[Mechanism of electroacupuncture on improving insulin resistance and intestinal mucosal barrier injury via the GLP-1R/PKA signaling pathway in type 2 diabetes mellitus rats].

Abstract

<p>OBJECTIVE: To observe the effect of electroacupuncture (EA) on the glucagon-like peptide-1 receptor (GLP-1R)/protein kinase A (PKA) signaling pathway in the colon tissue of type 2 diabetes mellitus (T2DM) rats and explore the mechanisms underlying EA's improvement of insulin resistance and intestinal mucosal barrier injury. METHODS: Among 24 SPF-grade male Wistar rats, 6 rats were randomly assigned to a blank group, and the remaining 18 rats were used to establish the T2DM model through a combination of high-sucrose/high-fat diet feeding and intraperitoneal injection of streptozotocin. Successfully modeled rats were randomly assigned to a model group, an EA group, and an EA+inhibitor group, with 6 rats per group. The EA group received EA at bilateral "Tianshu" (ST25), "Shangjuxu" (ST37), "Pishu" (BL20), and "Weiwanxiashu" (EX-B3); EA was applied to ipsilateral "Shangjuxu" (ST37) and "Weiwanxiashu" (EX-B3) using disperse-dense wave (2 Hz/50 Hz, 1-3 mA) for 20 min per session. The EA+inhibitor group received the same EA treatment combined with intraperitoneal injection of the GLP-1 receptor inhibitor Exenatide. All treatments were administered once daily, six times per week, for 6 weeks. Fasting blood glucose (FBG) was measured before modeling and before/after intervention. After the intervention, serum fasting insulin (FINS) and GLP-1 levels were measured by ELISA, and the homeostasis model assessment of insulin resistance (HOMA-IR) was calculated. Colon morphology was observed using HE staining. Positive expression of zonula occludens-1 (ZO-1) and Occludin in colon tissue was detected by immunohistochemistry. Protein expression of GLP-1R, PKA, GLP-1, ZO-1, and Occludin in colon tissue was analyzed by Western blot. RESULTS: After the intervention, compared with the blank group, the model group showed significantly increased FBG, FINS, and HOMA-IR (P&lt;0.01), while serum GLP-1 level was significantly decreased (P&lt;0.01); histologically, the colonic mucosal epithelial cells exhibited shedding, crypt atrophy was observed, the number of goblet cells was markedly reduced, and inflammatory cell infiltration was evident in the lamina propria; the positive expression of ZO-1 and Occludin in colon tissue was significantly reduced (P&lt;0.01, P&lt;0.05), and the protein expression of GLP-1R, PKA, GLP-1, ZO-1, and Occludin was also significantly decreased (P&lt;0.05, P&lt;0.01). Compared with the model group and the EA+inhibitor group, the EA group showed significantly lower levels of FBG, FINS, and HOMA-IR (P&lt;0.01), along with higher serum GLP-1 level (P&lt;0.01); the inflammatory cell infiltration in the colon was reduced, the number of goblet cells was increased, and the structure of the mucosal epithelium and crypts was more normalized; the positive expression of ZO-1 and Occludin in colon tissue was significantly increased (P&lt;0.01, P&lt;0.05), and the protein expression of GLP-1R, PKA, GLP-1, ZO-1, and Occludin was also significantly elevated (P&lt;0.05, P&lt;0.01). CONCLUSION: EA could reduce blood glucose levels, improve insulin resistance, and repair intestinal mucosal barrier injury in T2DM rats. The underlying mechanism may be related to the activation of the GLP-1R/PKA signaling pathway. 目的：观察电针对2型糖尿病（T2DM）大鼠结肠胰高血糖素样肽-1受体（GLP-1R）/蛋白激酶A（PKA）信号通路的影响，探讨电针改善T2DM胰岛素抵抗及肠黏膜屏障损伤的相关机制。 方法：24只SPF级雄性Wistar大鼠中，随机选取6只作为空白组，剩余18只采用高糖高脂饲料喂养联合链脲佐菌素腹腔注射制备T2DM大鼠模型，将造模成功的大鼠随机分为模型组、电针组与电针+抑制剂组，每组6只。电针组予电针干预，穴取双侧“天枢”“上巨虚”“脾俞”“胃脘下俞”，同侧“上巨虚”“胃脘下俞”连接电针仪，选择疏密波，频率2 Hz/50 Hz，电流1~3 mA，每次20 min。电针+抑制剂组予电针联合腹腔注射GLP-1R抑制剂艾塞那肽，电针干预同电针组。均每日1次，每周6次，干预6周。检测各组大鼠造模前及干预前后空腹血糖（FBG）。干预后，采用ELISA法检测大鼠血清空腹胰岛素（FINS）及胰高血糖素样肽-1（GLP-1）含量，并计算胰岛素抵抗指数（HOMA-IR）；HE染色法观察大鼠结肠组织形态；免疫组化法检测大鼠结肠闭锁小带蛋白-1（ZO-1）、闭锁蛋白（Occludin）阳性表达；Western blot法检测大鼠结肠GLP-1R、PKA、GLP-1、ZO-1、Occludin蛋白表达。 结果：干预后，与空白组比较，模型组大鼠FBG、血清FINS含量、HOMA-IR升高（P&lt;0.01），血清GLP-1含量降低（P&lt;0.01），结肠黏膜上皮细胞脱落，隐窝萎缩，杯状细胞数量明显减少，固有层可见炎性细胞浸润，结肠ZO-1、Occludin阳性表达降低 （P&lt;0.01，P&lt;0.05），结肠GLP-1R、PKA、GLP-1、ZO-1、Occludin蛋白表达降低（P&lt;0.05，P&lt;0.01）；与模型组和电针+抑制剂组比较，电针组大鼠FBG、血清FINS含量、HOMA-IR降低（P&lt;0.01），血清GLP-1含量升高 （P&lt;0.01），结肠炎性细胞浸润减轻，杯状细胞数量及黏膜上皮、隐窝结构趋于正常，结肠ZO-1、Occludin阳性表达升高（P&lt;0.01，P&lt;0.05），结肠GLP-1R、PKA、GLP-1、ZO-1、Occludin蛋白表达升高（P&lt;0.05，P&lt;0.01）。 结论：电针可以降低T2DM大鼠血糖水平，改善胰岛素抵抗，修复肠黏膜屏障损伤，其机制可能与激活GLP-1R/PKA信号通路有关。.</p>