Priming with magnesium-deficient media inhibits preadipocyte differentiation via potential upregulation of tumor necrosis factor-alpha.

The effect of priming stromal-vascular cells in primary cultures with magnesium-deficient (MgD) media on preadipocyte differentiation was studied. Cultures were derived from dorsal subcutaneous fat tissue of young pigs and maintained 3 d in serum-free control or MgD Dulbecco's modified Eagle's medium, 3 d in 10% fetal bovine serum and dexamethasone, and 6 d in insulin. At d 12 of culture, immunocytochemical and glycerophosphate dehydrogenase assays indicated depressed adipocyte differentiation in the MgD groups. Cultures were enriched for preadipocytes up to 50% of total cells. On the third day of treatment with control and MgD medium, total cell lysates were isolated and 50 microg of them were run on two-dimensional gel electrophoresis. The separated proteins from both treatment groups showed similar patterns. However, spots of proteins with predicted molecular weight in the range of 25.8-37.4 kDa and pI of 5.39-5.85 were sixfold denser from the MgD 10 groups than from the controls. These proteins migrate similarly to tumor necrosis factor-alpha (TNF-alpha). The amount of TNF-alpha in cell lysates from the MgD group was about 2.35 times greater than controls determined by TNF-alpha-ELISA. It is likely that proteins upregulated by MgD medium are TNF-alpha isoforms.