钠钾ATP酶/Src/Akt/mTOR信号通路介导足细胞损伤在狼疮性肾炎发病机制中的作用

The podocyte injury is the key factor in the pathogenesis of the lupus nephritis (LN), and the treatment depends on the immunosuppressive therapy has many side effects. Podocalyxin can bind with Na/K-ATPase (NKA) to maintain the stability of podocyte structures, and regulating the glomerular filtration barrier. The applicant found that the marinobufagenin (MBG) which was the ligand of the NKA could partly activate the signaling pathway of Src/Akt/mTOR in NKA knockdown mice (NKA+/-), which could modulate the shift from a state of survival to apoptosis, and mediate the apoptosis of cardiac myocyte; others had already proved that Akt/mTOR signaling pathway coulde be activated in LN. From all above, we presumed that the signaling pathway of NKA/Src/Akt/mTOR might mediate the apoptosis of podocyte in LN. In vivo, we will use the NZB/W.F1 and C57BL/6 mice as our animal model, inhibit the signal point of NKA, Src and mTOR in NKA/Src/Akt/mTOR signaling pathway. Then, we will detect the correlation between the signaling pathway of NKA/Src/Akt/mTOR and the podocyte injury or activity of LN; in vitro, we will establish the mice podocyte cell line with NKA knockdown, and detect the regulation mechanism of apoptosis under the MBG stimulation. The aim of our project is to seek a new approach for LN progression, we are looking forward to getting a new medicine for attenuating or replacing the immunosuppressive agent.

足细胞损伤是狼疮性肾炎(LN)发病过程中的关键因素，依赖于免疫抑制剂的治疗副作用大。足细胞标记蛋白可与钠钾ATP酶（NKA）维持足细胞骨架稳定，并调控肾小球滤过屏障。申请者前期研究提示NKA配体-海蟾蜍毒素（MBG）可部分活化NKA+/-小鼠Src/Akt/mTOR信号通路，使得细胞凋亡效应强于细胞存活效应，心肌细胞产生凋亡；既往报道称LN中Akt/mTOR存在信号通路活化；由此我们推测LN中可能存在NKA/Src/Akt/mTOR介导的足细胞凋亡。本课题拟在体内以NZB/W.F1和C57BL/6鼠为研究对象，阻断NKA、Src、mTOR信号位点，检测该信号通路与足细胞损伤、狼疮活动的关系；拟在体外实验中构建NKA基因沉默的小鼠足细胞系，在MBG干预下检测NKA表达不同的足细胞凋亡的调控机制。本研究旨在探索LN疾病进展的新途径，为寻找新型药物替代或减少免疫抑制剂的使用提供理论依据。

足细胞损伤是狼疮性肾炎(LN)发病过程中的关键因素，依赖于免疫抑制剂的治疗副作用大。足细胞标记蛋白可与钠钾ATP酶（NKA）维持足细胞骨架稳定，并调控肾小球滤过屏障。本课题拟在体内以NZB/W.F1和C57BL/6鼠为研究对象，阻断NKA、Src、mTOR信号位点，检测该信号通路与足细胞损伤、狼疮活动的关系；由于哇巴因（ouabain）和海蟾蜍毒素（MBG）均属于洋地黄类物质，MBG的来源由于地域问题无法保障，同时预实验中我们发现小管上皮细胞在疾病的发病机制中具有重要作用，因此实验调整为，在ouabain的干预下检测NKA表达不同的小管上皮细胞细胞凋亡的调控机制。