糖尿病氧化应激环境中α-Klotho对巨噬细胞-血管内皮细胞串扰的影响OACSTPCD

AIM:To investigate the effects of overexpressing α-Klotho(KL)in RAW264.7 cells stimulated by oxidative stress on the proliferation,migration,tube-formation and tight junction of human umbilical vein endothelial cells(HUVECs). METHODS:RAW264.7 cells were categorized into control,4-hydroxynonenal(4HNE),and 4HNE+KL groups,with F4/80 expression assessed via immunofluorescence staining.Three groups of conditional media were prepared for HUVECs and culture divided into Mø-NC,Mø-4HNE,and Mø-4HNE+KL groups.Cell proliferation was evaluated using CCK8 assay,while scratch test and Transwell assays were employed to measure cell migration.Additionally,tube-formation assay was conducted to assess cell tubule formation,and Western blot assay was utilized to detect the protein expression levels of Claudin 5,Occludin and ZO 1. RESULTS:The results of immunofluorescence staining showed that the fluorescence intensity of F4/80 of RAW264.7 cells in the 4HNE group was significantly enhanced compared with the control group,while that of F4/80 in the 4HNE+KL group was significantly decreased compared with the 4HNE group(all P＜0.05).The CCK8 assay results revealed a significant increase in the proliferation of HUVECs in the Mø-4HNE group compared with the Mø-NC group.Conversely,the proliferation of the Mø-4HNE+KL group exhibited a significant decrease compared with that in the Mø-4HNE group(all P＜0.01).The results of scratch test and Transwell assays demonstrated a significant increase in the migration of HUVECs in the Mø-4HNE group compared with the Mø-NC group,while the migration of the Mø-4HNE+KL group exhibited a significant decrease compared with the Mø-4HNE group(all P＜0.01).In the tube-formation assay,it was observed that the number of tubes formed by HUVECs in the Mø-4HNE group was significantly increased compared with the Mø-NC group,while that of tubes formed in the Mø-4HNE+KL group was significantly decreased compared with the Mø-4HNE group(all P＜0.01).Additionally,the Western blot results revealed a significant decrease in the relative expression levels of Claudin 5,Occludin,and ZO 1 in the Mø-4HNE group compared with the Mø-NC group.Conversely,in the Mø-4HNE+KL group,there was a significant increase in the relative expression levels of Claudin 5,Occludin,and ZO 1 compared to the Mø-4HNE group(all P＜0.01). CONCLUSIONS:KL inhibits the proliferation,migration,and tube-formation of HUVECs while enhancing the tight junction by changing the activation state of macrophages in the diabetic oxidative stress environment.