
Article
64Cu was injected in the form of CuCl2 either by subcutaneous or by intraperitoneal route, and its distribution inside different organs was analyzed in 5 different tumor models, 4 in mice and 1 in rats. In all organs tested (blood, liver, kidneys, spleen, intestine, muscle, and tumor) no significant differences were observed in the results obtained after either injection route. All tumors analyzed (Krebs ascite, intestinal Leiomyo sarcoma, human tumor, mammary adenocarcinoma, either spontaneous or chemically induced) contained a relatively high concentration of 64Cu. For all tumor models tested, the 64Cu distribution was altered as compared with that of the corresponding control animals.
Article
The first human tumor derived protein with in vivo angiogenic activity to be obtained in pure form has been isolated from serum-free supernatants of an established human adenocarcinoma cell line (HT-29) and named angiogenin. It was purified by cation-exchange and reversed-phase high-performance liquid chromatography; the yield was approximately 0.5 microgram/L of medium. Biological activity of angiogenin was monitored throughout purification by using the chick embryo chorioallantoic membrane assay. Statistical evaluation demonstrates that it displays activity in this system with as little as 35 fmol per egg. Moreover, only 3.5 pmol is required to induce extensive blood vessel growth in the rabbit cornea. The amino acid composition of this basic (isoelectric point greater than 9.5), single-chain protein of molecular weight approximately 14 400 has been determined. The amino terminus is blocked, and the carboxyl-terminal residue is proline.
Article
This article has no abstract; the first 100 words appear below. THE growth of solid neoplasms is always accompanied by neovascularization. This new capillary growth is even more vigorous and continuous than a similar outgrowth of capillary sprouts observed in fresh wounds or in inflammation.¹ Many workers have described the association between growing solid malignant tumors and new vessel growth.²³⁴⁵⁶ However, it has not been appreciated until the past few years that the population of tumor cells and the population of capillary endothelial cells within a neoplasm may constitute a highly integrated ecosystem. In this ecosystem the mitotic index of the two cell populations may depend upon each other. Tumor cells . . . Supported by a grant (5 RO1 CA08185–06) from the National Cancer Institute, a grant from the American Cancer Society, National Chapter (IC-28), and gifts from the Merck Company and the Alza Corporation. Source Information From the Department of Surgery, Children's Hospital Medical Center and Harvard Medical School, Boston, Massachusetts 02115.
Article
Copper's role in connective tissue is linked to the enzyme lysyl oxidase. From a biochemical perspective, copper is a cofactor for the enzyme and a determinant of its activity in connective tissues. Lysyl oxidase catalyses a post-translational oxidation of certain lysine and hydroxylysine residues. The peptidyl aldehydes so formed become active centres for the formation of cross-links in collagen and elastin. Less well understood is how copper controls the steady-state activity of lysyl oxidase; the enzyme fails in copper deficiency. Giving copper to a deprived animal increases lysyl oxidase activity in aortic tissue. Such activation in vivo appears to require caeruloplasmin. Suspending aortic tissue in a copper-enriched growth medium also activates lysyl oxidase provided that tissue structure is kept intact. Activation in vitro occurs with the binding of copper to a large-molecular-weight component, presumably the enzyme. Binding will not occur if protein synthesis is blocked. These studies clearly show that the synthesis of mature elastin and collagen can be controlled by the availability of copper. They further suggest that transport of copper to aortic tissue and its engagement to lysyl oxidase are linked to the synthesis or lysyl oxidase, an extracellular carrier, or both.
Article
The interstitial fluid of MTW9A and Walker carcinomas and their ethanol extract induced strong angiogenic response in the rabbit (New Zealand White) corneal test. The fluid collected in vivo was rich in E-type prostaglandins, and prostaglandin E1 (PGE1) in particular was strongly angiogenic at the lowest dose as compared with the angiogenic responses of prostaglandins E2, I2, and F2 alpha. Neoplastic fibroblasts also induced a strong angiogenic response, but in indomethacin-treated rabbits neovascularization failed to occur. Copper was concentrated in the cornea during PGE1-induced neovascularization, and copper-deficient rabbits were unable to mount an angiogenic response in the corneal test. Ceruloplasmin, the copper carrier of plasma, was found to be angiogenic at high doses. In indomethacin-treated rabbits, however, ceruloplasmin at the same high doses failed to induce angiogenesis. The experiments are interpreted to indicate that angiogenesis is the end result of a sequence of events, two of which are PGE1 production and copper mobilization in the tissue where neovascularization occurs.
Article
The ability to induce new formation of capillaries in the cornea was tested for ceruloplasmin, the copper carrier of serum, for fragments of the ceruloplasmin molecule with and without copper, for heparin, and for glycyl-L-histidyl-L-lysine, bound or not bound to copper ions. Male or female 2- to 3-kg New Zealand White rabbits were used. These experiments were prompted by the previous observation of copper accumulation in the cornea during angiogenesis and by the inability of copper-deficient rabbits to mount an angiogenic response. The results showed that the three different molecules were all able to induce angiogenesis provided that they were bound to copper. Fragments of the ceruloplasmin molecule also induced angiogenesis but only when copper was bound to the peptides. The data are interpreted to indicate that copper ions are involved in the sequence of events leading to angiogenesis and that the carrier molecules may be of quite a different nature.
Article
The objective of this work was to contribute to the interpretation of the mechanisms of capillary formation in the adult tissues. We have observed that the heparin-copper complex is angiogenic in vivo and stimulates migration of capillary endothelium in vitro. This effect is specific for capillary endothelium since aortic endothelium or fibroblasts from the rabbit cornea or human skin were unresponsive.
Article
SPARC is a transiently expressed extracellular matrix-binding protein that alters cell shape and regulates endothelial cell proliferation in vitro. In this study, we show that SPARC mRNA and protein are synthesized by endothelial cells during angiogenesis in vivo. SPARC and peptides derived from a cationic region of the protein (amino acids 113-130) stimulated the formation of endothelial cords in vitro; moreover, these peptides stimulated angiogenesis in vivo. Mapping of the active domain demonstrated that the sequence KGHK was responsible for most of the angiogenic activity; substitution of the His residue decreased the effect. We found that proteolysis of SPARC provided a source of KGHK, GHK, and longer peptides that contained these sequences. Although the Cu(2+)-GHK complex had been identified as a mitogen/morphogen in normal human plasma, we found KGHK and longer peptides to be potent stimulators of angiogenesis. SPARC113-130 and KGHK were shown to bind Cu2+ with high affinity; however, previous incubation with Cu2+ was not required for the stimulatory activity. Since a peptide from a second cationic region of SPARC (SPARC54-73) also bound Cu2+ but had no effect on angiogenesis, the angiogenic activity appeared to be sequence specific and independent of bound Cu2+. Thus, specific degradation of SPARC, a matrix-associated protein expressed by endothelial cells during vascular remodeling, releases a bioactive peptide or peptides, containing the sequence (K)GHK, that could regulate angiogenesis in vivo.
Article
Angiogenin is a potent inducer of blood-vessel formation with ribonucleolytic activity. Angiogenin binds to high affinity endothelial cell receptors and with lower affinity to extracellular matrix components. Here we report the effect of copper and zinc on these interactions. There was a 4.3-fold increase in angiogenin binding to calf pulmonary artery endothelial cells in the presence of Cu2+ in vitro. A 3.8-fold increase was observed with Zn2+, whereas Ni2+, Co2+, or Li+ had no effect. Specific angiogenin binding to the lower affinity matrix sites was increased by 2.7- and 1.9-fold in the presence of Cu2+ and Zn2+ respectively. Metal ion affinity chromatography and atomic absorption spectrometry were used to show the direct interaction of angiogenin with copper and zinc ions. Angiogenin bound 2.4 mol of copper per mole of protein. We suggest that copper, a modulator of angiogenesis in vivo, may be involved in the regulation of the biological activity of angiogenin.
Article
Copper ions stimulate proliferation of human umbilical artery and vein endothelial cells but not human dermal fibroblasts or arterial smooth muscle cells. Incubation of human umbilical vein endothelial cells for 48 h with 500 microM CuSO4 in a serum-free medium in the absence of exogenous growth factors results in a twofold increase in cell number, similar to the cell number increase induced by 20 ng/ml of basic fibroblast growth factor under the same conditions. Copper-induced proliferation of endothelial cells is not inhibited by 10% fetal bovine serum or by the presence of antibodies against a variety of angiogenic, growth, and chemotactic factors including angiogenin, fibroblast growth factors, epidermal growth factor, platelet-derived growth factor, tumor necrosis factor-alpha, transforming growth factor-beta, macrophage/monocyte chemotactic and activating factor, and macrophage inflammatory protein-1alpha. Moreover, despite the previous observations that copper increased total specific binding of 125I-angiogenin to endothelial cells, binding to the 170 kDa receptor is not changed; hence, the mitogenic activity of angiogenin is not altered by copper. Copper-induced proliferation, along with early reports that copper induces migration of endothelial cells, may suggest a possible mechanism for the involvement of copper in the process of angiogenesis.
Article
Fibronectin, a large extracellular matrix cell adhesion glycoprotein has diverse functions in wound repair including organization of matrix deposition and promotion of angiogenesis. We have previously shown that purified plasma fibronectin can be made into three-dimensional, fibrous materials, termed fibronectin mats (Fn-mat). The aim of this study was to examine means of increasing the stability of Fn-mats using a novel treatment with micromolar concentrations of copper ions which may be used to improve wound healing/nerve repair. Cytotoxicity of incorporated copper was determined using rat Schwann cells, rat tendon fibroblasts and human dermal fibroblasts. Dissolution of protein from the Fn-mat showed that treatment with the lowest concentration of copper used (1 microM) increased the stability of mats by 3-4 times at room temperature relative to control mats and twofold at 37 degrees C. Copper mediated increase in stability was dose dependent. Orientation of the Fn-fibres (within mats), monitored by scanning electron microscopy was retained with 1 microM copper but disappeared with higher concentrations. Schwann cells grew in culture with mats stabilized by 1 microM copper treatment without reduction in cell number but growth was inhibited at 10-200 microM Cu. All types of fibroblasts were unaffected by copper treatment upto 200 microM. Fn-mats can be successfully stabilized by this technique producing longer survival in vitro. The differential effects of copper on these cell types suggests that CuFn-mats may be used to select the type of cells which colonize these materials.
Article
Copper (Cu) accumulating in a form bound to metallothionein (MT) in the liver of Long-Evans rats with a cinnamon-like coat color (LEC rats), an animal model of Wilson disease, can be removed from the MT with tetrathiomolybdate (TTM). However, the insoluble Cu/TTM complex formed with excess TTM is known to be deposited in the liver. The metabolic fate of the insoluble Cu/TTM complex was investigated in the present study. LEC rats were injected with TTM at the dose of 10 mg/kg body weight for 8 consecutive days and were fed with a standard or low Cu diet for 14 days after the last injection. About 95% of the Cu in the liver became insoluble together with Mo. The concentration of Cu in the liver supernatants of rats fed with the standard diet increased significantly compared with that in rats dissected 24 h after the last injection (control rats), while the concentration in rats fed with the low Cu diet remained at a comparable level to that in the controls. The rate of Cu accumulation in the livers of rats fed with the standard diet did not differ before and after the treatment, suggesting that there was no rebound effect by treatment with TTM. These results suggest that the insoluble Cu/TTM complex is resolubilized in the liver, and that the solubilized complex is excreted into the bile and blood, i.e., the insoluble Cu/TTM complex is not the source of Cu re-accumulation in the form bound to MT in the liver after TTM treatment. It was concluded that, once Cu is complexed with TTM, the metal is excreted either immediately in the soluble form or slowly in the insoluble form into the bile and blood.
Article
Angiogenesis is now recognized as a crucial process in tumor development, including hepatocellular carcinoma (HCC). Since HCC is known as a hypervascular tumor, anti-angiogenesis is a promising approach to inhibit the HCC development. Trientine dihydrochloride (trientine) is used in clinical practice as an alternative copper (Cu)-chelating agent for patients with Wilson's disease of penicillamine intolerance. In our study, we examined the effect of Cu-chelating agents on tumor development and angiogenesis in the murine HCC xenograft model. Although both trientine and penicillamine in the drinking water suppressed the tumor development, trientine exerted a more potent inhibitory effect than penicillamine. In combination with a Cu-deficient diet, both trientine and penicillamine almost abolished the HCC development. Trientine treatment resulted in a marked suppression of neovascularization and increase of apoptosis in the tumor, whereas tumor cell proliferation itself was not altered. In vitro studies also exhibited that trientine is not cytotoxic for the tumor cells. On the other hand, it significantly suppressed the endothelial cell proliferation. These results suggested that Cu plays a pivotal role in tumor development and angiogenesis in the murine HCC cells, and Cu-chelators, especially trientine, could inhibit angiogenesis and enhance apoptosis in the tumor with consequent suppression of the tumor growth in vivo. Since trientine is already used in clinical practice without any serious side effects as compared to penicillamine, it may be an effective new strategy for future HCC therapy.
Article
Recent studies have revealed that copper is an important cofactor for several angiogenic agents. We examined the antiangiogenic effect against hepatocellular carcinoma (HCC) of the copper chelator trientine, especially focusing on the relationship between copper and interleukin-8 (IL-8), a potent angiogenic factor produced by hepatoma cells. HuH-7 hepatoma cells were transplanted into nude mice and the growth of xenograft tumors was compared to and without administration of trientine. Using the resected tumor, microvessel density, apoptotic potential and proliferative activity were analyzed histologically and IL-8 mRNA was semiquantified by RT-PCR. In addition, HuH-7 cells were cultured in control medium, medium supplemented with copper, medium supplemented with trientine and medium supplemented with both copper and trientine. Human IL-8 levels were measured in the supernatants by ELISA. Using the extracts from cultured cells, IL-8 mRNA was semiquantified by RT-PCR. Trientine suppressed the growth of xenograft tumors significantly. Histologically, apoptotic potential was increased significantly and microvessel density, decreased. The production of IL-8 from the tumor was suppressed by trientine. In vitro, IL-8 production by HuH-7 cells in copper-containing medium was significantly greater than that in copper-free medium, and this effect was weakened when trientine was added. However, no significant change was apparent when trientine was added to the medium alone. In conclusion, the chelating effect of trientine prevented copper from functioning as a cofactor in angiogenesis, which resulted in reduced IL-8 production from HuH-7 cells.