Abstract

The rat carboxylesterases, ES‐2 (serum esterase), ES‐3 (pI 5.6 esterase), ES‐4 (pI 6.2/6.4 esterase), ES‐10 (pI 6.0/6.1 esterase), and ES‐15 (pI 5.0/5.2 esterase) comprise a multi‐gene family, and function in hydrolysis of a variety of ester‐ and amide‐ containing chemicals to their respective free acids. Using realtime PCR, in the rat hepatic cell line McArdle‐RH7777 (McA) we observed ES‐4 mRNA had the highest level of expression compared to ES‐2 and ES‐3, while ES‐10 was not detectable. The subcellular location of ES‐4 in McA cells was investigated using confocal microscopy. ES4 co‐localized with ER Tracker dye, a maker for the endoplasmic reticulum (ER). Immunohistochemical studies with rat liver demonstrated the presence of ES‐4 within hepatocytes. ES‐4 also functions as a rat hepatic cholesteryl ester hydrolase. McA cells overexpressing ES‐4 showed decreased cholesteryl ester (CE) levels compared to control cells indicating that ES‐4 can hydrolyze CE in intact cells. In cells where ES‐4 levels were suppressed by 75% using siRNA there were increased CE levels. Confocal microscopy, using Oil Red O and ES‐4 antibodies, revealed areas of association of ES‐4 with the CE droplets. The results show ES‐4 is expressed in hepatocytes and specifically in endoplasmic reticulum. ES‐4 is directly associated with CE droplets and functions as cholesteryl ester hydrolase. Supported by NIH‐DK044498.

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