高糖高脂及肝素结合蛋白对HK-2/中性粒细胞共培养体系的炎症作用  

Inflammatory effects of high-glucose,high-free fatty acids,and heparin binding protein on HK-2/neutrophil co-culture system

作　　者：吴红 王凝瑞 邹海虹[2] 熊妍 戴子妍 聂益军[2] WU Hong;WANG Ningrui;ZOU Haihong;XIONG Yan;DAI Ziyan;NIE Yijun(School of Laboratory Medicine,Nanchang Medical College,Nanchang,Jiangxi 330052,China;Department of Clinical Laboratory,the First Affiliated Hospital,Medical College of Nanchang University,Nanchang,Jiangxi 330006,China)

机构地区：[1]南昌医学院检验医学院,江西南昌330052 [2]南昌大学第一附属医院检验科,江西南昌330006

出　　处：《中国热带医学》2024年第9期1100-1105,共6页China Tropical Medicine

基　　金：国家自然科学基金资助项目(No.81660144);南昌医学院博士科研启动基金项目(No.NYB22005);江西省卫健委基金项目(No.202310464,No.202410101);江西省中医药管理局基金项目(No.2023A0192)。

摘　　要：目的以人肾皮质近曲小管上皮(human kidney-2 cells,HK-2)细胞/中性粒细胞共培养体系为研究对象,探讨高糖高脂及肝素结合蛋白(heparin-binding protein,HBP)处理对共培养体系的炎症反应、氧化应激及细胞凋亡的影响。方法从正常人外周血中分离获得中性粒细胞,HK-2细胞与中性粒细胞悬液均以1×10^(5)/mL的浓度混匀建立共培养细胞体系。通过CCK-8试验确定脂多糖(lipopolysaccharides,LPS)和肝素钠(heparin sodium,Hep)的处理浓度,分别通过免疫荧光干式定量法、邻苯三酚自氧化法、基于细胞测量珠阵列技术及检测表达Annexin V-FITC荧光的流式细胞术分别检测HBP浓度、氧化物歧化酶(superoxide dismutase,SOD)含量、炎性细胞因子[白细胞介素(interleukin,IL)-1β、IL6和IL-8]浓度及细胞凋亡百分率。结果CCK-8试验确定LPS和Hep的处理浓度分别为10μg/mL和50μmol/L;高糖高脂处理的HBP(4.11±0.07)、SOD(2.67±0.06)、IL-1β(11.28±0.35)、IL-6(2.57±0.10)、IL-8(1.32±0.05)及细胞凋亡发生率(1.08±0.04)相对值均较正常对照组明显增加;LPS处理(刺激HBP的产生)可使HBP(7.09±0.08)、SOD(3.07±0.14)、IL1β(22.96±0.49)、IL-6(2.79±0.11)、IL-8(1.55±0.07)及细胞凋亡发生率(1.17±0.05)相对值升高;通过Hep处理(降低HBP的产生)可使由LPS引起的HBP(5.43±0.09)、SOD(2.76±0.07)、IL-1β(16.51±0.56)、IL-6(2.64±0.16)、IL-8(1.47±0.06)及细胞凋亡发生率(1.13±0.05)相对值降低。结论高糖高脂状态及HBP均可影响肾小管上皮细胞的功能,其机制可能与促进氧化应激和炎症反应有关。Objective To investigate the effect of high glucose,high free fatty acids(FFA),and heparin binding protein(HBP)treatment on the inflammatory response,oxidative stress,and cell apoptosis of the co-culture system established by human renal cortex proximal tubule epithelial cells(HK-2)/neutrophil cells.Methods Neutrophils were isolated from the peripheral blood of healthy individuals.Both HK-2 cells and neutrophil suspensions were mixed at a concentration of 10^(5)/mL to establish a co-cultured cell system.The treatment concentrations of lipopolysaccharide(LPS)and sodium heparin(Hep)were determined through the CCK-8 experiment.The concentrations of heparin binding protein(HBP),superoxide dismutase(SOD),inflammatory cytokines(IL-1β,IL-6,and IL-8),and the percentage of cells apoptosis was detected by quantitative immunofluorescence method,pyrogallol autoxidation method,the method based on the cytometric bead array(CBA)technology,and the detecting the expression of Annexin V-FITC fluorescence by flow cytometry,respectively.Results The treatment concentrations of LPS and Hep were determined respectively as 10μg/mL and 50μmol/L using the CCK-8 experiment.Compared with the normal group,treatment with high glucose and high FFA significantly increased the levels of HBP(4.11±0.07),SOD(2.67±0.06),IL-1β(11.28±0.35),IL-6(2.57±0.10),IL-8(1.32±0.05),and the incidence of cells apoptosis(1.08±0.04).LPS treatment(stimulation of HBP production)further exacerbated the relative levels of HBP(7.09±0.08),SOD(3.07±0.14),and IL-1β(22.96±0.49),IL-6(2.79±0.11),IL-8(1.55±0.07),and the incidence of cells apoptosis(1.17±0.05).Hep treatment(reducing HBP production)significantly reversed the effects induced by LPS on HBP(5.43±0.09),SOD(2.76±0.07),IL-1β(16.51±0.56),IL-6(2.64±0.16),IL-8(1.47±0.06),and the incidence of cells apoptosis(1.13±0.05).Conclusions Both the status of high glucose combined with high FFA,as well as HBP can affect the function of renal tubular epithelial cells.The underlying mechanisms may be related to

关 键 词：高糖高脂 肝素结合蛋白 人肾皮质近曲小管上皮细胞 炎症作用 

分 类 号：R5[医药卫生—内科学]