Endocrine Abstracts

Introduction: Nonalcoholic fatty liver disease (NAFLD) is the leading cause of chronic liver disorders. Endocan is a novel molecule of endothelial dysfuction which is expressed in liver. SGLT2i have been reported to improve NAFLD through amelioration of inflammation. While there are contradictory results regarding the serum endocan levels in NAFLD patients, data regarding endocan expression in liver tissue are limited.

Aim: Herein, we aimed to investigate the effect of SGLT2i empagliflozin on the expression of endocan in liver tissues as well as in hepatocyte cells and to delineate the underlying mechanism.

Material and methods: ApoE(-/-) mice fed a western high-fat diet were used as NAFLD model. At the age of 5 weeks, ApoE(-/-) male mice were switched from normal diet to HFD. After 5 weeks, mice were divided into two groups: Control-groups (HFD+vehicle), Empa-group (HFD+empagiflozin 10 mg/kg/day) for 5 further weeks. At the end of intervention, mice were sacrificed, whole blood was drawn by cardiac puncture and liver tissues were harvested. H&E staining was performed in all liver sections for histomorphometric analysis and IHC was performed to evaluate endocan levels. The mRNA levels of endocan, ICAM-1, VCAM-1, LFA-1, SGLT2, and IL-6 were measured by qRT-PCR and the protein levels of p65 and phospho-p65, were evaluated by immunoblotting. HepG2 cells were cultured in media supplemented with low(1 g/l), intermediate(2.25 g/l) and high(4.5 g/l) glucose concentrations. The expression of the abovementioned genes were evaluated after 24 and 48 h of treatment with empagliflozin (10^-6, 2×10^-6,10^-7, 5×10^-7 M).

Results: Biochemical tests revealed reduction in blood glucose, total cholesterol, LDL-cholesterol, and triglycerides after empagliflozin intervention for 5 weeks as compared to the Control-group. Additionally, empagliflozin administration resulted in reduced hepatic lipid accumulation and NA score (NAS). Empagliflozin significantly increased the endocan mRNA and protein levels while reduced IL-6 mRNA levels. Western blot analysis revealed that empagliflozin also regulates NF-κB pathway through phosphorylation of p65 subunit. Moreover, our in vitro data confirmed the in vivo results since 48 hours incubation of HepG2 cells with empagliflozin (10^-6M,2×10^-6M) at the presence of low and intermediate glucose levels, but not high glucose levels, increased endocan expression (P<0.01 and P<0.05, respectively). Interestingly, SGLT2 mRNA expression was detected in liver and HepG2 cells.

Conclusions: Our in vitro and animal study results indicate that empagliflozin ameliorates NAFLD through –among others- induction of endocan expression. Regulation of NF-κB pathway and IL-6 expression may mediate these effects.