The origin of brown adipocytes arising in white adipose tissue (WAT) after cold acclimatization is unclear. Here, we demonstrate that several UCP1-immunoreactive brown adipocytes occurring in WAT after cold acclimatization have a mixed morphology (paucilocular adipocytes). These cells also had a mixed mitochondrioma with classic “brown” and “white” mitochondria, suggesting intermediate steps in the process of direct transformation of white into brown adipocytes (transdifferentiation). Quantitative electron microscopy disclosed that cold exposure (6°C for 10 days) did not induce an increase in WAT preadipocytes. β₃-adrenoceptor-knockout mice had a blunted brown adipocyte occurrence upon cold acclimatization. Administration of the β₃-adrenoceptor agonist CL316,243 induced the occurrence of brown adipocytes, with the typical morphological features found after cold acclimatization. In contrast, administration of the β₁-adrenoceptor agonist xamoterol increased only the number of preadipocytes. These findings indicate that transdifferentiation depends on β₃-adrenoceptor activation, whereas preadipocyte recruitment is mediated by β₁-adrenoceptor. RT-qPCR experiments disclosed that cold exposure induced enhanced expression of the thermogenic genes and of genes expressed selectively in brown adipose tissue (iBAT) and in both interscapular BAT and WAT. β₃-adrenoceptor suppression blunted their expression only in WAT. Furthermore, cold acclimatization induced an increased WAT expression of the gene coding for C/EBPα (an antimitotic protein), whereas Ccna1 expression (related to cell proliferation) was unchanged. Overall, our data strongly suggest that the cold-induced emergence of brown adipocytes in WAT predominantly reflects β₃-adrenoceptor-mediated transdifferentiation.

Adipocyte precursor cells give raise to two major cell populations with different physiological roles: white and brown adipocytes. Here we demonstrate that the retinoblastoma protein (pRB) regulates white vs. brown adipocyte differentiation. Functional inactivation of pRB in wild-type mouse embryo fibroblasts (MEFs) and white preadipocytes by expression of simian virus 40 large T antigen results in the expression of the brown fat-specific uncoupling protein 1 (UCP-1) in the adipose state. Retinoblastoma gene-deficient ( Rb ^{– / –} ) MEFs and stem cells, but not the corresponding wild-type cells, differentiate into adipocytes with a gene expression pattern and mitochondria content resembling brown adipose tissue. pRB-deficient MEFs exhibit an increased expression of the Forkhead transcription factor Foxc2 and its target gene cAMP-dependent protein kinase regulatory subunit RIα, resulting in increased cAMP sensitivity. Suppression of cAMP-dependent protein kinase activity in Rb ^–/– MEFs blocked the brown adipocyte-like gene expression pattern without affecting differentiation per se . Immunohistochemical studies revealed that pRB is present in the nuclei of white but not brown adipocyte precursor cells at a developmental stage where both cell types begin to accumulate lipid and brown adipocytes express UCP-1. Furthermore, pRB rapidly undergoes phosphorylation upon cold-induced neodifferentiation and up-regulation of UCP-1 expression in brown adipose tissue. Finally, down-regulation of pRB expression accompanies transdifferentiation of white into brown adipocytes in response to β3-adrenergic receptor agonist treatment. We propose that pRB acts as a molecular switch determining white vs. brown adipogenesis, suggesting a previously uncharacterized function of this key cell cycle regulator in adipocyte lineage commitment and differentiation.

Leptin is a circulating hormone which plays an important role in the regulation of energy balance, haemopoiesis and reproduction. Leptin and its receptor (leptin-R) are localized in human placental tissue but their function is not known. In this study we have investigated the expression of leptin and leptin-R in the human placenta with particular attention to extravillous cytotrophoblastic cell islands and cell columns which play a pivotal role in trophoblast invasion and placental growth. We demonstrate that leptin-R immunoreactivity shows a strong expression in the distal extravillous cytotrophoblastic cells of cell columns invading the basal plate, whereas leptin expression is homogeneously expressed in all the cellular components of cell columns. Since the invasive ability of the distally located extravillous cytotrophoblast of cell columns is known to be regulated by a variety of proteases and some extracellular matrix molecules, we tested the influence of leptin on the in-vitro production of matrix metalloproteinase (MMP)-2, MMP-9 and fetal fibronectin (fFN) by cytotrophoblastic cells. We demonstrate that leptin increases, in a dose-dependent manner, the secretion of immunoreactive MMP-2 and fFN and enhances the activity of MMP-9 in cultured cytotrophoblastic cells. Our results suggest that leptin and leptin-R could have a role in the invasive processes of the extravillous cytotrophoblastic cells by modulating the expression of MMPs. In addition, these results provide a foundation for studying pathological conditions characterized by insufficient or excessive trophoblast invasion.

The uncoupling protein 1 (UCP1) is a hallmark of brown adipocytes and pivotal for cold- and diet-induced thermogenesis. Here we report that cyclooxygenase (COX) activity and prostaglandin E₂ (PGE₂) are crucially involved in induction of UCP1 expression in inguinal white adipocytes, but not in classic interscapular brown adipocytes. Cold-induced expression of UCP1 in inguinal white adipocytes was repressed in COX2 knockout (KO) mice and by administration of the COX inhibitor indomethacin in wild-type mice. Indomethacin repressed β-adrenergic induction of UCP1 expression in primary inguinal adipocytes. The use of PGE₂ receptor antagonists implicated EP₄ as a main PGE₂ receptor, and injection of the stable PGE₂ analog (EP_3/4 agonist) 16,16 dm PGE₂ induced UCP1 expression in inguinal white adipose tissue. Inhibition of COX activity attenuated diet-induced UCP1 expression and increased energy efficiency and adipose tissue mass in obesity-resistant mice kept at thermoneutrality. Our findings provide evidence that induction of UCP1 expression in white adipose tissue, but not in classic interscapular brown adipose tissue is dependent on cyclooxygenase activity. Our results indicate that cyclooxygenase-dependent induction of UCP1 expression in white adipose tissues is important for diet-induced thermogenesis providing support for a surprising role of COX activity in the control of energy balance and obesity development.