Endoglin and soluble endoglin in liver sinusoidal endothelial dysfunction in vivo.

Liver sinusoidal endothelial cells (LSECs) play a crucial role in regulating the hepatic function. Endoglin (ENG), a transmembrane glycoprotein, was shown to be related to the development of endothelial dysfunction. In this study, we hypothesized the relationship between changes in ENG expression and markers of liver sinusoidal endothelial dysfunction (LSED) during liver impairment. Male C57BL/6J mice aged 9-12 weeks were fed with 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet (intrahepatic cholestasis) or choline-deficient l-amino acid defined high-fat diet (CDAA-HFD) (non-alcoholic steatohepatitis (NASH)). Significant increases in liver enzymes, fibrosis, and inflammation biomarkers were observed in both cholestasis and NASH. Decreased p-eNOS/eNOS and VE-cadherin protein expression and a significant increase in VCAM-1 and ICAM-1 expression were detected, indicating LSED in both mouse models of liver damage. A significant reduction of ENG in the DDC-fed mice, while a significant increase of ENG in the CDAA-HFD group was observed. Both DDC and CDAA-HFD-fed mice showed a significant increase in MMP-14 protein expression, which is related to significantly increased levels of soluble endoglin (sENG) in the plasma. In conclusion, we demonstrated that intrahepatic cholestasis and NASH result in an altered ENG expression, predominantly in LSECs, suggesting a critical role of ENG expression for the proper function of liver sinusoids. Both pathologies resulted in elevated sENG levels, cleaved by MMP-14 expressed predominantly from LSECs, indicating sENG as a liver injury biomarker.

Monoclonal anti-endoglin antibody TRC105 (carotuximab) prevents hypercholesterolemia and hyperglycemia-induced endothelial dysfunction in human aortic endothelial cells.

Endoglin (Eng) is a co-receptor of the transforming growth factor ß superfamily playing an important role in endothelial dysfunction. TRC105 (carotuximab) is a monoclonal antibody that blocks Eng and its downstream Smad signaling pathway. Here we have investigated for the first time the effects of TRC105 treatment on the development of endothelial dysfunction induced by 7-ketocholesterol (7K) or high glucose (HG), focusing on Eng expression, signaling, and function. In the hypercholesterolemia study, human aortic endothelial cells (HAoECs) were treated with TRC105 (300 µg/ml) for 1 h, followed by the addition of 7K (10 µg/ml) for another 12 h. In the hyperglycemia study, HAoECs were exposed to HG (45 mM) for 60 h, followed by the addition of TRC105 for another 12 h, and cells treated with 5mM glucose and 40 mM mannitol served as control. Protein levels, adhesion, and transmigration of monocytes were assessed by flow cytometry, mRNA expression was measured by qRT-PCR. 7K and HG treatment increased protein levels of NF-κB and Eng and adhesion and transmigration of monocytes through HAoECs monolayer. TRC105 pretreatment reduced the 7K- or HG-induced Eng protein levels and pSmad1/5 and pSmad2/3 signaling. Despite increased protein levels of P-selectin and VCAM-1, TRC105 mediated blockage of Eng prevented 7K- and HG-induced adhesion and transmigration of monocytes through endothelial monolayers. These results suggest that TRC105-mediated Eng blockage can counteract the hypercholesterolemia- and hyperglycemia-induced endothelial dysfunction in HAoECs, suggesting that Eng might be a potential therapeutic target in disorders associated with elevated cholesterol and glucose levels.

Soluble Endoglin as a Potential Biomarker of Nonalcoholic Steatohepatitis (NASH) Development, Participating in Aggravation of NASH-Related Changes in Mouse Liver.

Nonalcoholic steatohepatitis (NASH) is characterized by hepatic steatosis with inflammation and fibrosis. Membrane endoglin (Eng) expression is shown to participate in fibrosis, and plasma concentrations of soluble endoglin (sEng) are increased in patients with hypercholesterolemia and type 2 diabetes mellitus. We hypothesize that NASH increases both hepatic Eng expression and sEng in blood and that high levels of sEng modulate cholesterol and bile acid (BA) metabolism and affect NASH progression. Three-month-old transgenic male mice overexpressing human sEng and their wild type littermates are fed for six months with either a high-saturated fat, high-fructose high-cholesterol (FFC) diet or a chow diet. Evaluation of NASH, Liquid chromatography-mass spectrometry (LC/MS) analysis of BA, hepatic expression of Eng, inflammation, fibrosis markers, enzymes and transporters involved in hepatic cholesterol and BA metabolism are assessed using Real-Time Quantitative Reverse Transcription Polymerase Chain reaction (qRT-PCR) and Western blot. The FFC diet significantly increases mouse sEng levels and increases hepatic expression of Eng. High levels of human sEng results in increased hepatic deposition of cholesterol due to reduced conversion into BA, as well as redirects the metabolism of triglycerides (TAG) to its accumulation in the liver, via reduced TAG elimination by ß-oxidation combined with reduced hepatic efflux. We propose that sEng might be a biomarker of NASH development, and the presence of high levels of sEng might support NASH aggravation by impairing the essential defensive mechanism protecting NASH liver against excessive TAG and cholesterol accumulation, suggesting the importance of high sEng levels in patients prone to develop NASH.