Expression of carboxylesterase and lipase genes in rat liver cell-types

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Abstract

Approximately 80% of the body vitamin A is stored in liver stellate cells with in the lipid droplets as retinyl esters. In low vitamin A status or after liver injury, stellate cells are depleted of the stored retinyl esters by their hydrolysis to retinol. However, the identity of retinyl ester hydrolase(s) expressed in stellate cells is unknown. The expression of carboxylesterase and lipase genes in purified liver cell-types was investigated by real-time PCR. We found that six carboxylesterase and hepatic lipase genes were expressed in hepatocytes. Adipose triglyceride lipase was expressed in Kupffer cells, stellate cells and endothelial cells. Lipoprotein lipase expression was detected in Kupffer cells and stellate cells. As a function of stellate cell activation, expression of adipose triglyceride lipase decreased by twofold and lipoprotein lipase increased by 32-fold suggesting that it may play a role in retinol ester hydrolysis during stellate cell activation.

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Materials and methods

Isolation of rat liver cells. The following protocol was approved by the Institutional Animal Care and Use Committee at Indiana University Purdue University at Indianapolis. The isolation of rat liver cells was adapted from published methods [18], [19], [20]. Male Wistar rats (Charles Rivers Labs, Wilmington, MA) weighing 500–600 g were used. In situ liver perfusion was performed via the portal vein, sequentially with 150 ml MEM without Ca2+ and Mg2+, 100 ml of 0.4% pronase (Roche, Indianapolis,

Isolation of liver cells

Disruption of the liver basement membrane with collagenase generates a mixture of hepatocytes and non-parenchymal cells. Hepatocytes are larger in size and spin down at 60g. Repeated washing of the hepatocyte pellet removes most of the smaller non-parenchymal cells. The typical yield of hepatocytes was 1.7 × 108 cells per liver. For isolation of non-parenchymal cells, which account for approximately 25–35% of liver cells, the liver was perfused with 0.4% pronase to digest most of the hepatocytes.

Discussion

The primary goal of this study was to identify the liver cell-specific expression of carboxylesterases and lipases that could be candidates for REH. Among different liver cell-types, hepatic lipase is exclusively expressed in the hepatocytes and is the only lipase expressed in these cells (Fig. 4B). Hepatic lipase can be secreted in the space of Disse by the hepatocytes where it can play a role in hydrolysis of chylomicron remnants (Fig. 1) and their uptake [1], [23]. In this study, we find

Acknowledgments

This work was supported by NIH Grant R01DK063141. We dedicate this work in fond memory of Dr. Judy White who had initiated the cell imaging work on this project.

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