ABSTRACT
AIM:To investigate the effect of angiotensin 1-7(Ang1-7)on the human glomerular endothelial cells(HGECs)injury induced by angiotensin Ⅱ(Ang Ⅱ)and its possible mechanism.METHODS: Cultured HGECs were divided into 6 groups randomly: control group, Ang Ⅱ group, Ang1-7 group, Ang Ⅱ +Ang1-7 group, Ang Ⅱ +Ang1-7+A779(an inhibitor of Mas receptor)group and A779 group.The apoptotic rate and reactive oxygen species (ROS)of HGECs were analyzed by flow cytometry and photographed by fluorescence microscopy.The levels of lactate de-hydrogenase(LDH),nitric oxide(NO),endothelin-1(ET-1),interleukin-6(IL-6),tumor necrosis factor-α(TNF-α), transforming growth factor-β(TGF-β),monocyte chemoattractant protein-1(MCP-1)and intercellular adhesion molecule-1(ICAM-1)in the supernatant of cell cultures were measured.RESULTS:Compared with the control group,the apoptot-ic rate and the average fluorescence intensity of ROS were increased in the Ang Ⅱ group,IL-6,TNF-α,TGF-β,ICAM-1 and MCP-1 in cell supernatants were also increased in the Ang Ⅱ group(P<0.05).Compared with the Ang Ⅱ group,the apoptotic rate,ROS level, and the above inflammatory factors were decreased in Ang Ⅱ +Ang1-7 group(P<0.05). Compared with the Ang Ⅱ +Ang1-7 group,adding A779 increased the cell apoptotic rate,ROS production and the releases of the above inflammatory factors in cell supernatants(P<0.05).Compared with the Ang Ⅱ group,adding Ang1-7 inhibi-ted the LDH leakage, ET-1 secretion and promoted the release of NO in a dose-dependent manner(P<0.05).CON-CLUSION:Ang1-7 attenuates the HGECs injury induced by Ang Ⅱ by inhibiting the Mas receptor.
ABSTRACT
Objective To observe the effect of TGF‐β1 in the endothelial‐mesenchymal transition EndM T of glomeruli ,and to explore the effect of TGF‐β/Smad signaling pathway mediated by integrin linked kinase(ILK ) in the progress of renal fibrosis . Methods Human glomerular endothelial cells (HGEnC) incubated in vitro were divided into blank control group ,TGF‐β1 (12 .5 , 25 .0 ,50 .0 ng/mL) group .TGF‐β1 50 .0 ng/mL receptor type one antagonist (LY364749)5 .0 μmol/L group ,ILK (QLT‐0267)5 .0μmol/L antagonist group .The mRNA level of P‐Smad 2/3 and ILK was determined by RT‐PCR ,and the protein level of P‐Smad 2/3 ,ILK ,E‐cadherin ,CD31 ,α‐SMA and FSP‐1 was determined by Western blot after 48 h and 72 h after incubation in each group un‐der the above‐mentioned condition .Results (1)TGF‐β1 could significantly increased the mRNA level of P‐Smad2/3 and ILK (P0 .05) .Conclusion TGF‐β1 as the effector molecule in downstream can promote endothelial‐mesenchymal transition of HGEnC ,and TGF‐β/Smad signaling pathway mediated integrin linked kinase participate in this process ,which probably play important role in the progress of renal fibrosis .
ABSTRACT
Whereas mesangial and epithelial cells from glomeruli are commonly grown in vitro, there has been major difficulties in developing homogenous cultures of human glomerular endothelial cells. This study defines the conditions for the reproducible isolation and growth of homogenous monolayers of human glomerular endothelial cells based on the method of Green DF et al published in 1992. Using the selective media and the sieving method, fibronectin was required as a surface matrix after adequate collagenase treatment, and endothelial cell growth factor and heparin was needed for the continuous growth of endothelial cells. The endothelial cell growth factor was isolated from the bovine hypothalamic extracts. Glomerular capillary endothelial cells exhibited a cobblestone morphology at confluence and stained homogenously with von Willebrand factor(factor VIII). The cytokeratin and the actin were not stained. This study might be helpful for in vitro study to know the biological characteristics of human glomerular endothelial cells under the predetermined condition.