Liver-Restricted Repin1 Deficiency Improves Whole-Body Insulin Sensitivity, Alters Lipid Metabolism, and Causes Secondary Changes in Adipose Tissue in Mice

Abstract

Replication initiator 1 (Repin1) is a zinc finger protein highly expressed in liver and adipose tissue and maps within a quantitative trait locus (QTL) for body weight and triglyceride (TG) levels in the rat. The QTL has further been supported as a susceptibility locus for dyslipidemia and related metabolic disorders in congenic and subcongenic rat strains. Here, we elucidated the role of Repin1 in lipid metabolism in vivo. We generated a liver-specific Repin1 knockout mouse (LRep1 2/2 ) and systematically characterized the consequences of Repin1 deficiency in the liver on body weight, glucose and lipid metabolism, liver lipid patterns, and protein/mRNA expression. Hyperinsulinemic-euglycemic clamp studies revealed significantly improved wholebody insulin sensitivity in LRep1 2/2 mice, which may be due to significantly lower TG content in the liver. Repin1 deficiency causes significant changes in potential downstream target molecules including Cd36, Pparg, Glut2 protein, Akt phosphorylation, and lipocalin2, Vamp4, and Snap23 mRNA expression. Mice with hepatic deletion of Repin1 display secondary changes in adipose tissue function, which may be mediated by altered hepatic expression of lipocalin2 or chemerin.Our findings indicate that Repin1 plays a role in insulin sensitivity and lipid metabolism by regulating key genes of glucose and lipid metabolism.Previously, we identified a quantitative trait locus (QTL) for body weight, serum fasting insulin, and triglycerides (TGs) on rat chromosome 4 (1-3). Replication initiator 1 (Repin1) emerged as a potential positional candidate gene within the QTL region considering associations of metabolic alterations in rats with a single nucleotide polymorphism (449C/T) in the Repin1 coding region and with the size of a triplet repeat in the 39-untranslated region of the Repin1 gene (4). Repin1 was initially discovered as replication initiation region protein 60 kDa (RIP60) in a study investigating DNA binding proteins involved in replication activation of the Chinese hamster dihydrofolate reductase gene (dhfr) (5). Repin1 binds to two ATT-rich sites in orib, a short region 39 to the dhfr gene, acting as an enhancer of DNA bending during initiation of DNA synthesis (6,7). Plasmid replication assays demonstrated

Tamoxifen affects glucose and lipid metabolism parameters, causes browning of subcutaneous adipose tissue and transient body composition changes in C57BL/6NTac mice

“…Body weight and food intake were recorded daily; naso-anal length and rectal body temperature (TH-5, Thermalert Monitoring Thermometer, Clifton, NJ, USA) were measured at the end at 18 weeks of age (n ¼ 10 per experimental group). Intraperitoneal insulin tolerance tests (ITTs) were performed at the age of 11 and 16 weeks as described previously [5]. Whole body composition (fat mass, lean mass, water) was determined in awake mice by using NMR technology with EchoMRI700™ instrument (Houston, TX, USA) at 11, 13, 15 and 17 weeks of age.…”

“…Whole body composition (fat mass, lean mass, water) was determined in awake mice by using NMR technology with EchoMRI700™ instrument (Houston, TX, USA) at 11, 13, 15 and 17 weeks of age. Indirect calorimetry was assessed by a Calorimetry Module (TSE Systems, Bad Homburg, Germany) at an age of 16 weeks as previously described [5].…”

“…Adipocytes were isolated from (EPI) and (SC) fat pads by 1 mg/ ml collagenase digestion and adipocyte size distribution was determined in 200 ml suspension in a Coulter Counter (Multisizer III; Beckman Coulter, Krefeld, Germany) as described [5]. Biopsies of SC and EPI AT were fixed in 10% buffered formalin and imbedded in paraffin.…”

“…Total RNA was isolated from SC and EPI AT and cDNA was subsequently amplified as described [5]. Customized primers (Biomers, Ulm, Germany) were used for the detection of Esr1, Ucp1, Ki67, 18Sr and L19r mRNA (Table 1).…”

“…Multiple sections were obtained from EPI and SC fat pads and analyzed systematically with respect to adipocyte size and number. The sections were stained with hematoxylin/eosin and UCP-1 (1:200, Abcam, ab10983, Cambridge, UK) immunohistochemistry was performed as previously described [5].…”