Related Experiment Videos
Wy-14,643 but not 2-ethylhexanol increases intracellular free calcium in cultured Kupffer cells
T Hijioka1, B J Keller, R G Thurman
1Department of Pharmacology, University of North Carolina, Chapel Hill 27599-7365.
Abstract:
Hypolipidemic drugs and phthalic ester plasticizers induce peroxisomes and cause hepatocellular carcinoma in rodents by mechanisms which remain unknown. Recent evidence from this laboratory suggests that many agents in this class of chemicals are uncouplers of mitochondrial oxidative phosphorylation both in vitro and in vivo. Uncoupling of oxidative phosphorylation decreases ATP required for ion pumps and could thereby indirectly increase intracellular free calcium. The goal of these experiments, therefore, was to compare the effect of the potent uncoupler and non-genotoxic carcinogen Wy-14,643 with the weaker agent 2-ethylhexanol on intracellular free calcium in cultured Kupffer cells. Kupffer cells, the resident hepatic macrophages, are activated by calcium and release a variety of mitogenic growth factors that may modulate cell proliferation. In this study, the cytosolic free calcium concentration in Fura-2-loaded cultured Kupffer cells was increased significantly from 78 +/- 11 to 838 +/- 112 nM following incubation with Wy-14,643 (1.25 mM). The increase in intracellular calcium due to Wy-14,643 was both time- and dose-dependent. At equimolar concentrations, ethylhexanol had no effect on intracellular calcium (65 +/- 20 nM). However, at higher concentrations (3 mM), ethylhexanol also increased intracellular calcium. These data suggest that elevation of intracellular calcium in Kupffer cells may be involved in the mechanism of action of this interesting class of non-genotoxic carcinogens.
Insights
Certain drugs and plasticizers may cause cancer by increasing intracellular calcium in liver cells. This study investigated how Wy-14,643 and 2-ethylhexanol affect calcium levels in Kupffer cells, potentially revealing mechanisms of non-genotoxic carcinogenesis.
Area of Science:
- Toxicology
- Cell Biology
- Carcinogenesis
Background:
- Hypolipidemic drugs and phthalic ester plasticizers are known to induce peroxisomes and cause liver cancer in rodents.
- The underlying mechanisms for this non-genotoxic carcinogenesis remain largely unknown.
- Recent evidence suggests these chemicals may act as uncouplers of mitochondrial oxidative phosphorylation, potentially affecting intracellular calcium.
Purpose of the Study:
- To investigate the role of intracellular free calcium in Kupffer cells (resident hepatic macrophages) in response to Wy-14,643 and 2-ethylhexanol.
- To compare the effects of a potent uncoupler (Wy-14,643) and a weaker agent (2-ethylhexanol) on intracellular calcium levels.
Main Methods:
- Primary cultured rat Kupffer cells were loaded with the calcium indicator Fura-2.
- Cells were incubated with varying concentrations of Wy-14,643 and 2-ethylhexanol.
- Cytosolic free calcium concentration was measured using fluorescence microscopy.
Main Results:
- Wy-14,643 (1.25 mM) significantly increased intracellular free calcium in Kupffer cells from 78 ± 11 nM to 838 ± 112 nM.
- The calcium increase induced by Wy-14,643 was time- and dose-dependent.
- 2-ethylhexanol had no effect at equimolar concentrations but increased intracellular calcium at higher concentrations (3 mM).
Conclusions:
- Elevation of intracellular free calcium in Kupffer cells may be a key mechanism by which Wy-14,643 and potentially other related non-genotoxic carcinogens exert their effects.
- Activated Kupffer cells release mitogenic factors that can modulate cell proliferation, suggesting a link between calcium signaling and carcinogenesis.