The effect of insulin on primary cultures of rat preadipocytes grown in fetal or postnatal pig serum

Abstract

Stromal vascular cell cultures, prepared from the inguinal pads of 50-g Sprague Dawley rats, were exposed to media with 10% fetal pig serum which is inherently low in insulin, for the first 3 to 5 d of culture. Insulin was supplemented to media for periods of 2 to 6 d. In cultures treated (2 to 4 d) with 10(-9), to 10(-10) or 10(-11) M insulin, differentiated cells (lipid and esterase staining) appeared 1.5 to 2 times wider than differentiated cells in control cultures. At 10(-9) M insulin (4 to 5 d), in cultures grown in the presence of fetal pig serum the number of esterase reactive cells was increased twofold to threefold. The percentage of total cells that were esterase reactive was elevated 50 to 300% relative to control cultures. Insulin-treated preadipocytes were more reactive for lipoprotein lipase activity (histochemical assay) compared with reactivity of control cells. Quantitative analysis of percentage of light transmittance (Zeiss photometer) through stained cells indicated an increase (P less than .001) in lipoprotein lipase staining at 10(-9), 10(-11) and 10(-13) M insulin (2 d). The specific activity of glycerol phosphate dehydrogenase was elevated twofold to threefold (P less than .05) and soluble protein elevated 50 to 100% (P less than .05) in cultures treated (3 to 6 d) with 10(-9) M insulin. Decreasing the cell plating density (50%) in cultures grown in the presence of pig serum reduced the elevation in enzyme activity induced by insulin in preadipocyte cultures. Physiological levels of insulin enhanced lipogenic enzyme activity in preadipocytes and may enhance the conversion of stromal cells to preadipocytes.