[Show abstract][Hide abstract] ABSTRACT: Imbalanced matrix metalloproteinase (MMP)-2 activity and transforming growth factor expression (TGF-β) are involved in vascular remodeling of hypertension. Atorvastatin and sildenafil exert antioxidant and pleiotropic effects that may result in cardiovascular protection. We hypothesized that atorvastatin and sildenafil alone or in association exert antiproliferative effects by down-regulating MMP-2 and TGF-β, thus reducing the vascular hypertrophy induced by two kidney, one clip (2K1C) hypertension. Sham and 2K1C rats were treated with oral atorvastatin 50mg/kg, sildenafil 45mg/kg, or both, daily for 8 weeks. Blood pressure was monitored weekly. Morphologic changes in the aortas were studied. TGF-β levels were determined by immunofluorescence. MMP-2 activity and expression were determined by in situ zymography, gel zymography, Western blotting, and immunofluorescence. The effects of both drugs on proliferative responses of aortic smooth muscle cells to PDGF and on on MMP-2 activity in vitro were determined. Atorvastatin, sildenafil, or both drugs exerted antiproliferative effects in vitro. All treatments attenuated 2K1C-induced hypertension and prevented the increases in the aortic cross-sectional area and media/lumen ratio in 2K1C rats. Aortas from 2K1C rats showed higher collagen deposition, TGF-β levels and MMP-2 activity and expression when compared with Sham-operated animals. Treatment with atorvastatin and/or sildenafil was associated with attenuation of 2K1C hypertension-induced increases in these pro-fibrotic factors. However, these drugs had no in vitro effects on hr-MMP-2 activity. Atorvastatin and sildenafil was associated with decreased vascular TGF-β levels and MMP-2 activity in renovascular hypertensive rats, thus ameliorating the vascular remodeling. These novel pleiotropic effects of both drugs may translate into protective effects in patients.
[Show abstract][Hide abstract] ABSTRACT: Results:
Compared with the first time point, all the samples collected at the second time point contained lower lead concentration (p<0.05), which suggested that the adopted measures effectively reduced exposure of the population to lead present in contaminated soil and dust. Statistically significant correlations only existed between lead in blood collected in the first year and lead in blood collected in the second year (Spearman's r=0.55; p<0.0001; n=62), and lead in house dust collected in the first year and lead in house dust collected in the second year (Spearman's r=0.5; p<0.0001; n=59).
Results support the validity of lead determination in blood and in house dust to assess lead exposure over time. However, lead in blood and lead in dust did not correlate with lead in serum or lead in saliva.
Environmental Research 07/2015; 142:337-344. DOI:10.1016/j.envres.2015.05.023 · 4.37 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Imbalanced matrix metalloproteinase (MMP) activity is involved in hypertensive cardiac hypertrophy. Pharmacological inhibition of nuclear factor kappaB (NF-кB) with pyrrolidine dithiocarbamate (PDTC) can prevent MMP up-regulation. We hypothesized that treatment with PDTC could prevent 2-kidney, 1-clip (2K1C) hypertension-induced left ventricular remodelling. Sham-operated controls or 2K1C rats with hypertension received either vehicle or PDTC (100 mg.Kg(-1) .day(-1) ) by gavage for eight weeks. Systolic blood pressure was monitored every week. Histological assessment of left ventricles was carried out with hematoxylin/eosin sections and fibrosis was quantified in picrosirius red-stained sections. Oxidative stress was evaluated in heart samples with the dihydroethidium probe. Cardiac MMP activity was determined by in situ zymography and cardiac MMP-2 was assessed by immunofluorescence. 2K1C surgery significantly increased systolic blood pressure in the 2K1C Vehicle. PDTC exerted antihypertensive effects after 2 weeks of treatment. Histology revealed increased left ventricle and septum wall thickness associated with augmented myocyte diameter in hypertensive rats, which were reversed by treatment with PDTC. Hypertensive rats developed pronounced cardiac fibrosis with increased interstitial collagen area, increased cardiac reactive oxygen species levels, gelatinase activity and MMP-2 expression. PDTC treatment decreased these alterations. These findings show that PDTC modulates myocardial MMP-2 expression and ameliorates cardiac remodelling in renovascular hypertension. These results suggest that interfering with MMPs expression at transcriptional level may be an interesting strategy in the therapy of organ damage associated with hypertension. This article is protected by copyright. All rights reserved.
This article is protected by copyright. All rights reserved.
[Show abstract][Hide abstract] ABSTRACT: Objectives
Investigate whether low-level laser therapy (LLLT) alters the expression and activity of MMP-2 and MMP-9 in the trigeminal ganglion (TG) during different stages of temporomandibular joint (TMJ) inflammation in rats. It also evaluated whether LLLT modifies mechanical allodynia and orofacial hyperalgesia.Materials and methodsWistar rats (± 250 g) were divided into groups that received saline (SAL) or complete Freund's adjuvant (CFA, 50 μL) in the TMJ, and that later underwent LLLT (20 J/cm2) at their TMJ or not (groups SAL, SAL+LLLT, CFA, and CFA+LLLT). LLLT was applied on days 3, 5, 7, and 9 after SAL or CFA. Mechanical allodynia was evaluated on days 1, 3, 5, 7, and 10; orofacial hyperalgesia was assessed on day 10. Gelatin zymography and in situ zymography aided quantification of MMPs in the TG.ResultsLLLT abolished the reduction in the mechanical orofacial threshold and the increase in orofacial rubbing during the orofacial formalin test induced by CFA. LLLT also decreased the CFA-induced rise in the levels of MMP-9 and MMP-2 as well as the gelatinolytic activity in the TG.ConclusionLLLT could constitute an adjuvant therapy to treat temporomandibular disorders and prevent inflammation-induced alterations in the levels of MMP-2 and MMP-9 and in the gelatinolytic activity in TGs.This article is protected by copyright. All rights reserved.
[Show abstract][Hide abstract] ABSTRACT: Background:
Abdominal aortic aneurysm (AAA) is characterized by chronic inflammation and degradation of the extracellular matrix, mediated by matrix metalloproteinases (MMPs). Doxycycline has been reported to control the progression of AAA by regulation of MMP. We hypothesized that doxycycline pretreatment in a rat model of AAA would cause reduction in gelatinolytic activity of MMP-2 and -9 and the inflammatory response in the wall of an aneurysm, consequently decreasing the formation and development of AAAs.
Male Wistar rats were divided into the following four groups: aneurysm (A); control (C); aneurysm+doxycycline (A+D) and control+doxycycline (C+D), with 24 animals per group subdivided into n=6 animals at different time points [1, 3, 7, and 15 days postsurgery (dps)]. The (A) and (A+D) groups simultaneously received the injury and extrinsic stenosis of the aortic wall. The (C) and (C+D) groups received sham operation. The treated animals received doxycycline via gavage (30 mg/kg/day) from 48 h before surgery until the end of experiment. At 1, 3, 7, and 15 dps, the animals were euthanized, and the aortas were collected for morphological analyses, immunohistochemistry, and zymography.
The animals from the (A) group developed AAAs. However, the animals treated with doxycycline showed a 85% decrease in AAA development, which was associated with a large reduction in gelatinolytic activity of MMP-2 and -9, and decreased inflammatory response (P<.05).
These results suggest that pretreatment with doxycycline before surgery inhibited the activity of MMP-2 and -9, as well as the inflammatory response, and may play an important role in the prevention of the development of AAAs.
[Show abstract][Hide abstract] ABSTRACT: Objective:
The present study evaluated the use of a reagent to stabilize the DNA extracted from human dental tissues stored under different temperature conditions and time intervals.
Material and methods:
A total of 161 teeth were divided into two distinct groups: intact teeth and isolated dental pulp tissue. The samples were stored with or without the product at different time intervals and temperature. After storage, DNA extraction and genomic DNA quantification were performed using real-time PCR; the fragments of the 32 samples that represented each possible condition were analyzed to find the four pre-selected markers in STR analysis.
The results of the quantification showed values ranging from 0.01 to 10,246.88 ng/μL of DNA. The statistical difference in the quantity of DNA was observed when the factors related to the time and temperature of storage were analyzed. In relation to the use of the specific reagent, its use was relevant in the group of intact teeth when they were at room temperature for 30 and 180 days. The analysis of the fragments in the 32 selected samples was possible irrespective of the amount of DNA, confirming that the STR analysis using an automated method yields good results.
The use of a specific reagent showed a significant difference in stabilizing DNA in samples of intact human teeth stored at room temperature for 30 and 180 days, while the results showed no justification for using the product under the other conditions tested.
Journal of applied oral science: revista FOB 07/2014; 22(4):331-335. DOI:10.1590/1678-775720130683 · 0.92 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Although disorders of the stomatognathic system are common, the mechanisms involved are unknown. Our objective was to study the changes in the masseter muscles after unilateral exodontia. Molar extraction was performed on Wistar
rats (left side), and the animals were sacrificed after either 14 or 26 days. The masseter muscle was processed for histological analysis, conventional and in situ zymography, and immunohistochemistry. The morphological analysis showed unique and specific characteristics for the experimental group. By conventional zymography no significant values of 72 kDa MMP-2 (P < 0.05) were found in both of the sides of masseter muscle after 14 and 26 days of unilateral extraction. The in situ zymography showed gelatinolytic activity on all deep masseter muscles, with significant increase on the contralateral side after 14 and 26 days (P < 0.05). The immunohistochemistry demonstrated greater expression of MMP-2 than MMP-9 and MMP-14 in all masseter muscles and there were few differences in the staining of 4 TIMPs. This knowledge about morphology and molecular masticatory muscle remodeling following environmental interventions can be used to develop clinically successful treatments.
BioMed Research International 06/2014; 2014:563463. DOI:10.1155/2014/563463 · 2.71 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Hypertension induces left ventricular hypertrophy (LVH) by mechanisms involving oxidative stress and imbalanced cardiac matrix metalloproteinase (MMP) activity. We hypothesized that β1-adrenergic receptor blockers with antioxidant properties (nebivolol) could reverse hypertension-induced LVH more effectively than conventional β1-blockers (metoprolol) when used at doses that exert similar antihypertensive effects. Two kidney one clip (2K1C) hypertension was induced in male Wistar rats. Six weeks after surgery, hypertensive and sham rats were treated with nebivolol (10mg.kg(-1).day(-1)) or metoprolol (20mg.kg(-1).day(-1)) for four weeks. Systolic blood pressure (SBP) was monitored weekly by tail-cuff plethysmography. LV structural changes and fibrosis were studied in hematoxylin/eosin and picrosirius stained sections, respectively. Cardiac MMP levels and activity were determined by in situ zymography, gel zymography, and by immunofluorescence. Dihydroethidium (DHE) and lucigenin-derived chemiluminescence assays were used to assess cardiac reactive oxygen species production. Nitrotyrosine levels were determined in LV samples by immunohistochemistry. green fluorescence, and was evaluated using the ImageJ software. Cardiac protein kinase B/Akt (AKT) phosphorilation state was assessed by Western blot. Both treatments exerted very similar antihypertensive effects. Both beta-blockers exerted similar antihypertensive effects and attenuated hypertension-induced cardiac remodeling. Both drugs reduced myocyte hypertrophy and collagen deposition in 2K1C rats. Those effects were associated with lower cardiac ROS and nitrotyrosine levels, and attenuation of hypertension-induced increases in cardiac MMP-2 levels and in situ gelatinolytic activity after treatment with both β-blockers. While hypertension increased AKT phosphorilation, no effects were found with β-blockers. In conclusion, we found evidence that two β1-blockers with different properties attenuate hypertension-induced LV hypertrophy and cardiac collagen deposition in association with significant cardiac antioxidant effects and MMP-2 dowregulation, thus suggesting a critical role for β1-adrenergic receptors in mediating those effects. Nebivolol is not superior to metoprolol, at least with respect to their capacity to reverse hypertension-induced LVH.
Free Radical Biology and Medicine 06/2014; 73. DOI:10.1016/j.freeradbiomed.2014.05.024 · 5.74 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: This MiniReview describes the essential biochemical and molecular aspects of matrix metalloproteinases (MMPs) and briefly discusses how they engage in different diseases, with particular emphasis on cardiovascular diseases. There is compelling scientific evidence that many MMPs, especially MMP-2, play important roles in the development of cardiovascular diseases; inhibition of these enzymes is beneficial to many cardiovascular conditions, sometimes precluding or postponing end-organ damage and fatal outcomes. Conducting comprehensive discussions and further studies on how MMPs participate in cardiovascular diseases is important, because inhibition of these enzymes may be an alternative or an adjuvant for current cardiovascular disease therapy.This article is protected by copyright. All rights reserved.
[Show abstract][Hide abstract] ABSTRACT: Shed teeth have been proposed as trace element biomarkers. This study determined variations in the spatial distribution of Ca, K, Zn, Pb, Mn, Cu, and Sr in four anatomical locations: superficial enamel (SE, 0-10 μm), subsuperficial enamel (SSE, 10-30 μm), primary dentine (PD), and secondary dentine (SD). Five primary incisors were analyzed by micro Synchrotron Radiation X-Ray Fluorescence (μ-SRXRF). Two teeth had low concentrations of lead in the SE (< 250 μg/g), while three contained very high lead concentrations in the SE (> 2,000 μg/g). Teeth were sliced, and five spot measurements (20 μm beam diameter) were accomplished in each location. The data are shown as absolute values and as the ratio between the different elements and Ca. The distribution of K was close to that of Ca. Zn was the third most abundant element, with the highest levels being found in the SE and SD and low levels detected in the PD. Increasing Sr levels were found progressing from the enamel to the dentine, with the highest levels being found in the SD, a distribution that was unique. Pb, Mn, and Cu exhibited a similar trend, with higher signals for these elements detected in the SE. This study provides preliminary data on the heterogeneous distribution of different elements in the tooth, highlighting the importance of the first 10 μm of the SE for determination of some elements, such as Zn, Pb, Mn, and Cu.
Journal of Trace Elements in Medicine and Biology 04/2014; 28(2). DOI:10.1016/j.jtemb.2014.01.007 · 2.37 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to see whether there would be differences in whole blood versus tibia lead concentrations over time in growing rats prenatally. Lead was given in the drinking water at 30 mg/L from the time the dams were pregnant until offspring was 28- or 60-day-old. Concentrations of lead were measured in whole blood and in tibia after 28 (28D) and 60 days (60D) in control (C) and in lead-exposed animals (Pb). Lead measurements were made by GF-AAS. There was no significant difference (P > 0.05) in the concentration of whole blood lead between Pb-28D (8.0 ± 1.1 μ g/dL) and Pb-60D (7.2 ± 0.89 μ g/dL), while both significantly varied (P < 0.01) from controls (0.2 μ g/dL). Bone lead concentrations significantly varied between the Pb-28D (8.02 ± 1.12 μ g/g) and the Pb-60D (43.3 ± 13.26 μ g/g) lead-exposed groups (P < 0.01), while those exposed groups were also significantly higher (P < 0.0001) than the 28D and 60D control groups (Pb < 1 μ g/g). The Pb-60D group showed a 25% decrease in tibia mass as compared to the respective control. The five times higher amount of lead found in the bone of older animals (Pb-60D versus Pb-28D), which reinforces the importance of using bone lead as an exposure biomarker.
[Show abstract][Hide abstract] ABSTRACT: Matrix metalloproteinase‐2 (MMP‐2) shares structural similarities with the angiotensin‐converting enzyme (ACE). ACE inhibitors have been described to inhibit MMP‐2, but this inhibitory potential was not shown using a highly purified MMP‐2. This study aimed to investigate the inhibitory potential of captopril and lisinopril regarding MMP‐2 activity. The first objective was to test the potential of captopril to change the pH of the buffer solution. The second objective was to test the direct inhibitory effect of captopril and lisinopril on plasma MMP‐2 and on recombinant human MMP‐2 (rhMMP‐2). The in vitro activity assays included gelatin zymography and a fluorimetric assay. Captopril solubilization significantly decreased the pH of the 50 mM Tris buffer solution at the following concentrations: 2 mM (p p p p p p p in vivo after drug administration. We also discuss possible pitfalls for gelatinase inhibitory assays (besides the obvious pH problem already cited). In conclusion, this study's data show that captopril and lisinopril did not inhibit MMP‐2 directly at the concentrations reached in vivo.
[Show abstract][Hide abstract] ABSTRACT: Nitric oxide (NO)-derived metabolites including the anion nitrite can recycle back to NO and thus complement NO formation independent of NO synthases. While nitrite is as a major vascular storage pool and source of NO, little is known about drugs that increase tissue nitrite concentrations. This study examined the effects of atorvastatin or sildenafil, or the combination, on vascular nitrite concentrations and on endothelial dysfunction in the 2 kidney-1 clip (2K1C) hypertension model. Sham-operated or 2K1C hypertensive rats were treated with vehicle, atorvastatin (50 mg/Kg), sildenafil (45 mg/Kg), or both for 8 weeks. Systolic blood pressure (SBP) was monitored weekly. Nitrite concentrations were assessed in the aortas and in plasma samples by ozone-based reductive chemiluminescence assay. Aortic rings were isolated to assess endothelium-dependent and independent relaxation. Aortic NADPH activity and ROS production were evaluated by luminescence and dihydroethidium, respectively, and plasma TBARS levels were measured. Aortic nitrotyrosine staining was evaluated to assess peroxynitrite formation. Atorvastatin and sildenafil, alone or combined, significantly lowered SBP by approximately 40 mmHg. Atorvastatin significantly increased vascular nitrite levels by 70% in hypertensive rats, whereas sildenafil had no effects. Both drugs significantly improved the vascular function, and decreased vascular NADPH activity, ROS, and nitrotyrosine levels. Lower plasma TBARS concentrations were found with both treatments. The combination of drugs showed no improved responses compared to each drug alone. These findings show evidence that atorvastatin, but not sildenafil, increases vascular NO stores, although both drugs exert antioxidant effects, improve endothelial function, and lower blood pressure in 2K1C hypertension.
[Show abstract][Hide abstract] ABSTRACT: Activated matrix metalloproteinases (MMPs) cause cardiomyocyte injury during acute pulmonary thromboembolism (APT). However, the functional consequences of this alteration are not known. We examined whether doxycycline (a MMP inhibitor) improves right ventricle function and the cardiac responses to dobutamine during APT. APT was induced with autologous blood clots (350 mg/kg) in anaesthetized male lambs pre-treated with doxycycline (Doxy, 10 mg/kg/day, intravenously) or saline. Non-embolized control lambs received doxycycline pre-treatment or saline. The responses to intravenous dobutamine (Dob, 1, 5, 10 μg/kg/min.) or saline infusions at 30 and 120 min. after APT induction were evaluated by echocardiography. APT increased mean pulmonary artery pressure and pulmonary vascular resistance index by ~185%. Doxycycline partially prevented APT-induced pulmonary hypertension (P < 0.05). RV diameter increased in the APT group (from 10.7 ± 0.8 to 18.3 ± 1.6 mm, P < 0.05), but not in the Doxy+APT group (from 13.3 ± 0.9 to 14.4 ± 1.0 mm, P > 0.05). RV dysfunction on stress echocardiography was observed in embolized lambs (APT+Dob group) but not in embolized animals pre-treated with doxycycline (Doxy+APT+Dob). APT increased MMP-9 activity, oxidative stress and gelatinolytic activity in the RV. Although doxycycline had no effects on RV MMP-9 activity, it prevented the increases in RV oxidative stress and gelatinolytic activity (P < 0.05). APT increased serum cardiac troponin I concentrations (P < 0.05), doxycycline partially prevented this alteration (P < 0.05). We found evidence to support that doxycycline prevents RV dysfunction and improves the cardiac responses to dobutamine during APT.
Journal of Cellular and Molecular Medicine 11/2013; 17(12). DOI:10.1111/jcmm.12163 · 4.01 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Increased matrix metalloproteinase (MMP) levels are involved in vascular remodeling of hypertension. In this study, we hypothesized that doxycycline (a MMP inhibitor) could exert antioxidant effects, reverse establish vascular remodeling, and lower blood pressure in spontaneously hypertensive rats (SHR). SHR and Wistar-Kyoto rats received either doxycycline at 30 mg/kg/day by gavage or vehicle. Systolic blood pressure (SBP) was assessed weekly by tail cuff. After 5 weeks of treatment, morphologic changes in the aortic wall were studied in hematoxylin/eosin sections. MMP activity and expression were determined by in situ zymography using DQ gelatin and immunofluorescence for MMP-2. Dihydroethidium was used to evaluate aortic reactive oxygen species (ROS) production by fluorescence microscopy. Doxycycline reduced SBP by 25 mmHg. However, the antihypertensive effects were not associated with significant reversal of hypertension-induced vascular hypertrophy. SHR showed increased aortic MMP-2 levels which co-localized with higher aortic MMP activity and ROS levels, and all those biochemical alterations associated with hypertension were blunted by treatment with doxycycline. These results show that MMP inhibition with doxycycline in SHR with established hypertension resulted in antioxidant effects, lower gelatinolytic activity, and antihypertensive effects which were not associated with reversal of hypertension-induced vascular remodeling.
[Show abstract][Hide abstract] ABSTRACT: The genetic-molecular methodology stands out as an accurate technique for human identification process and among the sources of biological evidence, the use of teeth is of great interest in Forensic Dentistry. Maintaining integrity of the material sent to laboratory is essential for success of the analysis, and one of the main difficulties is related to sample storage, which is usually carried out at low temperatures. This study evaluated the effectiveness of the Allprotect Tissue Reagent® (Qiagen, Germany) in stabilizing DNA extracted from human dental tissues stored under different conditions. Methods: In this study were used 160 teeth, distributed in two groups: intact teeth and isolated pulp tissue. The samples were stored with or without the product and varying the storage time and temperature. In addition to these groups, was formed a positive control group, composed by five teeth, which was stored at -20ºC for 180 days. After storage, DNA extraction, electrophoresis on agarose gel and genomic DNA quantification by Real-Time PCR were performed. The fragments of 32 samples representing every possible condition and five positive control group samples were analyzed to verify four pre-selected markers. Results: The agarose gel showed evidences of genomic DNA presence. Quantification results showed values ranging from 0.01 to 10,246.88 ng/μL of DNA. There was a decrease in DNA concentration in stored tooth samples at room temperature for 30 and 180 days compared to those stored for 1 and 7 days. Besides the time factor, temperature also influenced the DNA concentration, being higher in teeth that remained for 30 days and in tooth pulp maintained for 180 days, under refrigeration. Regarding the use of Allprotect Tissue Reagent® it showed a significant difference in stabilization of stored teeth at room temperature for 30 and 180 days. The analysis of fragments was possible in 37 selected samples, regardless of the DNA quantity variation, confirming that STR analysis using automated methods provides good results. Conclusion: The use of Allprotect Tissue Reagent® showed a significant difference in stabilizing DNA samples of intact human teeth stored at room temperature for 30 and 180 days, while in the other test conditions the results showed no justification for using this product.
The Journal of forensic odonto-stomatology 10/2013; 31 Suppl 1:134-5.