NAFLD is a highly prevalent condition currently lacking an approved pharmacological therapy. GABA, besides being a neurotransmitter, has been proposed to protect from liver toxicity in both in vivo an in vitro models. We aimed to determine if a novel positive allosteric modulator (PAM) of the GABAA receptor, the thioacrylamide-derivative HK4 which is devoid of blood brain barrier penetration, can protect human hepatocytes against lipotoxicity-induced injury.
Patch clamping in HEK-293 cells and calcium influx measurements in INS-1E cells proved HK4 as a selective PAM of the GABAA receptor. The expression of several main GABAA receptor subunits (α1, α3, α5, β2/3 and γ2) was demonstrated by Western blotting of HepG2 cells. Next generation sequencing was performed and an effect of HK4 on the gene expression profile in response to palmitate was observed. A preventive effect of HK4 on palmitate-induced apoptosis was demonstrated by reduced caspase 3/7 activity both in HepG2 (47.32 ± 6.92 % vs. 1065 ± 98.68 %) and human primary hepatocytes (57.13 ± 21.47 % vs. 155.1 ± 3.78 %) . This effect was further confirmed by the TUNEL assay (6.41 ± 1.42 % vs. 11.62 ± 0.99 % TUNEL positive cells) . This anti-apoptotic effect was mediated through reduced protein expression of cleaved PARP-1, reduced phosphorylation of NF-κB, and by upregulation of the ER chaperone PDI, as measured by Western blotting.
In conclusion, GABAergic signaling reduces lipotoxic-induced apoptosis in hepatocytes by preventing inflammation, DNA damage and ER stress. Therefore, we propose that HK4 may arise as an innovative pharmacological tool to treat or prevent NASH as a first-in-class drug.
E.Rohbeck: None. A.Romero: None. B.Knebel: None. B.Belgardt: None. M.Roden: Advisory Panel; Eli Lilly and Company, Research Support; Boehringer Ingelheim International GmbH, Nutricia, Speaker's Bureau; Novo Nordisk. T.Romacho: None. J.Eckel: Stock/Shareholder; CureDiab GmbH.