NGF通过抑制内质网应激途径保护SCs在高糖环境诱导的凋亡

已知神经生长因子 (nerve growth factor, NGF) 对糖尿病外周神经病变 (diabetic peripheral neuropathy, DPN) 患者具有良好的治疗效果，内质网应激 (endoplasmic reticulum stress, ERS) 在调控DPN 的发生发展方面扮演着重要的角色。然而，二者间的关系未知。本研究以30 mmol/L的高糖处理RSC-96大鼠雪旺细胞 (RSC96 Schwann cells, SCs)，模拟DPN患者外周神经的内环境。研究结果证实，在高糖条件下，NGF通过抑制SCs内 ERS的过度激活进而保护SCs的存活且这种抑制作用依靠P13K/AKT/GSK-3β和ERK1/2两条信号通路的调节实现。MTT检测细胞的存活率，结果显示高糖环境培养的SCs在24 h发生最佳程度的抑制，且此时间点加入的NGF浓度为50 ng/mL 时，其存活率最高。Western 印迹检测ERS和相关蛋白质的表达揭示，高糖能够过度激活SCs内ERS蛋白 (GRP-78、ATF-6、ATF-4、XBP-1、CHOP)，给予 50 ng/mL的NGF处理后，ERS蛋白的表达水平大幅下调并接近正常，且此时p-AKT、p-ERK1/2、p-GSK3β蛋白的检测水平明显升高。流式细胞术检测细胞凋亡显示，NGF能显著抑制SCs的早期凋亡。上述结果证明，高糖诱导SCs的凋亡增加是由于自身的ERS被过度激活，NGF可通过调节P13K/AKT/GSK-3β和ERK1/2两条信号通路来抑制ERS的过度激活，达到保护SCs存活的目的。此机制为临床治疗 DPN 提供新的理论基础。

Nerve Growth Factor Inhibits High Glucose-induced Schwann CellApoptosis by Regulating Endoplasmic Reticulum Stress Pathway

Nerve growth factor (NGF) has been proven to be an effective drug for treating diabetic peripheral neuropathy (DPN). Endoplasmic reticulum stress (ERS) plays a key role in the pathogenesis of DPN. However, the relationship between ERS disorder and NGF in DPN is still unclear. In the present study, we cultured RSC96 Schwann cells (SCs) in the medium containing 30 mmol/L glucose (higher glucose, HG) to imitate the condition of DPN in diabetic patients. Our results demonstrated that NGF decreased the rate of SCs death under HG conditions via moderating ERS, and this inhibitory effect was involved in regulating P13K/AKT/GSK-3β and ERK1/2 pathways. MTT assay for cell viability revealed that the optimal inhibition time of SCs under HG condition occurred at 24 h and the optimum therapeutic concentration of NGF was 50 ng/mL. Western blotting analysis revealed that the high level of ER stress-associated proteins of GRP-78, ATF-6, XBP-1, ATF-4 and CHOP induced by HG were significantly decreased by the treatment of NGF. Meanwhile, combination with NGF also increased expression of p-AKT, p-ERK1/2 and p-GSK-3β. Flow cytometry for apoptosis revealed that NGF decreased SCs apoptosis. Together, these data suggest that the neuroprotective role of NGF in promoting SCs survival under HG condition is related to the inhibition of ERS induced cell death, which was probably induced via the activation of upstream signals P13K/AKT/GSK-3β and ERK1/2. This may provide newinsights for the clinical treatment of diabetic neuropathy.