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Sphingolipids suppress preneoplastic rat hepatocytes in vitro and in vivo
Ilona Silins1, Mariann Nordstrand, Johan Högberg
1Institute of Environmental Medicine, Karolinska Institutet, Box 210, S-17177 Stockholm, Sweden.
Abstract:
Sphingolipids can modulate cell growth, differentiation and apoptosis. In the present investigation, selective death of hepatocytes localized in enzyme-altered foci (EAF hepatocytes) was shown to be induced by sphingolipids. Sphingosine (20 micro M) caused rapid cell death predominantly of EAF hepatocytes in vitro. During 4 h of such exposure, cytochrome c was released from the mitochondria into the cytoplasm and the number of cells demonstrating cleaved caspase-9 activity increased. The selective sensitivity of EAF cells to sphingolipid-induced death was attenuated by tumor necrosis factor-alpha. In previous studies we have demonstrated that EAF hepatocytes are resistant to Fas-mediated apoptosis, a resistance shown here to be reversed by low concentrations of sphingosine. Immunohistological staining revealed higher levels of glucosylated ceramide in EAF than in the surrounding tissue. Furthermore, an inhibitor of glucosylation enhanced the toxicity of ceramide towards EAF cells. TLC analysis suggested low levels of sphingosine in preneoplastic lesions. In in vivo experiments EAF-bearing rats were fed a diet supplemented with 0.1% sphingomyelin for 2 weeks. Sphingolipid feeding reduced the number of EAF and EAF area in the liver by 40-50% as compared with rats fed a control diet. These studies indicate that the turnover of sphingolipids in preneoplastic EAF hepatocytes is altered. This alteration may explain not only the increased sensitivity of EAF cells towards sphingolipid-induced cell death, but also the resistance of these hepatocytes to cell death involving sphingolipids as second messengers. Furthermore, sphingomyelin in the diet may prevent EAF development. It is suggested that the altered turnover of sphingolipids might be a target for chemoprevention of hepatocellular carcinoma.
Insights
Altered sphingolipid metabolism in preneoplastic liver cells (EAF hepatocytes) increases their sensitivity to sphingolipid-induced death. Dietary sphingomyelin reduced EAF development, suggesting a chemoprevention target for liver cancer.
Area of Science:
- Biochemistry
- Cell Biology
- Hepatology
Background:
- Sphingolipids play crucial roles in regulating cell growth, differentiation, and apoptosis.
- Enzyme-altered foci (EAF) hepatocytes exhibit altered sensitivity to apoptotic stimuli.
- Previous research indicated EAF hepatocytes are resistant to Fas-mediated apoptosis.
Purpose of the Study:
- To investigate the role of sphingolipids in selective EAF hepatocyte death.
- To explore the impact of sphingolipid metabolism on EAF development.
- To assess the potential of sphingolipids as a chemopreventive strategy for hepatocellular carcinoma.
Main Methods:
- In vitro exposure of EAF hepatocytes to sphingosine and analysis of apoptosis markers (cytochrome c, caspase-9).
- Assessment of EAF cell sensitivity modulation by tumor necrosis factor-alpha and sphingosine.
- Immunohistochemistry and thin-layer chromatography (TLC) to analyze sphingolipid levels in EAF.
- In vivo feeding studies with sphingomyelin-supplemented diets in EAF-bearing rats.
Main Results:
- Sphingosine induced selective death of EAF hepatocytes in vitro, involving mitochondrial release of cytochrome c and caspase-9 activation.
- EAF cell sensitivity to sphingolipids was modulated by tumor necrosis factor-alpha and sphingosine concentrations.
- Elevated glucosylated ceramide and reduced sphingosine levels were observed in EAF compared to normal tissue.
- Dietary sphingomyelin significantly reduced the number and area of EAF in the liver by 40-50%.
Conclusions:
- Sphingolipid turnover is altered in preneoplastic EAF hepatocytes, contributing to their unique apoptotic responses.
- Dietary sphingomyelin demonstrates potential in preventing EAF development, suggesting a role in chemoprevention of liver cancer.
- Targeting altered sphingolipid metabolism presents a promising strategy for the chemoprevention of hepatocellular carcinoma.