Adipose tissue represents an abundant source of stem cells. Along with anti-inflammatory effects, ASC secrete various factors that may modulate metabolism of extracellular matrix in osteoarthritic (OA) cartilage, suggesting that the presence of ASC could be advantageous for OA cartilage due to the recovery of homeostasis between matrix metalloproteinases (MMPs) and their tissue inhibitors of metalloproteinases (TIMPs). To evaluate these effects, cartilage explants (CE) were cocultured with ASC for 3 and 7 days under stimulation with or without IL-1β. The pattern of gene expression in CE was modified by ASC, including the upregulation of COL1A1 and COL3A1 and the downregulation of MMP13 and COL10A1. The production of MMP-1, MMP-3, and MMP-13 by ASC was not significant; moreover, cocultures with ASC reduced MMP-13 production in CE. In conclusion, active production of TIMP-1, TIMP-2, TIMP-3, IL-6, IL-8, and gelatinases MMP-2 and MMP-9 by ASC may be involved in the extracellular matrix remodelling, as indicated by the altered expression of collagens, the downregulated production of MMP-13, and the reduced chondrocyte apoptosis in the cocultured CE. These data suggest that ASC modulated homeostasis of MMPs/TIMPs in degenerated OA cartilage in vitro and might be favourable in case of the intra-articular application of ASC therapy for the treatment of OA.
Background and Objectives Balance between matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) ensures maintenance of healthy cartilage, whereas weighted damaging effects of the enzymes leads to the development of osteoarthritis (OA). To develop the adipose tissue derived stem cell (ASC) therapy in OA, we were seeking to evaluate paracrine and chondroprotective properties of the cells in recovering the homeostasis between MMPs/TIMPs in the joint. Methods To determine the effects of ASCs on OA cartilage, we performed short (3 & 7 days) and long (21 day) in vitro ASC cocultures with articular cartilage explants (CE). To further reproduce the OA conditions, cocultures were also stimulated with IL-1β. Production of TIMPs, MMPs, and factors, related to inflammation or chondrogenesis, was analysed in coculture supernatants by ELISA. CE were further processed for RT-qPCR and immunohistochemical analysis. Results ASC produced TIMP-1, -2 and -3, as well as gelatinases MMP-2 and -9 at the levels similar to those of CE. Gelatinases MMP-2 and -9 degrade denatured collagens, including type I. Cocultures with ASC resulted in up-regulation of COL1A1 gene. Although the increase in collagen type I production is generally considered as a sign of fibrosis, its enhancement has also been demonstrated during cartilage development, and might reflect early events of the repair. No or low production of MMP-1, MMP-3, and MMP-13 was determined in supernatants of ASC, whereas CE produced them abundantly. Noteworthy, cocultures with the cells resulted in significant suppression of MMP-13 on days 3 & 7, and in particular, day 21, as compared to CE alone at the both, mRNA and protein level. Stimulation with IL-1β differentially modulated production of various MMPs and TIMPs by ASCs and CE, however the effects of ASC seem to further be beneficial. Histological examination showed a tendency to extracellular matrix improvement in the osteoarthritic CE cocultured with ASC, further supporting chondroprotective effects of those cells in the both, presence or absence of IL-1β. Conclusions Production of TIMPs and gelatinases, associated with the increase in collagen I production in CE by ASC suggest their active participation in cartilage turnover, encompassing stimulation of remodelling processes. Absence of MMP-1, -3 or -13 production implies safety of the use of ASC for intraarticular clinical applications. Down-regulation of MMP13 in CE indicates possible antihypertrophic effects of ASC. Taken together, results of our study suggest beneficial properties of ASC in prevention of cartilage from degradation during OA, even in the presence of IL-1β.
Background Articular cartilage damage in osteoarthritis (OA) is characterized by degeneration of the extracellular matrix. Matrix metalloproteinases (MMPs) play a crucial role in the modulations of cell-matrix interactions and degradation of cartilage. MMPs are controlled through activation of proenzymes and the inhibition of active enzymes by tissue inhibitors of metalloproteinases (TIMPs). Adipose tissue derived stem cells (ADSCs) are close to mesenchymal stem cells from bone marrow in their anti-inflammatory and supportive for tissues repair effect, moreover, ADSCs are much easier available. Objectives To evaluate early and late effects of ADSCs for OA cartilage for the development of the cell therapy in OA, using their paracrine and chondroprotective effects. Objectives In vitro cocultures of ADSC with articular cartilage explants (CE) were performed for 3, 7 and 21 days. To further reproduce the OA conditions co-cultures were also stimulated with IL-1β. Secretion of TIMPs, MMPs, and other factors was determined in coculture supernatants by ELISA. Expression of genes in CE associated with chondrogenesis was analyzed by RT-qPCR. Methods we performed in vitro ADSC cocultures with articular cartilage explants (CE) for 3, 7 and 21 days. To further reproduce the OA conditions co-cultures were also stimulated with IL-1β. Secretion of TIMPs, MMPs, and other factors was analyzed in coculture supernatants by ELISA. Expression of genes associated with chondrogenesis was analyzed in CE by RT-qPCR. Results The both, ADSC and CE produced TIMP-1, TIMP-2 and TIMP-3 as well as gelatinases MMP-2 and MMP-9 at early stage (days 3 & 7) of cocultures and also later, on day 21. On the contrary to the other MMPs, suppressive role of MMP-2 in arthritis has been demonstrated. No or low production of VCAM-1, MMP-1, MMP-3, and MMP-13 was determined in supernatants of ADSC, whereas CE cocultures with the cells resulted in significant suppression of the both, mRNA and protein MMP-13 on days 3 & 7, but in particular on day 21. Cocultures with ADSC changed gene expression profile in CE, with the most pronounced up-regulation of COL1A1 gene on days 3&7, the effect being absent on day 21. Although increase in collagen type I production is generally considered as a sign of fibrosis, it has also been demonstrated during cartilage development and possibly may reflect early events of the repair. Stimulation with IL-1β differentially modulated gene expression and production of MMPs and TIMPs in cocultures. Histological examination has shown a tendency to extracellular matrix improvement in the osteoarthritic CE cocultured with ADSC, suggesting chondroprotective effects of those cells. Conclusions Secretory profile and results of histological analysis suggest possible beneficial effects of ADSC in prevention of cartilage from degeneration during OA. Production of MMP2 and MMP9 might be implicated in increased COL1A1 expression in CE. Down-regulation of MMP13 and COL10A1 genes in CE cocultured with ADSC implies antihypertrophic effects of ADSC...
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