瘦素对人脑血管平滑肌细胞活性及ROS表达的影响(1)
[摘要] 目的 探讨瘦素对人脑血管平滑肌细胞(HBVSMCs)活性及活性氧(ROS)表达水平的影响。
方法体外培养HBVSMCs，使用不同浓度瘦素干预HBVSMCs，采用CCK-8法检测瘦素对细胞增殖活性的影响，蛋白免疫印迹(Western blot)检测细胞中增殖细胞核抗原(PCNA)和Ki-67的表达水平，流式细胞术检测瘦素对细胞周期的影响，DCFH-DA探针法检测细胞内ROS水平。
结果 CCK-8检测显示，不同浓度瘦素处理组培养48和72 h后HBVSMCs的增殖活性明显升高(F=117.818、160.349，P<0.001);Western blot检测显示，不同浓度瘦素处理组培养48 h后HBVSMCs中PCNA和Ki-67的表达水平明显上调(F=290.000、898.769，P<0.001);流式细胞术检测显示，不同浓度瘦素处理组G0/G1期细胞比例降低(F=264.180，P<0.001)，S期和G2/M期细胞比例升高(F=160.033、380.234，P<0.001);DCFH-DA探针法检测显示，不同浓度瘦素处理组HBVSMCs內ROS水平明显升高(F=394.025，P<0.001);以上各指标变化均具有浓度依赖性。
结论 瘦素能够促进HBVSMCs的增殖活性，并诱导细胞中ROS的表达。
[关键词] 瘦素;肌细胞，平滑肌;细胞增殖;活性氧;颅内动脉硬化
[中图分类号] R977.1;R329.24
[文献标志码] A
[文章编号] 2096-5532(2020)03-0270-05
doi：10.11712/jms.2096-5532.2020.56.105
[开放科学(资源服务)标识码(OSID)]
[网络出版] http：//kns.cnki.net/kcms/detail/37.1517.R.20200528.0935.004.html;2020-05-28 12：03
EFFECT OF LEPTIN ON THE ACTIVITY OF HUMAN BRAIN VASCULAR SMOOTH MUSCLE CELLS AND THE EXPRESSION OF REACTIVE OXYGEN SPECIES
＼ LI Yangyang， WANG Xiaoxue， ZHANG Fengju， XU Hongxia， LIU Xinxin
＼ (Department of Geriatric Neurology， the First Affiliated Hospital of Henan University， Kaifeng 475000， China)
[ABSTRACT]＼ Objective＼ To investigate the effect of leptin on the activity of human brain vascular smooth muscle cells (HBVSMCs) and the expression of reactive oxygen species (ROS).
＼ Methods＼ HBVSMCs were cultured in vitro and then treated with different concentrations of leptin. CCK-8 assay was used to observe the effect of leptin on the proliferative activity of cells; Western blot was used to measure the expression of proliferating cell nuclear antigen (PCNA) and Ki-67; flow cytometry was used to observe the effect of leptin on cell cycle; the DCFH-DA probe method was used to measure the level of ROS in cells.
＼ Results
The results of CCK-8 assay showed that HBVSMCs treated with different concentrations of leptin had a significant increase in proliferative activity after 48 and 72 hours of culture (F=117.818，160.349;P<0.001). The results of Western blot showed that HBVSMCs treated with different concentrations of leptin had significantly upregulated expression of PCNA and Ki-67 after 48 hours of culture (F=290.000，898.769;P<0.001). The results of flow cytometry showed that HBVSMCs treated with different concentrations of leptin had a significant reduction in the proportion of cells in G0/G1 phase (F=264.180，P<0.001) and a significant increase in the proportion of cells in S phase or G2/M phase (F=160.033，380.234;P<0.001). The results of the DCFH-DA probe method showed that HBVSMCs treated with different concentrations of leptin had a significant increase in the level of ROS (F=394.025，P<0.001). All the changes above were concentration-dependent., http://www.100md.com(李阳阳 王晓雪 张峰菊 许宏侠 刘欣欣)
方法体外培养HBVSMCs，使用不同浓度瘦素干预HBVSMCs，采用CCK-8法检测瘦素对细胞增殖活性的影响，蛋白免疫印迹(Western blot)检测细胞中增殖细胞核抗原(PCNA)和Ki-67的表达水平，流式细胞术检测瘦素对细胞周期的影响，DCFH-DA探针法检测细胞内ROS水平。
结果 CCK-8检测显示，不同浓度瘦素处理组培养48和72 h后HBVSMCs的增殖活性明显升高(F=117.818、160.349，P<0.001);Western blot检测显示，不同浓度瘦素处理组培养48 h后HBVSMCs中PCNA和Ki-67的表达水平明显上调(F=290.000、898.769，P<0.001);流式细胞术检测显示，不同浓度瘦素处理组G0/G1期细胞比例降低(F=264.180，P<0.001)，S期和G2/M期细胞比例升高(F=160.033、380.234，P<0.001);DCFH-DA探针法检测显示，不同浓度瘦素处理组HBVSMCs內ROS水平明显升高(F=394.025，P<0.001);以上各指标变化均具有浓度依赖性。
结论 瘦素能够促进HBVSMCs的增殖活性，并诱导细胞中ROS的表达。
[关键词] 瘦素;肌细胞，平滑肌;细胞增殖;活性氧;颅内动脉硬化
[中图分类号] R977.1;R329.24
[文献标志码] A
[文章编号] 2096-5532(2020)03-0270-05
doi：10.11712/jms.2096-5532.2020.56.105
[开放科学(资源服务)标识码(OSID)]
[网络出版] http：//kns.cnki.net/kcms/detail/37.1517.R.20200528.0935.004.html;2020-05-28 12：03
EFFECT OF LEPTIN ON THE ACTIVITY OF HUMAN BRAIN VASCULAR SMOOTH MUSCLE CELLS AND THE EXPRESSION OF REACTIVE OXYGEN SPECIES
＼ LI Yangyang， WANG Xiaoxue， ZHANG Fengju， XU Hongxia， LIU Xinxin
＼ (Department of Geriatric Neurology， the First Affiliated Hospital of Henan University， Kaifeng 475000， China)
[ABSTRACT]＼ Objective＼ To investigate the effect of leptin on the activity of human brain vascular smooth muscle cells (HBVSMCs) and the expression of reactive oxygen species (ROS).
＼ Methods＼ HBVSMCs were cultured in vitro and then treated with different concentrations of leptin. CCK-8 assay was used to observe the effect of leptin on the proliferative activity of cells; Western blot was used to measure the expression of proliferating cell nuclear antigen (PCNA) and Ki-67; flow cytometry was used to observe the effect of leptin on cell cycle; the DCFH-DA probe method was used to measure the level of ROS in cells.
＼ Results
The results of CCK-8 assay showed that HBVSMCs treated with different concentrations of leptin had a significant increase in proliferative activity after 48 and 72 hours of culture (F=117.818，160.349;P<0.001). The results of Western blot showed that HBVSMCs treated with different concentrations of leptin had significantly upregulated expression of PCNA and Ki-67 after 48 hours of culture (F=290.000，898.769;P<0.001). The results of flow cytometry showed that HBVSMCs treated with different concentrations of leptin had a significant reduction in the proportion of cells in G0/G1 phase (F=264.180，P<0.001) and a significant increase in the proportion of cells in S phase or G2/M phase (F=160.033，380.234;P<0.001). The results of the DCFH-DA probe method showed that HBVSMCs treated with different concentrations of leptin had a significant increase in the level of ROS (F=394.025，P<0.001). All the changes above were concentration-dependent., http://www.100md.com(李阳阳 王晓雪 张峰菊 许宏侠 刘欣欣)
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