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目的 :研究ClC-3氯通道在氧糖剥夺所致小胶质细胞表型转化中的作用。方法:应用小胶质细胞株(BV-2)制备氧糖剥夺模型,分别给予氯通道阻断剂DIDS和NPPB预处理BV-2细胞。通过MTT活性测定确定BV-2细胞损伤及药物的有效浓度;通过实时荧光定量PCR法测定细胞表型相关分子,如M1型相关分子[肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白介素-1β(interleukin 1β,IL-1β)、CD86]和M2型相关分子[转化生长因子β(transforming growth factorβ,TGF-β)、CD206]m RNA水平的表达;通过RNA干扰的方法下调ClC-3的表达,再给予氯通道阻断剂DIDS和NPPB干预,进一步检测细胞的MTT活性,观察药物作用效果。结果:预先给予DIDS(1和10μmol/L)和NPPB(1μmol/L)能够改善氧糖剥夺所诱导的BV-2细胞MTT活性下降,抑制M1型相关分子如TNF-α、IL-1β和CD86的表达并促进M2型相关分子(TGF-β和CD206)的表达;通过RNA干扰下调ClC-3的表达,能够消除DIDS和NPPB的作用。结论:ClC-3氯通道参与调控氧糖剥夺所致小胶质细胞的表型转化,阻断ClC-3氯通道抑制氧糖剥夺诱导小胶质细胞向M1型转化。
AIM: To investigate the role of ClC-3 chloride channel in the phenotypic transformation of microglia induced by oxygen-glucose deprivation. Methods: Oxygen-glucose deprivation model was established by using microglia cell line (BV-2), and BV-2 cells were pretreated with chloride channel blockers DIDS and NPPB respectively. The MTT activity was used to determine the BV-2 cell injury and the effective concentration of the drug. The cell phenotype-related molecules such as M1 (tumor necrosis factor-α, TNF-α ), Interleukin 1β (IL-1β), CD86 and M2-related molecules [transforming growth factorβ (TGF-β), CD206] m RNA were detected by RNA interference ClC-3 expression, and then given chloride channel blockers DIDS and NPPB intervention to further test the MTT activity of cells observed the effect of drugs. Results: Pretreatment with DIDS (1 and 10 μmol / L) and NPPB (1 μmol / L) decreased the MTT activity of BV-2 cells induced by oxygen glucose deprivation and inhibited the expression of M1-related molecules such as TNF-α, IL-1β and CD86 And promote the expression of M2-related molecules (TGF-β and CD206). The effect of DIDS and NPPB can be eliminated by down-regulating the expression of ClC-3 by RNA interference. Conclusion: ClC-3 chloride channels are involved in the regulation of microglial phenotypes induced by oxygen-glucose deprivation and ClC-3 chloride channel inhibition of glucose-deprivation induced microglial cells to M1 type transformation.