The Expression of Peroxisome Proliferator‐Activated Receptor γ in Pig Fetal Tissue and Primary Stromal‐Vascular Cultures

Abstract

The expression of peroxisome proliferator-activated receptor ␥ in pig fetal tissue and primary stromalvascular cultures. Obes Res. 2000:8:83-88. Objective: This study was designed to determine when peroxisome proliferator-activated receptor ␥ (PPAR␥) is expressed in developing fetal adipose tissue and stromalvascular adipose precursor cells derived from adipose tissue. In addition we examined developing tissue for CCAAT/enhancer-binding protein ␤ (C/EBP␤) expression to see if it was correlated with PPAR␥ expression. Pituitary function and hormones involved with differentiation (dexamethasone and retinoic acid) were also tested for their effects on PPAR␥ expression to determine if hormones known to affect differentiation also effect PPAR␥ expression in vivo and in cell culture. Research Methods and Procedures: Developing subcutaneous adipose tissues from the dorsal region of the fetal pig were collected at different gestation times and assayed using Western blot analysis to determine levels of PPAR␥ and C/EBP␤. Hypophysectomy was performed on 75-day pig fetuses and tissue samples were then taken at 105 days for Western blot analysis. Adipose tissue was also taken from postnatal pigs to isolate stromal-vascular (S-V) cells. These adipose precursor cells were grown in culture and samples were taken for Western blot analysis to determine expression levels of PPAR␥. Results: Our results indicate that PPAR␥ is expressed as early as 50 days of fetal development in adipose tissue and continues through 105 days. Expression of PPAR␥ was found to be significantly enhanced in adipose tissue from hypophysectomized fetuses at 105 days of fetal development (p Ͻ 0.05). C/EBP␤ was not found in 50-or 75-day fetal tissues and was found only at low levels in 105-day tissues. C/EBP␤ was not found in hypophysectomized (hypoxed) 105-day tissue where PPAR␥ was elevated. S-V cells freshly isolated from adipose tissue of 5-to 7-day postnatal pigs showed the expression of PPAR␥1. When S-V cells were cultured, both PPAR␥1 and 2 were expressed after the first day and continued as cells differentiated. High concentrations of retinoic acid decreased PPAR␥ expression in early S-V cultures (p Ͻ 0.05). Discussion: Our data indicate that PPAR␥ is expressed in fetal adipose tissue very early before distinct fat cells are observed and can be expressed without the expression of C/EBP␤. The increase in PPAR␥ expression after hypophysectomy may explain the increase in fat cell size under these conditions. Adipose precursor cells (S-V cells) from 5-to 7-day postnatal pigs also express PPAR␥ in the tissue before being induced to differentiate in culture. Thus S-V cells from newborn pig adipose tissue are probably more advanced in development than the 3T3-L1 cell model. S-V cells may be in a state where PPAR␥ and C/EBP␣ are expressed but new signals or vascularization are needed before cells are fully committed and lipid filling begins.

The adipose conversion process: regulation by extracellular and intracellular factors

“…There is still very few data that have emerged from primary cultures despite of their interest, as demonstrated by the variations that have been observed concerning the regulation of adipose conversion by hormonal agents and second messengers. In addition, it has been shown recently that, when compared to cell lines, porcine adipose precursor cells express C/EBPα and PPARγ very early [111,137]. The authors then suggested that porcine preadipocytes are probably more advanced in development than 3T3-L1 cells.…”

Stromal Vascular Cells and Adipogenesis: Cells within Adipose Depots Regulate Adipogenesis

“…Expression of C/EBPα protein was increased and, unlike before adipogenesis, C/EBPα reactive cells are clustered in vivo and in vitro . No change in PPARγ protein was evident in vivo but there was an increase in PPARγ reactive cells in vitro 15,16. Microarray studies indicated that S-V cells expressed over 50 genes encoding for secreted factors detected in vitro (fetal S-V cell cultures) at 90 d of fetal development [Hausman unpublished data], and S-V cells expressed over 150 genes encoding regulatory proteins in vitro at 90 d. Clearly, fetal S-V cells expressed and secreted numerous factors that may be involved in the maintenance of subcutaneous adipose tissue S-V developmental gradients.…”

“…In fetal adipose tissue after adipogenesis five IGFBP proteins were detected in vivo and in vitro : IGFBP-1-5 4. Moreover, IGF-1 was secreted in vitro and IGF-1 mRNA was detected in small adipocytes, stromal cells and endothelial cells 15. Furthermore, TGFβ1 protein and mRNA was detected in adipocytes and stromal cells around developing blood vessels 14.…”

“…Transcription factor expression included PPARγ protein expression in the nucleus of a small number of cells in vitro (fetal S-V cell cultures) and PPARγ protein expression in vivo despite the absence of lipid-filled adipocytes 15,16. Additionally, C/EBPα protein was detected in vivo and C/EBPα reactive cells were present in vitro , but not yet clustered 16,17.…”

Enhancing Effect of Troglitazone on Porcine Adipocyte Differentiation in Primary Culture: A Comparison with Dexamethasone

“…Adipocyte differentiation and expression of C/EBPα and PPARγ transcripts and proteins have been studied during in vitro differentiation of porcine S‐V cells (7, 8, 9). Experiments under conditions similar to ours showed that the development of C/EBPα immunoreactive cells and preadipocytes (AD‐3 reactive cells) preceded lipid accretion (9).…”