导管素相关抗菌肽在高糖损伤的心脏微血管内皮细胞中的作用  

Effect of catheterin-related antimicrobial peptides on the high glucose induced cardiac microvascular endothelial cell injury

作　　者：张正伟[1] 李亚彭[2] 党瑜华[2] 陈魁[2] Zhang Zhengwei;Li Yapeng;Dang Yuhua;Chen Kui(Department of Cardiology,Anyang People's Hospital,Anyang 455000,China;Department of Cardiology,the First Affiliated Hospital,Zhengzhou University,Zhengzhou 450052,China)

机构地区：[1]安阳市人民医院心内科,安阳455000 [2]郑州大学第一附属医院心内科,郑州450052

出　　处：《中华老年医学杂志》2022年第8期975-980,共6页Chinese Journal of Geriatrics

摘　　要：目的探讨导管素相关抗菌肽(CRAMP)对心脏微血管内皮细胞损伤的影响。方法分离培养小鼠成体心脏微血管内皮细胞, 采用高糖刺激微血管内皮细胞损伤模型;采用不同浓度小鼠重组CRAMP(0.15、0.5 mg/L)蛋白处理微血管内皮细胞;采用细胞计数试剂盒(CKK-8)染色检查细胞增殖活性;采用酶联免疫吸附法(ELISA)试剂盒检测微血管内皮细胞炎症因子的分泌、活性氧检测试剂盒检测细胞活性氧的水平;采用凋亡试剂盒检测细胞凋亡、基质胶检测小管形成和小管数量、一氧化氮(NO)检测试剂盒检测NO的水平、免疫印迹法检测细胞一氧化氮合成酶(eNOS)的表达。结果高糖组细胞增殖活性明显低于对照组[(52.2±5.4)%比(100.0±7.3)%], 0.15 mg/L CRAMP组和0.5 mg/L CRAMP组细胞增殖活性高于高糖组[(72.0±3.4)%比(84.2±5.8)%比(52.2±5.4)%(F=75.300, P<0.001)]。高糖组细胞肿瘤坏死因子-α的表达明显高于对照组和0.5 mg/L CRAMP组[(239.1±32.1)μg/L比(22.1±3.7)μg/L比(84.6±9.4)μg/L](F=197.300, P<0.001)。高糖组细胞活性氧的水平明显高于对照组和0.5 mg/L CRAMP组[(20.8±2.4)比(4.8±1.7)比(10.2±1.5)](F=105.700, P<0.001)。高糖组凋亡细胞数量明显高于对照组和0.5 mg/L CRAMP组[(21.2±3.1)%比(2.2±0.6)%比(9.5±1.2)%](F=141.900, P<0.001)。高糖组小管长度和数量低于对照组和0.5 mg/L CRAMP组[(87.8±9.1)μm比(337.0±37.2)μm比(206.5±16.3)μm(F=160.800, P<0.001)及(9.1±1.9)个/视野比(22.0±3.4)个/视野比(16.8±2.2)个/视野(F=36.200, P<0.001)]。高糖组细胞一氧化氮水平低于对照组, 0.5 mg/L CRAMP组细胞一氧化氮水平高于高糖组[(0.25±0.05)比(1.05±0.16)比(0.75±0.06)](F=83.200, P<0.001)。高糖组细胞内皮型eNOS的蛋白表达和mRNA转录水平低于对照组, 0.5 mg/L CRAMP组细胞内皮型eNOS的蛋白表达和mRNA水平高于高糖组[(0.07±0.03)比(0.81±0.05)比(0.54±0.07)(F=275.700, P<0.001)及(0.11±0.07)比(1.00±0.22)比(0.57±0.12)(F=50.600, PObjective To investigate the effect of catheterin-related antimicrobial peptides(CRAMP)on the damage of cardiac microvascular endothelial cells induced by high glucose.Methods Adult mouse heart microvascular endothelial cells were isolated and cultured.A model of microvascular endothelial cell injury was established by high glucose culture.The endothelial cells were randomly divided into 4 groups as the following.In the control group,27.5 mmol/L mannitol was given as isoosmotic control as compared with the high glucose group.In the high glucose group(HG group),cells were cultured with 33.3 mmol/L high glucose for 48 h,and then treated without CRAMP.In 0.15 mg/L CRAMP treatment group,cells were cultured with 33.3 mmol/L high glucose for 48 h,followed by 0.15 mg/L CRAMP treatmen for 24 h.In the 0.5 mg/L CRAMP treatment group,cells were cultured with 33.3 mmol/L high glucose treatment for 48 h,and then treated with 0.5 mg/L CRAMP for 24 h.Cell proliferation was examined by staining with CKK-8 cell counting kit.The secretion of inflammatory factors in microvascular endothelial cells was detected by ELISA kit.Reactive oxygen species assay kit detects the level of reactive oxygen species in cells.Cell apoptosis was detected by apoptosis kit.Tubule formation and tubule number were measured by cells cultured on the matrix glue membrane,then detected by microscopic observation.The nitric oxide(NO)test kit measures levels of NO.The expression of nitric oxide synthase(eNOS)was detected by western blotting.Results The cell proliferation activity was significant lower in the HG group than in control group[(52.2±5.4)%vs.(100.0±7.3)%].The cell proliferation activity was higher in the 0.15 and 0.5 mg/L CRAMP groups than in the HG group[(72.0±3.4)%vs.(52.2±5.4)%;and(84.2±5.8)%vs.(52.2±5.4)%(F=75.300,P<0.001)].The expression of tumor necrosis factor-αwas significantly higher in the HG group than in the control group and in 0.5 mg/L CRAMP group[HG group of(239.1±32.1)μg/L,the control of(22.1±3.7)μg/L,0.5 mg/L CRAMP of(84

关 键 词：内皮 血管 一氧化氮合酶 导管素相关抗菌肽 

分 类 号：R587.2[医药卫生—内分泌]