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Limited availability of the organs donors has facilitated the establishment of xenogeneic organ sources for transplantation. Numerous studies have decellularized several organs and assessed their implantability in order to provide such organs. Among all the decellularized organs studies for xenotransplantation, the pancreas has garnered very limited amount of research. The presently offered alternatives for pancreas transplantation are unable to liberate patients from donor dependence. The rat and mice pancreas are not of an accurate size for transplantation but can only be used for in-vitro studies mimicking in-vivo immune response in humans, while the porcine pancreas can cause zoonotic diseases as it carries porcine endogenous retrovirus (PERV- A/B/C). Therefore, we propose caprine pancreas as a substitute for these organs, which not only reduces donor dependence but also poses no risk of zoonosis. Upon decellularization the extracellular matrix (ECM) of different tissues responds differently to the detergents used for decellularization at physical and physiological level; this necessitates a comprehensive analysis of each tissue independently. This study investigates the impact of decellularization by ionic (SDS and SDC), non-ionic (Triton X-100 and Tween-20), and zwitterionic detergents (CHAPS). All these five detergents have been used to decellularize caprine pancreas via immersion (ID) and perfusion (PD) set-up. In this study, an extensive comparison of these two configurations (ID and PD) with regard to each detergent has been conducted. The final obtained scaffold with each set-up has been evaluated for the left-over cytosolic content, ECM components like sGAG, collagen, and fibronectin were estimated via Prussian blue and Immunohistochemical staining respectively, and finally for the tensile strength and antimicrobial activity. All the detergents performed consistently superior in PD than in ID. Conclusively, PD with SDS, SDC, and TX-100 successfully decellularizes caprine pancreatic tissue while retaining ECM architecture and mechanical properties. This research demonstrates the viability of caprine pancreatic tissue as a substitute scaffold for porcine organs and provides optimal decellularization protocol for this xenogeneic tissue. This research aims to establish a foundation for further investigations into potential regenerative strategies using this ECM in combination with other factors.

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Intervertebral disc degeneration (IDD)-induced low back pain significantly influences the quality of life, placing a burden on public health systems worldwide. Currently available therapeutic strategies, such as conservative or operative treatment, cannot effectively restore intervertebral disc (IVD) function. Decellularized matrix (DCM) is a tissue-engineered biomaterial fabricated using physical, chemical, and enzymatic technologies to eliminate cells and antigens. By contrast, the extracellular matrix (ECM), including collagen and glycosaminoglycans, which are well retained, have been extensively studied in IVD regeneration. DCM inherits the native architecture and specific-differentiation induction ability of IVD and has demonstrated effectiveness in IVD regeneration in vitro and in vivo. Moreover, significant improvements have been achieved in the preparation process, mechanistic insights, and application of DCM for IDD repair. Herein, we comprehensively summarize and provide an overview of the roles and applications of DCM for IDD repair based on the existing evidence to shed a novel light on the clinical treatment of IDD.

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The 1,9-dimethylmethylene blue (DMMB) assay enables the detection of sulfated glycosaminoglycans (sGAGs). This assay can be used to quickly quantify the sGAG content in a large number of samples using spectrophotometry. While this widespread assay appears straightforward, there are certain pitfalls that need to be considered.

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The described research focused on the diagnostic usefulness of sulfated glycosaminoglycans (sGAG), hyaluronan (HA), and extracellular part of syndecan-1 (sCD138) as new markers related to extracellular matrix (ECM) remodeling in the intestine during the two most common forms of inflammatory bowel diseases (IBD), i.e., ulcerative colitis (UC) and Crohn' disease (CD). Inflammatory markers belonging to ECM components were assessed in serum of patients with IBD using an immunoenzymatic method (HA and sCD138) and a method based on the reaction with dimethylmethylene blue (sulfated GAG). Measurements were carried out twice: at baseline and after one year of therapy with prednisone (patients with CD) or adalimumab (patients with UC). No quantitative changes were observed in serum sGAG, HA, and sCD138 concentrations between patients newly diagnosed with CD and the healthy group. In the case of patients with UC, the parameter which significantly differentiated healthy subjects and patients with IBD before biological therapy was HA. Significant correlation between serum HA level and inflammation activity, expressed as Mayo score, was also observed in patients with UC. Moreover, the obtained results have confirmed that steroid therapy with prednisone significantly influenced the circulating profile of all examined ECM components (sGAG, HA, and sCD138), whereas adalimumab therapy in patients with UC led to a significant change in only circulating sGAG levels. Moreover, the significant differences in serum HA levels between patients with UC and CD indicate that quantification of circulating HA may be useful in the differential diagnosis of CD and UC.

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Determining the influence of tissue composition on the osmotic swelling stress of articular cartilage and meniscus fibrocartilage is important to enhance our understanding of physiology and disease. This osmotic swelling stress is critical for the load-bearing capability of both tissues and results in part due to the interactions between the negatively charged sulfated glycosaminoglycan (sGAG) chains and the ionic interstitial fluid. Changes in sGAG content, as those occurring during the progression of degenerative joint disease, alter such interactions. Here, we compare the time-varying effects of altered osmotic environments on the confined compression swelling behavior of bovine tissues spanning a range of sGAG concentrations: juvenile articular cartilage, juvenile and adult meniscus, and juvenile cartilage enzymatically degraded to reduce its sGAG content. The transient response to changes in bath conditions was evaluated for explants assigned to one of three compressive offsets (5%, 10%, or 15% strain) and one of three bath conditions (0.1X, 1X, or 10X phosphate-buffered saline). Our results show that relative responses to alterations to the osmotic environment are consistent across native tissues but differ for degraded juvenile cartilage, demonstrating that changes in sGAG do not completely recapitulate the native swelling behaviors. Further, we found a strong correlation between aggregate modulus and sGAG/collagen, as well as between sGAG and collagen contents across native tissue types, suggesting some conservation of composition-function relationships across a range of tissue types with varying sGAG concentrations. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:785-792, 2020.

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BACKGROUND: Anterior cruciate ligament tears can lead to posttraumatic osteoarthritis. In addition to biomechanical factors, changes in biochemical profiles within the knee joint after injury and anterior cruciate ligament reconstruction (ACLR) may play a role in accelerating joint degeneration. Hypothesis/Purpose: It was hypothesized that cartilage matrix composition after ACLR is associated with the degree of inflammatory response after initial injury. This study evaluated the association between the inflammatory response after injury-as indicated by cytokine, metalloproteinase, and cartilage degradation marker concentrations in synovial fluid-and articular cartilage degeneration, measured by T1ρ and T2 quantitative magnetic resonance imaging up to 3 years after ACLR. STUDY DESIGN: Cohort study; Level of evidence, 2. METHODS: Twenty-six subjects from a longitudinal cohort study who underwent ACLR at a mean 8.5 weeks after injury (range, 4-19 weeks) had synovial fluid aspirated at the time of surgery. Immunoassays quantified biomarkers in synovial fluid. T1ρ and T2 values of articular cartilage were calculated with magnetic resonance scans acquired prior to surgery and at 6 months and 1, 2, and 3 years after surgery. Pearson correlation coefficients were calculated among the various biomarkers. K-means clustering was used to group subjects with similar biomarker profiles. Generalized estimating equations were used to find the overall differences in T1ρ and T2 values throughout these first 3 years after surgery between the clusters while controlling for other factors. RESULTS: Significant and strong correlations were observed between several cytokines (interleukin 6 [IL-6], IL-8, IL-10, and tumor necrosis factor α) and 2 matrix metalloproteinases (MMP-1 and MMP-3) ( P < .05). Moderate correlations were found among combinations of C-terminal crosslinked telopeptide type II collagen, N-terminal telopeptide, cartilage oligomeric matrix protein, and sulfated glycosaminoglycan ( P < .05). Two clusters were generated, 1 of which was characterized by lower concentrations of cytokines (IL-6, IL-8, IL-10, tumor necrosis factor α) and MMP-1 and MMP-3 and higher sulfated glycosaminoglycan. This cluster was associated with significantly higher T1ρ and T2 values in the medial tibial and patellar cartilage over the first 3 years after ACLR. CONCLUSION: At the time of ACLR surgery, profiles of synovial fluid inflammatory cytokines, degradative enzymes, and cartilage breakdown products show promise as predictors of abnormal cartilage tissue integrity (increased T1ρ and T2 values) throughout the first 3 years after surgery. CLINICAL RELEVANCE: The results suggest an intricate relationship between inflammation and cartilage turnover, which can in turn be influenced by timing after injury and patient factors.

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BACKGROUND: Patients who sustain an acute anterior cruciate ligament (ACL) rupture are at increased risk to develop posttraumatic arthritis (PTA) in the injured knee whether the ACL is reconstructed or treated nonoperatively. Inflammatory cytokines and cartilage degradation biomarkers are elevated at the time of acute injury and postoperatively. This suggests that one mechanism for PTA may be an inflammatory degradative process initiated on the acute injury and sustained for some length of time independent of whether adequate joint stability is restored. HYPOTHESIS: Inflammatory cytokines and biomarkers of cartilage degradation are elevated in the synovial fluid several years after reconstruction of the ACL, indicating an ongoing imbalance between extracellular matrix destruction and repair. STUDY DESIGN: Cross-sectional study; Level of evidence, 3. METHODS: In 11 patients who had undergone ACL reconstruction 8 years earlier, knee synovial fluid was aspirated from the operated knee and the contralateral nonoperated knee. The synovial fluid was analyzed for interleukin (IL)-1ß, IL-6, tumor necrosis factor (TNF)-α, sulfated glycosaminoglycans (sGAG), aggrecan neoepitope fragment (ARGS-aggrecan), and cartilage oligomeric matrix protein (COMP). At follow-up, the patients underwent bilateral weightbearing radiographs and bilateral MRIs of their knees. RESULTS: No significant differences between the operated and the contralateral knee were found for the synovial fluid concentrations of IL-1ß, IL-6, TNF-α, sGAG, ARGS-aggrecan, or COMP. There were significantly more radiographically visible osteoarthritic changes in the operated knees compared with the contralateral knees. MRIs revealed that all grafts and all contralateral ACLs were intact and, furthermore, that there was significantly more meniscal and cartilage damage in the index knees than the contralateral knees. CONCLUSION: Eight years after ACL reconstruction, there were no significant differences in inflammatory cytokines and biomarkers for cartilage degeneration between the nonoperated and the ACL-reconstructed knee, even though there were more osteoarthritic changes and meniscal and cartilage damage in the operated knee, as seen on weightbearing radiographs and MRI.

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OBJECTIVES: To observe the age-related changes of sulfated glycosaminoglycan (sGAG) content of hip joint cartilage of elderly people based on Equilibrium Partitioning of an Ionic Contrast Agent (EPIC) micro-CT. METHODS: Seventy human hip cartilage-bone samples were collected from hip-fracture patients (ages 51-96) and divided into five groups (10 years in an age group). They were first immersed in 20% concentration of the contrast agent Meglumine Diatrizoate (MD) for 6 h at 37 °C, and then scanned by micro-CT. Following scanning, samples were stained for sGAG with toluidine blue. The X-ray attenuation and sGAG optical density were calculated by image processing. The correlation between X-ray attenuation and sGAG optical density was then analyzed. RESULTS: The X-ray mean attenuation of the cartilage increased by 18.81% from the 50-80 age groups (p < 0.01), but decreased by 7.15% in the 90 age group compared to the 80 age group. The X-ray mean attenuation of the superficial layer and middle layer increased by 31.60 % and 44.68% from the 50-80 age groups, respectively (p < 0.01), but reduced by 4.67% and 6.05% separately in the 90 age group. However, the deep layer showed no significant change with aging. The sGAG optical density showed a linear correlation (r = -0.91, p < 0.01) with the X-ray attenuation. CONCLUSION: The sGAG content of hip joint cartilage varied with aging in elderly people. The changes in superficial layer and middle layer were more evident than deep layer.

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