中药药理与临床

目的：探讨八味如意丸对坐骨神经痛大鼠的作用及其对环状鸟苷酸-腺苷酸合成酶/刺激物分子表达基因（cGAS/STING）信号通路的调节作用。方法：选用48只SPF级SD雄性大鼠，随机分为假手术对照组、模型对照组、吲哚美辛7.5 mg/kg组、八味如意丸720 、360 mg/kg组以及八味如意丸720 mg/kg联合2，5-己酮可可碱（DMXAA）25 mg/kg组。除假手术对照组外，造模大鼠通过坐骨神经环扎建立坐骨神经痛模型，给药大鼠灌胃给予相应的药物处理，持续14 d。通过冯·佛雷（Von Frey）测试法和热刺激测试评估大鼠的机械缩爪阈值（PWT）和热缩爪潜伏期（PWL）；采用免疫荧光染色（IF）、苏木素-伊红（HE）染色、定量实时聚合酶链反应（qRT-PCR）和蛋白印迹法（Western Blot）等方法检测坐骨神经的病理变化、小胶质细胞数、cGAS/STING信号通路相关mRNA和蛋白表达。结果：与模型对照组相比，八味如意丸720 mg/kg组大鼠的机械缩爪阈值（PWT）从1.6±0.9 g显著提高至20.5±3.7 g（P<0.01），PWL从5.0±1.5 s显著延长至13.0±2.0 s（P<0.01）；坐骨神经组织病理学观察显示，八味如意丸720 mg/kg组大鼠坐骨神经的髓鞘结构破坏明显减轻，神经纤维排列更为紧密有序，炎症细胞浸润显著减少；脊髓小胶质细胞免疫荧光染色结果显示，八味如意丸720 mg/kg组小胶质细胞荧光强度从模型对照组的2.85±0.36显著降低至1.48±0.19（P<0.01）；分子水平检测发现，八味如意丸720 mg/kg组坐骨神经组织中Cgas、Sting、干扰素β（Ifn-β）、CXC趋化因子配体10（Cxcl10）及相关炎症因子肿瘤坏死因子-α（Tnf-α）、白细胞介素-1β（Il-1β）和Il-6的mRNA表达水平分别较模型对照组下调（P<0.01）；同时，cGAS、STING、IFN-β、CXCL10、p-IRF3及p-TBK1的蛋白表达水平分别较模型对照组下调3（P<0.01）。此外，联合DMXAA给药可显著逆转八味如意丸的上述有益效应，表现为PWT和PWL值降低、坐骨神经损伤加重、小胶质细胞活化增加以及cGAS/STING通路相关因子表达的上调（P<0.05或P<0.01），这证实了cGAS/STING通路是八味如意丸发挥治疗作用的关键靶点。结论：八味如意丸可能通过抑制cGAS/STING信号通路以减轻神经炎症反应并改善坐骨神经痛的症状。

Objective: To investigate the therapeutic effects of Bawei Ruyi (八味如意) Pills on sciatica in rats and the regulatory effect on cyclic GMP-AMP synthase (cGAS)/stimulator of interferon genes (STING) signaling pathway. Methods: Forty-eight SPF-grade male Sprague Dawley (SD) rats were randomly allocated into a sham operation group， a model control group， an indomethacin (7.5 mg/kg) group， Bawei Ruyi Pills (720 mg/kg and 360 mg/kg) groups， and a Bawei Ruyi Pills (720 mg/kg) + 5，6-dimethylxanthenone-4-acetic acid (DMXAA， 25 mg/kg) group. The rats were modeled for sciatica via sciatic nerve ligation in other groups except the sham operation group. Rats were administrated with corresponding drugs by gavage for 14 consecutive days. The mechanical paw withdrawal threshold (PWT) and thermal paw withdrawal latency (PWL) were assessed by the Von Frey test and thermal stimulation test， respectively. Pathological changes of the sciatic nerve， microglial counts， and the mRNA and protein levels of cGAS/STING signaling pathway-related molecules were analyzed via immunofluorescence (IF)， hematoxylin-eosin (H&E) staining， qRT-PCR， and Western blotting， respectively. Results: Compared with the model control group， Bawei Ruyi Pills at 720 mg/kg increased the PWT from (1.6±0.9) g to (20.5±3.7) g (P<0.01)， prolonged the PWL from (5.0±1.5) s to (13.0±2.0) s (P<0.01)， alleviated the destruction of myelin sheath structure， with densely and orderly arranged nerve fibers and reduced inflammatory cell infiltration. IF staining of spinal microglia showed that the fluorescence intensity of microglia in the Bawei Ruyi Pills (720 mg/kg) group decreased from 2.85±0.36 in the model control group to 1.48±0.19 (P<0.01). Molecular detection revealed that compared with model control group， the Bawei Ruyi Pills (720 mg/kg) group showed down-regulated mRNA levels of Cgas， Sting， interferon-beta (Ifnβ)， CXC chemokine ligand 10 (Cxcl10)， and related inflammatory factors tumor necrosis factor-alpha (Tnfα)， interleukin-1 beta (Il1β)， and Il6 in the sciatic nerve (P<0.01)， as well as down-regulated protein levels of cGAS， STING， IFN-β， CXCL10， p-IRF3， and p-TBK1 (P<0.01). Furthermore， co-administration with DMXAA significantly reversed the beneficial effects of Bawei Ruyi Pills， manifested as decreased PWT and PWL values， aggravated sciatic nerve injury， increased microglial activation， and up-regulation of cGAS/STING pathway-related factors (P<0.05 or P<0.01)， confirming that the cGAS/STING pathway was a key target for the therapeutic effects of Bawei Ruyi Pills. Conclusion: Bawei Ruyi Pills alleviate neuroinflammation and symptoms of sciatica by suppressing the cGAS/STING signaling pathway.