Tongqiao Mingmu formula alleviates retinal ganglion cell autophagy through PI3K/AKT/mTOR pathway

Glaucoma is a severe blindness-causing optic nerve disease characterized by a loss of retinal ganglion cells (RGCs). Previous studies have shown that the Tongqiao Mingmu (TQMM) formula can reduce retinal and optic nerve damage, but its mechanism of action requires further elucidation. In this study, an RGC injury model was prepared using glutamate and then treated with serum-containing drug from the TQMM formula (hereafter called "TQMM formula serum"). In the glutamate-induced RGC injury model, cell viability decreased with an increase in glutamate concentration, whereas the expression of autophagy-related biomarkers LC3 and Belicin-1 increased. An adenovirus transfection experiment revealed that glutamate markedly promoted autophagic flux in RGCs. Notably, TQMM formula serum inhibited the expression of autophagy-related biomarkers, reduced autophagy flux, and reversed the damage caused by glutamate to RGCs. Furthermore, the PI3K inhibitor LY294002 was used to intervene in the RGC autophagy model and was found to suppress the PI3K/AKT/mTOR pathway and enhance RGC autophagy. However, TQMM formula serum could generate an opposite effect and upregulate the expressions of the PI3K/AKT/mTOR pathway genes and proteins. In conclusion, the TQMM formula can prevent glutamate-induced autophagy in RGCs, possibly by activating the PI3K/AKT/mTOR pathway and reducing the expression of autophagy-related biomarkers LC3 and Belicin-1 to attenuate autophagy and maintain RGC viability.青光眼是一种严重的致盲性视神经疾病, 具有视网膜节细胞 (RGCs) 丢失的特点。前期研究发现通窍明目方能够减轻视网膜和视神经损害, 但其作用机制仍有待进一步阐明。本研究采用谷氨酸制备 RGC 损伤模型, 并通过含有通窍明目方的血清 (以下称 "通窍明目方血清") 处理。在谷氨酸诱导的 RGC 损伤模型中, 我们观察到细胞活性随谷氨酸浓度的增加而降低, 而自噬相关指标LC3 和 Belicin-1 的表达则升高。腺病毒转染实验显示, 谷氨酸明显地促进了 RGCs 的自噬通量。值得注意的是, 通窍明目方血清能够抑制 RGCs 自噬相关指标的表达, 减少 RGCs 的自噬通量, 并逆转谷氨酸对 RGCs 的伤害。随后, 采用 PI3K 通路抑制剂 LY294002 干预 RGC 自噬模型, 发现其能够抑制 PI3K/AKT/mTOR 通路, 增强了RGC的自噬。然而, 通窍明目方血清可以产生与 LY294002 相反的效果, 它可以上调 PI3K/AKT/mTOR 基因和蛋白的表达。综上所述, 通窍明目方能够防止谷氨酸诱导的 RGCs 自噬, 其作用可能是通过激活 PI3K/AKT/mTOR 信号通路, 减少自噬相关指标 LC3 和 Belicin-1 的表达, 从而减轻 RGCs 自噬, 维持 RGC 细胞活力。.