Biochemical and Biophysical Research Communications, Vol.513, No.4, 1055-1062, 2019
High glucose/ox-LDL induced hepatic sinusoidal capillarization via alpha v beta 5/FAK/ERK signaling pathway
Objective: The main purpose of this study is to explore the role of integrin alpha v beta 5 in hepatic sinusoidal capillarization under high glucose/ox-LDL conditions. Methods: Establish rat model of diabetic fatty liver disease. LSECs were extracted from tissue obtained from rats of control group, cultured and treated with media containing glucose (25 mM, 24 h)/ox-LDL (100 mu g/ml, 24 h) in different inhibitors. The expression of integrin alpha v beta 5, FAK, ERK, VN in LSECs were detected by RT-PCR and Western blot. Hematoxylin-eosin staining and gomori methenaminutese silver stain was used to observe the basement membrane histopathological features of the liver tissue. Immunohistochemical to detected the protein expression of integrin alpha v beta 5 and VN in liver tissue. Using scanning electron microscopy to visualise the fenestration frequency and fenestration diameter. Protein expression of VN was also testified by immunofluorescence assay. Results: 1. The expression of integrin alpha v, integrin beta 5, FAK, ERK, VN increased in a time- and concentration-dependent manner under high glucose or oxLDL. This effect is the most significant when they co-exist (P < 0.05). 2. The expression of ERK, FAK and VN was down-regulated when LSECs were treated with.the integrin alpha v beta 5 inhibitor RGDfv. DF228, inhibitor of FAK only suppressed expression of ERK and VN. Furthermore, VN expression was down-regulated by intervention of LSECs with the ERK inhibitor PD98059, while the expression of integrin alpha v beta 5 and FAK was not significantly changed (P < 0.05). 3. We observed that high glucose and oxLDL can lead to decreasing in the porosity, diameter and number of fenestrae compared with the control group, which is more significant in the combination of high glucose and oxLDL (P < 0.05). 4. Result of gomori methenaminutese silver stain showed that the T2DM + NAFLD group had more obvious and darker brown coloration in basement membrane and sinus space. HE staining showed that in the T2DM + NAFLD group, the hepatocytes were loose in structure and disordered in arrangement, with a large number of lipid droplets in some cytoplasm, and most hepatocytes showed steatosis. In immunohistochemical staining, the positive areas of integrin alpha v beta 5 and VN showed a increasing trend from the control group to T2DM group, NAFLD group and T2DM + NAFLD group (P < 0.05). Conclusions: lntegrin alpha v beta 5 which induces LSECs dysfunction, promoting hepatic sinusoidal capillarization regulates VN expression via ERK/FAK pathway under high glucose/ox-LDL, may be a potential target for prevention and treatment of T2DM with fatty liver disease. (C) 2019 Elsevier Inc. All rights reserved.