汉黄芩苷干预高糖刺激RAW264.7巨噬细胞KLF4/NF-κB信号通路  

Effect of wogonoside on KLF4/NF-κB signal pathway in RAW264.7 macrophages stimulated by high glucose

作　　者：谢姗 徐兴欣 Xie Shan;Xu Xingxin(Dept of Nephrology,The Fourth Affiliated Hospital of Anhui Medical University,Hefei 230001;Dept of Nephrology,The First Affiliated Hospital of Anhui Medical University,Hefei 230022)

机构地区：[1]安徽医科大学第四附属医院肾内科,合肥230001 [2]安徽医科大学第一附属医院肾内科,合肥230022

出　　处：《安徽医科大学学报》2022年第10期1523-1528,共6页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:81600568)。

摘　　要：目的探究汉黄芩苷(WG)对高糖刺激下RAW264.7巨噬细胞的KLF4/NF-κB信号通路的影响。方法培养RAW264.7巨噬细胞,CCK-8法检测WG对细胞无活性影响的浓度范围。将细胞随机分为LG组(5.5 mmol/L葡萄糖培养基)、D组(30 mmol/L甘露醇培养基)、HG组(30 mmol/L葡萄糖培养基)、LG+WG50组(5.5 mmol/L葡萄糖+50μmol/L WG)、HG+WG12.5组(30 mmol/L葡萄糖+12.5μmol/L WG)、HG+WG25组(30 mmol/L葡萄糖+25μmol/L WG)、HG+WG50组(30 mmol/L葡萄糖+50μmol/L WG);免疫荧光检测各组细胞iNOS的表达,Western blot检测各组细胞NF-κB p65、NF-κB pp65、KLF4、iNOS、IL-1β、TNF-α蛋白表达,qRT-PCR检测各组细胞KLF4、iNOS、TNF-α、IL-1β的mRNA表达量,ELISA法检测各组细胞上清液中TNF-α、IL-1β的分泌水平。结果与LG组相比,HG组细胞中iNOS表达高(P<0.01),与HG组相比,HG+W50组细胞中iNOS表达降低(P<0.01)。与LG组相比,高糖可以抑制KLF4的表达(P<0.01),上调NF-κB pp65、iNOS、IL-1β、TNF-α的表达(P<0.01),增加上清液中IL-1β、TNF-α的水平(P<0.01);与HG组相比,WG组各炎症指标明显下降(P<0.01),KLF4的表达量反而剂量依赖性增高(P<0.01)。结论WG可以抑制高糖刺激巨噬细胞分泌炎症因子及相关蛋白的表达,有提高保护性因子KLF4的治疗作用。Objective To explore the effect of wogonoside(WG)on the Krüppel-like factor4(KLF4)/nuclear factorκB(NF-κB)pathway of RAW264.7 macrophages subject to high glucose stimulation.Methods RAW264.7 macrophages were cultured and the concentration range of inactive effect of WG on cells was detected by CCK-8.Macrophages randomly grouped into LG group(medium for 5.5 mmol/L of glucose),D group(30 mmol/L mannitol medium),HG group(medium for 30 mmol/L glucose),LG+WG50 group(5.5 mmol/L glucose+50μmol/L WG),HG+WG12.5 group(30 mmol/L glucose+12.5μmol/L WG),HG+WG25 group(30 mmol/L glucose+25μmol/L WG),HG+WG50 group(30 mmol/L glucose+50μmol/L WG).The expression of inducible nitric oxide synthase(iNOS)in cells of each group was determined by immunofluorescence.NF-κB p65,NF-κB pp65,KLF4,iNOS,interleukin-1β(IL-1β),tumor necrosis factor alpha(TNF-α)expression in each group was detected by Western blot.mRNA expression of KLF4,iNOS,IL-1β,TNF-αin each group was detected by qRT-PCR.IL-1β,TNF-αsecretion of cells in each group was detected by ELISA.Results Compared with LG group,the expression of iNOS in HG group was higher(P<0.01),and the expression of iNOS in HG+W50 group was lower than that in HG group(P<0.01).Compared with the LG group,high glucose can inhibit KLF4 expression(P<0.01),stimulate NF-κB pp65,iNOS,IL-1β,TNF-αexpression(P<0.01),and increase the IL-1β,TNF-αlevel in the supernatant(P<0.01);However,the inflammatory index in the WG groups significantly decreased(P<0.01),but expression of KLF4 dose-dependent increased(P<0.01).Conclusion WG inhibits the expression of inflammatory factors and associated proteins secreted by macrophages by high glucose,and has the therapeutic effect of improving the protective factor-KLF4.

关 键 词：糖尿病肾病 巨噬细胞 汉黄芩苷 KLF4 

分 类 号：R587.2[医药卫生—内分泌] R392.9[医药卫生—内科学]