Inactivation of the Rcan2 Gene in Mice Ameliorates the Age- and Diet-Induced Obesity by Causing a Reduction in Food Intake

Obesity is a serious international health problem that increases the risk of several diet-related chronic diseases. The genetic factors predisposing to obesity are little understood. Rcan2 was originally identified as a thyroid hormone-responsive gene. In the mouse, two splicing variants that harbor distinct tissue-specific expression patterns have been identified: Rcan2-3 is expressed predominately in the brain, whereas Rcan2-1 is expressed in the brain and other tissues such as the heart and skeletal muscle. Here, we show that Rcan2 plays an important role in the development of age- and diet-induced obesity. We found that although the loss of Rcan2 function in mice slowed growth in the first few weeks after birth, it also significantly ameliorated age- and diet-induced obesity in the mice by causing a reduction in food intake rather than increased energy expenditure. Rcan2 expression was most prominent in the ventromedial, dorsomedial and paraventricular hypothalamic nuclei governing energy balance. Fasting and refeeding experiment showed that only Rcan2-3 mRNA expression is up-regulated in the hypothalamus by fasting, and loss of Rcan2 significantly attenuates the hyperphagic response to starvation. Using double-mutant (Lepob/ob Rcan2−/−) mice, we were also able to demonstrate that Rcan2 and leptin regulate body weight through different pathways. Our findings indicate that there may be an Rcan2-dependent mechanism which regulates food intake and promotes weight gain through a leptin-independent pathway. This study provides novel information on the control of body weight in mice and should improve our understanding of the mechanisms of obesity in humans

Aristaless-related homeobox plays a key role in hyperplasia of the pancreas islet α-like cells in mice deficient in proglucagon-derived peptides.

Defects in glucagon action can cause hyperplasia of islet α-cells, however, the underlying mechanisms remain largely to be elucidated. Mice homozygous for a glucagon-GFP knock-in allele (Gcg(gfp/gfp) ) completely lack proglucagon-derived peptides and exhibit hyperplasia of GFP-positive α-like cells. Expression of the transcription factor, aristaless-related homeobox (ARX), is also increased in the Gcg(gfp/gfp) pancreas. Here, we sought to elucidate the role of ARX in the hyperplasia of α-like cells through analyses of two Arx mutant alleles (Arx(P355L/Y) and Arx ([330insGCG]7/Y) ) that have different levels of impairment of their function. Expression of Gfp and Arx genes was higher and the size and number of islets increased in the Gcg(gfp/gfp) pancreas compared to and Gcg(gfp/+) pancreas at 2 weeks of age. In male Gcg(gfp/gfp) mice that are hemizygous for the Arx(P355L/Y) mutation that results in a protein with a P355L amino acid substitution, expression of Gfp mRNA in the pancreas was comparable to that in control Gcg(gfp/+)Arx(+/Y) mice. The increases in islet size and number were also reduced in these mice. Immunohistochemical analysis showed that the number of GFP-positive cells was comparable in Gcg(gfp/gfp) Arx(P355L/Y) and Gcg(gfp/+)Arx(+/Y) mice. These results indicate that the hyperplasia is reduced by introduction of an Arx mutation. Arx(P355L/Y) mice appeared to be phenotypically normal; however, Arx ([330insGCG]7/Y) mice that have a mutant ARX protein with expansion of the polyalanine tract had a reduced body size and shortened life span. The number of GFP positive cells was further reduced in the Gcg(gfp/gfp) Arx ([330insGCG]7/Y) mice. Taken together, our findings show that the function of ARX is one of the key modifiers for hyperplasia of islet α-like cells in the absence of proglucagon-derived peptides

Pancreatic Neuroendocrine Tumors in Mice Deficient in Proglucagon-Derived Peptides.

Animal models with defective glucagon action show hyperplasia of islet α-cells, however, the regulatory mechanisms underlying the proliferation of islet endocrine cells remain largely to be elucidated. The Gcggfp/gfp mice, which are homozygous for glucagon/green fluorescent protein knock-in allele (GCGKO), lack all proglucagon-derived peptides including glucagon and GLP-1. The present study was aimed to characterize pancreatic neuroendocrine tumors (panNETs), which develop in the GCGKO mice. At 15 months of age, macroscopic GFP-positive tumors were identified in the pancreas of all the GCGKO mice, but not in that of the control heterozygous mice. The tumor manifested several features that were consistent with pancreatic neuroendocrine tumors (panNETs), such as organoid structures with trabecular and cribriform patterns, and the expression of chromogranin A and synaptophysin. Dissemination of GFP-positive cells was observed in the liver and lungs in 100% and 95%, respectively, of 15-month-old GCGKO mice. To elucidate the regulatory mechanism for tumor growth, PanNET grafts were transplanted into subrenal capsules in GCGKO and control mice. Ki-67 positive cells were identified in panNET grafts transplanted to GCGKO mice 1 month after transplantation, but not in those to control mice. These results suggest that humoral factors or conditions specific to GCGKO mice, are involved in the proliferation of panNETs. Taken together, GCGKO mice are novel animal model for studying the development, pathogenesis, and metastasis panNETs

Expression of <i>Gfp</i>, <i>glucagon</i>, <i>insulin</i> and <i>Arx</i> mRNAs in the pancreas of <i>Gcg/Arx</i> double mutant mice.

<p>Relative mRNA levels for <i>Gfp</i> (A) <i>glucagon</i> (B), <i>insulin1</i> (C), and <i>Arx</i> (D) in the pancreas of 2 weeks old mice are shown. Number of animals is indicated in parenthesis in each column. ‘aa’ vs <i>Gcg^gfp/+Arx^+/Y</i>, P<0.01; ‘bb’ vs <i>Gcg^gfp/gfpArx^+/Y</i>, P<0.01; ‘cc’ vs <i>Gcg^gfp/+Arx^PL/Y</i>, P<0.01; ‘dd’ vs <i>Gcg^gfp/gfpArx^PL/Y</i>, P<0.01; ‘ee’ vs <i>Gcg^gfp/+Arx^7/Y</i>, P<0.01; ‘f’ vs <i>Gcg^gfp/gfpArx^7/Y</i>, P<0.05; ‘ff’ vs <i>Gcg^gfp/gfpArx^7/Y</i>, P<0.01.</p