CAJ | 학술논문

目的观察辛伐他汀对脓毒症内皮细胞的保护作用并探讨其机制.方法盲肠结扎穿孔法制作脓毒症雄性SD大鼠模型,6h后收集脓毒症SD大鼠血清备用.将体外培养的人脐静脉内皮细胞(human umbilical vein endothelial cells,HUVECS)株(ECV-304)随机(随机数字法)分为3组:对照组、脓毒症组和辛伐他汀组.予相应条件培养液处理24h后,采用四唑盐比色法检测HUVECS的生长情况,Hoechst染色法、吖啶橙荧光染色法观察细胞凋亡形态变化及流式细胞术检测细胞凋亡率,RT-PCR检测细胞Bcl-2和Bax基因的表达情况.细胞相对生长率比较采用两独立样本t检验,组间吸光度值及细胞凋亡率比较采用one-way ANOVA方差分析.结果 (1)与脓毒症组比较,辛伐他汀组HUVECS相对生长率(％)显著增加[(64.06±1.70)％ vs.(37.59±2.13)％,t=-16.846,P=0.000].(2) Hoechst染色法与吖啶橙荧光染色法均显示,与对照组比较,脓毒症组凋亡细胞显著增加；而辛伐他汀组较脓毒症组显著减少.流式细胞术凋亡检测发现与对照组比较,脓毒症组细胞凋亡率显著增加[(4.07±1.55)％vs.(30.20±8.94)％,P=0.001]；辛伐他汀组细胞凋亡率较脓毒症组显著减少[(16.50±4.26)％vs.(30.20±8.94)％,P=0.027].(3)脓毒症大鼠血清处理后,HUVECS中Bcl-2基因表达降低,而Bax基因表达增高；与脓毒症组比较,辛伐他汀组两者呈反向改变.结论辛伐他汀通过上调Bcl-2和下调Bax基因表达抑制脓毒症内皮细胞凋亡,发挥其保护血管内皮细胞的作用；这可能是辛伐他汀治疗脓毒症的机制之一.
목적 관찰신벌타정대농독증내피세포적보호작용병탐토기궤제.방법 맹장결찰천공법제작농독증웅성SD대서모형,6h후수집농독증SD대서혈청비용.장체외배양적인제정맥내피세포(human umbilical vein endothelial cells,HUVECS)주(ECV-304)수궤(수궤수자법)분위3조:대조조、농독증조화신벌타정조.여상응조건배양액처리24h후,채용사서염비색법검측HUVECS적생장정황,Hoechst염색법、아정등형광염색법관찰세포조망형태변화급류식세포술검측세포조망솔,RT-PCR검측세포Bcl-2화Bax기인적표체정황.세포상대생장솔비교채용량독립양본t검험,조간흡광도치급세포조망솔비교채용one-way ANOVA방차분석.결과 (1)여농독증조비교,신벌타정조HUVECS상대생장솔(％)현저증가[(64.06±1.70)％ vs.(37.59±2.13)％,t=-16.846,P=0.000].(2) Hoechst염색법여아정등형광염색법균현시,여대조조비교,농독증조조망세포현저증가；이신벌타정조교농독증조현저감소.류식세포술조망검측발현여대조조비교,농독증조세포조망솔현저증가[(4.07±1.55)％vs.(30.20±8.94)％,P=0.001]；신벌타정조세포조망솔교농독증조현저감소[(16.50±4.26)％vs.(30.20±8.94)％,P=0.027].(3)농독증대서혈청처리후,HUVECS중Bcl-2기인표체강저,이Bax기인표체증고；여농독증조비교,신벌타정조량자정반향개변.결론 신벌타정통과상조Bcl-2화하조Bax기인표체억제농독증내피세포조망,발휘기보호혈관내피세포적작용；저가능시신벌타정치료농독증적궤제지일.
Objective To observe the protective effect of simvastatin on endothelial cells in sepsis in order to investigate its mechanism.Methods The SD rat sepsis model was established by cecal ligation and puncture,and the serum was collected 6 h later.A strain of human umbilical vein endothelial cells (HUVECs) cultivated in vitro,ECV-304,were randomly divided into a control group,rat sepsis serum intervention group (abbreviated:sepsis group) and simvastatin + rat sepsis serum intervention group (abbreviated:simvastatin group).The relative growth rate of HUVECS in each group was detected by MTT (methyl thiazolyl tetrazolium) method.The morphological changes of apoptotic HUVECS were detected by using Hoechst assay and Hoechst33258 assay after 24 h incubated with corresponding culture medium,respectively.At the same time,the apoptotic rate of HUVECS in each group was detected by flow cytometry.Moreover,the expressions of Bcl-2 and Bax genes of the HUVECS in each group were detected by RT-PCR,respectively.The relative growth rate of HUVECS in each group was analyzed by using independent-samples t-test.The absorbance values and the apoptotic rate of HUVECS were analyzed by using One-Way ANOVA.Results 1) Compared with the sepsis group,the HUVECS in the simvastatin group had a higher relative growth rate [(64.06±1.70)％ vs (37.59±2.13)％,t values was-16.846,P=0.000].2) Both Hoechst assay and AO assay showed that compared with the control group,the apoptosis of HUVECS was significantly increased in the sepsis group,but decreased significantly in the simvastatin group.The apoptotic rate of HUVECS in the sepsis group was higher than that in the control group [(4.07 ± 1.55)％vs (30.20 ±8.94)％,P =0.001],but compared with the sepsis group,it was significantly lower in the simvastatin group [(16.50 ± 4.26) ％ vs (30.20 ± 8.94) ％,P =0.027].3) The expression of Bcl-2 gene was decreased,but the expression of Bax gene was increased in the HUVECS 24 hours after incubation with sepsis serum,and the expressions of Bcl-2 and Bax genes in the simvastatin group were contrary to those in the sepsis group.Conclusions The mechanisms of inhibiting apoptosis of vascular endothelial cells by simvastatin treating sepsis may attributed to up-regulating Bcl-2 and down-regulating Bax gene expressions.