Article

A Hyaluronic Acid-Salmon Calcitonin Conjugate for the Local Treatment of Osteoarthritis: Chondro-protective effect in a rabbit model of early OA.

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Abstract

Osteoarthritis (OA) is characterized by chronic degeneration of joints, involving mainly the articular cartilage and the underlying bone, and severely impairing the quality of life of the patient. Although with limited efficacy, currently available pharmacological treatments for OA aim to control pain and to retard disease progression. Salmon calcitonin (sCT) is a drug which has been shown to have therapeutic effects in experimental arthritis by inhibiting both bone turnover and cartilage degradation and reducing the activities of matrix metalloproteinases (MMP). High molecular weight hyaluronic acid (HA) is used as a lubricant in OA therapy, and, interestingly, HA polymers may normalize the levels of MMP-1, -3 and -13. We demonstrated that sCT rapidly clears from the knee joint of rat animal model, after intra-articular (i.a.) administration, and it induces systemic effects. Here, sCT was conjugated to HA (200kDa) with the aim of prolonging the residence time of the polypeptide in the joint space by reducing its clearance. An aldehyde derivative of HA was used for N-terminal site-selective coupling of sCT. The activity of sCT was preserved, both in vitro and in vivo, after its conjugation and the i.a. injection of HA-sCT did not trigger any systemic effects in rats. The efficacy of HA-sCT treatment was tested in a rabbit OA model and clear chondro-protective effect was proven by macro- and microscopic assessments and histological findings. Our results indicate that HAylation of sCT increases the size of the polypeptide in a stable covalent manner and delays its passage into the blood stream. We conclude that HA conjugation prolongs the anti-catabolic effects of sCT in joint tissues, including the synovial membrane and cartilage.

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... Academic and industrial researchers have presented and investigated new polymers for protein delivery such as hydroxy-ethyl-starch [31], polyoxazoline [32][33][34], dextrin [35,36] polysialic acid [37,38], hyaluronic acid [39][40][41][42], polypeptides (XTEN technology [43], PASylation [44]), etc. While biodegradable polymers are desirable for drug delivery, biodegradability is not easily controlled. ...
... Later, more sophisticated conjugates were proposed by exploiting HA not only as a simple inert carrier but also as an active ingredient whose biological activity could be combined to that of a conjugated drug to achieve synergic therapeutic actions. A HA-salmon calcitonin (HA-sCT) conjugate was, for example, designed to treat osteoarthritis [40], a disease characterized by the chronic deterioration of the articular cartilage of joints and the underlying bone. Proposed as a drug, sCT showed therapeutic effects in experimental arthritis by inhibiting both bone turnover and cartilage degradation and reducing matrix metalloproteinases (MMP) activities. ...
... In view of the facts that high molecular weight HA is already used as a viscosupplement in osteoarthritis and has been shown to normalize the levels of MMP-1, -3 and -13, HAylation of sCT was considered a reasonable approach to combine the two synergic actions and as a means to delay the rapid clearance of sCT from knee articular space after intraarticular injections of the free form. As the conjugate showed good preservation of sCT activity and prolonged residence time in the joint space, it proved possible to avoid systemic exposure to free sCT [40]. A promising chondro-protective effect has also been demonstrated in a rabbit model of osteoarthritis. ...
Article
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Drug and protein delivery techniques and methods, which, in most cases, are concurrently investigated from the very time a new molecular entity is proposed, have become valuable tools for the development of numerous therapeutic molecules. Attracting the attention of academia and industrial researchers, several polymer conjugation methodologies have been shown to have a wide range of applications and a success rate that have been confirmed by the numerous conjugates already in clinical use or under investigation in on-going clinical trials. Having attained a high level of sophistication, this technology is able to generate homogeneous conjugates by exploiting specifically designed coupling strategies (based, for example, on the use of enzymes), which ensure good product characterization, high activity retention, and batch-to-batch reproducibility. Poly(ethylene glycol) (PEG), which is the current gold standard for stealth polymers in the emerging field of polymer-based drug delivery, is in competition with other polymers that have convenient features such as biodegradability. PEG still holds first place as far as PEGylated derivatives under investigation in clinical trials are concerned, and it will presumably remain the polymer of choice even in the future as further improvements such as the development of new copolymers based on PEG or PEG derivatives will overcome for example the hurdle of biodegradability.
... Evaluated on model proteins (RNase A, trypsin and insulin), the new hyaluronan-aldehyde derivative for protein HAylation produced some impressive results, such as, for example, good retention of enzymatic activity (in the range of 60-80%) and increased thermal stability of proteins [45,46]. We recently described the advantages of HA conjugation with respect to salmon calcitonin (sCT) for the local treatment of osteoarthritis and demonstrated that the conjugate prolonged the anti-catabolic effects of sCT in joint tissues, including the synovial membrane and cartilage, while it prevented sCT clearance from the joint cavity thus hindering undesired sCT systemic effects [47]. ...
... None of these treatments are, however, able to delay disease progression and patients often find themselves facing total joint replacement surgery [51]. As explained above, we recently demonstrated that HA-sCT conjugate can be considered a valid approach to improve the management of osteoarthritis because it combines sCT's activity in inhibiting bone turnover and cartilage degradation with HA's action in reducing MMPs expression [47]. ...
... In vivo studies confirmed that hGH was quickly cleared from the joint space while HA-hGH was retained at the injection site. In agreement with a study focusing on HA-salmon calcitonin conjugate [47], these findings, although preliminary are important to identifying a local hGH-based therapeutic approach for OA. ...
Article
Hyaluronic acid (HA) is a natural polysaccharide primarily present in the vitreous humor and in cartilages where it plays a key structural role in organizing the cartilage extracellular matrix. HA is used in a wide range of applications including treatment of arthritis (as a viscosupplementation agent for joints) and in a variety of cosmetic injectable products. Its safety profile is thus well established. Thanks to its high biocompatibility and targeting properties, HA has also been investigated for use as a carrier of anticancer drugs and, recently, also of proteins. Its role in the last case is a particularly challenging one as dedicated coupling chemistries are required to preserve the protein's conformation and activity. This study focuses on the state of the art on protein HAylation. New data from our laboratory on the local delivery of specific biologics to joints will also be outlined.
... Therefore, most of the SF proteins will probably be metabolized or replaced within a few hours. Particles with higher molecular weights, such as high molecular weight HA, can have a turnover rate greater than 10 hours [9,10]. When high molecular weight HA is injected into the knee joints, these particles may remain undegraded for a longer period of time. ...
... Although continuous release of PDGF was also observed in the liquid-form PRP, it is likely to be metabolized in a short period of time as the SF turnover rate in the knee joint is about 1 hour [9]. When the liquid-form PRP was heated up to 95 degrees Celsius, significant decreases in PDGF concentrations were observed in the paste-like PRP end product as compared with other heating temperatures. ...
Article
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Objectives: The texture of the autologous platelet-rich plasma (PRP) that is used in treating degenerative joint diseases such as knee osteoarthritis (OA) is usually in liquid form. However, the turnover rate of protein metabolism in the knee synovial fluid (SF) is less than one hour. This study examined the feasibility of the thermal oscillation technique in converting the liquid-form PRP into an injectable viscous paste-like PRP that may delay the degradation of PRP and continuously release growth factors in the knee joint for a longer period of time. Methods: This study was conducted in the rehabilitation department of a tertiary hospital. A total of 10 elderly patients with an average age of greater than 65 years and diagnosed with moderate degree of knee OA were recruited. The RegenPRP (RegenLab, Le Mont-sur-Lausanne, Switzerland) test tube chamber was used for PRP generation. A total of 60 milliliters (mL) of blood was drawn from each patient. 10 mL of blood was injected into each PRP test tube chamber. As a result, a total of 6 test tube chambers were obtained and each chamber was centrifuged for 15 minutes. Approximately 5 mL of PRP supernatant (the liquid-form end product) was aspirated and sent for thermal oscillation treatment. Five temperatures were tested: 55, 65, 75, 85, and 95 degrees Celsius. Oscillation was set at 200 revolutions per minute (rpm) for 15 minutes. The enzyme-linked immunosorbent assay (ELISA) was applied in measuring the concentration of platelet-derived growth factor (PDGF) in picogram/milliliter (pg/mL). Repeated measures ANOVA followed by the Bonferroni post hoc test was used to compare the PDGF concentrations between each testing condition. Results: Under 75 degrees Celsius of heating, the resultant paste-like PRP end product had the highest concentration of PDGF in picograms per milliliter (pg/mL) as compared with other heating conditions (p < 0.05). The viscosity of the paste-like PRP was measured to be 70,000 centipoise (cP), which is similar to the viscosity of a toothpaste. The paste-like PRP end product was able to release PDGF continuously for about 14 days, with the highest concentration achieved on the 8th day with an average of 35646 ± 2499 pg/mL. In nonthermally treated liquid-form PRP sample, the highest number of PRP was observed on the 4th day with an average value of 8444 ± 831 pg/mL. Under the heating conditions of 55 and 95 degrees Celsius, the highest concentration of PDGF was observed on the 5th day (13346 ± 764 pg/mL and 3440 ± 303 pg/mL, respectively). Under the heating conditions of 65 and 85 degrees Celsius, the highest concentration of PDGF was observed on the 7th day (15468 ± 744 pg/mL and 20432 ± 1118 pg/mL, respectively). Conclusion: Through thermal oscillation, liquid-form PRP can be converted to paste-like PRP end product with a viscosity similar to that of a toothpaste. The best heating condition was discovered to be 75 degrees Celsius. The paste-like PRP was able to release PDGF continuously for about 2 weeks, with the highest concentration obtained on the 8th day. The findings in this study suggested that paste-like PRP may be a viable option in treating degenerative knee joint diseases.
... During OA, the biochemical status impairment leads to the destruction of subchondral bone and joint cartilage. Some studies indicate that synovial uid (SF), synovium, knee tendon, and ligament were also involved in the pathogenesis and development of OA [2]. In addition, age is one of primary risk factors of OA, since chondrocytes have a reduced capacity to restore and maintain the extracellular matrix in cartilage in elder patients [2]. ...
... Some studies indicate that synovial uid (SF), synovium, knee tendon, and ligament were also involved in the pathogenesis and development of OA [2]. In addition, age is one of primary risk factors of OA, since chondrocytes have a reduced capacity to restore and maintain the extracellular matrix in cartilage in elder patients [2]. ...
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Background: Hyaluronic acid (HA) is the most common intra-articular therapy used to treat mild to moderate osteoarthritis (OA). However, the mechanism involved in this treatment is still not fully understood. The aim of the present study was to examine the effect and the possible mechanism of intra-articular HA (IAHA) injection in patients with knee osteoarthritis (OA). Methods: Twenty-eight patients with Kellgren–Lawrence scale II to III were enrolled in this study. All patients underwent ultrasound-guided injection using three consecutive weekly IAHA. Functional ability and pain were determined by the Western Ontario and McMaster University Index (WOMAC) questionnaire and visual analog scale (VAS). Further, the levels of HA, metalloproteinase (MMP)-1, MMP-3, MMP-13, interleukin (IL)-1β and IL-6 in synovial fluid were determined weekly before HA injection. Results: Functional improvement and pain relief were observed 4 weeks after treatment. At week 4, a significant increase of HA concentration was found, and the concentration of inflammatory cytokines including IL-1β, and IL-6, as well as matrix MMP-3 and MMP-13 significantly decreased. However, no significant difference was observed in MMP-1 level. Conclusion: These results suggest that increasing HA accumulation in synovial fluid may be associated with disease relief after weekly IAHA injection in patients with knee OA.
... Corticosteroids are the most investigated agents, since they are commonly used as rapid analgesics for pain management in osteoarthritic patients [69][70][71][72][73][74]. Other drugs are under research like antioxidant molecules [89][90][91][92][93][94] and the so called disease modifying drugs for OA (DMOAD) [95][96][97][98]. DMOAD include drugs like calcitonin and bisphosphonates that are thought to slow down or even inhibit the disease progression [99,100]. ...
... However, its rapid elimination from the body presents limitations for its administration. In order to prevent the joint clearance of calcitonin, Mero et al. prepared HA-salmon calcitonin (HA-sCT) covalent conjugates using an aldehyde derivative of HA for N-terminal site-selective coupling of calcitonin [96]. Calcitonin activity was maintained after its coupling to HA in vitro and in vivo and HA-sCT conjugates did not trigger any systemic effect as calcitonin did. ...
Article
Objective: Osteoarthritis is a chronic, painful and disabling disease which prevalence is increasing in developing countries. Patients with osteoarthritis present a reduced synovial fluid viscoelasticity due to a reduction in concentration and molecular weight of hyaluronic acid. Currently, the main treatment used to restore the compromised rheological properties of synovial fluid is the viscosupplementation by hyaluronic acid injections that can be combined with oral anti-inflammatory drugs for pain relief. Combination of viscosupplements with chemical agents or drugs is emerging as a new strategy to provide a double action of synovial fluid viscoelasticity recovery and the therapeutic effect of the bioactive principle. Methods: In this review, we present the latest research on the combination of viscosupplements with active molecules. We conducted a literature review of articles published in different web search engines and categorized according to the active molecule introduced into the viscosupplement. Results: Generally, the introduction of anti-inflammatory molecules have shown to improve pain relief although some cytotoxicity has been demonstrated especially for non-steroidal anti-inflammatory drugs. Other molecules such as antioxidant or disease modifying osteoarthritis drugs have been reported to improve viscosupplementation action. Drug delivery systems combined with hyaluronic acid could enhance the activity of the encapsulated molecules and provide better control over the drug release. Finally, biological approaches such as the use of stem cells or platelet-rich plasma seem to be the most promising strategies for cartilage recovery. Conclusions: Combination therapy of viscosupplements with therapeutic agents, drug delivery systems or regenerative therapies can improve viscosupplementation outcome in terms of pain relief and joint functionality. However, further research is needed in order to reach more conclusive results.
... When the tibial plateau was observed, OA lesions in this area were mild or moderate. This is a phenomenon that takes place in the ACLT model in rabbits, in agreement with a study by Mero et al. [45]. Given that the lesion is mild or moderate, the tibial plateau appears to be an unreasonable indicator for assessing cartilage degeneration. ...
... Unlike other animals, rabbits tend to be loaded on the lateral aspect of the stifle joint [46]. Mild early arthritic changes begin to appear from the 4th week after ACLT, and severe cartilage degeneration first occurs in the lateral femoral condyle, followed by the medial femoral condyle and meniscus [45]. Therefore, the lateral compartment in the stifle joint of a rabbit is the site for confirming early cartilage changes [47]. ...
Article
Full-text available
We investigated the effects of intra-articular injections of alginate-microencapsulated adipose tissue-derived mesenchymal stem cells (ASCs) during osteoarthritis (OA) development in a rabbit model of anterior cruciate ligament transection (ACLT). We induced OA in mature New Zealand white rabbits by bilateral ACLT. Stifle joints were categorised into four groups according to intra-articular injection materials. Alginate microbeads and microencapsulated ASCs were prepared using the vibrational nozzle technology. Two weeks after ACLT, the rabbits received three consecutive weekly intra-articular injections of 0.9% NaCl, alginate microbeads, ASCs, or microencapsulated ASCs, into each joint. Nine weeks after ACLT, we euthanised the rabbits and collected bilateral femoral condyles for macroscopic, histological, and immunohistochemical analyses. Macroscopic evaluation using the modified OA Research Society International (OARSI) score and total cartilage damage score showed that cartilage degradation on the femoral condyle was relatively low in the microencapsulated-ASC group. Histological analysis of the lateral femoral condyles indicated that microencapsulated ASCs had significant chondroprotective effects. Immunohistochemically, the expression of MMP-13 after the articular cartilage damage was relatively low in the microencapsulated-ASC-treated stifle joints. During the development of experimental OA, as compared to ASCs alone, intra-articular injection of microencapsulated ASCs significantly decreased the progression and extent of OA.
... Bilateral ACLT was performed as described by other authors. 36 The animal's anterior instability was manually confirmed by an anterior drawer test. The joint was flushed with sterile saline solution, and the joint capsule and the incised skin were su-tured. ...
... weeks from ACLT. 12,36,37 To this end, the needle was inserted into the knee joint posterior to the lateral edge of the patella at the junction between femur and tibia to avoid damage to the articular cartilage. For local biodistribution studies, cells were previously labeled with 6 mM chloromethyl-benzamido-1,1-dioctadecyl-3,3,3¢3¢-tetra-methyl-indocarbocyanine per-chlorate (CM-DiI) dye (Molecular Probes, Carlsbad, CA) as indicated by the manufacturer, 41 before their combination with PS or with HA, subsequent IA injections into the OA knee joints, and final evaluation after 7 and 14 days from the treatments. ...
Article
Full-text available
The evaluation of key factors modulating cell homing following transplantation can provide new insights in the comprehension of unsolved biological questions about the use of cell therapies for osteoarthritis (OA). The main purpose of this in vivo study was to investigate the biodistribution of an intra-articular (IA) injection of mesenchymal stromal cells (MSCs) and bone marrow concentrate (BMC) in a rabbit OA model, and whether the additional use of sodium hyaluronate (HA) could modulate their migration and promote joint repair. OA was surgically induced in adult male New Zealand rabbits. A group of animals was used to test the biodistribution of labelled-cells alone or with HA at 7 and 14 days to investigate cell migration. The efficacy of treatments was evaluated in other experimental groups at 2 months. Histology and immunohistochemistry for markers identifying anabolic and catabolic processes in the cartilage and meniscus, or macrophage subset population in the synovial membrane, were performed. Kruskall Wallis test, followed by post-hoc Dunns test and Spearman's rank-order correlation method, were used. MSCs and BMC preferentially migrate towards tissue areas showing OA features in the meniscus and cartilage, and in detail near inflammatory zones in the synovial membrane. The combination with HA contributed to boost cell migration towards articular cartilage. In general, both labelled-cells combined with HA were found near cell cluster and fissures in the cartilage and meniscus respectively, and close to areas of synovial membrane showing mainly anti-inflammatory macrophages. A promotion of joint repair was observed at different levels for all treatments, although BMC-HA treatment resulted the best strategy to support joint repair. This last, owning a high number of platelets, displayed a good protein expression of type II collagen in the cartilage, as well as the presence of anti-inflammatory macrophages in the synovial membrane at 2 months from the treatment. Studies tracking cell biodistribution indicate that priming progenitor cells with HA modulated cell homing favoring not only attachment but also their integration within articular cartilage.
... sCT nevertheless shows a rapid clearance from the joint thus preventing prolonged local action and causes undesired systemic effects. In view of the fact that high molecular weight HA is already being used as a viscosupplement in osteoarthritis and has been shown to normalize the levels of MMP-1, -3 and -13, we designed a HA-salmon calcitonin (HA-sCT) conjugate for the local treatment of osteoarthritis [51]. HAylation of sCT was considered a reasonable approach to combine the two synergic actions and to delay the rapid clearance of sCT from the knee articular space after intra-articular injections. ...
... HAylation of sCT was considered a reasonable approach to combine the two synergic actions and to delay the rapid clearance of sCT from the knee articular space after intra-articular injections. The conjugate showed good preservation of sCT activity and prolonged residence time in the joint space, reducing systemic exposure to sCT [51]. A promising chondroprotective effect has also been demonstrated in a rabbit model of osteoarthritis. ...
Chapter
Full-text available
Originally considered simply another naturally occurring component of a number of tissues, breakthrough findings later disclosed hyaluronan’s (HA) extraordinary biological properties and paved the way for a new era of HA-associated medical applications. HA is, in fact, recognized by many cellular receptors, it can mediate cell migration, proliferation, cell-cell aggregation, it has been shown to promote angiogenesis, and the list goes on. This plethora of activities initially moved attention away from a polymer that was considered a simple carrier of biomolecules and towards its potential use in both treating many diseases and conditions and being involved in drug delivery. Given these premises, medical applications exploiting HA’s different roles have been developed. The focus of this chapter is directed towards the chemical conjugation of HA with small drugs, peptides and proteins. Reviewing the vast body of literature dedicated to this field, an extraordinary range of applications will be outlined. Although HA cannot be considered a polymer that is appropriate for all uses, conjugates specifically designed to exploit some of its biological properties and for numerous specific applications will no doubt enjoy an advantage over other polymeric conjugates.
... A novel functionalised hydrogel with an integrated drug delivery system was developed to stimulate articular cartilage regeneration (19). The hyaluronic acidsalmon calcitonin conjugate was used for the local treatment of osteoarthritis and had chondro-protective effect in a rabbit model of early OA (20). Although the development of drugs in animals and humans is promising in preventing the cartilage macromolecules from breakdown and effectively halting further progression of OA, the long natural history of OA (approximately 10-20 years in humans) validating the efficacy of these drugs requires a noninvasive technique directly evaluating their effect on molecular changes in association with early stages of cartilage degeneration prior to the emergence of distinct morphological changes (13,21,22). ...
... Residual transverse magnetization is then dephased by a crusher gradient. The magnetization stored along the z-axis can then be read out by different pulse sequences such as fast spin-echo, single-shot fast spin-echo, and 3D gradient-echo (20,26). These different pulse sequences were performed in the spin-lock pulse cluster in order to improve the spin-locking robustness to the B 0 and B 1 inhomogeneity. ...
Article
Magnetic resonance imaging (MRI) offers the direct visualization of the human musculoskeletal (MSK) system, especially all diarthrodial tissues including cartilage, bone, menisci, ligaments, tendon, hip, synovium, etc. Conventional MRI techniques based on T1 - and T2 -weighted, proton density (PD) contrast are inconclusive in quantifying early biochemically degenerative changes in MSK system in general and articular cartilage in particular. In recent years, quantitative MR parameter mapping techniques have been used to quantify the biochemical changes in articular cartilage, with a special emphasis on evaluating joint injury, cartilage degeneration, and soft tissue repair. In this article we focus on cartilage biochemical composition, basic principles of T1ρ MRI, implementation of T1ρ pulse sequences, biochemical validation, and summarize the potential applications of the T1ρ MRI technique in MSK diseases including osteoarthritis (OA), anterior cruciate ligament (ACL) injury, and knee joint repair. Finally, we also review the potential advantages, challenges, and future prospects of T1ρ MRI for widespread clinical translation. J. Magn. Reson. Imaging 2014. © 2014 Wiley Periodicals, Inc.
... However, there are still some inevitable problems in current therapeutic approaches. First, the retention time of drugs is short in vivo due to decomposition before reaching the zone of action [27]. Additionally, the protection of drugs is limited. ...
Article
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Bone-related diseases refer to a group of skeletal disorders that are characterized by bone and cartilage destruction. Conventional approaches can regulate bone homeostasis to a certain extent. However, these therapies are still associated with some undesirable problems. Fortunately, recent advances in nanomaterials have provided unprecedented opportunities for diagnosis and therapy of bone-related diseases. This review provides a comprehensive and up-to-date overview of current advanced theranostic nanomaterials in bone-related diseases. First, the potential utility of nanomaterials for biological imaging and biomarker detection is illustrated. Second, nanomaterials serve as therapeutic delivery platforms with special functions for bone homeostasis regulation and cellular modulation are highlighted. Finally, perspectives in this field are offered, including current key bottlenecks and future directions, which may be helpful for exploiting nanomaterials with novel properties and unique functions. This review will provide scientific guidance to enhance the development of advanced nanomaterials for the diagnosis and therapy of bone-related diseases.
... The short maintain time of calcitonin in the knee joint limited its clinical application. After conjugation with hyaluronic acid, the anti-catabolic effects of salmon calcitonin in joint tissues was prolonged by reducing its clearance and chondro-protective effect was preserved without any systematic effect in rat and rabbit model (Mero et al., 2014). Short plasma half-life is one of the main problems for application of bioactive peptides including calcitonin in clinical studies. ...
Article
Background Bioactive peptides from different food protein have been reported to have a lot of biological activities, such as anti-inflammation and anti-oxidation, making them beneficial for chronic disease. Osteoarthritis (OA) is one of the major diseases affecting human health worldwide. The main treatment methods of OA are physiotherapy, medication and surgery. Many drug, such as adrenocortical hormone, have serious side effects on the patients. Therefore, it is necessary to search for treatment with less side effects. Recently, biologically active peptides from natural foods have been extensively reported to show beneficial effects on alleviating symptom of osteoarthritis, which make it potential for industrial application in the functional foods. Scope and approach In this review, we focus on biological characteristics of food-derived peptides, especially their therapeutic effect in osteoarthritis, and the underlying mechanism by which these peptides exert the biologically function in osteoarthritis. Key findings and conclusions Peptides from bovine, chicken, deer are the predominant species for improvement of osteoarthritis in in vitro, in vivo and clinical studies. Marine food is another important protein source for production of bioactive peptide that showed beneficial effect in the treatment of osteoarthritis. A few peptides from plant protein such as soybean also showed positive effect on chondrocytes. However, many bioactive peptides that showed the beneficial effect on osteoarthritis were in the form of mixture. Finding out the exact peptide that exerts the positive effect in the osteoarthritis is necessary for the industrial application in the future.
... In the study, three injections of sCT/HA-sCT once a week were administered, with the first injection at 10 days after surgery. It was observed that both the systems significantly reduced the loss of the superficial layer and erosion of the cartilage and imparted protective effects against the OA-like degenerative changes in the articular cartilage, but the HA-sCT was more potent than the bare sCT (Mero et al., 2014). Dong et al. reported similar observations when Celecoxib liposomes (CLX-Lip) were developed to improve OA therapy and to reduce CLX associated cardiovascular adverse effects. ...
Article
Full-text available
With the change in lifestyle and aging of the population, osteoarthritis (OA) is emerging as a major medical burden globally. OA is a chronic inflammatory and degenerative disease initially manifesting with joint pain and eventually leading to permanent disability. To date, there are no drugs available for the definitive treatment of osteoarthritis and most therapies have been palliative in nature by alleviating symptoms rather than curing the disease. This coupled with the vague understanding of the early symptoms and methods of diagnosis so that the disease continues as a global problem and calls for concerted research efforts. A cascade of events regulates the onset and progression of osteoarthritis starting with the production of proinflammatory cytokines, including interleukin (IL)‐1β, IL‐6, tumor necrosis factor (TNF)‐α; catabolic enzymes, such as matrix metalloproteinases (MMPs)‐1, ‐3, and ‐13, culminating into cartilage breakdown, loss of lubrication, pain, and inability to load the joint. Although intra‐articular injections of small and macromolecules are often prescribed to alleviate symptoms, low residence times within the synovial cavity severely impair their efficacy. This review will briefly describe the factors dictating the onset and progression of the disease, present the current clinically approved methods for its treatment and diagnosis, and finally elaborate on the main challenges and opportunities for the application of nano/micromedicines in the treatment of osteoarthritis. Thus, future treatment regimens will benefit from simultaneous consideration of the mechanobiological, the inflammatory, and tissue degradation aspects of the disease. This article is categorized under: Nanotechnology Approaches to Biology > Nanoscale Systems in Biology Implantable Materials and Surgical Technologies > Nanotechnology in Tissue Repair and Replacement
... Sophisticated technologies now offer an unprecedented number of site selective conjugation methods based on chemical, enzymatic, or genetic approaches ( Figure 5), but applications have been limited to improving protein pharmacokinetics or in some cases to reducing the immunogenicity of non-human proteins. In the future, the polymer may be able to contribute to the protein's pharmacological activities (Mero et al., 2014) or it may modulate the pharmacodynamic activity of a protein, as has already been reported, for example, Pegvisomat or Bempegaldesleukin, a drug in a phase II clinical trial for treatment of urothelial cancer (Table 1). Research will hopefully be able to develop "smart" conjugates that are able to respond to specific conditions permitting the protein to carry out its pharmacological activity. ...
Article
Polymer conjugation can be considered one of the leading approaches within the vast field of nanotechnology-based drug delivery systems. In fact, such technology can be exploited for delivering an active molecule, such as a small drug, a protein, or genetic material, or it can be applied to other drug delivery systems as a strategy to improve their in vivo behavior or pharmacokinetic activities such as prolonging the half-life of a drug, conferring stealth properties, providing external stimuli responsiveness, and so on. If on the one hand, polymer conjugation with biotech drug is considered the linchpin of the protein delivery field boasting several products in clinical use, on the other, despite dedicated research, conjugation with low molecular weight drugs has not yet achieved the milestone of the first clinical approval. Some of the primary reasons for this debacle are the difficulties connected to achieving selective targeting to diseased tissue, organs, or cells, which is the main goal not only of polymer conjugation but of all delivery systems of small drugs. In light of the need to achieve better drug targeting, researchers are striving to identify more sophisticated, biocompatible delivery approaches and to open new horizons for drug targeting methodologies leading to successful clinical applications. This article is categorized under: Therapeutic Approaches and Drug Discovery > Emerging Technologies Therapeutic Approaches and Drug Discovery > Nanomedicine for Oncologic Disease Toxicology and Regulatory Issues in Nanomedicine > Regulatory and Policy Issues in Nanomedicine.
... Nonetheless, many researchers are investigating alternative polymers (some biodegradable and others not) such as polyglutamic acid (a negatively charged and biodegradable polymer [12]), polysarcosine (a biodegradable polypeptoid based on the amino acid sarcosine, i.e. N-methyl glycine, [13][14][15]), hyaluronic acid (a biodegradable polysaccharide composed of glucuronic acid and N-acetyl glucosamine [16][17][18]), polysialic acid (a biodegradable polysaccharide of N-acetylneuraminic acid [19][20][21]), poly-2-ethyl-2-oxazoline (a non-biodegradable polymer, [22][23][24][25]), etc. ...
Article
Poly(L-glutamic acid)-co-poly(ethylene glycol) block copolymers (PLE-PEG) are here investigated as polymers for conjugation to therapeutic proteins such as granulocyte colony stimulating factor (G-CSF) and human growth hormone (hGH). PLE-PEG block copolymers are able to stabilize and protect proteins from degradation and to prolong their residence time in the blood stream, features that are made possible thanks to PEG's intrinsic properties and the simultaneous presence of the biodegradable anionic PLE moiety. When PLE-PEG copolymers are selectively tethered to the N-terminus of G-CSF and hGH, they yield homogeneous monoconjugates that preserve the protein's secondary structure. During the current study the pharmacokinetics of PLE10-PEG20k-G-CSF and PLE20-PEG20k-G-CSF derivatives and their ability to induce granulopoiesis were, respectively, assessed in Sprague-Dawley rats and in C57BL6 mice. Our results show that the bioavailability and bioactivity of the derivatives are comparable to or better than those of PEG20k-Nter-G-CSF (commercially known as Pegfilgrastim). The therapeutic effects of PLE10-PEG20k-hGH and PLE20-PEG20k-hGH derivatives tested in hypophysectomized rats demonstrate that the presence of a negatively charged PLE block enhances the biological properties of the conjugates additionally with respect to PEG20k-Nter-hGH.
... Salmon calcitonin has a poweful anti-resorptive effect by direct binding to osteoclasts [7] . chondrocytes in articular cartilage respond directly to calcitonin as they have calcitonin receptors on their surfaces [8] . ...
... Comparable in vitro and in vivo activity to the free peptide was observed when Mero et al. conjugated HA-aldehyde (200 kDa HA; 7.3 mol% modification) to the N-terminal amine of salmon calcitonin (sCT) (28% w/w sCT; 3.6 sCT:1 HA molar ratio) for the treatment of osteoarthritis [55]. The combination of HA with sCT is advantageous because HA's ability to reduce matrix metalloproteinase (MMP)-1, MMP-3, and MMP-13 expression in osteoarthritic subchondral bone may complement salmon calcitonin's inhibitory effect on bone turnover and cartilage degradation. ...
Chapter
Since the 1970s, polysaccharides such as dextran, dextrin, hyaluronic acid, hydroxyethyl starch and polysialic acid have been investigated as carriers for therapeutic protein and peptide drugs in a wide range of clinical applications. Polysaccharides possess many favorable features for the targeted delivery of protein and peptide drugs since they are biodegradable, hydrophilic and non-toxic. Given the reported potential of non-biodegradable polymers, such as polyethylene glycol (PEG), to induce lysosomal storage disease and induce antibody formation, the use of polysaccharides for the delivery of therapeutic proteins and peptides has become increasingly attractive. This Chapter presents a historical overview of the field and explores the progress that has been made to date, using examples of conjugates from the literature. The available techniques to activate polysaccharides and purify and characterize polysaccharide-protein conjugates are reviewed. The current status, future possibilities and barriers to the clinical translation of polysaccharide-protein conjugates are also discussed. Book chapter available here: https://www.sciencedirect.com/science/article/pii/B978044464081900019X depending on your credentials. Available on request.
... Several HA-drug conjugates have been tested for cancer treatment, with payloads including butyric acid, PTX, SN38, doxorubicin, cisplatin, 5-fluorouracil, and methotrexate (Campisi and Renier, 2011;Coradini et al., 2004;Dong et al., 2013;Lee et al., 2012;Oommen et al., 2014;Tripodo et al., 2015;Venable et al., 2012;Yang et al., 2014). HA has also been conjugated with several anti-inflammatory drugs for intra-articular arthritis treatment, such as dexamethasone, hydrocortisone, fluorocortisone, betamethasone, corticosterone, prednisone, and prednisolone (Pouyani and Prestwich, 1994) and with peptides, including epidermal growth factor, salmon calcitonin, Exendin, anti-FLT1 peptide (Kong et al., 2010;Mero et al., 2014) and proteins, such as interferon alpha, hGH, insulin, ovalbumin, and antibodies (Ferguson et al., 2010;Friedrich et al., 2014;Lee et al., 2012;Mero et al., 2013Mero et al., , 2015Montagner et al., 2016). ...
Chapter
The targeted delivery of bioactive molecules to the appropriate site of action, one of the critical focuses of pharmaceutical research, improves therapeutic outcomes and increases safety at the same time; a concept envisaged by Ehrlich over 100 years ago when he described the “magic bullet” model. In the following decades, a considerable amount of research effort combined with enormous investment has carried selective drug targeting into clinical practice via the advent of monoclonal antibodies (mAbs) and antibody-drug conjugates derivatives. Additionally, a deeper understanding of physiopathological conditions of disease has permitted the tailored design of targeted drug delivery platforms that carry drugs, many copies of the same drug, and different drugs in combination to the appropriate site of action least selectively or preferentially. The acquired know-how has provided the field with the design rationale to develop a successful delivery system that will provide new and improved means to treat many intractable diseases and disorders. In this review, we discuss a wide range of molecular platforms for drug delivery, and focus on those with more success in the clinic, given their potential for targeted therapies.
... Due to the importance of MMPs and TIMPs in cartilage physiology, and because they are an evaluation factor in OA, the dosage of MMP-3, MMP-13, TIMP-1, and TIMP-3, which are the main ones involved in OA, was measured. 28,29 The data found were consistent with the literature on the subject 18,29 : MMP-3 and MMP-13 presented increased staining values in the P group, while TIMP-1 and TIMP-3 values decreased in relation to the other groups, demonstrating an increased catabolic activity, i.e., an ongoing cartilaginous destruction. In the S (Synvisc ® ) and PR (Polireumin ® ) groups, a decrease in the specific staining of MMP-3 and MMP-13 and an increase in TIMP-1 and TIMP-3 was observed, suggesting a lower cartilaginous catabolic action. ...
Article
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Objective To analyze, from the immunohistochemical perspective, the effects of hyaluronic acid of different molecular weights in an experimental model of osteoarthritis in rabbits. Methods Forty-four male California rabbits were randomly assigned to three different groups (PR, S, and P) and submitted to the resection of the anterior cruciate ligament of the right knee. Three weeks after the surgical procedure, three intra-articular weekly injections were carried out with low-molecular-weight native hyaluronic acid (Hyalgan®) to PR group, high molecular weight branched chain hyaluronic acid (Synvisc®) to group S, and saline solution 0.9% to group P. All animals were sacrificed 12 weeks after the surgical procedure, and the tibial plateaus of the infiltrated knees were then dissected. Histological sections of cartilage from the tibial plateau support areas were stained with immunohistochemical markers in order to investigate the amount of metalloproteases (MMPs 3 and 13) and their inhibitors (TIMPs 1 and 3). The staining intensity was quantified on a Zeiss Imager.Z2 Metasystems microscope and analyzed by Metafer4 Msearch software. Results The chondroprotective effect of the hyaluronic acids used in the study was demonstrated when compared to the control group. However, the comparison between them presented no significant statistical difference regarding chondroprotection. Conclusion The injection of saline solution demonstrated signs of OA development, while adding native hyaluronic acid of low molecular weight (Hyalgan®) and hyaluronic acid of high molecular weight (Synvisc®) protected the articular cartilage in this model of OA.
... Thus, a similar therapeutic effect can be achieved with a reduced dose compared to the systemic route [17]. IA injection of intermediate molecular weight hyaluronic acid or glucocorticosteroids has been exploited to relieve knee pain associated with OA [18][19][20]. Despite the promising advantages of IA injection, challenges do exist that limit the clinical application. ...
Article
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Tetramethylpyrazine (TMP) is a traditional Chinese herbal medicine with strong anti-inflammatory and cartilage protection activities, and thus a promising candidate for treating osteoarthritis. However, TMP is rapidly cleared from the joint cavity after intra-articular injection and requires multiple injections to maintain efficacy. The aim of this study was to encapsulate TMP into poly (lactic-co-glycolic acid) (PLGA) microspheres to enhance the TMP retention in the joint, reducing injection frequencies and decreasing dosage. TMP microspheres were prepared by emulsion/solvent evaporation method. The intra-articular retention of the drug was assessed by detecting the drug concentration distributed in the joint tissue at different time points. The therapeutic effect of TMP microspheres was evaluated by the swelling of knee joints and histological analysis in papain-induced OA rat model. The prepared freeze-dried microspheres with a particle size of about 10 µm can effectively prolong the retention time of the drug in the articular cavity to 30 d, which is 4.7 times that of the TMP solution. Intra-articular injection of TMP microspheres efficiently relieved inflammatory symptoms, improved joint lesions and decreased the depletion of proteoglycan. In conclusion, Intra-articular injection of TMP loaded microspheres was a promising therapeutic method in the treatment of OA.
... [32][33][34] In animal ACLT models, the greatest OA severity occurs in the MFC and LFC. [35][36][37][38][39] In our study, we transected only the cranial cruciate ligament instead of removing both ligaments to avoid instability and minimize the risk of disease. In the previously study by Lippiello et al., both cruciate ligaments and medial meniscus were transected, which could have led to instability and rapid passage of disease. ...
Article
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Aim: The aim was to study whether oral glucosamine hydrochloride (GlcN.HCl) or mucopolysaccharide protein (MucoP) has a structure-modifying effect on an anterior cruciate ligament transection (ACLT) rabbit model of osteoarthritis (OA). Methods: OA was surgically induced in the right knees of rabbits by transection of the ACLT. The left knees served as a sham-operated control. The animals were divided into four groups (n = 6 each): negative control (phosphate buffered saline, orally), positive control (oral celecoxib 10 mg/kg body weight/day), GlcN.HCl (oral 100 mg/kg/day) and MucoP (oral 100 mg/kg/day). Experimental animals were sacrificed after 8 weeks of treatment and the distal femur was removed for macroscopic examination, histological assessment, and terminal deoxynucleotidyl transferase-mediated nick-end labeling (TUNEL) assay of the OA rabbits. Results: On gross morphology, severe lesions were observed in articular cartilage in the negative control group. In the GlcN.HCl and MucoP treatment groups, fibrillations and cartilaginous lesions were significantly (P < 0.05) decreased compared to the negative control group. In particular, degenerative changes in cartilage and chondrocyte cellularity were significantly reduced (P < 0.05) in the positive control (celecoxib) group, GlcN.HCl treatment group and MucoP treatment group compared with the negative control group. TUNEL assay showed that apoptotic chondrocytes were significantly suppressed in the celecoxib group. Similar significant (P < 0.05) results were seen in the GlcN.HCl group and MucoP group but apoptosis of chondrocytes were high in the negative control group. Conclusion: These data suggest that the protective effects of GlcN.HCl and MucoP may play a useful role in the clinical treatment of OA.
... Devido à importância das MMP e das TIMP na fisiologia da cartilagem e por serem um fator de avaliação na osteoartrite, a dosagem das MMP-3 e MMP-13, TIMP-1 e TIMP-3, as principais envolvidas na osteoartrite, foi mensurada. 28,29 Os dados encontrados obtiveram conformidade com a literatura específica do assunto, 18 30 em um estudo feito em coelhos, concluíram que os ácidos hialurônicos de baixo peso molecular foram superiores aos de maior peso molecular. Não se encontrou nesse estudo a dose usada nas infiltraç ões para melhor comparação com o presente estudo. ...
Article
Full-text available
Analisar do ponto de vista imuno‐histoquímico os efeitos do ácido hialurônico de diferentes pesos moleculares em modelo experimental em coelhos.
... Interleukin-4 decreases nitric oxide production by chondrocytes and ameliorates subsequent destruction of cartilage in a rat OA model [4]. A hyaluronic acid-salmon calcitonin conjugate showed a chondro-protective effect in the local treatment of OA in a rabbit model of early OA [5]. Intra-articular injection of N-[N-(3,5-diflurophenylacetate)-L-alanyl]-(S)-phenylglycine t-butyl ester (DAPT), a small compound Notch inhibitor, in a surgical mouse OA model caused suppression of cartilage degradation [6]. ...
Article
Full-text available
Hyaluronic acid (HA) is used clinically to treat osteoarthritis (OA), but its pharmacological effects under hypoxic conditions remain unclear. Articular chondrocytes in patients with OA are exposed to a hypoxic environment. This study investigated whether hypoxia could potentiate the anabolic effects of exogenous HA in rat articular cartilage and whether these mechanisms involved HA receptors. HA under hypoxic conditions significantly enhanced the expression of extracellular matrix genes and proteins in explant culture, as shown by real-time reverse transcription-polymerase chain reaction (RT-PCR), Western blotting, and dimethylmethylene blue (DMMB) assays. Staining with Safranin-O and immunohistochemical staining with antibody to type II collagen were also enhanced in pellet culture. The expression of CD44 was increased by hypoxia and significantly suppressed by transfection with siRNAs targeting hypoxia-inducible factor 1 alpha (siHIF-1α). These findings indicate that hypoxia potentiates the anabolic effects of exogenous HA by a mechanism in which HIF-1α positively regulates the expression of CD44, enhancing the binding affinity for exogenous HA. The anabolic effects of exogenous HA may increase as OA progresses.
... Osteoarthritis (OA) is the most common and disability joint disease [1,2]. Treatments of OA are divided into non-surgical interventions for patients in early and moderate stages and surgery interventions for patients in advanced stages according to the severity of OA [3][4][5]. OA therapy was mainly to reduce pain and increase life quality of the OA patient [6]. Oral non-steroidal antiinflammatory drugs (NSAIDs), the most efficacious and widely used treatments for knee OA, have shown some risks of systemic adverse events including gastrointestinal or cardiovascular abnormalities [7]. ...
Article
Full-text available
Gold nanoparticles (AuNPs) have been widely used in biomedical science including antiarthritic agents, drug loading, and photothermal therapy. In this report, we studied the effects of AuNPs with diameters of 3, 13, and 45 nm, respectively, on rabbit articular chondrocytes. AuNPs were capped with citrate and their diameter and zeta potential were measured by dynamic light scattering (DLS). Cell viability was evaluated by Cell Counting Kit-8 (CCK-8) assay after the rabbit articular chondrocytes were pre-incubated with 3, 13, and 45 nm AuNPs, respectively, for 24 h. Flow cytometry (FCM) analysis with annexin V/propidium iodide (PI) double staining and fluorescence imaging with Hoechst 33258 staining were used to determine the fashion of AuNPs-induced chondrocyte death. Further, 13 nm AuNPs (2 nM) significantly induced chondrocyte death accompanying apoptotic characteristics including mitochondrial damage, externalization of phosphatidylserine and nuclear concentration. However, 3 nm AuNPs (2 nM) and 45 nm (0.02 nM) AuNPs did not induce cytotoxicity in chondrocytes. Although 13 nm AuNPs (2 nM) increased the intracellular reactive oxygen species (ROS) level, pretreatment with Nacetyl cysteine (NAC), a ROS scavenger, did not prevent the cytotoxicity induced by 13 nm AuNPs, indicating that 13 nm AuNPs (2 nM) induced ROS-independent apoptosis in chondrocytes. These results demonstrate the size-dependent cytotoxicity of AuNPs in chondrocytes, which must be seriously considered when using AuNPs for treatment of osteoarthritis (OA).
... Regarding the effect of other neuropeptides on ADAMTS and consequently on ECM maintenance, data are scarce, even more in OA pathology. Only one study described that calcitonin decreases ADAMTS-4 expression in a mouse model of OA [55], having a chondroprotective effect in a rabbit model of early OA [56]. We have demonstrated that VIP and CRF are able to decrease ADAMTS expression and function in OA-SF. ...
Article
Full-text available
ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) family is known to play an important role in the pathogenesis of osteoarthritis (OA), working on aggrecan degradation or altering the integrity of extracellular matrix (ECM). Thus, the main purpose of our study was to define the role of vasoactive intestinal peptide (VIP) and corticotrophin-releasing factor (CRF), as immunoregulatory neuropeptides, on ADAMTS production in synovial fibroblasts (SF) from OA patients and healthy donors (HD). OA- and HD-SF were stimulated with pro-inflammatory mediators and treated with VIP or CRF. Both neuropeptides decreased ADAMTS-4, -5, -7 and -12 expressions, aggrecanase activity, glycosaminoglycans (GAG), and cartilage oligomeric matrix protein (COMP) degradation after stimulation with fibronectin fragments (Fn-fs) in OA-SF. After stimulation with interleukin-1β, VIP reduced ADAMTS-4 and -5, and both neuropeptides decreased ADAMTS-7 production and COMP degradation. Moreover, VIP and CRF reduced Runx2 and β-catenin activation in OA-SF. Our data suggest that the role of VIP and CRF on ADAMTS expression and cartilage degradation could be related to the OA pathology since scarce effects were produced in HD-SF. In addition, their effects might be greater when a degradation loop has been established, given that they were higher after stimulation with Fn-fs. Our results point to novel OA therapies based on the use of neuropeptides, since VIP and CRF are able to stop the first critical step, the loss of cartilage aggrecan and the ECM destabilization during joint degradation.
... Articular chondrocytes express cal- citonin receptors, and calcitonin has been shown to reduce carti- lage degradation [191]. A local delivery approach was studied in a rabbit osteoarthritis model by injecting calcitonin conjugated to hyaluronic acid into a knee joint [192]. The conjugation to hyal- uronic acid successfully reduced the clearance and prolonged the protective effect of calcitonin in the joint. ...
Conference Paper
Decreased bone strength and healing capacity make treatment of osteoporotic bone challenging. We aim to use a thermoreversible injectable hydrogel for the delivery of osteogenic drugs to stimulate bone healing locally and to increase implant stability in osteoporotic bone. Thermoreversible semi-synthetic hyaluronan-poly(N-isopropyl acrylamide) (HA-PNIPAM) with 15% w/v composition was designed to be a flowing liquid at room temperature and a stable physical gel when exposed to body temperature. The viscoelastic properties, gelling temperature and % of PNIPAM grafting onto HA dimers were characterized by rheology and 1H NMR spectroscopy. The in vitro release studies of osteogenic factors, bone morphogenetic protein-2 (BMP-2), strontium ranelate and icariin showed a high retention of the drugs in the gel, with a release of 10-35% within 14 days. Cell culture studies with human mesenchymal stem cells (hMSCs) were conducted to define the effective concentrations of BMP-2, strontium ranelate and icariin to achieve osteogenic differentiation. Cell culture studies also confirmed the bioactivity of BMP-2 released from the hydrogel. Animal studies using the HA-PNIPAM hydrogel around a screw in osteoporotic rat tibia showed no adverse reaction to bone turnover as assessed by a longitudinal in vivo microCT. Further animal studies have been planned to test HA-PNIPAM gel for drug delivery in the same model. To conclude, HA-PNIPAM gel is a versatile release system for the study of osteogenic factors in vitro and presents no adverse effects on bone turnover in vivo, and is therefore a promising drug carrier to study influence of osteogenic factors in osteoporotic model.
... In settings of arthritis, for instance, following a therapy with a biologic and methotrexate to keep under control the florid phase of the disease [74], a resolutionbased therapy could be applied for maintenance and promotion of tissue (synovia, cartilage) repair. Indeed, experimental work demonstrates that proresolving mediators are able to 'revert' or prevent cartilage damage [75,76]. Interesting tissue-reparative properties have been reported for AnxA1 [38], in analogy to recent studies on tissue regeneration with maresins [77]. ...
Article
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Current medicines for the clinical management of inflammatory diseases act by inhibiting specific enzymes or antagonising specific receptors or blocking their ligands. In the past decade, a new paradigm in our understanding of the inflammatory process has emerged with the appreciation of genetic, molecular, and cellular mechanisms that are engaged to actively resolve inflammation. The 'resolution of acute inflammation' is enabled by counter-regulatory checkpoints to terminate the inflammatory reaction, promoting healing and repair. It may be possible to harness this knowledge for innovative approaches to the treatment of inflammatory pathologies. Here we discuss current translational attempts to develop agonists at proresolving targets as a strategy to rectify chronic inflammatory status. We reason this new approach will lead to the identification of better drugs that will establish a new branch of pharmacology, 'resolution pharmacology'.
... The available treatments for OA predominantly target symptomatic relief without addressing the fundamental mechanism underlying OA, which is the destruction of articular cartilage [3]. More studies are identifying and characterizing potential disease-modifying therapeutic agents for OA, including calcitonin [4], bone morphogenetic protein-7 [5], lubricin [6], trichostatin A [7] and mesenchymal stem cells [8]. ...
Article
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Background: Osteoarthritis (OA) is likely to become an increasing burden in the coming decades. Various agents have been developed to slow the progression of OA, and are collectively known as 'disease-modifying drugs', however, there is still little reliable evidence that such agents will be successful. Dehydroepiandrosterone (DHEA), a sex hormone precursor, has been recently proven as protective agent against OA, but the exact mechanism is still unkown. In the current study, the effects of weekly intra-articular injections of DHEA in preventing the progression of existing cartilage degeneration in an OA rabbit model were evaluated. The aim of the current study is to demonstrate the feature of its disease-modifying efficacy during OA progression. Methods: Thirty male New Zealand white rabbits were used in this study. An anterior cruciate ligament transection (ACLT) model was used to create a progressive OA model in twenty rabbits. The animals were treated with DHEA or a placebo and were necropsied at 9 and 16 weeks. Ten rabbits receiving sham operations served as controls. The articular cartilage of the medial femoral condyle (MFC), lateral femoral condyle (LFC), medial tibial plateau (MTP) and lateral tibial plateau (LTP) was evaluated macroscopically and histologically. Results: In the joints of the sham-operated rabbits, few histological changes were detected on the articular surfaces of the femoral condyles and tibial plateaus. ACLT obviously induced erosive changes on the cartilage surfaces. Compared to the placebo group, the macroscopic and Mankin score analyses demonstrated that the DHEA treatment markedly reduced the cartilage lesions and delayed cartilage degeneration in the four regions of the knee at 9 weeks after operation (macroscopic score: MFC P = 0.013; LFC P = 0.048; MTP P = 0.045; LTP P = 0.02, Mankin score: MFC P = 0.012; LFC P = 0.034; MTP P = 0.016; LTP P = 0.002). At 16 weeks, DHEA demonstrated chondroprotective effects on the lateral compartment of the knee compared to the placebo group, whereas the cartilage degeneration at the medial compartment of the knee did not differ among the groups (macroscopic score: LFC P = 0.046; LTP = 0.034, Mankin score: LFC P = 0.005; LTP P = 0.002). Conclusion: The disease-modifying efficacy of DHEA aganist OA is time-specific and site-dependent. DHEA could be used as a disease-modifying strategy to limit the progression of OA, especially in the middle stage.
... Mediators Potential therapeutics a Autophagy and cell survival mTOR [85,98]; AKT/Fox03/mTOR [99]; PPARg [87,178,179] Rapamycin, polyamines, v-6 polyunsaturated fatty acids; glucosamine CXCR2 [180 ] Chemokine antagonists or blocking antibodies Oral or intraarticular calcitonin [189][190][191] NFATc2/c2 [192] Calcitonin [193] Circadian clock Bmal1 [194 ] REV-ERB agonists [195] allow us to define the different regulatory requirements for stress-related phenotypes of OA chondrocytes [156]. However, although therapies that target DNA or histone methylation or miRNAs have been considered for cancer, their use in a non-life-threatening disease such as OA will require better understanding of gene and target cell specificities to avoid cytotoxicity and off-target effects. ...
Article
Osteoarthritis (OA) is a destructive joint disease in which the initiation may be attributed to direct injury and mechanical disruption of joint tissues, but the progressive changes are dependent on active cell-mediated processes that can be observed or inferred during the generally long time-course of the disease. Based on clinical observations and experimental studies, it is now recognized a that it is possible for individual patients to exhibit common sets of symptoms and structural abnormalities due to distinct pathophysiological pathways that act independently or in combination. Recent research that has focused on the underlying mechanisms involving biochemical cross talk among the cartilage, synovium, bone, and other joint tissues within a background of poorly characterized genetic factors will be addressed in this review. Copyright © 2015. Published by Elsevier Ltd.
... Articular chondrocytes express calcitonin receptors, and calcitonin has been shown to reduce cartilage degradation [191]. A local delivery approach was studied in a rabbit osteoarthritis model by injecting calcitonin conjugated to hyaluronic acid into a knee joint [192]. The conjugation to hyaluronic acid successfully reduced the clearance and prolonged the protective effect of calcitonin in the joint. ...
Article
Fragility fractures can cause significant morbidity and mortality in patients with osteoporosis and inflict considerable medical and socioeconomic burden. Moreover, treatment of an osteoporotic fracture is challenging due to the decreased strength of the surrounding bone and suboptimal healing capacity, predisposing both to fixation failure and non-union. Whereas a systemic osteoporosis treatment acts slowly, local release of osteogenic agents in osteoporotic fracture would act rapidly to increase bone strength and quality as well as to reduce the bone healing period and prevent development of a problematic non-union. The identification of agents with potential to stimulate bone formation and improve implant fixation strength in osteoporotic bone has raised hope for the fast augmentation of osteoporotic fractures. Stimulation of bone formation by local delivery of growth factors is an approach already in clinical use for the treatment of non-unions, and could be utilized for osteoporotic fractures as well. Small molecules have also gained ground as stable and inexpensive compounds to enhance bone formation and tackle osteoporosis. The aim of this paper is to present the state of the art on local drug delivery in osteoporotic fractures. Advantages, disadvantages and underlying molecular mechanisms of different active species for local bone healing in osteoporotic bone are discussed. This review also identifies promising new candidate molecules and innovative approaches for the local drug delivery in osteoporotic bone.
Article
Intraarticular (IA) injections provide the opportunity to deliver biologics directly to their site of action for a local and efficient treatment of osteoarthritis. However, the synovial joint is a challenging site of administration since the drug is rapidly eliminated across the synovial membrane and has limited distribution into cartilage, resulting in unsatisfactory therapeutic efficacy. In order to rationally develop appropriate drug delivery systems, it is essential to thoroughly understand the unique biopharmaceutical environments and kinetics in the joint to adequately simulate them in relevant experimental models. This review presents a detailed view on articular kinetics and drug-tissue interplay of IA administered drugs and summarizes how these can be translated into reasonable formulation strategies by identification of key factors through which the joint residence time can be prolonged and specific structures can be targeted. In this way, pros and cons of the delivery approaches for biologics will be evaluated and the extent to which biorelevant models are applicable to gain mechanistic insights and ameliorate formulation design is discussed.
Article
The repair of cartilage lesions faces challenges in the integration of biological and mechanical cues into scaffolds. Herein, injectable hyaluronic acid (HA) hydrogels were developed with inoculation of bone marrow mesenchymal stem cell (MSC) spheroids and short fiber fillers. Kartogenin (KGN)-loaded short fibers (Fk) are assembled into cell spheroids (Fk-CS) as cell growth substrates and depots of cell differentiation modulators. Celecoxib (CXB)-loaded short fibers (FC) in HA hydrogels (HA-FC) benefit mechanical reinforcement and sustained release of anti-inflammatory agent. After inoculation in hydrogels, Fk-CS up-regulate differentiation-related gene expressions (aggrecan, collagen II, and Sox9) and the sustained release of KGN promotes chondrogenic differentiation and extracellular matrix (ECM) production in the entire spheroids. Osteochondral defects and osteoarthritis model are surgically created and treated with intraarticular injection of Fk-CS-loaded hydrogels. Osteochondral defects are fully repaired with integrated surrounding tissues, and the chondrocyte morphology, ECM (sulfated glycosaminoglycane and collagen II) deposition, and compressive moduli are comparable to those of normal cartilage. Osteoarthritis-derived cartilage lesions are repaired by regenerated ECMs, and osteoarthritis progression is retarded by downregulating pro-inflammatory cytokines and proteinase expressions. Thus, hierarchically structured injectable hydrogels and cell spheroids provide biochemical and mechanical cues for MSC differentiation, ECM deposition, cartilage regeneration, and inflammation removal in the treatment of osteochondral defects and osteoarthritis.
Article
Bone is a connective tissue that support the entire body and protect the internal organs. However, there are great challenges on curing intractable skeletal diseases such as hypercalcemia, osteoporosis and osteoarthritis. To address these issues, calcitonin (CT) therapy is an effective treatment alternative to regulate calcium metabolism and suppress inflammation response, which are closely related to skeletal diseases. Traditional calcitonin formulation requires frequent administration due to the low bioavailability resulting from the short half-life and abundant calcitonin receptors distributed through the whole body. Therefore, long-term and targeted calcitonin delivery systems (LCDS and TCDS) have been widely explored as the popular strategies to overcome the intrinsic limitations of calcitonin and improve the functions of calcium management and inflammation inhibition in recent years. In this review, we first explain the physiological effects of calcitonin on bone remodeling: (i) inhibitory effects on osteoclasts and (ii) facilitated effects on osteoblasts. Then we summarized four strategies for spatiotemporally controlled delivery of calcitonin: micro−/nanomedicine (e.g. inorganic micro−/nanomedicine, polymeric micro−/nanomedicine and supramolecular assemblies), hydrogels (especially thermosensitive hydrogels), prodrug (PEGylation and targeting design) and hybrid biomaterials. Subsequently, we discussed the application of LCDS and TCDS in treating hypercalcemia, osteoporosis, and arthritis. Understanding and analyzing these advanced calcitonin delivery applications are essential for future development of calcitonin therapies toward skeletal diseases with superior efficacy in clinic.
Article
Serious injuries of the endometrium (the innermost lining layer of the uterus) or naturally thin endometrium can cause uterine dysfunction and subsequently predispose patients to infertility or miscarriage. In this context, many stem cell researchers have devoted effort to the regeneration of injured or thin endometrium by administering various types of stem cells. Despite some promising initial results, regenerating the endometrium with stem cells is very challenging, as the lack of a 3D microenvironment for the transplanted stem cells leads to weak therapeutic effects. Therefore, as an alternative, we developed a novel endometrial stem cell-laden 3D artificial endometrium by combining various endometrial cellular components and natural biodegradable polymers to mimic the multilayered endometrial structure and its microenvironment. The artificial endometrium constitutes a triple-layered structure that recapitulates the structural and physiological features of the human endometrium. The cell viability and biological characteristics of various types of encapsulated endometrial cells are well maintained within the artificial endometrium, which exhibits similar behavior to the human endometrium by properly responding to steroid hormones and actively secreting various growth factors. Remarkably, severe degenerative changes were significantly relieved by artificial endometrium transplantation. More importantly, successful pregnancy and subsequent successful live birth without any phenotypic or genetic abnormalities can be achieved by transplanting our artificial endometrium into endometrial ablation mice.
Article
Elderly patients with knee osteoarthritis (OA) are often troubled with joint soreness, swelling, weakness, and pain. Knee intra-articular (IA) injection of autologous platelet rich plasma (PRP) is one of the options that can be used in treating knee OA. However, there are doubts on the effect of PRP when used in treating elderly patients with moderate to severe degrees of knee OA. In this study, 47 elderly patients with moderate degree of knee OA combined with supra-patellar bursitis were recruited. Musculoskeletal ultrasound was used for accurate needle placement for the aspiration of bursa synovial fluid SF followed by PRP injections. In group 1, 23 patients received conventional liquid-form IA PRP injections. In group 2, 24 patients received IA injections of thermal oscillation prepared PRP injectants. It was discovered that when the liquid-form PRP was heated up to a temperature of 75 °C and simultaneously oscillated under 200 revolutions per minute (rpm) for 15 min, the end product became paste-form like and with a viscosity similar to that of an anti-cough syrup. Under incubation, the highest number of platelet-derived growth factor (PDGF) was obtained on the 8th day. At one month after the completion of PRP injections, group 2 patients revealed significant decreases in SF total protein concentrations, SF volumes, and Lequesne index values. Proteins associated with inflammation, such as apolipoprotein A-I, haptoglobin, immunoglobulin kappa chain, transferrin, and matrix metalloproteinase also decreased significantly. Therefore, the thermal oscillation preparation method can augment the effectiveness of autologous PRP in treating elderly patients with moderate knee OA. It may be recommended that the PRP injectant should be prepared first using the thermal oscillation method to increase its viscosity, enabling prolonged release of growth factors once it is injected into the knee joint.
Article
Osteoarthritis (OA) is a degenerative disease of the joints and a leading cause of physical disability in adults. Intra‐articular (IA) therapy is a popular treatment strategy for localized, single‐joint OA; however, small‐molecule drugs such as corticosteroids do not provide prolonged relief. One possible reason for their lack of efficacy is high clearance rates from the joint through constant lymphatic drainage of the synovial tissues and synovial fluid and also by their exchange via the synovial vasculature. Advanced drug delivery strategies for extended release of therapeutic agents in the joint space is a promising approach to improve outcomes for OA patients. Broadly, the basic principle behind this strategy is to encapsulate therapeutic agents in a polymeric drug delivery system (DDS) for diffusion‐ and/or degradation‐controlled release, whereby degradation can occur by hydrolysis or tied to relevant microenvironmental cues such as pH, reactive oxygen species (ROS), and protease activity. In this review, the development of clinically tested IA therapies for OA and recent systems which have been investigated preclinically are highlighted. DDS strategies including hydrogels, liposomes, polymeric microparticles (MPs) and nanoparticles (NPs), drug conjugates, and combination systems are introduced and evaluated for clinical translational potential.
Article
Pharmacotherapy towards hypercalcemia treatment mainly caused by osteoporosis and bone tumor is an effective method to regulate in vivo calcium equilibrium. As a clinical therapeutic peptide, salmon calcitonin (sCT) is considered as a quick-acting medicine while limited by the short half-life. To address this challenge, we designed an injectable thermo-sensitive hydrogel based on hydroxypropyl chitin (HPCH) and incorporated complex of sCT and hyaluronic acid (HA) (sCT-HA) with high association efficiency up to 96.84 ± 7.25%. This composite hydrogel showed tunable biodegradable property. In vitro sCT release profiles revealed that this hydrogel can achieve long-term sustained sCT release (28 days) with considerable structure stability. The cell study illustrated outstanding compatibility and osteoconductive potential of this multi-component hydrogel according to the higher ALP activity (2.10-fold), calcium expression (2.30-fold) and deposition (1.10-fold) compared to that of sCT group. In vivo sCT release confirmed that this hydrogel system realized sustained sCT release and continuous hypocalcemic effect as long as 28 days and there were no inflammation and immune response according to the histological evaluations (H&E and IgG staining). These findings demonstrate this osteoconductive hydrogel system can provide a promising method for therapy of bone related disease.
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Despite major advances in the treatment of rheumatoid arthritis (RA) led by the success of biologic therapies, the lack of response to therapy in a proportion of patients, as well as therapy discontinuation owing to systemic toxicity, are still unsolved issues. Unchecked RA might develop into progressive structural joint damage, loss of function and long-term disability, disorders which are associated with a considerable health-economic burden. Therefore, new strategies are required to actively target and deliver therapeutic agents to disease sites in order to promote in situ activity and decrease systemic toxicity. Polymer-drug conjugates can improve the pharmacokinetics of therapeutic agents, conferring desirable properties such as increased solubility and tissue penetration at sites of active disease. Additionally, nanotechnology is an exciting modality in which drugs are encapsulated to protect them from degradation or early activation in the circulation, as well as to reduce systemic toxicity. Together with the targeting capacity of antibodies and site-specific peptides, these approaches will facilitate selective accumulation of therapeutic agents in the inflamed synovium, potentially improving drug efficacy at disease sites without affecting healthy tissues. This Review aims to summarize key developments in the past 5 years in polymer conjugation, nanoparticulate drug delivery and antibody or peptide-based targeting-strategies that might constitute the platform for the next generation of RA therapeutics.
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Prolonged inappropriate inflammatory responses contribute to the pathogenesis of rheumatoid arthritis (RA) and to aspects of osteoarthritis (OA). The orphan nuclear receptor, NR4A2, is a key regulator and potential biomarker for inflammation and represents a potentially valuable therapeutic target. Both salmon calcitonin (sCT) and hyaluronic acid (HA) attenuated activated mRNA expression of NR4A1, NR4A2, NR4A3, and matrix metalloproteinases (MMPs) 1, 3 and 13 in three human cell lines: SW1353 chondrocytes, U937 and THP-1 monocytes. Ad-mixtures of sCT and HA further down-regulated expression of NR4A2 compared to either agent alone at specific concentrations, hence the rationale for their formulation in nanocomplexes (NP) using chitosan. The sCT released from NP stimulated cAMP production in human T47D breast cancer cells expressing sCT receptors. When NP were injected by the intra-articular (I.A.) route to the mouse knee during on-going inflammatory arthritis of the K/BxN serum transfer model, joint inflammation was reduced together with NR4A2 expression, and local bone architecture was preserved. These data highlight remarkable anti-inflammatory effects of sCT and HA at the level of reducing NR4A2 mRNA expression in vitro. Combining them in NP elicits anti-arthritic effects in vivo following I. A. delivery.
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The interest in the relationship between articular cartilage and the structural and functional properties of peri-articular bone relates to the intimate contact that exists between these tissues in joints that are susceptible to the development of osteoarthritis (OA). The demonstration in several animal models that osteoporosis and decreased bone tissue modulus leads to an increased propensity for the development of post-traumatic OA is paradoxical in light of the extensive epidemiological literature indicating that individuals with high systemic bone mass, assessed by bone mineral density, are at increased risk for OA. These observations underscore the need for further studies to define the pathophysiological mechanisms involved in the interaction between subchondral bone and articular cartilage and for applying this information to the development of therapeutic interventions to improve the outcomes in patients with OA.
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The primary goal of this body of work is to suggest a standardized system for histopathological assessment of experimental surgical instability models of osteoarthritis (OA) in rabbits, building on past experience, to achieve comparability of studies from different centres. An additional objective is to review methodologies that have been employed in the past for assessing OA in rabbits with particular reference to the surgical anterior cruciate ligament transection (ACLT) model. A panel of scientists and clinician-scientists with recognized expertise in assessing rabbit models of OA reviewed the literature to provide a critical appraisal of the methods that have been employed to assess both macroscopic and microscopic changes occurring in rabbit joint tissues in experimental OA. In addition, a validation of the proposed histologic histochemical grading system was performed. The ACLT variant of the surgical instability model in skeletally mature rabbits is the variation most capable of reproducing the entire range of cartilage, synovial and bone lesions recognized to be associated with OA. These lesions can be semiquantitatively graded using macroscopic and microscopic techniques. Further, as well as cartilage lesions, this ACLT model can produce synovial and bone lesions similar to that of human OA. The ACLT variant of the surgical instability model in rabbits is a reproducible and effective model of OA. The cartilage lesions in this model and their response to therapy can be graded according to an adapted histological and histochemical grading system, though also this system is to some extent subjective and, thus, neither objective nor entirely reproducible.
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Osteoporosis (OP) increases cartilage damage in a combined rabbit model of OP and osteoarthritis (OA). Accordingly, we assessed whether microstructure impairment at subchondral bone aggravates cartilage damage in this experimental model. OP was induced in 20 female rabbits, by ovariectomy and intramuscular injections of methylprednisolone hemisuccinate for four weeks. Ten healthy animals were used as controls. At week 7, OA was surgically induced in left knees of all rabbits. At 22 weeks, after sacrifice, microstructure parameters were assessed by micro-computed tomography, and osteoprotegerin (OPG), receptor activator of nuclear factor-κB ligand (RANKL), alkaline phosphatase (ALP) and metalloproteinase 9 (MMP9) protein expressions were evaluated by Western Blot at subchondral bone. In addition, cartilage damage was estimated using the histopathological Mankin score. Mann-Whitney and Spearman statistical tests were performed as appropriate, using SPSS software v 11.0. Significant difference was established at P < 0.05. Subchondral bone area/tissue area, trabecular thickness and polar moment of inertia were diminished in OPOA knees compared with control or OA knees (P < 0.05). A decrease of plate thickness, ALP expression and OPG/RANKL ratio as well as an increased fractal dimension and MMP9 expression occurred at subchondral bone of OA, OP and OPOA knees vs. controls (P < 0.05). In addition, the severity of cartilage damage was increased in OPOA knees vs. controls (P < 0.05). Remarkably, good correlations were observed between structural and remodelling parameters at subchondral bone, and furthermore, between subchondral structural parameters and cartilage Mankin score. Microstructure impairment at subchondral bone associated with an increased remodelling aggravated cartilage damage in OA rabbits with previous OP. Our results suggest that an increased subchondral bone resorption may account for the exacerbation of cartilage damage when early OA and OP coexist simultaneously in same individuals.
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Calcitonin has been demonstrated to have chondroprotective effects under pre-clinical settings. It is debated whether this effect is mediated through subchondral-bone, directly on cartilage or both in combination. We investigated possible direct effects of salmon calcitonin on proteoglycans and collagen-type-II synthesis in osteoarthritic (OA) cartilage. Human OA cartilage explants were cultured with salmon calcitonin [100 pM-100 nM]. Direct effects of calcitonin on articular cartilage were evaluated by 1) measurement of proteoglycan synthesis by incorporation of radioactive labeled 35SO4 [5 microCi] 2) quantification of collagen-type-II formation by pro-peptides of collagen type II (PIINP) ELISA, 3) QPCR expression of the calcitonin receptor in OA chondrocytes using four individual primer pairs, 4) activation of the cAMP signaling pathway by EIA and, 5) investigations of metabolic activity by AlamarBlue. QPCR analysis and subsequent sequencing confirmed expression of the calcitonin receptor in human chondrocytes. All doses of salmon calcitonin significantly elevated cAMP levels (P < 0.01 and P < 0.001). Calcitonin significantly and concentration-dependently [100 pM-100 nM] induced proteoglycan synthesis measured by radioactive 35SO4 incorporation, with a 96% maximal induction at 10 nM (P < 0.001) corresponding to an 80% induction of 100 ng/ml IGF, (P < 0.05). In alignment with calcitonin treatments [100 pM-100 nM] resulted in 35% (P < 0.01) increased PIINP levels. Calcitonin treatment increased proteoglycan and collagen synthesis in human OA cartilage. In addition to its well-established effect on subchondral bone, calcitonin may prove beneficial to the management of joint diseases through direct effects on chondrocytes.
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Salmon calcitonin (sCT) was conjugated via its N-terminal cysteine to a comb-shaped end-functionalized poly(poly(ethylene glycol) methyl ether methacrylate) (PolyPEG, 6.5 kDa), and to linear PEG (5 kDa). Conjugate molecular weight and purity was assessed by SEC-HPLC and MALDI-TOF MS. Bioactivity of conjugates was measured by cyclic AMP assay in T47D cells. Calcium and calcitonin levels were measured in rats following intravenous injections. Stability of conjugates was tested against serine proteases, intestinal and liver homogenates and serum. Cytotoxicity of conjugates was assessed by lactate dehydrogenase (LDH) assay and by haemolytic assay of rat red blood cells. Results showed that the two conjugates were of high purity with molecular weights similar to predictions. Both conjugates retained more than 85% bioactivity in vitro and had nanomolar EC(50) values similar to sCT. While both sCT-PolyPEG(6.5 K) and sCT-PEG(5 K) were resistant to metabolism by serine proteases, homogenates and serum, PolyPEG (6.5 K) was more so. Although both conjugates reduced serum calcium to levels similar to those achieved with sCT, PolyPEG(6.5 K) extended the T(1/2) and AUC of serum sCT over values achieved with sCT-PEG and sCT itself. None of PolyPEG, PEG or methacrylic acid displayed significant cytotoxicity. PolyPEG may therefore have potential to improve pharmacokinetic profiles of injected peptides.
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The peptide hormone calcitonin is widely used therapeutically in the treatment of bone disorders such as Paget's disease, osteoporosis, and the hypercalcemia of some malignancies. However, emerging evidence suggests the actions of calcitonin via its G protein-coupled receptor, the calcitonin receptor, may not be limited to bone. Calcitonin receptors have also been identified in the central nervous system, testes, skeletal muscle, lymphocytes, and the placenta. We are now becoming aware that the complexity of the calcitonin response mediated by the calcitonin receptor can be influenced by accessory proteins, receptor isoforms, genetic polymorphisms, developmental and/or transcriptional regulation, feedback inhibition, and the specific cellular or tissue background. This article discusses what is known about the molecular and pharmacological actions of the calcitonin receptor and highlights areas of current research.
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Background/rationale: Introducing new or testing existing drugs in an attempt to modify the progress of osteoarthritis (OA) is of paramount importance. Questions/purposes: This study aims to determine the effect exerted by Calcitonin on the progress of early-stage osteoarthritic lesions. Methods: We used 18, skeletally mature, white, female, New Zealand rabbits. OA was operatively induced in the right knee of each animal by the complete dissection of the anterior cruciate ligament, complete medial meniscectomy and partial dissection of the medial collateral ligament. Postoperatively, animals were divided into two groups. Starting on the ninth postoperative day and daily thereafter, group A animals (n = 9) received 10 IU oculus dexter (o.d.) of synthetic Calcitonin IntraMuscularly (I.M.); group B animals (n = 9) received equal volume of saline o.d. Three animals from each group were sacrificed at 1, 2 and 3 months following treatment's initiation. The extent and the grade of OA were assessed macroscopically, histologically and by radiographs, Computed Tomography (CT) and Magnetic Resonance Imaging (MRI)-scans. The Osteoarthritis Research Society International (OARSI) score, incorporating histological and macroscopic information, was calculated for each knee. Results: Osteoarthritic changes in group A animals were less severe and progressed less rapidly when compared with those of group B animals (sham). This difference was statistically significant in the first and second month (P = 0.05), but not in the third month (P = 0.513). Conclusions: I.M. administration of Calcitonin seems to delay the progress of early-stage osteoarthritic lesions induced by mechanical instability in a rabbit experimental model.
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Polymer conjugation has been widely exploited to prolong half-life and reduce immunogenicity of therapeutic proteins. Here, the potentials of hyaluronic acid (HA) have been investigated by studying the conjugates with two model enzymes, trypsin and RNase A, and with insulin. As the direct coupling of proteins to the HA's carboxylic groups can cause cross-linking problems, a hyaluronan-aldehyde derivative has been synthesized for N-terminal site-selective conjugation. HA conjugation, termed HAylation, preserved the activities of enzymes and their thermal stabilities. Insulin HAylation was studied by preparing two conjugates with different peptide loadings (32% and 17%, w/w). Noticeably, the conjugate with the lower loading showed the greater effect on blood glucose level. The 17% HA-insulin conjugate showed a lowering effect on blood glucose level for up to 6h, while free insulin exhausted its action after 1h. This study highlights the potentials of hyaluronan-aldehyde for protein delivery.
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There has been a recent interest in calcitonin as a potential treatment for osteoarthritis, based on its metabolic activities in both bone turnover and cartilage. The aim of this study was to evaluate the effects of nasal form calcitonin on knee osteoarthritis and quality of life in women who receive calcitonin treatment for postmenopausal osteoporosis. Two hundred and twenty postmenopausal women, aged between 55 and 65 years with knee pain and knee osteoarthritis, graded II-III by using Kellgren-Lawrence radiographic scoring system, were included. Western Ontario and McMaster Universities (WOMAC) osteoarthritis index, the quality of life questionnaire of the European Foundation for Osteoporosis (QALEFFO-41) and visual analog scale were used for the algofunctional assessments. Need of rescue analgesic was recorded. Pain (P < 0.001), stiffness (P < 0.05), functional capability (P < 0.05) and total score of WOMAC (P < 0.05) revealed statistically significant improvements after 3 months of the treatment and remained consistent throughout 1 year of the treatment period. Participants experienced significant reductions in WOMAC perceptions of pain (-53 %), joint stiffness (-44 %) and limitations in physical function (-49 %) at the end of 1 year of calcitonin treatment. Need of rescue analgesic intake was reported to have decreased approximately by 60 % at the end of the 1-year treatment period. QUALEFFO_41 scores improved: 37.6 (baseline), 30.9 (3 months), 28.0 (6 months) and 24.4 (1 year). In conclusion, nasal calcitonin treatment provided dual action on osteoporosis and osteoarthritis with significant improvements in quality of life and algofunctional results in knee osteoarthritis.
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Calcitonin is well-known for its inhibitory actions on bone-resorbing osteoclasts and recently potential beneficial effects on cartilage were shown. We investigated effects of salmon calcitonin (sCT) on the articular cartilage and bone, after destabilization of the medial meniscus (DMM) in normal and sCT over-expressing mice. Bone phenotype of transgenic (TG) C57Bl/6 mice over-expressing sCT at 6 months and 12 months was investigated by (1) serum osteocalcin and urinary deoxypyridinoline and (2) dynamic and normal histomorphometry of vertebrae bodies. In subsequent evaluation of cartilage and subchondral bone changes, 44 10-week old TG or wild-type (WT) mice were randomized into four groups and subjected to DMM or sham-operations. After 7 weeks animals were sacrificed, and knee joints were isolated for histological analysis. Trabecular bone volume (BV/TV) increased 150% after 6 months and 300% after 12 months in sCT-expressing mice when compared to WT controls (P<0.05). Osteoblast number, bone formation rate and osteocalcin measurements were not affected in TG mice over-expressing sCT. In WT animals, a 5-fold increase in the quantitative erosion index was observed after DMM, and the semi-quantitative OARSI score showed over 400% (P<0.001) increase, compared to sham-operated WT mice. DMM-operated TG mice were protected against cartilage erosion and showed a 65% and 64% (P<0.001) reduction, respectively, for the two histopathological evaluation methods. sCT over-expressing mice had higher bone volume, and were protected against cartilage erosion. These data suggest that increased levels of sCT may hamper the pathogenesis of osteoarthritis (OA). However more studies are necessary to confirm these preliminary results.
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Modification of therapeutic proteins and peptides by polyethylene glycol (PEG) conjugation is a well-known approach to improve the pharmacological properties of drugs. Several chemical procedures of PEG coupling are already in use but an alternative method based on microbial transglutaminase (mTGase) was recently devised. The enzyme catalyzes the link of mPEG-NH(2) to glutamines (Gln) of a substrate protein. In this case the advantage resides in the fact that usually only few Gln(s) in a protein are substrate of mTGase. In order to further restrict the selectivity of the enzyme, we investigated a new approach leading to the formation of a single conjugate isomer as well as for those proteins containing two or more Gln(s) as mTGase substrates. It was found that the addition of co-solvents in the reaction mixture influenced both the secondary structure of the targeted protein and the mTGase activity. The enzymatic PEGylation under these conditions yielded only mono- and selectively modified conjugates. The method was investigated with salmon calcitonin (sCT) and human growth hormone (hGH). In the case of sCT we also demonstrated the importance of site-selective conjugation for the preservation of in vivo activity.
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Osteoarthritis (OA) is a prevalent and disabling condition for which few safe and effective therapeutic options are available. Current approaches are largely palliative and in an effort to mitigate the rising tide of increasing OA prevalence and disease impact, modifying the structural progression of OA has become a focus of drug development. This Review describes disease modification and discusses some of the challenges involved in the discovery and development of disease-modifying OA drugs (DMOADs). A variety of targeted agents are in mature phases of development; specific agents that are beyond preclinical development in phase II and III trials and show promise as potential DMOADs are discussed. A research agenda with respect to disease modification in OA is also provided, and some of the future challenges we face in this field are discussed.
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Subchondral bone is a candidate for treatment of osteoarthritis (OA). We investigated the effects of intra-articular injection of hyaluronan (IAI-HA) on subchondral bone in rabbit OA model. OA was induced by anterior cruciate ligament transection, with some rabbits receiving IAI-HA. OA was graded morphologically, and expression of mRNA was assessed by real-time RT-PCR. Tissue sections were stained with hyaluronan-binding protein, and penetration of fluorescent hyaluronan was assessed. The in vitro inhibitory effect of hyaluronan on MMP-13 was analyzed in human osteoarthritic subchondral bone osteoblasts (OA Ob) by real-time RT-PCR and ELISA. Binding of hyaluronan to OA Ob via CD44 was assessed by immunofluorescence cytochemistry. Expression of MMP-13 and IL-6 mRNA in cartilage and subchondral bone, and morphological OA grade, increased over time. IAI-HA ameliorated the OA grade and selectively suppressed MMP-13 mRNA in subchondral bone. IAI-HA enhanced the hyaluronan staining of subchondral bone marrow cells and osteocyte lacunae. Fluorescence was observed in the subchondral bone marrow space. In OA Ob, hyaluronan reduced the expression and production of MMP-13, and anti-CD44 antibody blocked hyaluronan binding to OA Ob. These findings indicate that regulation of MMP-13 in subchondral bone may be a critical mechanism during IAI-HA.
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Oral delivery of proteins has been hampered by an array of difficulties. However, promising novel oral delivery systems have been developed. 5-CNAC, formulated with the peptide salmon calcitonin, is in phase III clinical trials for the treatment of osteoporosis or osteoarthritis and could become the first marketed oral peptide. This article reviews key findings and implications from studies undertaken to date with this oral formulation. Findings include these: (1) the optimal calcitonin tablet dose is 0.8 mg; (2) 0.8 mg of oral calcitonin is rapidly absorbed, reaching maximum concentration in 15 to 30 minutes, and is eliminated from plasma with a short half-life-9 to 15 minutes; (3) the 0.8-mg tablet is more highly absorbed than the marketed nasal formulation, with biomarker levels indicating significantly greater efficacy in suppression of bone resorption; (4) drug absorption is increased with dosing at least 10 minutes before a meal rather than postprandially and also with 50 mL of water; (5) the optimal timing of dosing for osteoporosis therapy is in the evening to mitigate the circadian peak in bone resorption; and (6) the oral formulations of synthetic and recombinant calcitonin have similar pharmacokinetic and pharmacodynamic properties. These key findings may aid researchers in the development of other oral formulations.
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Following the introduction of mandatory Canadian folic acid flour fortification in mid-1997, the incidence of selected childhood cancers that declined in Ontario prior to and subsequent to this public policy initiative was examined. A population-based cohort study of all incident cases of childhood malignancy in Ontario between the years 1985 and 2006 was conducted. Participants were identified from a database provided by the Pediatric Oncology Group of Ontario and included children 0 to 4 years of age and 5 to 9 years of age who were diagnosed with cancer. Among children aged 0 to 4 years, the incidence rate of Wilms' tumor declined from 1.94 to 1.43 per 100,000 (incidence rate ratio 0.74, 95% confidence interval, 0.57-0.95). No significant change was seen in the prefortification vs postfortification time periods for acute lymphoblastic leukemia, brain cancers, or embryonal cancers among the 0- to 4-year or 5- to 9-year age groups. There was an approximately 30% reduction in risk of Wilms' tumor following introduction of the initiative. This corroborates a recent case-control study from Germany. These data may also provide some reassurance that universal flour fortification does not heighten the risk of pediatric cancer.
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The articular surface plays an essential role in load transfer across the joint, and conditions that produce increased load transfer or altered patterns of load distribution accelerate the development of osteoarthritis (OA). Current knowledge segregates the risk factors into two fundamental mechanisms related to the adverse effects of "abnormal" loading on normal cartilage or "normal" loading on abnormal cartilage. Although chondrocytes can modulate their functional state in response to loading, their capacity to repair and modify the surrounding extracellular matrix is limited in comparison to skeletal cells in bone. This differential adaptive capacity underlies the more rapid appearance of detectable skeletal changes, especially after acute injuries that alter joint mechanics. The imbalance in the adaptation of the cartilage and bone disrupts the physiological relationship between these tissues and further contributes to OA pathology. This review focuses on the specific articular cartilage and skeletal features of OA and the putative mechanisms involved in their pathogenesis.
Article
To update evidence for available therapies in the treatment of hip and knee osteoarthritis (OA) and to examine whether research evidence has changed from 31 January 2006 to 31 January 2009. A systematic literature search was undertaken using MEDLINE, EMBASE, CINAHL, AMED, Science Citation Index and the Cochrane Library. The quality of studies was assessed. Effect sizes (ESs) and numbers needed to treat were calculated for efficacy. Relative risks, hazard ratios (HRs) or odds ratios were estimated for side effects. Publication bias and heterogeneity were examined. Sensitivity analysis was undertaken to compare the evidence pooled in different years and different qualities. Cumulative meta-analysis was used to examine the stability of evidence. Sixty-four systematic reviews, 266 randomised controlled trials (RCTs) and 21 new economic evaluations (EEs) were published between 2006 and 2009. Of 51 treatment modalities, new data on efficacy have been published for more than half (26/39, 67%) of those for which research evidence was available in 2006. Among non-pharmacological therapies, ES for pain relief was unchanged for self-management, education, exercise and acupuncture. However, with new evidence the ES for pain relief for weight reduction reached statistical significance, increasing from 0.13 [95% confidence interval (CI) -0.12, 0.36] in 2006 to 0.20 (95% CI 0.00, 0.39) in 2009. By contrast, the ES for electromagnetic therapy which was large in 2006 (ES=0.77, 95% CI 0.36, 1.17) was no longer significant (ES=0.16, 95% CI -0.08, 0.39). Among pharmacological therapies, the cumulative evidence for the benefits and harms of oral and topical non-steroidal anti-inflammatory drugs, diacerhein and intra-articular (IA) corticosteroid was not greatly changed. The ES for pain relief with acetaminophen diminished numerically, but not significantly, from 0.21 (0.02, 0.41) to 0.14 (0.05, 0.22) and was no longer significant when analysis was restricted to high quality trials (ES=0.10, 95% CI -0.0, 0.23). New evidence for increased risks of hospitalisation due to perforation, peptic ulceration and bleeding with acetaminophen >3g/day have been published (HR=1.20, 95% CI 1.03, 1.40). ES for pain relief from IA hyaluronic acid, glucosamine sulphate, chondroitin sulphate and avocado soybean unsponifiables also diminished and there was greater heterogeneity of outcomes and more evidence of publication bias. Among surgical treatments further negative RCTs of lavage/debridement were published and the pooled results demonstrated that benefits from this modality of therapy were no greater than those obtained from placebo. Publication of a large amount of new research evidence has resulted in changes in the calculated risk-benefit ratio for some treatments for OA. Regular updating of research evidence can help to guide best clinical practice.
Article
The aim of this study was to investigate the pharmacokinetic and pharmacodynamic parameters of oral salmon calcitonin (oSCT) administered over 14 days to men and women presenting with osteoarthritis (OA). The study was a phase-I, 2-week, placebo-controlled, double-blind, double-dummy, randomized, gender-stratified study including 73 subjects aged 57-75 years. Patients had painful OA with a Kellgren and Lawrence index score of I-III. Treatment allocations were; 0.6 mg, 0.8 mg of oSCT, or placebo. Treatment was given twice daily for 14 days. The morning dose was administered between 07:00 and 08:00 at least 30 min before breakfast. The second dose was administered 30 min before evening dinner. On treatment day 1 and 14, the morning dose was followed by 5h of fasting, and blood samples and urine were collected immediately prior to dosing and according to the protocol. Study parameters were: plasma sCT levels, bone resorption by CTX-I (serum C-terminal telopeptide of collagen type I), bone formation by osteocalcin (serum OC), and cartilage degradation by CTX-II (urine C-terminal telopeptide of collagen type II) (clinicaltrials.gov identifier: NCT00486369). Doses of 0.8 mg compared with 0.6 mg produced significantly higher C(max) and AUC(0-4 hrs), of calcitonin, P=0.03. This resulted in significant reductions in CTX-I and CTX-II, [P<0.0001; P=0.007]. No differences were observed between baseline and follow-up at day 14 in pharmacokinetic and pharmacodynamic parameters. Gender had no observable influence on results. oSCT given twice daily with a pre-dinner and morning fasting dosing resulted in reductions in markers of bone resorption and cartilage degradation.
Article
Intra-articular drug delivery is very useful for treating local disease flare-ups, synovitis and pain in joints. However, the effectiveness of drugs following intra-articular administration is limited by drug delivery issues. This review addresses critical drug delivery parameters that influence the biocompatibility, tolerability and efficacy of intra-articular administrations and offers an opinion on aspects of formulation design. The relevant literature was reviewed, focusing on factors influencing tissue targeting, safety and effectiveness of particulate formulations. Therapeutic applications of novel drug delivery systems for the localized treatment of joints have lagged significantly. Future innovations in the field will require the discovery of new therapeutic agents for regional delivery, combination regimens, novel biomaterials as drug carriers and targeting carriers to specific molecules.
Article
To assess whether calcitonin exerts an influence on cartilage, three models of arthropathies in rabbits--representing three different modes of cartilage destruction--were used: (1) corticosteroid administration (endocrinological disturbances model); (2) meniscectomy (mechanical stress model); and (3) immobilization of the hind leg (nutritional disorder model). After 12 weeks of methylprednisolone (MP) administration, the rabbit femur heads displayed cartilage erosions, marked decrease of glycosaminoglycans (GAG) content, and narrowing of joint spaces. Elevation of serum uronic acid, activity of alkaline phosphatase, and calmodulin content was evident. All these changes were minimal--close to normal--in the group treated for 12 weeks with MP + salmon calcitonin (sCT). Partial meniscectomy and hind leg immobilization caused statistically significant loss of GAG from the cartilage and narrowing of the knee joint space during the same experimental period, 12 weeks. In both these models the groups of rabbits treated simultaneously with sCT showed only insignificantly smaller joint spaces and GAG content. These results support our hypothesis of a chondroprotective property of calcitonin. However, the mechanism through which calcitonin influences joint cartilage remains unknown. A direct effect of calcitonin on cultivated chondrocytes, as well as the role of calmodulin, beta-endorphins, calcium, and interleukin-1 in the process are discussed.
Article
We have previously demonstrated that a cultured porcine kidney cell, LLC-PK(1), maintains the characteristics of a polar renal epithelial cell in culture, and responds to salmon calcitonin and [arginine]vasopressin by increasing cyclic AMP content. To demonstrate the usefulness of this cell line as a model for the study of the biochemical events distal to cyclic AMP production, the activation of cyclic AMP-dependent protein kinase was examined. Intact cells in monolayer demonstrated progressive increases in cyclic AMP content and activation of protein kinase in response to [arginine]vasopressin (2-200nm) and salmon calcitonin (0.03-30nm) with both hormones fully activating the enzyme at a cell cyclic AMP content of 35pmol/mg of protein. Of the total cyclic AMP-dependent protein kinase activity, 80% was found in the 27000g supernatant fraction of sonicated cell material, and this soluble protein kinase could be fully activated by hormone. Conversely, the 27000g pellet contained a significant proportion of cyclic AMP-independent protein kinase and only 20% of total cell cyclic AMP-dependent protein kinase; the latter showed little response to hormone. On the basis of DEAE-cellulose chromatography, type II protein kinase was the predominant isoenzyme in both soluble and particulate fractions of the LLC-PK(1) cells and the soluble fractions of rat and guinea-pig renal medulla. Thus, the LLC-PK(1) cell line can serve as a model for hormonal modulation of protein kinase and as a potential source for defining the endogenous substrates for these enzymes.