Absolute rates of sterol synthesis estimated from [3H]water for bovine lens epithelial cells in culture

Document Type

Article

Publication Title

Journal of Lipid Research

Abstract

All cells of the vascular ocular lens derive from a monolayer of epithelial cells located on only the anterior surface of this organ. The source of the cholesterol required for the growth and division of these cells was studied by using cultures of bovine lens epithelial cells. Cells were in active growth during the third to fourth day of subculture following seeding. Absolute rates of cholesterol synthesis were estimated for the cultured cells from incorporation of [3H]water. Rates were estimated on the assumption that 0.81 atoms of 3H of [3H]water were incorporated into cholesterol per carbon atom of cholesterol, a situation where all of the NADPH would be generated by oxidative enzymatic processes. We tested this assumption by measuring the changes in sterol mass per dish of cells grown in lipoprotein-deficient media over day 3 to 4 of subculture and by simultaneously measuring the rates of incorporation of [3H]water into sterols during this period. In this situation, the increases in sterol mass should be attributable solely to de novo sterol synthesis. We calculated that an average of 0.79 atoms of 3H of [3H]water were incorporated by these cells into cholesterol per carbon atom of cholesterol. Sterol synthesis was only modestly decreased (about 30%) when the cells were cultured in media prepared with whole calf serum. Growth rates of the cells were also little affected by the absence of lipoproteins. In spite of the capacity to furnish its sterol requirements by the novo synthesis, the lens epithelial cells readily degraded 125I-labeled bovine LDL, and LDL greatly decreased sterol synthesis when added to the media at low levels. 125I-Labeled HDL3 was not significantly metabolized by the cultured lens cells, and HDL3 stimulated sterol synthesis when added to the media, apparently by removing sterol synthesis from the cells. In conclusion, bovine lens epithelial cels in culture incorporated 0.79 atoms of 3H of [3H]water into synthesized cholesterol per carbon atom of cholesterol.

First Page

1455

Last Page

1463

Publication Date

12-1-1985

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