摘要
目的 探讨高迁移率族蛋白1(HMGB1)对低氧状态下肺动脉平滑肌细胞(PASMC)的影响。方法本实验时间为2022年3月—2023年9月。将PASMC分为对照组、低氧组、HMGB1中和抗体(HMGB1 Ab)组、HMGB1Ab+单钠尿酸盐(MSU)组,对照组:PASMC置于37℃、21%O_(2)、5%CO_(2)培养箱中常氧培养24 h;低氧组:PASMC置于37℃、3%O_(2)、5%CO_(2)培养箱中低氧培养24 h;HMGB1 Ab组:PASMC置于37℃、3%O_(2)、5%CO_(2)培养箱中低氧培养24 h,加入终浓度为20μg/ml的HMGB1 Ab培养3 h;HMGB1 Ab+MSU组:PASMC置于37℃、3%O_(2)、5%CO_(2)培养箱中低氧培养24 h,加入终浓度为20μg/ml的HMGB1 Ab培养3 h,然后加入终浓度为200 g/L的MSU培养3 h。采用CCK-8法检测各组细胞增殖活力,细胞划痕实验检测各组细胞迁移能力,免疫荧光染色观察各组PASMC中α-平滑肌肌动蛋白(α-SMA)表达情况,实时荧光定量反转录聚合酶链式反应(qRT-PCR)检测各组PASMC中HMGB1及细胞焦亡相关因子[NLRP3、含半胱氨酸的天冬氨酸蛋白水解酶1(Caspase-1)、gasdermin D(GSDMD)、白介素1β(IL-1β)、白介素18(IL-18)]的mRNA相对表达量,Western blot法检测各组PASMC中HMGB1及细胞焦亡相关因子相对表达量。结果 低氧组细胞增殖活力高于对照组,HMGB1 Ab组细胞增殖活力低于低氧组,HMGB1 Ab+MSU组细胞增殖活力高于HMGB1 Ab组(P<0.05)。低氧组划痕愈合率高于对照组,HMGB1 Ab组划痕愈合率低于低氧组,HMGB1 Ab+MSU组划痕愈合率高于HMGB1 Ab组(P<0.05)。免疫荧光染色结果显示,与对照组比较,低氧组细胞荧光染色强度明显增强,α-SMA表达增加;与低氧组比较,HMGB1 Ab组细胞荧光染色强度明显减弱,α-SMA表达减少;与HMGB1 Ab组比较,HMGB1 Ab+MSU组细胞荧光染色强度明显增强,α-SMA表达增加。低氧组PASMC中HMGB1、NLRP3、Caspase-1、GSDMD、IL-1β、IL-18 mRNA及蛋白相对表达量高于对照组,HMGB1 Ab组PASMC中HMGB1、NLRP3、Caspase-1、GSDMD、IL-1β、IL-18 mRNA及蛋白相对表达量低于�
Objective To investigate the effect of high mobility group box 1(HMGB1)on pulmonary arterial smooth muscle cell(PASMC)under hypoxic conditions.Methods This experiment was conducted from March 2022 to September 2023.PASMC was divided into control group,hypoxia group,HMGB1 neutralizing antibody(HMGB1 Ab)group,HMGB1 Ab+monosodium urate(MSU)group.Control group:PASMC was incubated at 37℃,21%O_(2),and 5%CO_(2) incubator under normal oxygen for 24 hours;hypoxia group:PASMC was incubated at 37℃,3%O_(2),and 5%CO_(2) in a low oxygen incubator for 24 hours;HMGB1 Ab group:PASMC was incubated at 37℃,3%O_(2),and 5%CO_(2) in a low oxygen incubator for 24 hours,add a final concentration of 20μg/ml HMGB1 Ab cultivate for 3 hours;HMGB1 Ab+MSU group:PASMC was incubated at 37℃,3%O_(2),and 5%CO_(2) in a low oxygen incubator for 24 hours,add a final concentration of 20μg/ml HMGB1 Ab cultivate for 3 hours,then add a final concentration of 200 g/L MSU cultivate for 3 hours.The cell proliferation activity of each group was detected by CCK-8,the cell migration ability of each group was detected by cell scratch assay,the expression ofα-smooth muscle actin(α-SMA)in PASMC of each group was observed by immunofluorescence staining,the mRNA relative expression levels of HMGB1 and pyroptosis related factors[NLRP3,cysteinyl aspartate specific proteinase 1(Caspase-1),gasdermin D(GSDMD),interleukin-1β(IL-1β)and interleukin 18(IL-18)]in PASMC of each group were detected by real time quantity polymerase chain reaction(qRT-PCR),the relative expression levels of HMGB1 and pyroptosis related factors in PASMC of each group were detected by Western blot.Results The cell proliferation activity of hypoxia group was higher than that of control group,the cell proliferation activity of HMGB1 Ab group was lower than that of hypoxia group,and the cell proliferation activity of HMGB1Ab+MSU group was higher than that of HMGB1 Ab group(P<0.05).The scratch healing rate of hypoxia group was higher than that of control group,the scratch healing rate of H
作者
李鸣远
李倩
武云
LI Mingyuan;LI Qian;WU Yun(Department of General Medicine,the First Affiliated Hospital of Xinjiang Medical University,Urumqi 830054,China)
出处
《实用心脑肺血管病杂志》
2024年第2期63-69,共7页
Practical Journal of Cardiac Cerebral Pneumal and Vascular Disease
基金
新疆维吾尔自治区自然科学基金面上项目(2022D01C476)。
关键词
肺动脉高压
肌细胞
平滑肌
低氧
高迁移率族蛋白1
细胞焦亡
Pulmonary arterial hypertension
Myocytes,smooth muscle
Hypoxia
High mobility group protein 1
Pyroptosis
