ABSTRACT
The release of Ca(2+) from the endoplasmic reticulum (ER) and subsequent replenishment of ER Ca(2+) by Ca(2+) entry through store-operated Ca(2+) channels (SOCE) play critical roles in the regulation of liver metabolism by adrenaline, glucagon and other hormones. Both ER Ca(2+) release and Ca(2+) entry are severely inhibited in steatotic hepatocytes. Exendin-4, a slowly-metabolised glucagon-like peptide-1 (GLP-1) analogue, is known to reduce liver glucose output and liver lipid, but the mechanisms involved are not well understood. The aim of this study was to determine whether exendin-4 alters intracellular Ca(2+) homeostasis in steatotic hepatocytes, and to evaluate the mechanisms involved. Exendin-4 completely reversed lipid-induced inhibition of SOCE in steatotic liver cells, but did not reverse lipid-induced inhibition of ER Ca(2+) release. The action of exendin-4 on Ca(2+) entry was rapid in onset and was mimicked by GLP-1 or dibutyryl cyclic AMP. In steatotic liver cells, exendin-4 caused a rapid decrease in lipid (half time 6.5min), inhibited the accumulation of lipid in liver cells incubated in the presence of palmitate plus the SOCE inhibitor BTP-2, and enhanced the formation of cyclic AMP. Hormone-stimulated accumulation of extracellular glucose in glycogen replete steatotic liver cells was inhibited compared to that in non-steatotic cells, and this effect of lipid was reversed by exendin-4. It is concluded that, in steatotic hepatocytes, exendin-4 reverses the lipid-induced inhibition of SOCE leading to restoration of hormone-regulated cytoplasmic Ca(2+) signalling. The mechanism may involve GLP-1 receptors, cyclic AMP, lipolysis, decreased diacylglycerol and decreased activity of protein kinase C.
Subject(s)
Calcium Signaling/drug effects , Fatty Liver/pathology , Glucagon-Like Peptide 1/analogs & derivatives , Hepatocytes/metabolism , Intracellular Space/metabolism , Peptides/pharmacology , Venoms/pharmacology , Animals , Bucladesine/pharmacology , Calcium/pharmacology , Cyclic AMP/metabolism , Exenatide , Glucagon-Like Peptide 1/metabolism , Hormones/pharmacology , Intracellular Space/drug effects , Palmitic Acid/pharmacology , Rats, ZuckerABSTRACT
Lipid accumulation in hepatocytes can lead to non-alcoholic fatty liver disease (NAFLD), which can progress to non-alcoholic steatohepatitis (NASH) and Type 2 diabetes (T2D). Hormone-initiated release of Ca²âº from the endoplasmic reticulum (ER) stores and subsequent replenishment of these stores by Ca²âº entry through SOCs (store-operated Ca²âº channels; SOCE) plays a critical role in the regulation of liver metabolism. ER Ca²âº homoeostasis is known to be altered in steatotic hepatocytes. Whether store-operated Ca²âº entry is altered in steatotic hepatocytes and the mechanisms involved were investigated. Lipid accumulation in vitro was induced in cultured liver cells by amiodarone or palmitate and in vivo in hepatocytes isolated from obese Zucker rats. Rates of Ca²âº entry and release were substantially reduced in lipid-loaded cells. Inhibition of Ca²âº entry was associated with reduced hormone-initiated intracellular Ca²âº signalling and enhanced lipid accumulation. Impaired Ca²âº entry was not associated with altered expression of stromal interaction molecule 1 (STIM1) or Orai1. Inhibition of protein kinase C (PKC) reversed the impairment of Ca²âº entry in lipid-loaded cells. It is concluded that steatosis leads to a substantial inhibition of SOCE through a PKC-dependent mechanism. This enhances lipid accumulation by positive feedback and may contribute to the development of NASH and insulin resistance.
