RXR antagonism induces G0 /G1 cell cycle arrest and ameliorates obesity by up-regulating the p53-p21(Cip1) pathway in adipocytes.
ABSTRACT The peroxisome proliferator activated receptor-γ (PPARγ) agonist, pioglitazone (PIO), exerts anti-diabetic properties associated with increased fat mass, whereas the retinoid X receptor (RXR) antagonist HX531 demonstrates anti-obesity and anti-diabetic effects with reduced body weight and fat pad mass. The cell cycle abnormality in adipocytes has not been well-investigated in obesity or during treatment with modulators of nuclear receptors. We therefore investigated cell size and cell cycle distributions of adipocytes in vivo and examined the expression of cell cycle regulators in cultured human visceral preadipocytes. The cell size distribution and cell cycle analyses of in vivo adipocytes derived from OLETF rats demonstrated that HX531 brought about G0/G1 cell cycle arrest associated with the inhibition of cellular hypertrophy, which resulted in the reduction of fat pad mass. In contrast, PIO promoted proliferation activities associated with the increase in M + late M:G0 + G1 ratio and the appearance of both small and hypertrophied adipocytes. In cultured human visceral preadipocytes HX531 up-regulated cell cycle regulators, p53, p21(Cip1), cyclin D1, Fbxw7 and Skp2, which are known contributors towards G0 /G1 cell cycle arrest. The knockdown of p53 with a shRNA lentivirus reversed the HX531-induced up-regulation of p21(Cip1), which is one of the major p53-effector molecules. We conclude that HX531 exerts anti-obesity and anti-diabetes properties by up-regulating the p53-p21(Cip1) pathway, resulting in G0/G1 cell cycle arrest and the inhibition of cellular hypertrophy of adipocytes.
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ABSTRACT: Early diabetic nephropathy is characterized by glomerular hypertrophy. Previous studies in vitro have demonstrated that mesangial cells exposed to high glucose are arrested in the G1-phase of the cell cycle and express increased levels of the cyclin-dependent kinase inhibitor p27Kip1. The present study was performed to investigate the renal expression of p27Kip1 in db/db mice, a model of diabetes mellitus type II. Glomerular p27Kip1 protein, but not mRNA expression, was strongly enhanced in diabetic db/db mice compared with non-diabetic db/+ littermates. Immunohistochemical studies revealed that this stimulated expression was mainly restricted to the nuclei of mesangial cells and podocytes, but glomerular endothelial cells occasionally also stained positively. Quantification of p27Kip1 positive glomerular cells showed a significant increase of these cells in db/db mice compared with non-diabetic db/+ animals. Although tubular cells revealed a positive staining for p27Kip1 protein, there was no difference between db/+ and db/db mice. Immunoprecipitation experiments revealed that p27Kip1 protein associates with Cdk2 and Cdk4, but not with Cdk6. To test for the influence of hyperglycemia on cell cycle arrest and p27Kip1 expression, mesangial cells were isolated from db/+ and db/db mice. There was a similar basal proliferation when these cells were grown in normal glucose-containing medium (100 mg/dl). However, raising the glucose concentration to 275 to 450 mg/dl induced cell cycle arrest in db/+ as well as db/db mesangial cells. Increasing the medium osmolarity with D-mannitol failed to induce p27Kip1 expression in mesangial cells. Transfection of cells with p27Kip1 antisense, but not missense, phosphorothioate oligonucleotides facilitated cell cycle progression equally well in db/+ and db/db mesangial cells. Furthermore, p27Kip1 expression was comparable in both cell lines in normal glucose, but increased in high glucose medium. Our studies demonstrate that p27Kip1 expression is enhanced in diabetic db/db animals. This induction appears to be due to hyperglycemia. Expression of p27Kip1 may be important in cell cycle arrest and hypertrophy of mesangial cells during early diabetic nephropathy.Kidney International 04/1998; 53(4):869-79. · 7.92 Impact Factor
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ABSTRACT: Regulation of the exit of cells from the cell cycle is important in the development of multicellular organisms and is also implicated in the maintenance of stem cells. Furthermore, defects in cell cycle exit are thought to be a major cause of cancer. However, the mechanisms responsible for regulation of cell cycle exit have remained largely unknown. Fbxw7 is the F-box protein subunit of an SCF-type ubiquitin ligase complex that targets positive regulators of the cell cycle-including cyclin E, c-Myc, Notch and c-Jun-for ubiquitylation and subsequent degradation by the 26S proteasome in order to promote cell cycle exit. Consistent with such a function, mutations of the Fbxw7 gene have been detected in various human malignancies. We have recently generated conventional and conditional Fbxw7 knockout mice and examined stem cells, progenitor cells and differentiated cells in the mutant animals for cell cycle defects. Here we summarize the pleiotropic phenotypes of Fbxw7 deficiency in various cell types including T cells, hematopoietic stem cells and embryonic fibroblasts. Such phenotypes have provided insight into the biological roles of Fbxw7 in cell cycle exit, stem cell maintenance and oncosuppression.Cell cycle (Georgetown, Tex.) 12/2008; 7(21):3307-13. · 5.24 Impact Factor
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ABSTRACT: All-trans-retinoic acid (RA) treatment of acute promyelocytic leukemia (APL) cases expressing the t(15;17) product, PML/RARalpha, is a successful example of differentiation therapy. Uncovering RA target genes is of considerable interest in APL. This study comprehensively examines in APL cells transcriptional and post-transcriptional regulation of the novel candidate RA target gene, G0S2, the G0/G1 switch gene. Reverse transcription (RT)-polymerase chain reaction (PCR) and heteronuclear PCR assays performed +/- treatment with the protein synthesis inhibitor cycloheximide (CHX) revealed G0S2 induction within 3 h of RA-treatment. Treatment with the RNA synthesis inhibitor actinomycin D did not implicate G0S2 transcript stabilization in the RA-mediated increase of G0S2 mRNA expression. Promoter elements of G0S2 were cloned into a reporter plasmid and retinoic acid receptor (RAR) co-transfection assays confirmed transcriptional activation after RA-treatment. Consistent with G0S2 being a direct RA target gene, retinoic acid response element (RARE) half-sites were found in this promoter. Mutation of these sites blocked RA-transcriptional activation of G0S2. To extend analyses to the protein expression level, a polyclonal anti-G0S2 antibody was derived and detected murine and human G0S2 species. G0S2 protein was rapidly induced in cultured NB4-S1 human APL cells and in APL transgenic mice treated with RA. An RAR pan-antagonist confirmed dependence on RARs for this induction. That these findings are clinically relevant was shown by analyses of APL cells derived directly from patients. These leukemic cells induced both a prominent increase in the cellular differentiation marker nitrotetrazolium blue (NBT) staining and marked increase in G0S2 expression. Taken together, these findings indicate G0S2 is an RA target gene. The functional role of G0S2 in retinoid response of APL warrants further study.International Journal of Oncology 08/2008; 33(2):397-404. · 2.66 Impact Factor