Differentiation Of Rat Preadipocytes Research Articles

Effects of ATP-sensitive potassium channels on the expression of P21, P27 and leptin

This study investigated the effects of ATP-sensitive potassium channels on the expression of P21, P27 and leptin. The expression of receptor of ATP-sensitive potassium channels (sulphonylurea receptor, SUR) mRNA in the preadipocytes and leptin mRNA was detected by PCR after rat preadipocytes were treated with the opener (diazoxide) or inhibitor (glibenclamide) of ATP-sensitive potassium channels during the process of inducing differentiation. The expression of P21 and P27 in preadipocytes treated with diazoxide or glibenclamide was assayed by Western blot. The results showed that the expression of SUR2, not SUR1 was detected in adipose tissue, preadipocytes and adipocytes. After treatment of preadipocytes with diazoxide, the expression levels of P21 and P27 were obviously higher than those in control group, but the expression levels of P21 and P27 in glibenclamide-treated group were lower than those in control group. During the process of inducing differentiation, the expression of leptin mRNA in preadipocytes treated with diazoxide was increased greatly, but the expression of leptin mRNA in glibenclamide-treated group decreased obviously. It was concluded that ATP-sensitive potassium channels might be involved in the proliferation and differentiation of rat preadipocytes by changing the expression of P21, P27 and leptin.

Prenatal Protein Restriction Does Not Affect the Proliferation and Differentiation of Rat Preadipocytes

Poor development in utero may favor the development of obesity in adulthood. Animal studies showed that embryo manipulation in vitro or nutritional insults during the embryonic and fetal stages of development may lead to obesity in adult life. We studied the in vitro proliferation and differentiation of adipocytes to investigate whether early protein restriction may program cell growth and development. In a series of experiments, 2 different low-protein diet protocols were compared. In both cases, pregnant rats were fed a diet with a high (18-20%) or low (8-9%) protein content during gestation and/or lactation. Preadipocytes were isolated from the fetuses, neonates, and weanling offspring. Moderate protein restriction, imposed during either gestation and/or lactation, did not affect the capacity of preadipose cells to divide or store fat. Because previous studies showed that early protein restriction alters the metabolism of sulfur amino acids, we also investigated the effects of methionine, taurine, and homocysteine on proliferation and differentiation of preadipocytes. The supplementation of the diet with methionine or the addition of homocysteine and taurine to the culture media did not influence the development of preadipocytes. We obtained no evidence for the direct reprogramming of the precursor or stem cells and suggest that the subsequent alteration in fat accretion may therefore reflect a change in the neuroendocrine environment.

The role of ghrelin and growth hormone secretagogues receptor on rat adipogenesis.

Recent research progress indicates a close link between ghrelin, a natural ligand of GH secretagogues receptor (GHS-R), and both the metabolic balance and body composition. To clarify the involvement of ghrelin and GHS-R in the process of adipogenesis, we measured the expression of GHS-R and peroxisome proliferator-activated receptor gamma 2 (PPAR-gamma 2) mRNA in rat adipocytes using semiquantitative RT-PCR methods. The levels of GHS-R mRNA increased by up to 4-fold in adipose tissue from epididymal and parametrial regions as the rat aged from 4-20 wk and were significantly elevated during the differentiation of preadipocytes in vitro. Ghrelin (10(-8) M for 10 d) stimulated the activity of glycerol-3-phosphate dehydrogenase and the differentiation of rat preadipocytes in vitro. Ghrelin treatment also significantly increased the levels of PPAR-gamma 2 mRNA in primary cultured rat differentiated adipocytes. In addition, isoproterenol (10(-8) M, 40 min)-stimulated lipolysis was significantly reduced by simultaneous ghrelin treatment in a dose-dependent manner in vitro. In conclusion, the expression of GHS-R in rat adipocytes increases with the age and during adipogenesis. Ghrelin in vitro stimulates the differentiation of preadipocytes and antagonizes lipolysis. Ghrelin may therefore play an important role in the process of adipogenesis in rats.

Differentiation of rat preadipocytes is accompanied by expression of thyrotropin receptors.

To investigate the regulation of expression of the TSH receptor (TSHR) in extrathyroidal tissues, the level of TSHR messenger RNA (mRNA) and TSH-dependent signal transduction were determined in isolated rat adipocytes and cultured preadipocytes. The epididymal, sc, and perirenal, but not the interscapular brown adipose tissues, possessed TSHR mRNA and increased cAMP responses to TSH and were thus used as the source of preadipocytes. Morphological analysis revealed that the combination of insulin and T3 most effectively caused the differentiation of rat preadipocytes. These differentiated preadipocytes exhibited increased cAMP production in response to TSH. The addition of FCS to the culture medium inhibited the differentiation of rat preadipocytes as well as TSH-stimulated production of cAMP. The stimulation of differentiation was associated with an increased expression of TSHR mRNA levels, whereas the inhibition of differentiation was associated with a decreased expression of TSHR mRNA, as detected by Northern blot analysis. The results indicate that the expression and function of the TSHR in cultured rat preadipocytes are closely related to cellular differentiation. Cultured rat preadipocytes appear to provide a useful system for studying the mechanism of extrathyroidal expression of TSHR.

Differentiation of rat preadipocytes is accompanied by expression of thyrotropin receptors

To investigate the regulation of expression of the TSH receptor (TSHR) in extrathyroidal tissues, the level of TSHR messenger RNA (mRNA) and TSH-dependent signal transduction were determined in isolated rat adipocytes and cultured preadipocytes. The epididymal, sc, and perirenal, but not the interscapular brown adipose tissues, possessed TSHR mRNA and increased cAMP responses to TSH and were thus used as the source of preadipocytes. Morphological analysis revealed that the combination of insulin and T3 most effectively caused the differentiation of rat preadipocytes. These differentiated preadipocytes exhibited increased cAMP production in response to TSH. The addition of FCS to the culture medium inhibited the differentiation of rat preadipocytes as well as TSH-stimulated production of cAMP. The stimulation of differentiation was associated with an increased expression of TSHR mRNA levels, whereas the inhibition of differentiation was associated with a decreased expression of TSHR mRNA, as detected by Northern blot analysis. The results indicate that the expression and function of the TSHR in cultured rat preadipocytes are closely related to cellular differentiation. Cultured rat preadipocytes appear to provide a useful system for studying the mechanism of extrathyroidal expression of TSHR.

Development of insulin and epidermal growth factor receptors during the differentiation of rat preadipocytes in primary culture

An homogeneous cell population isolated from the inguinal tissue of 3-day-old rats is able to proliferate in primary culture. In the presence of a physiological concentration of insulin (1.5 nM) it converts into cells exhibiting the morphology and the biochemical characteristics of adipocytes. Insulin and epidermal growth factor (EGF) receptors were studied during both the exponential growth and the adipose conversion phases of these cells. Binding experiments with 125I-labelled peptides were performed directly in the culture dishes. The number of high affinity insulin binding sites increased, during the entire culture period studied, reaching 18 days after plating the value of 10 600 × 2360. Control cells (cultured in the presence of anti-insulin antibody) exhibited an increase of the concentration of insulin binding sites from no more than 500 sites/cell to 6880 ± 1710 sites/cell between dat 0 and 9 (corresponding to the exponential growth phase); this increase was followed by a rapid reduction in insulin receptors during the stationary phase. The density of EGF binding sites increased between day 0 and 4 (one cell cycle), whether the cells were maintained or not with insulin, and plateaued therafter. Mature adipocytes freshly isolated from the inguinal tissue of 3-day-old rats had no detectable EGF binding sites, but their content in high affinity binding sites for insulin was similar to that of cells complete adipocyte conversion in primary culture.