ABSTRACT
OBJECTIVE: To investigate the effects of T-2 toxin on expressions of Fas, p53, Bcl-xL, Bcl-2, Bax and caspase-3 on human chondrocytes. METHODS: Human chondrocytes were treated with T-2 toxin (1-20 ng/ml) for 5 d. Fas, p53 and other apoptosis-related proteins such as Bax, Bcl-2, Bcl-xL, caspase-3 were determined by Western blot analysis and their mRNA expressions were determined by reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: Increases in Fas, p53 and the pro-apoptotic factor Bax protein and mRNA expressions and a decrease of the anti-apoptotic factor Bcl-xL were observed in a dose-dependent manner after exposures to 1-20 ng/ml T-2 toxin, while the expression of the anti-apoptotic factor Bcl-2 was unchanged. Meanwhile, T-2 toxin could also up-regulate the expressions of both pro-caspase-3 and caspase-3 in a dose-dependent manner. CONCLUSION: These data suggest a possible underlying molecular mechanism for T-2 toxin to induce the apoptosis signaling pathway in human chondrocytes by regulation of apoptosis-related proteins.
Subject(s)
Chondrocytes/drug effects , Chondrocytes/metabolism , T-2 Toxin/toxicity , Apoptosis/drug effects , Base Sequence , Blotting, Western , Caspase 3/genetics , Caspase 3/metabolism , Cell Proliferation/drug effects , Cells, Cultured , Chondrocytes/cytology , DNA Primers/genetics , Gene Expression/drug effects , Humans , Proto-Oncogene Proteins c-bcl-2/genetics , Proto-Oncogene Proteins c-bcl-2/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Signal Transduction/drug effects , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Protein p53/metabolism , bcl-2-Associated X Protein/genetics , bcl-2-Associated X Protein/metabolism , bcl-X Protein/genetics , bcl-X Protein/metabolism , fas Receptor/genetics , fas Receptor/metabolismABSTRACT
OBJECTIVE: To investigate the relationship of T-2 toxin-induced chondrocytes apoptosis with nitric oxide(NO) and Fas apoptosis pathway. METHODS: Human chondrocytes cultured in vitro were treated with different concentrations of T-2 toxin at different time (1-5 d). Cell viability of the treated cells was measured by MTT assay. Apoptotic ultrostructural changes of the treated cells were observed with electron microscopy. Biological changes of apoptosis were detected by annexin V/PI Flow cytometer (FCM). The levels of NO in culture media were detected by colorimetric method of Griess assay. Nitric oxide synthase (iNOS) and Fas protein were measured by Western blot. RESULTS: In this study the results shown the dose-dependent and time-dependent effects of T-2 toxin, within a range of concentration (1-2000 ng/mL) and a period of time (1-5 d), on the T-2 toxin-treated chondrocytes. Apoptotic body was found in T-2 toxin-treated chondrocytes by electron microscopy. Early-stage apoptosis rate and late-stage apoptosis rate were both increased in T-2 toxin-treated cells when compared with non-treated cells in a dose-dependent manner. The levels of NO in T-2 toxin-treated culture media were higher than that of normal control. Over-expressions of iNOS and Fas protein were detected in T-2 toxin-treated cells. T-2 toxin-induced apoptosis was noted to be significtnly correlated with the level of NO production and the levels of iNOS and Fas protein expression. CONCLUSION: T-2 toxin can enhance NO production and upregulate the expression of iNOS and Fas protein. Both NO and Fas signaling pathway are involved in T-2 toxin-induced apoptosis.
Subject(s)
Apoptosis/drug effects , Chondrocytes/cytology , Nitric Oxide/biosynthesis , T-2 Toxin/toxicity , fas Receptor/biosynthesis , Cells, Cultured , Chondrocytes/metabolism , Humans , Nitric Oxide Synthase Type II/biosynthesisABSTRACT
OBJECTIVE: To study the inhibitory effect of T-2 toxin on the expression of aggrecan and collagen II in chondrocytes and the protection of selenium against this effect. METHODS: Human chondrocytes cultured in vitro were treated with T-2 toxin at different concentrations for varied time periods (1-5 days), and the cell viability was measured by MTT assay. Aggrecan expression was detected by toluidine blue staining and collagen II expression by immunostaining using monoclonal antibody of collagen. Aggrecan and collagen II mRNA expressions were measured by semiquantitative RT-PCR. RESULTS: T-2 toxin dose- and time-dependently affected chondrocyte viability within the concentration range of 0.001-2 mg/L, the prolonged treatment time further enhanced the dose dependence of the inhibitory effect. T-2 toxin lowered aggrecan and collagen II synthesis in the chondrocytes and reduced their mRNA expressions. Selenium could partly attenuate the inhibitory effects of T-2 toxin on aggrecan mRNA expression, but showed no such effect against T-2-induced collagen II expression. CONCLUSION: T-2 toxin can obviously inhibit aggrecan and collagen II synthesis in human chondrocytes, and selenium can partly antagonize the inhibitory effects of T-2 toxin on aggrecan.
