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Measuring microscale strain fields in articular cartilage during rapid impact reveals thresholds for chondrocyte death and a protective role for the superficial layer.

ABSTRACT: Articular cartilage is a heterogeneous soft tissue that dissipates and distributes loads in mammalian joints. Though robust, cartilage is susceptible to damage from loading at high rates or magnitudes. Such injurious loads have been implicated in degenerative changes, including chronic osteoarthritis (OA), which remains a leading cause of disability in developed nations. Despite decades of research, mechanisms of OA initiation after trauma remain poorly understood. Indeed, although bulk cartilage mechanics are measurable during impact, current techniques cannot access microscale mechanics at those rapid time scales. We aimed to address this knowledge gap by imaging the microscale mechanics and corresponding acute biological changes of cartilage in response to rapid loading. In this study, we utilized fast-camera and confocal microscopy to achieve roughly 85 µm spatial resolution of both the cartilage deformation during a rapid (~3 ms), localized impact and the chondrocyte death following impact. Our results showed that, at these high rates, strain and chondrocyte death were highly correlated (p<0.001) with a threshold of 8% microscale strain norm before any cell death occurred. Additionally, chondrocyte death had developed by two hours after impact, suggesting a time frame for clinical therapeutics. Moreover, when the superficial layer was removed, strain - and subsequently chondrocyte death - penetrated deeper into the samples (p<0.001), suggesting a protective role for the superficial layer of articular cartilage. Combined, these results provide insight regarding the detailed biomechanics that drive early chondrocyte damage after trauma and emphasize the importance of understanding cartilage and its mechanics on the microscale.

SUBMITTER: Bartell LR

PROVIDER: S-EPMC4592793 | biostudies-literature | 2015 Sep

REPOSITORIES: biostudies-literature

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Measuring microscale strain fields in articular cartilage during rapid impact reveals thresholds for chondrocyte death and a protective role for the superficial layer.

Bartell Lena R LR Fortier Lisa A LA Bonassar Lawrence J LJ Cohen Itai I

Journal of biomechanics 20150612 12

Articular cartilage is a heterogeneous soft tissue that dissipates and distributes loads in mammalian joints. Though robust, cartilage is susceptible to damage from loading at high rates or magnitudes. Such injurious loads have been implicated in degenerative changes, including chronic osteoarthritis (OA), which remains a leading cause of disability in developed nations. Despite decades of research, mechanisms of OA initiation after trauma remain poorly understood. Indeed, although bulk cartilag ...[more]

PMID: 26150096

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Project description:Background/objectives:Articular cartilage erosion probably plays a substantial role in osteoarthritis (OA) initiation and development. Studies demonstrated that umbilical cord-derived mesenchymal stem cells (UCMSCs) could delay chondrocytes apoptosis and ameliorate OA progression in patients, but the detailed mechanisms are largely uncharacterised. In this study, we aimed to study the effects of UCMSCs on monosodium iodoacetate (MIA)-induced rat OA model, and explore the cellular mechanism of this effect. Methods:Intra-articular injection of 0.3 ?mg MIA in 50 ??L saline was performed on the left knee of the 200 ?g weight male Sprague-Dawley rat to induce rat knee OA. A single dose of 2.5 ?× ?105 undifferentiated UCMSCs one day after MIA or three-time intra-articular injection of 2.5 ?× ?105 UCMSCs on Days 1, 7 ?and 14 were given, respectively. Four weeks after MIA, joints were harvested and processed for paraffin sections. Safranine-O staining, haematoxylin and eosin staining ?and immunohistochemistry of MMP-13, ADAMTS-5, Col-2, CD68 ?and CD4 were performed to observe cartilage erosion and synovium. For in vitro ?studies, migration ability of cartilage superficial layer cells (SFCs) by UCMSCs were accessed by transwell assay. Furthermore, catabolism change of MIA-induced SFCs by UCMSCs was performed by real-rime polymerase chain reaction of Col-X and BCL-2 genes. CCK-8 assay was performed to check proliferation ability of SFCs by UCMSCs-conditioned media. Result:In this study, we locally injected human UCMSCs, which is highly proliferative and noninvasively collectible, into MIA-induced rat knee OA. An important finding is on obviously ameliorated cartilage erosion and decreased OA Mankin score by repeated UCMSCs injection after MIA injection compared with single injection, both of which attenuated OA progression compared with vehicle. Interestingly, we observed significantly increased number of SFCs on the articular cartilage surface, probably related to elevated proliferation, mobilisation and inhibited catabolism marker: Col-X and BCL-2 gene expression of cultured SFCs by UCMSCs-conditioned media treatment in vitro. In addition to the change of unique SFCs, catabolism markers of ADAMTS-5 and MMP-13 were substantially upregulated in the whole cartilage layer chondrocytes as well. Strikingly, MIA-induced inflammatory cells infiltration, on both CD4+ Th cells and CD68+ macrophages, and hyperplasia of the synovium, which was alleviated by repeated UCMSCs injection. Conclusion:Our study demonstrated a critical role of repeated UCMSCs dosing on preserving SFCs function, cartilage structure and inhibiting synovitis during OA progression, and thus provided mechanistic proof of evidence for the use of UCMSCs on OA patients in the future. The translational potential of this article:UCMSCs are a relatively "young" stem cell, and noninvasively collectible. In our study, we clearly demonstrated that it could effectively delay OA progression, possibly through reserving SFCs function and inhibiting synovitis. Therefore, it could be a new promising therapeutic cell source for OA after further clinical trials.

Project description:The degradation of proteoglycan was examined in cultured slices of pig articular cartilage. Pig leucocyte catabolin (10 ng/ml) was used to stimulate the chondrocytes and induce a 4-fold increase in the rate of proteoglycan loss from the matrix for 4 days. Material in the medium of both control and depleted cultures was mostly a degradation product of the aggregating proteoglycan. It was recovered as a very large molecule slightly smaller than the monomers extracted with 4M-guanidinium chloride and lacked a functional hyaluronate binding region. The size and charge were consistent with a very limited cleavage or conformational change of the core protein near the hyaluronate binding region releasing the C-terminal portion of the molecule intact from the aggregate. The 'clipped' monomer diffuses very rapidly through the matrix into the medium. The amount of proteoglycan extracted with 4M-guanidinium chloride decreased during culture from both the controls and depleted cartilage, and the average size of the molecules initially remained the same. However, the proportion of molecules with a smaller average size increased with time and was predominant in explants that had lost more than 70% of their proteoglycan. All of this material was able to form aggregates when mixed with hyaluronate, and glycosaminoglycans were the same size and charge as normal, indicating either that the core protein had been cleaved in many places or that larger molecules were preferentially released. A large proportion of the easily extracted and non-extractable proteoglycan remained in the partially depleted cartilage and the molecules were the same size and charge as those found in the controls. There was no evidence of detectable glycosidase activity and only very limited sulphatase activity. A similar rate of breakdown and final distribution pattern was found for newly synthesized proteoglycan. Increased amounts of latent neutral metalloproteinases and acid proteinase activities were present in the medium of depleted cartilage. These were not thought to be involved in the breakdown of proteoglycan. Increased release of proteoglycan ceased within 24h of removal of the catabolin, indicating that the effect was reversible and persisted only while the stimulus was present.

Dataset Information

Measuring microscale strain fields in articular cartilage during rapid impact reveals thresholds for chondrocyte death and a protective role for the superficial layer.

Publications

Measuring microscale strain fields in articular cartilage during rapid impact reveals thresholds for chondrocyte death and a protective role for the superficial layer.

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