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Distribution and concentration of beta-adrenergic receptor (betaAR) subtype transcripts (beta1AR, beta2AR, and beta3AR) were investigated in porcine and rat tissues. Reverse transcription (RT) coupled with PCR indicated the presence of beta1AR and beta2AR transcripts in porcine left ventricle, lung, longissimus muscle, and subcutaneous adipose tissue. The RT-PCR indicated that beta3AR transcripts were present in porcine subcutaneous adipose tissue, and transcripts were suggested in left ventricle and lung, but they were not detected in longissimus muscle. Quantitative ribonuclease protection assays indicated that the porcine beta1AR, beta2AR, and beta3AR transcript concentrations (amol/microg of total RNA) were, respectively as follows: heart left ventricle = 6.1, 2.4, and .02; lung = 3.8, 1.9, and .01; liver = 1.7, 2.1, and undetectable; skeletal muscle = 1.7, 1.1, .02; subcutaneous adipose tissue = 1.3, .4, .14. The proportions of porcine betaAR subtypes (beta1AR:beta2AR:beta3AR) were as follows: heart left ventricle = 72:28:.25; lung = 67:33:.2; liver = 45:55:0; skeletal muscle = 60:39:.7; and subcutaneous adipose = 73:20:7. Normalization of data to beta-actin transcript concentration did not change these relationships. Rat perigonadal adipose tissue beta1AR and beta3AR transcript concentrations were, respectively, .59 and 1.84 amol/microg of total RNA. The rat perigonadal adipose tissue concentration of beta3AR transcript was tenfold that of porcine subcutaneous adipose tissue. The predominant betaAR transcript in rat adipose tissue was beta3AR, and the predominant betaAR transcript in pig adipose tissue was beta1AR.

Conjugated linoleic acids (CLAs) reduce fat deposition in several mammalian species. Among the proposed mechanisms for this effect are reduced preadipocyte proliferation and differentiation. We measured proliferation and differentiation of cultured human preadipocytes treated with CLAs. Preadipocytes were differentiated with insulin, hydrocortisone, transferrin, and 10% fetal bovine serum, with isobutyl-methylxanthine included for the first 2 d. The differentiation medium contained 200 microM oleic acid (C18:1), 50 microM cis-9,trans-11-CLA (9,11-CLA), or 50 microM trans-10,cis-12-CLA (10,12-CLA); the negative control medium contained no added fatty acid, and the cells did not differentiate. Cell number increased three to four times during the 17 d of differentiation, but was 30-35% lower in the CLA-treated cells than in the negative control cells. Compared with the negative control cells, differentiation was increased in the cells treated with C18:1 (increased Oil Red O-stained material [OROSM], triacylglycerol, glycerol 3-phosphate dehydrogenase activity [GPDH], peroxisome proliferator-activated receptor-gamma [PPAR gamma] messenger ribonucleic acid [mRNA], and lipoprotein lipase [LPL] mRNA). In effect, the C18:1-treated cells act as a positive control to demonstrate the differentiation capacity of each cell lot. Both 9,11-CLA- and 10,12-CLA-treated cells had increased differentiation (increased OROSM, triacylglycerol, GPDH, PPAR gamma, and LPL) compared with the negative control cells. The data suggest that early in differentiation when de novo fatty acid (FA) synthesis is limited and competition for FAs by membrane and triacylglycerol synthetic pathways is great, human preadipocytes do not differentiate unless a PPAR gamma ligand is added. Either CLA isomer or C18:1 can provide such a ligand.

The transcription factor, sterol regulatory binding protein 1c (also called adipocyte determination and differentiation-dependent factor 1), stimulates transcription of the messenger ribonucleic acids (mRNAs) for lipid synthesis enzymes. Hepatic ADD1 transcripts are reduced by polyunsaturated fatty acids (PUFAs). The ADD1 transcripts are expressed to a considerable extent in porcine adipocytes. Consequently, it was of interest to examine the effects of several PUFAs on ADD1 in a tissue wherein several long-chain fatty acids (FAs) increase adipocyte differentiation. The effects of arachidonic acid (C20:4), docosahexaenoic acid (C22:6), and cis 9, trans 11-conjugated linoleic acid (9,11-CLA) on differentiating preadipocyte ADD1 mRNA and protein and on preadipocyte differentiation were determined. Porcine stromal-vascular cells were plated in serum-containing medium and differentiated in serum-free medium containing insulin, hydrocortisone, and transferrin +/- an individual FA. After 24-h differentiation +/- FA, plates were stained with Oil Red O as an indicator of differentiation or total RNA was extracted or a nuclear fraction was isolated for protein measurement. Addition of C20:4 or 9,11-CLA increased the number of Oil Red O-stained cells or the Oil Red O-stained material, whereas C22:6 did not. Addition of C20:4, C22:6, or 9,11-CLA decreased the concentration of the mRNA and protein for ADD1. Thus, although all three FAs decreased the ADD1 mRNA and protein concentrations, C20:4 and 9,11-CLA increased differentiation, measured by Oil Red O staining, whereas C22:6 did not. The data suggest that the regulation of differentiation and mRNAs by individual FAs may involve distinct mechanisms.

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