基于海马小胶质细胞P2X7/NLRP3 信号通路探讨左归降糖解郁方对糖尿病合并抑郁症模型大鼠海马神经突触可塑性作用及机制

王婷婷; 余晓诗; 郭海鹏; 张瑶; 唐璎娟; 杨松; 苏海龙; 邹蔓姝; 韩远山; 王宇红

doi:10.13748/j.cnki.issn1007-7693.20251148

基于海马小胶质细胞P2X7/NLRP3 信号通路探讨左归降糖解郁方对糖尿病合并抑郁症模型大鼠海马神经突触可塑性作用及机制

Exploring the Effects and Mechanisms of Zuogui Jiangtang Jieyu Formula on Hippocampal Synaptic Plasticity in a Rat Model of Diabetes Mellitus Complicated with Depression Based on the Hippocampal Microglial P2X7/NLRP3 Signaling Pathway

摘要

摘要:
目的基于嘌呤能受体P2X7(P2X purinoceptor 7，P2X7)/核苷酸结合寡聚化结构域样受体蛋白3(nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3，NLRP3)轴，探讨左归降糖解郁方改善糖尿病合并抑郁症(diabetes mellitus complicated with depression，DD)模型大鼠神经炎症及增强神经保护作用的相关机制。
方法通过4周高脂饲料饲养联合链脲佐菌素(streptozotocin，STZ)腹腔注射，再辅以4周慢性温和不可预知应激联合孤笼饲养，建立DD大鼠模型，实验分为正常组，模型组，阳性药组(二甲双胍0.18 g·kg⁻¹+氟西汀1.8 mg·kg⁻¹)，左归降糖解郁方低、中、高剂量组和左归降糖解郁方中剂量+P2X7受体特异性拮抗剂亮蓝 G(brilliant blue G，BBG)组。采用旷场试验、强迫游泳试验和Morris水迷宫检测大鼠抑郁样行为和记忆认知功能；ELISA检测血清中去甲肾上腺素(norepinephrine，NE)、5-羟色胺(5-hydroxyteyptamine，5-HT)、多巴胺(dopamine，DA)含量；免疫荧光检测海马P2X7/Iba-1、NLRP3/Iba-1、突触后致密蛋白-95(postsynaptic density protein-95，PSD95)、突触蛋白-1(synapsin1，SYN1)；苏木素-伊红(hematoxylin-eosin，HE)染色和尼氏染色检测海马神经元损伤情况；免疫印迹法检测P2X7/NLRP3轴相关蛋白P2X7、NLRP3、凋亡相关斑点样蛋白(apoptosis-associated spot-like protein，ASC)、胱天蛋白酶-1(Caspase-1)、白细胞介素-1β(interleukin-1β，IL-1β)和脑源性神经营养因子(brain-derived neurotrophic factor，BDNF)的表达情况。
结果与模型组比较，左归降糖解郁方可以改善DD大鼠抑郁行为，增强神经保护作用；显著升高PSD95、SYN1、BDNF表达(P<0.05或P<0.01)，降低P2X7/Iba-1、NLRP3/Iba-1、ASC、Caspase-1和IL-1β表达(P<0.05或P<0.01)。
结论左归降糖解郁方可能通过抑制P2X7/NLRP3通路减轻神经炎症，进而上调PSD95/SYN1/BDNF表达，发挥神经保护作用。

Abstract:
OBJECTIVE To explore the related mechanisms of Zuogui Jiangtang Jieyu formula in ameliorating neuroinflammation and enhancing neuroprotective effects in model rats with diabetes mellitus complicated with depression(DD) based on the P2X purinoceptor 7(P2X7)/nucleotide-binding oligomerization domain-like receptor protein 3(NLRP3) axis.
METHODS A DD rat model was established by feeding high-fat diet for 4 weeks combined with intraperitoneal injection of streptozotocin(STZ), followed by 4-week chronic unpredictable mild stress combined with single-housing. The rats were divided into normal group, model group, positive drug group(metformin 0.18 g·kg⁻¹+fluoxetine 1.8 mg·kg⁻¹), low-, medium-, and high-dose groups of Zuogui Jiangtang Jieyu formula, as well as medium-dose Zuogui Jiangtang Jieyu formula+P2X7 receptor specific antagonist(brilliant blue G, BBG) group. The open field experiment, forced swimming experiment, and Morris water maze were used to detect depression like behavior and memory cognitive function in rats; ELISA was used to detect the levels of norepinephrine(NE), 5-hydroxytryptamine(5-HT), and dopamine(DA) in serum; immunofluorescence detection of hippocampal P2X7/Iba-1, NLRP3/Iba-1, postsynaptic density protein-95(PSD95), and synapsin-1(SYN1); hematoxylin eosin(HE) staining and Nissl staining were used to detect hippocampal neuronal damage; Western blotting was used to detect the expression levels of proteins related to the P2X7/NLRP3 axis, including P2X7, NLRP3, apoptosis-associated spot-like protein(ASC), Caspase-1, interleukin-1β(IL-1β) and brain-derived neurotrophic factor(BDNF).
RESULTS Compared with the model group, Zuogui Jiangtang Jieyu formula could improve depressive behaviors and enhance neuroprotective effects in DD rats; it significantly upregulated the expression levels of PSD95, SYN1, and BDNF(P<0.05 or P<0.01), and decreased the expression levels of P2X7/Iba-1, NLRP3/Iba-1, ASC, Caspase-1, and IL-1β(P<0.05 or P<0.01).
CONCLUSION Zuogui Jiangtang Jieyu formula may alleviate neuroinflammation by inhibiting the P2X7/NLRP3 pathway, thereby upregulating the expression of PSD95/SYN1/BDNF and exerting neuroprotective effects.

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