Regulation of autophagy in rat hepatocytes treated in vitro with low concentration of mercury

Several proteins are implicated in the regulation of autophagy in cells under various physiological and pathological conditions. Recently it was found that in vitro initiation of autophagy in 5 μM mercuric chloride (HgCl₂) treated rat hepatocytes occurred within 30 min of incubation. The aim of this study was to monitor the autophagy-ubiquitination link to determine the regulatory mechanisms underlying autophagy at a concentration of 5 μM HgCl₂. Autophagic cell death was evidenced by the presence of LC3B positive hepatocytes throughout the incubation period of 4 hr. Autophagosome maturation occurred following 1 hr of metal treatment accompanied by an enhanced expression of several regulatory proteins such as p62, Keap1, and caspase-8, which drive the hepatocytes toward autophagy. In addition, the rate of increase in down-regulation in a p38 expression was significantly higher than that of extracellular signal-regulated kinase (ERK) at 1 hr incubation, followed by a significant decline in the p38 level at 2 and 4 hr following metal treatment. In contrast, the expression of ERK remained higher than that of p38 at 2 and 4 hr. Data indicate that intracellular activation of the major regulatory proteins such as LC3B, caspase-8, Keap1, p62, ERK, and p38 modulates autophagy through ubiquitination in rat hepatocytes exposed to low concentrations of Hg.