儿茶素抑制p38 MAPK磷酸化减轻脂多糖诱导的大鼠心肌细胞凋亡、炎症及氧化损伤OACSTPCD
Catechin Alleviates Apoptosis,Inflammation and Oxidative Damage of Rat Cardiomyocytes Induced by Lipopolysaccharide by Inhibiting p38 MAPK Phosphorylation
为了研究儿茶素对脂多糖(LPS)诱导的大鼠心肌细胞(H9C2)氧化损伤、炎症和凋亡的影响以及对p38丝裂原活化蛋白激酶(MAPK)信号通路的调控作用,本研究体外培养H9C2细胞,并将其分为对照组(不做干预处理)、LPS组(10 μg/mL LPS处理)、不同浓度儿茶素组(在LPS组基础上以20、40、80、160 nmol/L儿茶素处理)、SB203580组(10 μg/mL LPS+1 μmol/L SB203580处理)、抑制剂组(10 μg/mL LPS+160 nmol/L儿茶素+1 μmol/L p38 MAPK通路抑制剂SB203580处理)和激活剂组(10 μg/mL LPS+160 nmol/L儿茶素+10 μmol/L p38 MAPK通路激活剂C16-PAF处理).药物干预处理24 h后,用细胞计数试剂盒-8(CCK-8)测定细胞活力;酶联免疫吸附试验(ELISA)检测炎症因子白细胞介素-6(IL-6)和白细胞介素-10(IL-10)的含量;丙二醛(MDA)和超氧化物岐化酶(SOD)试剂盒检测氧化应激因子MDA和SOD在细胞上清液中的表达水平;Hoechst 33258染色法测定细胞凋亡率;蛋白免疫印迹法测定天冬氨酸特异性半胱氨酸蛋白酶-3(Caspase-3)和p38 MAPK通路相关蛋白表达水平.与对照组相比,LPS组细胞活力显著降低(P<0.05);与LPS组相比,添加160 nmol/L儿茶素后细胞活力显著升高(P<0.05),最终选择160 nmol/L儿茶素作为儿茶素组进行后续试验.后续试验中,与对照组相比,LPS组SOD、IL-10含量显著降低(P<0.05),MDA、IL-6含量、细胞凋亡数、Caspase-3、p-p38 MAPK蛋白表达显著升高(P<0.05);与LPS组相比,儿茶素组和SB203580组显著扭转了上述指标的变化(P<0.05);与儿茶素组相比,抑制剂组中的SB203580增强了上述指标的变化,而激活剂组中的C16-PAF则减弱了儿茶素对LPS诱导的H9C2细胞的作用(P<0.05).本研究表明,儿茶素可显著抑制LPS诱导的大鼠心肌H9C2细胞的凋亡、炎症和氧化应激损伤,其作用机制或与抑制p38 MAPK通路的信号转导相关.
To investigate the effects of catechins on oxidative damage,inflammation and apoptosis of rat cardiomyocytes(H9C2)induced by lipopolysaccharide(LPS),as well as the regulation of p38 mitogen-activated protein kinase(MAPK)sig-naling pathway,H9C2 cells were cultured in vitro and divided into the control group which is without intervention,LPS group with 10 μg/mL LPS treatment,catechin with different concentrations(20,40,80,160 nmol/L catechin treatment based on LPS group)and SB203580 group(10 μg/mL LPS+1 μmol/L SB203580 treatment),inhibitor group(10 μg/mL LPS+160 nmol/L cat-echin+1 μmol/L p38 MAPK pathway inhibitor SB203580 treatment)and activator group(10 μg/mL LPS+160 nmol/L catechin+10 μmol/L p38 MAPK pathway activator C16-PAF).Cell viability was measured with cell counting kit-8(CCK-8)after 24 h treatment.The levels of interleukin-6(IL-6)and interleukin-10(IL-10)were detected by enzyme-linked immunosorbent assay(ELISA).Malondialdehyde(MDA)and superoxide dismutase(SOD)kits were used to detect the expression levels of oxidative stress factors MDA and SOD in cell supernatant.The apoptosis rate was determined by Hoechst 33258 staining.The expres-sion levels of Caspase-3 and p38 MAPK pathway-related proteins were determined by Western blot.Compared with the control group,the cell viability of LPS group significantly decreased(P<0.05).Compared with LPS group,the cell viability signifi-cantly increased after 160 nmol/L catechin supplementation(P<0.05).Finally,160 nmol/L catechin group was selected as cat-echin group for follow-up experiment.In the follow-up experiment,compared with the control group,SOD and IL-10 contents in LPS group significantly decreased(P<0.05),MDA and IL-6 contents,apoptosis number,Caspase-3 and P-P38 MAPK pro-tein expression significantly increased(P<0.05);compared with LPS group,catechin group and SB203580 group significant-ly reversed the changes of the above indexes(P<0.05);compared with the catechin group,SB203580 in the inhibitor group enhanced the changes in the above indicators,while C16-PAF in the activator group attenuated the effect of catechin on LPS-induced H9C2 cells(P<0.05).In this study,catechins can significantly inhibit the apoptosis,inflammation and oxidative stress injury of H9C2 cells induced by LPS,and the mechanism of action may be related to the inhibition of p38 MAPK pathway sig-nal transduction.
邢建华;孙鹿璐;李哲贤
河北中石油中心医院老年医学科,廊坊 065000
临床医学
儿茶素心肌细胞脂多糖p38丝裂原活化蛋白激酶信号通路氧化损伤
catechinscardiomyocyteslipopolysaccharidep38 mitogen-activated protein kinase signaling pathwayoxida-tive damage
《激光生物学报》 2024 (003)
259-266 / 8
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