目的 探究槲皮素通過調(diào)節(jié)沉默信息調(diào)控因子1(SIRT1)/NOD樣受體蛋白3(NLRP3)信號通路對實驗性巨結(jié)腸大鼠腸損傷的影響。方法 采用苯扎溴銨滴注法誘導(dǎo)建立大鼠實驗性巨結(jié)腸模型,并隨機分為模型組,槲皮素低(5 mg/kg)、高(50 mg/kg)劑量組, SIRT1抑制劑(EX527)組,槲皮素+EX527組,每組15只,另取15只作為假手術(shù)組。各組大鼠連續(xù)給藥2周, 1次/d,末次給藥12 h后,觀察大鼠排便和腹脹情況,開腹觀察結(jié)腸變化。免疫熒光法檢測肽能神經(jīng)遞質(zhì)蛋白基因產(chǎn)物9.5(PGP9.5)、M2型巨噬細胞標(biāo)記抗體-CD206表達水平。蘇木精-伊紅(HE)觀察結(jié)腸病理變化,免疫組化法檢測乙酰膽堿酯酶(AChE)、NLRP3陽性表達變化。Western blotting檢測結(jié)腸組織SIRT1、NLRP3、炎性因子[腫瘤壞死因子(TNF)-α、白細胞介素(IL)-1β]、促神經(jīng)恢復(fù)相關(guān)蛋白[骨形成蛋白-2(BMP-2)、骨形成蛋白受體(BMPR) ]、神經(jīng)節(jié)細胞相關(guān)因子[乙酰膽堿(ACh)、神經(jīng)巢蛋白(Nestin) ]表達。結(jié)果 與假手術(shù)組相比,模型組大鼠處理段結(jié)腸組織神經(jīng)節(jié)細胞減少、結(jié)腸黏膜炎癥損傷加重, SIRT1以及M2型巨噬細胞介導(dǎo)的抗炎和促神經(jīng)恢復(fù)蛋白表達降低, NLRP3炎性反應(yīng)升高(P<0.05)。與模型組相比,槲皮素5、50 mg/kg組大鼠結(jié)腸組織炎癥損傷緩解, SIRT1以及M2型巨噬細胞介導(dǎo)的抗炎和促神經(jīng)恢復(fù)蛋白表達升高, NLRP3炎性反應(yīng)降低(P<0.05),且槲皮素高劑量組改善效果優(yōu)于低劑量組(P<0.05);EX527組可減弱槲皮素的上述作用(P<0.05)。結(jié)論 槲皮素可通過上調(diào)SIRT1表達,抑制NLRP3促炎反應(yīng),提高M2型巨噬細胞介導(dǎo)的抗炎和促神經(jīng)恢復(fù)作用,改善實驗性巨結(jié)腸大鼠結(jié)腸炎性損傷。

Objective To investigate the effect of quercetin on intestinal injury in experimental hirscholon rats through regulating silencing message modulator 1 (SIRT1)/Nod-like receptor protein 3 (NLRP3) signaling pathway.Methods Benzalkonium bromide infusion was used to induce the establishment of rat experimental megacolon models, and they were randomly divided into model group, quercetin low(5 mg/kg) and high (50 mg/kg) dose groups, SIRT1 inhibitor (EX527) group, quercetin + EX527 group, 15 per group, another 15 rats in the sham operation group. Rats in each group were administered for 2 weeks, once daily, 12 h after the last administration, the rat's defecation and abdominal distension were observed, and the abdomen was opened to observe changes in the colon. Immunofluorescence method was used to detect the expression levels of peptidergic neurotransmitter protein gene product 9.5 (PGP9.5) and M2 macrophage marker antibody-CD206. HE staining was used to observe the pathological changes of colon, immunohistochemical method was used to detect the positive expression of acetylcholinesterase (AChE) and NLRP3. Western blotting was used to detect the expression of SIRT1, NLRP3, tumor necrosis factor (TNF)-α, interleukin (IL)-1β, neurorestoration-related protein (BMP-2, BMPR), acetylcholine (ACh) and Nestin in colon tissues.Results Compared with the sham operation group, the rats in the model group had fewer ganglion cells in the treated colon tissue and more severe colonic mucosal inflammation, the activity of SIRT1 and the expression of anti-inflammatory and neurorestorative proteins mediated by M2 macrophages decreased, and the inflammatory response of NLRP3 increased (P < 0.05). Compared with the model group, the inflammation in the colon tissue of rats in quercetin 5 and 50 mg/kg groups was alleviated, the activity of SIRT1 and the expression of anti-inflammatory and neurorestorative proteins mediated by M2 macrophages increased, and the inflammatory response of NLRP3 decreased (P < 0.05), and the improvement effects of the high-dose quercetin group were better than those of the low-dose group (P < 0.05). EX527 could attenuate the abovementioned effects of quercetin (P < 0.05).Conclusion Quercetin can up-regulate the expression of SIRT1, inhibit the proinflammatory response of NLRP3, enhance the anti-inflammatory and neurorestorative effects mediated by M2 macrophages, and improve the intestinal inflammatory injury in experimental hirscholon rats.

R285.5

河南省醫(yī)學(xué)科技攻關(guān)計劃聯(lián)合共建項目(LHGJ20200906)