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T-2 toxin is one of the most toxic and common mycotoxins in grains and related products. It is considered a risk factor for Kashin-Beck disease (KBD), an endemic osteoarthritis. Both in vitro and in vivo studies have shown that T-2 toxin can cause extracellular matrix degradation; however, the underlying mechanism is unclear. Integrins have been found to regulate the expression of matrix metalloproteinases (MMPs), the 'scissors' of matrix proteins. In this study, we investigated whether integrin αv played a role in T-2 toxin-induced matrix degradation. Results from our study showed that the expression of integrin αv in the cartilage of rats fed T-2 toxin was reduced compared to that in rats fed a normal diet. Integrin αv was downregulated in T-2 toxin-treated C28/I2 chondrocytes, and selenium was found to have a protective effect. The expression of MMP-1, -3, -10, and -13 increased whereas that of type II collagen (Col II) protein decreased in C28/I2 cells treated with an integrin αv inhibitor. In conclusion, T-2 toxin can downregulate integrin αv expression in chondrocytes. Reduced integrin αv signalling could induce the release of MMPs, leading to matrix degradation., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Recent evidence suggests a role of type II collagen in Kashin-Beck disease (KBD) degeneration. We aimed to assess the abnormal expression of heat shock protein 47 (HSP47) which is associated with a decrease in type II collagen and an increase in cartilage degradation in KBD. Hand phalange cartilages were collected from KBD and healthy children. Rats were administered with T-2 toxin under the selenium (Se)-deficient diet. ATDC5 cells were seeded on bone matrix gelatin to construct engineered cartilaginous tissue. C28/I2 and ATDC5 cells and engineered tissue were exposed to different concentrations of T-2 toxin with or without Se. Cartilage degeneration was determined through histological evaluation. The distribution and expression of type II collagen and HSP47 were investigated through immunohistochemistry, western blotting, and real-time PCR. KBD cartilages showed increased chondronecrosis and extracellular matrix degradation in deep zone with decreased type II collagen and HSP47 expression. The low-Se + T-2 toxin animal group showed a significantly lower type II collagen expression along with decreased HSP47 expression. Decreased type II collagen and HSP47 in C28/I2 and ATDC5 cells induced by T-2 toxin showed a dose-dependent manner. Hyaline-like cartilage with zonal layers was developed in engineered cartilaginous tissues, with decreased type II collagen and HSP47 expression found in T-2 toxin-treated group. Se-supplementation partially antagonized the inhibitory effects of T-2 toxin in chondrocytes and cartilages. HSP47 plays a role in the degenerative changes of KBD and associated with T-2 toxin-induced decreased type II collagen expression, further promoting matrix degradation.

Kashin-Beck disease (KBD) is an endemic degenerative osteoarthropathy of uncertain etiology. Our study sought to identify a correlation between small proteoglycans decorin and biglycan expression and Kashin-Beck Disease. Immunohistochemistry was used to assess the decorin and biglycan levels in cartilage specimens from both child KBD patients, and rats fed with T-2 toxin under a selenium-deficient condition. Real-time PCR and Western blot were used to assess mRNA and protein levels of decorin and biglycan in rat cartilages, as well as in C28/I2 chondrocytes stimulated by T-2 toxin and selenium in vitro. The result showed that decorin was reduced in all zones of KBD articular cartilage, while the expression of biglycan was prominently increased in KBD cartilage samples. Increased expression of biglycan and reduced expression of decorin were observed at mRNA and protein levels in the cartilage of rats fed with T-2 toxin and selenium- deficiency plus T-2 toxin diet, when compared with the normal diet group. Moreover, In vitro stimulation of C28/I2 cells with T-2 toxin resulted in an upregulation of biglycan and downregulation of decorin, T-2 toxin induction of biglycan and decorin levels were partly rescued by selenium supplement. This study highlights the focal nature of the degenerative changes that occur in KBD cartilage and may suggest that the altered expression pattern of decorin and biglycan have an important role in the onset and pathogenesis of KBD.

The purpose of this paper was to investigate chondrocyte distribution and death in the cartilage in Kashin-Beck disease (KBD). Apoptotic chondrocytes were detected by TUNEL assay. Ultrastructural changes were examined by transmission electron microscope (TEM). Biochemical markers associated with apoptosis (eg, caspase-3) and necroptosis (eg, RIP3) were investigated by immunohistochemistry. In KBD cartilage chondrocyte death was characterized by paler staining of the cells. Multiple chondral cell clusters surrounded the areas lacking cells in the deep zone. The per cent of TUNEL-positive and RIP3-positive chondrocytes were higher in the middle zones of KBD samples; however, there was some positive staining for TUNEL but negative staining for caspase-3. Immunohistochemistry failed to detect significant differences in caspase-3 levels in KBD children compared to controls, suggesting that beside apoptosis necroptosis dominates as a cell death mechanism in the middle zone of cartilage from KBD children. To clarify further the presence of chondrocyte necroptosis in KBD, we performed TUNEL, caspase-3 and RIP3 staining in a rat KBD model which is based upon T-2 toxin treatment under selenium-deficient conditions. Apoptosis and necroptosis co-existed in the middle zone in this rat KBD model. Ultrastructural analysis of chondrocyte from deep cartilage revealed abnormal cells with numerous morphological changes, such as plasma membrane breakdown, generalized swelling of the cytoplasm and loss of identifiable organelles. Chondrocyte death by necrosis in the deep zone of cartilages in KBD may be a result of exposure to T-2 toxin from bone marrow or bloodstream under selenium-deficient nutrition status in KBD endemic areas. Chondrocyte death plays a key role in either the initiation or the progression of KBD pathogenesis., (© 2019 The Authors. International Journal of Experimental Pathology © 2019 International Journal of Experimental Pathology.)

Chondrocyte is targeted for disruption in Osteoarthritis (OA) and Kashin-Beck Disease (KBD), and chondrocyte death in cartilage may contribute to the progression of OA and KBD. Oxidative stress leads to increased risk for OA. Previous work in our laboratory implicates oxidative stress as a potential mediator in children with KBD. While these studies suggest a role for oxidative stress in the modulation of OA and KBD, the direct effects of reactive oxygen species/reactive nitrogen species (ROS/RNS) on the stability of this domain remain unclear. Here, we demonstrate that oxidative stress, as induced through treatment with 3-morpholinosydnonimine (SIN-1), a spontaneous ROS/RNS generator, decreased the cell viability in hypertrophic chondrocytes in a dose- and time- dependent manner. SIN-1 induced necrosis in hypertrophic chondrocytes, whereas triggered apoptosis in non-hypertrophic cells of non-differentiated ATDC5 cells and C28/I2 cells. Ultrastructural analysis of hypertrophic chondrocyte treated with SIN-1 revealed morphological changes, such as plasma membrane breakdown, generalized swelling of the cytoplasm and organelles, even to disappearance. Moreover, SIN-1 induced chondronecrosis in the deep zone of engineered cartilage tissue, such as cell-free vacancy and "red ghost" cells. Overall, we demonstrate for the first time that oxidative stress, as induced through exogenous ROS/RNS, leads to necrosis in hypertrophic chondrocytes. Oxidative stress-mediated necrotic cell death contributes to chondronecrosis in the deep zone of cartilage in both OA and KBD., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Objective: The aim of this study was to investigate FGF8 and FGFR3 expression in clinical samples of Kashin-Beck disease (KBD), an endemic osteochondropathy found in China, as well as in pre-clinical models of this disease., Method: Cartilage was collected from the hand phalanges of five patients with KBD and from five healthy children. Sprague-Dawley rats were administered a selenium-deficient diet for four weeks prior to exposure to the T-2 toxin. ATDC5 cells were differentiated into hypertrophic chondrocytes for twenty-one days, and then treated with 3-morpholinosydnonimine (SIN-1) (0, 1, 3, or 5 mM) for 24 h. FGF8 and FGFR3 were visualized using immunohistochemistry; protein levels were assessed by western blotting, and mRNA levels were determined by real-time RT-PCR., Results: Increased staining of FGF8 and FGFR3 was observed in the cartilage of children with KBD compared to normal children. Both increased FGF8 and FGFR3 staining, as well as protein levels, were also observed in the cartilage of rats fed normal or Se-deficient diets plus T-2 toxin exposure, compared to those in rats fed with normal or Se-deficient diets alone. SIN-1 treatment of hypertrophic chondrocytes (ATCD5 cells) increased FGF8 and FGFR3 protein and mRNA levels in a dose-dependent manner., Conclusion: Our data indicate that SIN-1 induces FGF8 and FGFR3 overexpression and this is involved in the abnormal terminal differentiation and degradation of the ECM in cartilage. FGF8 and FGFR3 may therefore play an important role in the onset of deep zone necrosis and pathogenesis in KBD in adolescent children., (Copyright © 2018. Published by Elsevier Inc.)

Objective To identify the osteogenesis genes whose expression is altered in hypertrophic chondrocytes treated with H 2 O 2. Methods Murine chondrogenitor cells (ATDC5) were differentiated into hypertrophic chondrocytes by Insulin-Transferrin-Selenium (ITS) treatment, and then treated with H 2 O 2. Suitable conditions (concentration, time) were determined by using the MTT assay. After total RNA isolation and cDNA synthesis, the levels of 84 genes were determined using the PCR array, whereas quantitative RT-PCR was carried out to validate the PCR array data. Results We identified 9 up-regulated genes and 12 down-regulated genes, encoding proteins with various functions, such as collagen proteins, transcription factors, proteins involved in skeletal development and bone mineral metabolism, as well as cell adhesion molecules. Quantitative RT-PCR confirmed the altered expression of 5 down-regulated genes (Smad2, Smad.4, transforming growth factor β receptor 1, transforming growth factor β receptor 3, and matrix metalloproteinase 10). Conclusions H 2 O 2 significantly changed the expression of several genes involved in a variety of biological functions. Because of the link between oxidative damage and Kashin-Beck disease, these genes may also be involved in the deep-zone necrosis of the cartilage observed in Kashin-Beck disease. [ABSTRACT FROM AUTHOR]