基于NF-κB/NLRP3/Caspase-1信号轴探究白藜芦醇对痛风性肾病模型大鼠肾脏的保护作用机制

目的 基于NF-κB/NLRP3/Caspase-1信号轴探究白藜芦醇对痛风性肾病模型大鼠的肾脏的保护作用机制。方法 将60只SD雄性大鼠随机分为对照组、模型组、秋水仙碱（阳性对照，0.03 mg · kg^-1）组和白藜芦醇高、中、低剂量（1 000、500、250 mg·kg^-1）组，连续7 d ig给药，给药过程中，除对照组外，其余各组使用氧嗪酸钾合并尿酸钠的方法制备大鼠痛风性肾病模型。ELISA法检测大鼠血清中白细胞介素（IL）-1β、IL-18、尿酸、肌酐（SCr）、尿素氮（BUN）水平，肾脏组织匀浆中肿瘤坏死因子-α（TNF-α）、单核细胞趋化蛋白-1（MCP-1）、环氧合酶-2（COX-2）水平；HE、Masson染色观察肾脏组织细胞形态变化；PAS染色检测大鼠肾组织中肾小球损伤情况，TUNEL观察肾脏组织细胞DNA损伤情况；实时荧光定量PCR （qRT-PCR）、免疫组化法检测肾脏组织中核因子-κB （NF-κB）、NOD样受体热蛋白结构域相关蛋白3（NLRP3）、半胱氨酸蛋白酶-1（Caspase-1） mRNA以及蛋白的表达量，最后采用分子对接研究白藜芦醇与NF-κB、NLRP3、Caspase-1的结合情况。结果 与模型组比较，秋水仙碱组及白藜芦醇各给药组大鼠血清中IL-1β、IL-18、SCr、BUN及肾脏TNF-α、MCP-1、COX-2水平显著降低（P<0.01），白藜芦醇高剂量组尿酸、中和高剂量组BUN显著降低（P<0.05）；白藜芦醇各剂量组不同程度降低肾组织中胶原纤维化面积、肾小球阳性率以及肾组织细胞TUNEL染色阳性率，减缓病理损伤情况，其中高剂量组作用最显著（P<0.05）；qRT-PCR、免疫组化结果表明，白藜芦醇各给药组均抑制肾脏组织细胞中NF-κB、NLRP3、Caspase-1mRNA和蛋白的表达，其中高剂量组作用最显著（P<0.05）；分子对接结果进一步表明，白藜芦醇与NF-κB、NLRP3、Caspase-1结合状态良好，即白藜芦醇对NF-κB、NLRP3、Caspase-1具有良好的靶向调控作用。结论 白藜芦醇对痛风性肾病模型大鼠的肾脏保护作用可能为抑制NF-κB信号通路，进而抑制NLRP3的激活从而阻断Caspase-1招募IL-1β、IL-18，降低其分泌，遏制肾脏细胞程序性死亡的初始阶段细胞焦亡的发生，从而逆转痛风性肾病大鼠肾组织的炎症损伤。

Exploring protective mechanism of resveratrol on rat kidney in gouty nephropathy model based on NF-κB/NLRP3/Caspase-1 signaling axis

Objective Exploring the protective mechanism of resveratrol on rat kidney in gouty nephropathy model based on NF-κB/NLRP 3/Caspase-1 signaling axis. Method Sixty male SD rats were randomly divided into control group, model group, colchicine (positive control, 0.03 mg· kg^-1) group, and resveratrol high, medium, and low dose (1 000, 500, 250 mg· kg^-1) group. The rats were administered ig for seven consecutive days. During the administration process, except for the control group, the other groups were prepared with potassium oxazinate combined with sodium urate to create a rat model of gouty nephropathy. Level of interleukin (IL)-1β, IL-18, uric acid, creatinine (SCr), urea nitrogen (BUN) in serum, and tumor necrosis factor-α (TNF-α), monocyte chemotactic protein-1 (MCP-1) and cyclooxygenase-2 (COX-2) levels in renal tissue homogenate were detected by ELISA method. The morphological changes of kidney tissues were observed by HE and Masson staining. The glomerular injury in rat renal tissue was detected by PAS staining. The DNA damage in kidney tissue cells was observed by TUNEL. The expression levels of NF-κB, NLRP3, Caspase-1 mRNA and protein in kidney tissues were detected by qRT-PCR and immunohistochemistry. Finally, molecular docking technology was used to study the combination between resveratrol and NF-κB, NLRP3, Caspase-1. Result Compared with model group, the levels of IL-1β, IL-18, SCr, and BUN in the serum and and TNF-α, MCP-1 and COX-2 in renal of rats treated with colchicine and resveratro were significantly reduced (P< 0.01). While uric acid in the high-dose group of resveratrol, and BUN in medium and high dose group showed a significant decrease (P< 0.05). Each dose group of resveratrol reduced the collagen fibrosis area, glomerular positivity rate, and TUNEL staining positivity rate of renal tissue cells to varying degrees, slowing down pathological damage, and the high-dose group had the most significant effect among them (P< 0.05). qRT-PCR and immunohistochemistry results showed that all treatment groups of resveratrol inhibited NF-κB, NLRP3, and Caspase-1 mRNA and protein in renal tissue cells, with the high-dose group having the most significant effect (P< 0.05). The molecular docking results further indicate that resveratrol interacts with NF-κB. The binding state of NLRP3 and Caspase-1 is good, indicating that resveratrol has an effect on NF-κB. NLRP3 and Caspase-1 have good targeted regulatory effects. The molecular docking results further indicated that, resveratrol binds well to NF-κB, NLRP3, and Caspase-1, resveratrol showed good targeted regulation of NF-κB, NLRP3 and Caspase-1. Conclusion The renal protective effect of resveratrol in GN model of gouty arthritis with hyperuricemia is related to the inhibition of inflammatory factor secretion. Its anti-inflammatory mechanism may be through the inhibition of NF-κB signaling pathway, and then NLRP3 activation to block Caspase-1 recruitment of IL-1 IL-1 and IL-18, reduce the secretion of inflammatory factors such as IL-1β and IL-18, and curb the occurrence of cell pyroptosis in the initial stage of programmed renal cell death, thus reversing the inflammatory injury of renal tissue in GN.