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Cadmium is a toxic element to which man can be exposed at work or in the environment. Cd's most salient toxicological property is its exceptionally long half-life in the human body. Once absorbed, Cd accumulates in the human body, particularly in the liver. The cellular actions of Cd are extensively documented, but the molecular mechanisms underlying these actions are still not resolved. The liver manages the cadmium to eliminate it by a diverse mechanism of action. Still, many cellular and physiological responses are executed in the task, leading to worse liver damage, ranging from steatosis, steatohepatitis, and eventually hepatocellular carcinoma. The progression of cadmium-induced liver damage is complex, and it is well-known the cellular response that depends on the time in which the metal is present, ranging from oxidative stress, apoptosis, adipogenesis, and failures in autophagy. In the present work, we aim to present a review of the current knowledge of cadmium toxicity and the cellular response in the liver.

Metabolic factors are the major risk of non-alcoholic fatty liver disease, although other factors may contribute steatosis. Cadmium exposure produces histopathological and molecular changes in liver, which are consistent with steatosis. In the present study, we describe the effect of low cadmium acute treatment on hepatocytes obtained from mice fed with a high cholesterol diet. Our data suggest that hepatocytes with cholesterol overload promote an adaptive response against cadmium-induced acute toxicity by up-regulating anti-apoptotic proteins, managing ROS overproduction, increasing GSH synthesis and MT-II content to avoid protein oxidation. Cadmium treatment increases lipid content in cholesterol-fed mice hepatocytes because of an impaired autophagy process. Our data suggest an essential function of macroautophagy in the regulation of lipid storage induced by Cd on hepatocytes, that implies that alterations in this pathway may be a mechanism that aggravates hepatic steatosis.

The mechanism of cadmium-mediated hepatotoxicity has been the subject of numerous investigations, principally in hepatocytes. Although, some uncertainties persist, sufficient evidence has emerged to provide a reasonable account of the toxic process in parenchymal cells. However, there is no information about the effect of cadmium in other hepatic cell types, such as stellate cells (fat storing cells, Ito cells, perisinusoidal cells, parasinusoidal cells, lipocytes). Hepatic stellate cells (HSC) express a quiescent phenotype in a healthy liver and acquire an activated phenotype in liver injury. These cells play an important role in the fibrogenic process. The objective of this study was to investigate the effect of a 24 h treatment of low Cd concentrations in glutathione content, lipid peroxidation damage, cytosolic free Ca, antioxidant enzyme activities: glutathione peroxidase, glutathione reductase, superoxide dismutase and catalase along with the capacity of this heavy metal to induce metallothionein II and alpha(1)collagen (I) in an hepatic stellate cell line (CFSC-2G). Cd-treated cells increased lipid peroxidation and the content of cytosolic free calcium, decreased glutathione content and superoxide dismutase, glutathione peroxidase and catalase activity. Cd was able to induce the expression of the metallothionein II and alpha(1)collagen (I) gene, that was not described in this cell type. Cadmium may act as a pro-fibrogenic agent in the liver probably by inducing oxidative damage by enhancing lipid peroxidation and altering the antioxidant system of the cells. Although, the exact role metallothionein induction plays in this process is unknown, it probably, provides a cytosolic pool of potential binding sites to sequester ionic Cd, thereby decreasing its toxicity.

Inflammatory mediators, including cytokines, growth factors, and reactive oxygen species, are associated with the pathology of chronic liver disease. In the liver, cytokine and growth factor secretion are usually associated with nonparenchymal cells, particularly Kupffer cells. In the present studies, the effect of 24 and 72 h administration of ethanol (50 mM). acetaldehyde (175 microM), and LPS (1 microg/ml) were studied on the expression and secretion of TNF-alpha, IL-1beta, IL-6, and TGF-beta3, lipid peroxidation damage and glutathion content in HepG2 cell cultures. A 24 h exposure to ethanol induced the expression of TNF-alpha and TGF-beta1, and the secretion of IL-1beta and TGF-beta1. With the same period of treatment, acetaldehyde markedly increased TNF-alpha expression, and stimulated IL-1beta secretion, while LPS exposure induced the expression of TNF-alpha, IL-6, and TGF-beta1, and the secretion of IL-1beta, IL-6, and TGF-beta1. A reduced in TNF-alpha response and TGF-beta1 expression were observed after 72 h exposure to ethanol. A 72 h acetaldehyde exposure decreased markedly TNF-alpha expression and stimulated a previously absent TGF-beta1 response. With the same time of exposure, LPS reduced slightly TGF-beta1 expression, and decreased its secretion. IL-1beta and IL-6 were not detected under 72 h exposure conditions. Lipid peroxidation damage was increased in all treatments, but higher values were found in 72 h treatments. Glutathion content diminished in all treatments. These findings suggest that HepG2 cells, independent of other cells such as Kupffer or macrophages, participate in a differential cytokine, growth factor and oxidative stress response, which differs according to the toxic agent and the time of exposure.

The effects of acute ethanol and acetaldehyde treatment on cell proliferation, cell adhesion capacity, neutral red incorporation into lysosomes, glutathione content, protein sulfhydryl compounds, lipid peroxidation, inner mitochondrial membrane integrity (MTT test), lactate dehydrogenase activity (LDH) and ultrastructural alterations were investigated in a human fetal hepatic cell line (WRL-68 cells). WRL-68 cells were used, due to the fact that, although this cell line expresses some hepatic characteristics, it does not express alcohol dehydrogenase or cytochrome P450 activity, so it could be a good model to study the effect of the toxic agents per se. Cells were exposed during 120 min with 200 mM ethanol or 10 mM acetaldehyde. Under these conditions, cells presented 100% viability and no morphological alteration was observed by light microscopy. Acetaldehyde-treated cells reduced their proliferative capacity drastically while the ethanol-treated ones presented no difference with control cells. Cell adhesion to substrate, measured as time required to adhere to the substrate and time required to detach from the substrate, was diminished in acetaldehyde WRL-68-treated cells. Cytotoxicity measures as neutral red and MTT test showed that acetaldehyde-treated cells presented more damage than ethanol-treated ones. Cellular respiratory capacity was compromised by acetaldehyde treatment due to 40% less oxygen consumption than control cells. Lipid peroxidation values, measured as malondialdehyde production, were higher in ethanol-treated WRL-68 cells (127%) than in acetaldehyde-treated ones (60%) to control cell values. Lactate dehydrogenase activity (LDH) in extracellular media of ethanol-treated cells presented the highest values. GSH content was reduced 95% and thiol protein content was diminished severely in acetaldehyde-treated cells. Transmission electron microscopy showed more ultrastructural alterations in cells treated with acetaldehyde. The results indicate that acetaldehyde, like ethanol, produced damage at cellular level, although more damage could be observed in acetaldehyde WRL-68-treated cells.

Toxic metals appear to use the transport pathways that exist for biologically essential metals. Calcium uptake in cells occurs through specific membrane channels. Since cadmium inhibits calcium uptake, this study was carried on to elucidate the mechanism of Cd interference with calcium transport using the fetal hepatic cell line WRL-68 as an in vitro model. Ca accumulation by WRL-68 cells presented an initial rapid phase, followed by a sustained phase of slower accumulation over a 60 min period. A concentration of 50 microM CdCl2 produced 39% inhibition of the uptake of CaCl2 (100 microM), while 100 microM nifedipine or verapamil decreased Ca accumulation by 35 and 63%, respectively. All Cd concentrations tested produced significant decrease in Ca uptake in a concentration-dependent manner at 1 min and thereafter, although with 10 microM CdCl2 no significant difference was found after 30 min of incubation. From the Lineweaver-Burk plot, we found that Cd exerted a competitive inhibition on Ca uptake, since there was no significant effect on the Vmax but an increased K(m). A second order rate constant of Cd inactivation of 0.061 mM-1.s-1 was determined from the course of Ca uptake during Cd inhibition. SH groups seemed to play an essential role in Ca inhibition uptake by Cd because the inhibition of Ca accumulation by 50 microM Cd was practically reversed after the addition of dithiothreitol.

The toxic effects of cadmium (Cd) and mercury (Hg), as chloride salts, were studied using an hepatic human fetal cell line (WRL-68 cells). From viability curves and the proliferative capacity of the cell in the presence of the metal, three different cell treatments were chosen, (1) 0.5 microM of the metal chloride for 24 h (acute low dose treatment), (2) 0.5 microM of the metal chloride for 7 days (chronic treatment), and (3) 5 microM of the metal chloride for 24 h (acute high dose treatment). WRL-68 cells grown in the presence of Cd exhibited the same proliferative curve as control cells, whereas in the case of Hg, the cells increased their proliferative capacity. Both metals produced ultrastructural alterations in different degrees, mainly observed as mitochondrial and RER structural changes, depending of the treatment and concentration of the metal used. Cytotoxicity was assessed by measuring the release of lactate dehydrogenase from the cells. Acutely high dose-treated cells showed the highest value for this parameter, and Cd-treated cells presented higher lactate dehydrogenase release than the Hg-treated ones. Cell damage was also measured by alanine aminotransferase (ALAT) and aspartate aminotransferase (ASAT) activities. Acute high dose Cd treatment caused the highest value of enzymatic release. Lipid peroxidation was significantly different with respect to control cells in chronic and acute high dose treatments with both metals. Metallothionein (MT) induction in response to Hg treatment was not detected. However, a dramatic induction of this protein occurred in Cd-treated cells. WRL-68 cells differentially respond to Cd and Hg making this hepatic fetal human cell line a useful tool in investigating the mechanism of toxicity of these heavy metals.

Cadmium (Cd) has been regarded as one of the inflammation-related xenobiotics. Cd has been extensively studied in many cellular systems, but a lot of parameters have been evaluated in different experimental conditions. This study was undertaken to examine the effects of low cadmium concentrations in HepG2 cells in the oxidative stress produced, the IL-1beta, tumor necrosis factor (TNF-alpha), IL-6, and IL-8 expression, production of heat shock protein 70 (Hsp70) and the activation of nuclear factors activation protein-1 (AP-1) and NF-kappaB under the same experimental conditions. Also, the participation of TNF-alpha and oxidative stress in AP-1 activation was evaluated. Lipid peroxidation damage increased 1.5 times after the first hour of Cd treatment and increased 1.9 times after 2h. Similar values were maintained until 6h. Reduced glutathione (GSH) diminished 65% after 6h CdCl(2) treatment. N-acetylcysteine (NAC) pre-treatment increased 332% GSH in Cd-treated cells. RNA was isolated from HepG2 cells after 0.5, 1, 3, or 6h incubation with 1, 5, or 10 microM CdCl(2). TNF-alpha and IL-1beta presented a maximum response after 1h treatment, while IL-6 and IL-8 maximum response was after 3h treatment. The Hsp70, determined by Western blot, was constitutively produced, and it increased after 3h Cd treatment. NF-kappaB activation, determined by EMSA, was not increased as a result of Cd treatment. DNA binding of AP-1 was detected and increased, with time up to 4h with an increment of 24 times control value with 5 microM CdCl(2). The HepG2 cells were pretreated with anti-TNF-alpha antibody or 1mM N-acetylcysteine 1h before Cd treatment. Anti-TNF-alpha treatment reduced 67% AP-1 activation, while NAC 47.5%. These data indicate that, Cd-induced TNF-alpha and IL-1beta, that probably, activate AP-1 transcription factor and IL-6 and IL-8 were induced. Anti-TNF-alpha and NAC partially inhibited AP-1 activation. All imply that, a number of factors participate in AP-1 cadmium-induced activation. The Hsp70 is produced by the HepG2 cells after cadmium treatment, and probably has a role in the non-participation of NF-kappaB in the cellular response.

A hepatic human cell line (WRL-68 cells) was employed to investigate the uptake of the toxic heavy metal cadmium. Cd accumulation in WRL-68 cells is a time-, temperature- and concentration-dependent process. A rapid initial phase of uptake was followed by a second slower phase. The transport does not require energy and 55% of Cd transport occurs by temperature-insensitive processes, possibly by diffusion. The rest of Cd transport (45%) occurs by temperature-sensitive processes, probably ion channels and carriers, that involve interaction with sulfhydryl groups. The calcium channel blockers nifedipine and verapamil inhibit the uptake of cadmium, with an inhibition of 35% after 30 min incubation with 100 microM verapamil and 10 microM Cd. These data suggest that about one third of the Cd enters WRL-68 cells through the calcium channels. The toxic metals appear to use the transport pathways that exist for biologically essential metals. Our results in human hepatic cells are very similar to those reported in cultured rat hepatocytes. It appears that transport pathways available for Cd uptake are similar and independent of the species of hepatocyte origin. Moreover, the WRL-68 cell line seems to be an excellent in vitro model to study the mechanism of cell damage due to Cd.