格列苯脲对活化T细胞凋亡和接触性皮炎的调控作用研究

发布时间：2018-09-02 06:27

【摘要】：T淋巴细胞过度活化和凋亡不足与许多自身免疫疾病息息相关。与能量代谢相关的分子很可能在免疫性疾病的病理过程中发挥重要的作用,但是其具体机制还不清楚。我们在研究中发现格列苯脲的一种新的药理作用机制,即它选择性诱导活化T细胞凋亡,进而抑制T细胞介导的炎症反应。在给予Con A,anti-CD3/anti-CD28或者PMA等刺激时,T细胞会启动它的活化和增殖。格列苯脲对静息状态的T淋巴细胞几乎无影响,但对T细胞活化增殖72小时后的细胞数呈现剂量依赖式的抑制。对T细胞活化过程检测发现,格列苯脲对T细胞活化marker无影响,活化24小时产生的细胞因子水平无影响,但CFSE染色结果显示增殖后的细胞数减少。说明格列苯脲选择性地作用于活化后的T淋巴细胞。我们发现,格列苯脲能诱导活化T淋巴细胞凋亡,表现为上调cleaved-caspases、cleaved-PARP等;增加Fas的表达、下调细胞型FLICE抑制蛋白的表达;引起活化的T细胞的细胞周期阻滞。机制研究发现,格列苯脲对诱导活化T细胞的凋亡的机制与其经典作—抑制ATP敏感性K+通道没有关系,而是通过抑制另一个细胞内重要能量平衡分子—ATP依赖的蛋白激酶(Adenosine 5'-monophosphate(AMP)-activated protein kinase,AMPK)的磷酸化。最后,我们利用T细胞相关动物实验,三硝基氯苯(picryl chloride,PCl)诱发的接触性皮炎小鼠模型,证实了格列苯脲具有治疗免疫性疾病的功能。因此,本论文说明格列苯脲能够选择性地诱导活化T淋巴细胞凋亡并且治疗自身免疫性疾病,同时也提示了抑制细胞内AMPK磷酸化水平可作为自身免疫性疾病的治疗策略。
[Abstract]:Over-activation and insufficient apoptosis of T lymphocytes are closely related to many autoimmune diseases. The molecules related to energy metabolism may play an important role in the pathological process of immune diseases, but its mechanism is not clear. We have found a new pharmacological mechanism of glibenclamide which selectively induces apoptosis of activated T cells and thus inhibits the inflammatory response mediated by T cells. The T cells activated and proliferated after stimulation of Con anti-CD3 / anti-CD28 or PMA. Glibenclamide had little effect on resting T lymphocytes, but showed a dose-dependent inhibition on the number of T lymphocytes after 72 hours of activation and proliferation. The results of T cell activation showed that glibenclamide had no effect on T cell activation marker, and had no effect on cytokines produced after 24 hours of activation, but CFSE staining showed that the number of proliferated cells decreased. The results suggest that glibenclamide selectively acts on activated T lymphocytes. We found that glibenclamide could induce apoptosis of activated T lymphocytes, such as up-regulating cleaved-caspases,cleaved-PARP, increasing the expression of Fas, down-regulating the expression of cell-type FLICE inhibitor protein, and inducing the cell cycle arrest of activated T cells. The mechanism of glibenclamide induced apoptosis of activated T cells was not related to its classic inhibition of ATP sensitive K channels. Instead, it inhibits the phosphorylation of Adenosine 5'-monophosphate (AMP)-activated protein kinase,AMPK, an important energy balance molecule in another cell. Finally, we demonstrated that glibenclamide has the ability to treat immune diseases in contact dermatitis mouse model induced by trinitrochlorobenzene (picryl chloride,PCl). Therefore, this paper suggests that glibenclamide can selectively induce apoptosis of activated T lymphocytes and treat autoimmune diseases. It also suggests that inhibition of intracellular AMPK phosphorylation can be used as a therapeutic strategy for autoimmune diseases.
【学位授予单位】：南京大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R96

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