虎杖苷调控JAK-STAT3信号通路对心肌缺血再灌注损伤的保护作用及机制
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摘要
目的研究虎杖苷对氧糖剥夺/再复氧(oxygen-glucosed/deprivation,OGD/R)条件下对心肌细胞的保护作用及机制研究。方法 H9C2细胞在常氧和OGD/R条件下培养,采用20、40、80μmol/L虎杖苷和渥曼青霉素(Wortmannin)处理。细胞计数试剂盒(cell counting kit,CCK)-8检测H9C2细胞在不同条件下的存活能力,酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)检测培养细胞上清肿瘤坏死因子(tumor necrosis factor,TNF-α)和白细胞介素6(interleukin-6,IL-6)的浓度,蛋白免疫印迹(Western Blot)检测总STAT3、JAK、磷酸化STAT3和磷酸化JAK的蛋白浓度。结果 (1)虎杖苷明显改善了OGD/R条件下细胞的存活能力,并且抑制了OGD/R条件下诱导产生的TNF-α和IL-6的分泌;(2)虎杖苷诱导H9C2细胞JAK-STAT3信号通路;(3)抑制JAK-STAT3信号通路会抵消虎杖苷抗炎和促进细胞生存能力的作用。结论 OGD/R条件下,虎杖苷提高了H9C2细胞的存活率,同时其可以通过磷酸化JAK和STAT3蛋白表达激活JAK-STAT3信号通路,从而对心肌缺血再灌注损伤起到保护作用。
Objectives To study the protective effect and mechanism of polydatin on myocardial cells under oxygen glucose deprivation/reoxygenation(OGD/R)conditions. Methods H9C2 cells were cultured under normoxia and OGD/R conditions and treated with 20,40,80 μmol/L of polydatin and wortmannin. Cell counting kit(CCK)-8 was used to detect the survival ability of H9C2 cells under different conditions. The concentration of tumor necrosis factor(TNF)-αand interleukin-6(IL-6)were detected by enzyme-linked immunosorbent assay(ELISA). The total STAT3,JAK and phosphorylation of JAK and STAT3 protein expression were detected by Western Blot(WB). Results(1)Polydatin significantly improved the viability of cells under OGD/R conditions and inhibited the secretion of TNF-α and IL-6 induced by OGD/R conditions.(2)JAK-STAT3 signaling path way was induced by polydatin.(3)Inhibition of JAKSTAT3 signaling pathway by Hooks in the H9C2 cells counteracted the anti-inflammatory and cell viability of polydatin.Conclusions Under OGD/R conditions,polydatin increases the survival rate of H9C2 cells,and can activate JAKSTAT3 signaling through phosphorylation of JAK and STAT3 protein expression,thus protecting myocardial ischemia/reperfusion injury.
Objectives To study the protective effect and mechanism of polydatin on myocardial cells under oxygen glucose deprivation/reoxygenation(OGD/R)conditions. Methods H9C2 cells were cultured under normoxia and OGD/R conditions and treated with 20,40,80 μmol/L of polydatin and wortmannin. Cell counting kit(CCK)-8 was used to detect the survival ability of H9C2 cells under different conditions. The concentration of tumor necrosis factor(TNF)-αand interleukin-6(IL-6)were detected by enzyme-linked immunosorbent assay(ELISA). The total STAT3,JAK and phosphorylation of JAK and STAT3 protein expression were detected by Western Blot(WB). Results(1)Polydatin significantly improved the viability of cells under OGD/R conditions and inhibited the secretion of TNF-α and IL-6 induced by OGD/R conditions.(2)JAK-STAT3 signaling path way was induced by polydatin.(3)Inhibition of JAKSTAT3 signaling pathway by Hooks in the H9C2 cells counteracted the anti-inflammatory and cell viability of polydatin.Conclusions Under OGD/R conditions,polydatin increases the survival rate of H9C2 cells,and can activate JAKSTAT3 signaling through phosphorylation of JAK and STAT3 protein expression,thus protecting myocardial ischemia/reperfusion injury.
引文
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[4]CHEN B P,WOLFGANG C D,HAI T.Analysis of ATF3,a transcription factor induced by physiological stresses and modulated by gadd153/Chop10[J].Mol Cell Biol,1996,16(3):1157-1168.
[5]NOBORI K,ITO H,TAMAMORI-ADACHI M,et al.ATF3inhibits doxorubicin-induced apoptosis in cardiac myocytes:a novel cardioprotective role of ATF3[J].J Mol Cell Cardiol,2002,34(10):1387-1397.
[6]BLYDT-HANSEN T D,KATORI M,LASSMAN C,et al.Gene transfer-induced local heme oxygenase-1 overexpression protects rat kidney transplants from ischemia/reperfusion injury[J].J Am Soc Nephrol,2003,14(3):745-754.
[7]ALAM J,KILLEEN E,GONG P,et al.Heme activates the heme oxygenase-1 gene in renal epithelial cells by stabilizing Nrf2[J].Am J Physiol Renal Physiol,2003,284(4):F743-F752.
[8]MIYAKE S,MAKIMURA M,KANEGAE Y,et al.Efficient generation of recombinant adenoviruses using adenovirus DNA-terminal protein complex and a cosmid bearing the full-length virus genome[J].Proc Natl Acad Sci U S A,1996,93(3):1320-1324.
[9]YOSHIDA T,TANG S S,HSIAO L L,et al.Global analysis of gene expression in renal ischemia-reperfusion in the mouse[J].Biochem Biophys Res Commun,2002,291(4):787-794.
[10]SUPAVEKIN S,ZHANG W,KUCHERLAPATI R,et al.Differential gene expression following early renal ischemia/reperfusion[J].Kidney Int,2003,63(5):1714-1724.
[11]葛鹏,杨波,李小飞,等.蛇床子素在大鼠肠缺血再灌注肺损伤模型中的肺保护机制研究[J].现代生物医学进展,2013,13(28):5401-5404.
[12]XU X,LI J,SUN X,et al.Tumor suppressor NDRG2 inhibits glycolysis and glutaminolysis in colorectal cancer cells by repressing c-Myc expression[J].Oncotarget,2016,6(28):26161-26176.
[13]LI Y,YANG J,LI S,et al.N-myc downstream-regulated gene2,a novel estrogen-targeted gene,is involved in the regulation of Na+/K+-ATPase[J].J Biol Chem,2011,286(37):32289-32299.
[14]COBELENS P M,VAN PUTTE B P,KAVELAARS A,et al.Inflammatory consequences of lung ischemia-reperfusion injury and low-pressure ventilation[J].J Surg Res,2017,153(2):295-301.
[15]WU G,WILSON G,GEORGE J,et al.Modulation of Notch signaling as a therapeutic approach for liver cancer[J].Curr Gene Ther,2015,15(2):171-181.
[16]GEISLER F,STRAZZABOSCO M.Emerging roles of Notch signaling in liver disease[J].Hepatology,2015,61(1):382-392.
[17]朱玉菡,沈晓旭,韩晴晴,等.参附注射液辅助治疗心肌梗死伴心力衰竭临床疗效的Meta分析[J].中国循证心血管医学杂志,2018,10(4):402-406.
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[19]陈恒星.舒芬太尼对大鼠心肌缺血再灌注心肌组织超氧化物歧化酶活性及丙二醛水平的影响[J].新乡医学院学报,2015,29(3):180-181.
[20]刘爱华,石孟琼,杨文雁,等.珠子参总皂甙对大鼠心肌缺血/再灌注损伤的保护作用及机制研究[J].中国临床药理学与治疗学,2013,18(11):1224-1232.