Optimization of in vitro conditions for bovine subcutaneous and intramuscular preadipocyte differentiation

¹ Department of Animal Science, Michigan State University, East Lansing, MI 48824-1225
² Departments of Animal Science and Food Science and Human Nutrition, Michigan State University, East Lansing, MI 48824-1225

^* To whom correspondence should be addressed. E-mail: buskirk{at}msu.edu.

	Abstract

The objective of these experiments was to develop an in vitro cell culture system for differentiation of bovine preadipocytes, which will permit examination of differences in differentiation between intramuscular (i.m.) and subcutaneous (s.c.) bovine preadipocytes. Stromal-vascular cells from bovine i.m. and s.c. adipose depots were isolated and cultured. Clonally derived s.c. preadipocytes were used to determine the ability of insulin, bovine serum lipids, octanoate, acetic acid, dexamethasone (DEX), and troglitazone (TRO) to elicit differentiation of these cells when added to serum-free medium. Addition of 10 and 20 µL/mL of a commercially available serum lipids supplement to low glucose Dulbecco's modified Eagle's medium containing 280 nM insulin increased glycerol-3-phosphate dehydrogenase (GPDH) activity (P < 0.01). Inclusion of 1.25 to 10 µM TRO to medium containing 280 nM insulin and 20 µL/mL serum lipids supplement also increased GPDH activity (P < 0.001) compared with 0 µM TRO. The combination of 280 nM insulin, 1 mM octanoate, and 10 mM acetic acid, with 48 h exposure to 0.25 µM DEX caused morphological differentiation in a small number of cells but did not stimulate GPDH activity (P = 0.99). When used together, 280 nM insulin, 20 µL/mL serum lipids supplement, 40 µM TRO, and 0.25 µM DEX stimulated differentiation compared with the aforementioned treatment (P < 0.001). Omission of TRO or insulin from this medium reduced GPDH activity by 68% (P < 0.001), while removal of DEX tended to reduce activity (P = 0.06). Preadipocytes from s.c. (n = 3) and i.m. (n = 2) adipose tissues of three steers were used to determine the effects of TRO on differentiation using the established conditions. Forty to 60 µM TRO enhanced differentiation compared with 0 µM TRO (P < 0.02) in both depots. No depot differences in response to TRO were detected (P = 0.32). These data demonstrate that bovine preadipocytes are capable of differentiation in response to combinations of insulin, serum lipids, DEX, and TRO. Although TRO enhanced differentiation of bovine preadipocytes, no differential effects of TRO on the differentiation of s.c. and i.m. cells were detected.

Key Words: bovine, differentiation, glycerol-3-phosphate dehydrogenase, preadipocyte, thiazolidinedione