Preparation and Metabolism of Isolated Cells from Bovine Adipose Tissue1,2

Abstract

A procedure for isolating bovine adipose cells was developed, and glucose and acetate metabolism and effects of insulin were studied.Acetate oxidation to CO2 and conversion to fatty acid in isolated bovine fat cells was stimulated greatly by minimal concentrations of glucose. Insulin enhanced glucose uptake. Responsiveness to insulin was lower than reported for adipose tissue of young rats. Intensive transfer of reducing equivalents between NADPH2 and NADH2 and between mitochondria and cytoplasm was indicated and may have occurred via the isocitrate-α-ketoglutarate and oxalacetatemalate shuttles. Pentose phosphate pathway activity was not affected by presence and absence of fatty acid synthesis from acetate. Thls lack of feedback response may be due to high rates of lactate release and glyceride-glycerol formation and possible transfer of reducing equivalents between cytoplasmic NADP and mitochondrial NAD. These factors would prevent drastic changes in cytoplasmic concentrations of free reduced or oxidized NADP. Fatty acid synthesis in bovine adipose cells was limited by low concentrations of extracellular acetate. When acetate is not limiting, fatty acid synthesis may be limited by glucose availability. If transfer of reducing equivalents from mitochondria to cytoplasm via the proposed isocitrate-α-ketoglutarate shuttle is sufficiently active, the NADPH2 required for fatty acid synthesis in presence of excess acetate could be furnished by minimal quantities of glucose required to supply Kreb's cycle intermediates. Fatty acid synthesis might be limited then by lack of α-glycerol phosphate for fatty acid esterification. Relative glucose carbon flows via pentose phosphate and Embden-Meyerhof pathways were altered by epinephrine. This probably was mediated through changes in phosphofructokinase activity and intracellular free fatty acid concentrations.