目的 探討白藜蘆醇對動脈粥樣硬化（AS）中血管鈣化的影響及機制。方法 將雄性健康SD大鼠隨機分為對照組、模型組、白藜蘆醇（5 mg·kg^-1）組，每組10只。造模前ip預給藥21 d，每天1次。模型組、白藜蘆醇組均制備AS模型：sc 5 mg·kg^-1維生素D₃5 d后，分離并結扎左頸動脈，0.12 mmol·L^-1的CaCl₂溶液濕敷30 min，隨即縫合，對照組濕敷生理鹽水。造模后繼續(xù)給藥，1周后所有大鼠同時禁食24 h后處死，取大鼠左頸動脈進行蘇木精-伊紅（HE）染色和Von Kossa染色。體外鈣化培養(yǎng)基誘導CRL-1999細胞鈣化，給予白藜蘆醇（10μmol·L^-1）干預，茜素紅S染色檢測鈣鹽沉積；實時熒光定量PCR （qRT-PCR）和Western blotting法檢測鈣化指標骨形態(tài)發(fā)生蛋白-2（BMP2）、侏儒相關轉(zhuǎn)錄因子2（Runx2），炎性小體NOD樣受體熱蛋白結構域相關蛋白3（NLRP3），細胞焦亡相關指標天冬氨酸蛋白水解酶-1（Caspase-1）、Gasdermin-D （GSDMD）、白細胞介素（IL）-1β、IL-18蛋白和mRNA表達變化；免疫熒光檢測Runx2、NLRP3、Caspase-1、GSDMD、IL-1β蛋白表達；分子對接驗證白藜蘆醇與靶蛋白NLRP3、Caspase-1、GSDMD、BMP2、Runx2結合活性。結果 HE與VonKossa染色顯示模型組血管壁結構紊亂、鈣鹽沉積，白藜蘆醇緩解鈣鹽沉積。體外實驗表明鈣化結節(jié)可被白藜蘆醇緩解；與對照組比較，模型組BMP2、Runx2、NLRP3、Caspase-1、GSDMD、IL-1β、IL-18 mRNA和蛋白表達均顯著上升（P<0.05、0.01、0.001）；經(jīng)白藜蘆醇處理后，上述指標的mRNA和蛋白表達均顯著下降（P<0.05、0.01、0.001）。白藜蘆醇與Runx2、NLRP3分別形成2、3個氫鍵，對接活性較好。結論 白藜蘆醇抑制AS中血管鈣化，機制可能與抑制NLRP3/Caspase-1信號通路，進而抑制細胞焦亡有關。

Objective To investigate the effect of resveratrol on vascular calcification in atherosclerosis(AS) and its mechanism.Methods Male healthy SD rats were randomly divided into control group, model group and resveratrol(5 mg·kg^-1) group, with 10 rats in each group.Resveratrol was ip preadministered for 21 days, once a day, before modeling.AS model was prepared in the model group and the resveratrol group:sc 5 mg·kg^-1 vitamin D3 for five days, the left carotid artery was isolated and ligated, and0.12 mmol·L^-1 CaCl₂ solution was wet applied for 30 min, and then sutured.The control group was wet applied normal saline.One week later, all rats were sacrificed after fasting for 24 h at the same time.The left carotid arteries of the rats were taken for HE staining and Von Kossa staining.The calcification of CRL-1999 cells was induced by calcification medium in vitro, and resveratrol(10 μmol·L^-1) was administered.Alizarin red S staining was used to detect calcium salt deposition.The expression of calcium markers BMP2, Runx2, inflammasome NLRP3, pyroptosis related indicators aspartic acid proteolytic enzyme 1(Caspase-1), Gasdermin-D(GSDMD), interleukin(IL)-1β and IL-18 were detected by Quantitative real-time PCR(qRT-PCR) and Western blotting.The expressions of Runx2, NLRP3, Caspase-1, GSDMD and IL-1β were detected by immunofluorescence.The binding activity of resveratrol to target protein was verified by molecular docking.Results HE and Von Kossa staining showed vascular wall structure disorder and calcium salt deposition in the model group, which was alleviated by resveratrol pretreatment.VSMCs in vitro showed that calcified nodules could be alleviated by resveratrol.The expressions of BMP2, Runx2, inflammasome NLRP3, Caspase-1, GSDMD, IL-1β and IL-18 in model group were increased(P<0.05, 0.01 and 0.001).After resveratrol treatment, the mRNA and protein expressions of the above indexes were decreased(P<0.05, 0.01 and 0.001).Resveratrol formed two and three hydrogen bonds with Runx2 and NLRP3, respectively, showing good docking activity.Conclusion Resveratrol inhibits vascular calcification in AS, and the mechanism may be related to the inhibition of NLRP3/Caspase-1 signaling pathway and the inhibition of pyroptosis.

R285.5

國家自然科學基金面上項目（8207131747）；湖北省衛(wèi)生健康委中醫(yī)藥科研立項項目（ZY2021F007）