钙调神经磷酸酶抑制剂对过氧化氢诱导的干细胞凋亡的影响简

目的比较钙调神经磷酸酶（CAN）抑制剂FK-506和环孢菌素A（CsA）保护脂肪组织来源干细胞（AT-DSCs）抗凋亡作用及可能机制。方法采用过氧化氢（H2O2）[分浓度0（对照组）、50、100、200、300、400μmo/LH2O26组]体外诱导AT-DSCs凋亡模型．凋亡模型的评价采用磷脂酰丝氨酸结合蛋白-异硫氰酸荧光素（AnnexinV-FITC）／碘化丙啶（PI）双染流式细胞术和线粒体内跨膜电位测定法；观察AT-DSCs凋亡细胞形态；AT-DSCs细胞活性检测采用CCK-8试剂；细胞色素C的表达采用Westernblot分析。结果与对照组比较，引起细胞凋亡率最高的是浓度100μmol／LH20，（15．60％±8．50％±4．50％±0．88％；p〈0．01）；但CaN抑制剂FK-506（质量浓度1mg／mL）（9．8％±1．3％Vs17．0％±1．6％，P〈0．01）和CsA（质量浓度1mg／mL）（12．2％±1．2％Vs17．0％4±1．6％，P〈0．01）预处理后细胞凋亡率均明显下降，细胞存活率明显提高（P〈0．01）。Western blot结果显示．CaN抑制剂FK-506（质量浓度1mg／mL）和CsA（质量浓度1mg／mL）可完全抑制H2O2诱导的细胞色素C释放。结论CaN抑制剂能保护H2O2诱导的干细胞凋亡。可能机制是CaN抑制剂能抑制细胞凋亡，提高细胞存活率，下调细胞色素C的释放。

Inhibitor of calcineurin protects stem cells from H2O2-induced apoptosis

Objective To investigate the effect of inhibitor of calcineurin（CaN） in protecting the adipose tissue-derived stem cells （AT-DSCs） against apoptosis and the possible mechanisms. Methods A model of hydrogen peroxide（H202）-induced apoptosis was assessed by Annexin V-FITC/PI double staining cytometry and measurement of mitoehondria membrane potential. The morphology of apoptotic cells was observed. Cell activity was determined by CCK-8 assay. Expression of cytochrome C was investigated by Western blot method. Results Compared to the control, evidences showed that 100 txmol/L H202 could induce highest degree of apoptotic change in AT-DSCs（15.60 % ± 8.50 % vs 4.50 % ± 0.88 %, P〈 0.01）. This effect was inhibited by pre- treatment with calcineurin inhibitor FK-506 （1 mg/mL）（9.8 % ± 1.3 % vs 17.0 % ± 1.6 %, P〈 0.01） and CsA （1 mg/mL）（12.2 % ± 1.2 % vs 17.0 % ± 1.6 %, P 〈 0.01）, induced to significantly increased survival rate. Furthermore, Western blot analysis showed that FK-506（1 mg/mL） and CsA（1 mg/mL） completely inhibited the H~Og-induced release of cytochrome C. Conclusion These resuhs suggest that the inhibitor of calcineurin could protect the H202-induced stem cell apoptosis. The possible mechanisms may be related to the decreased apoptosis rate, increase cell activity and down -regulation of the release of cytochrome C.