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Abstract Dexamethasone (DEX) is known to induce diabetes and dyslipidemia. We have compared fasting triacylglycerol and cholesterol concentrations across 20 lipoprotein fractions and glucose tolerance in control (standard diet) and DEX-treated 7-month-old males of two rat strains, Brown Norway (BN) and congenic BN.SHR-(Il6-Cd36)/Cub (BN.SHR4). These two inbred strains differ in a defined segment of chromosome 4, originally transferred from the spontaneously hypertensive rat (SHR) including the mutant Cd36 gene, a known target of DEX. Compared to BN, the standard-diet-fed BN.SHR4 showed higher cholesterol and triacylglycerol concentrations across many lipoprotein fractions, particularly in small VLDL and LDL particles. Total cholesterol was decreased by DEX by more than 21% in BN.SHR4 contrasting with the tendency to increase in BN (strain*DEX interaction p = 0.0017). Similar pattern was observed for triacylglycerol concentrations in LDL. The LDL particle size was significantly reduced by DEX in both strains. Also, while control BN and BN.SHR4 displayed comparable glycaemic profiles during oral glucose tolerance test, we observed a markedly blunted DEX induction of glucose intolerance in BN.SHR4 compared to BN. In summary, we report a pharmacogenetic interaction between limited genomic segment with mutated Cd36 gene and dexamethasone-induced glucose intolerance and triacylglycerol and cholesterol redistribution into lipoprotein fractions.

Dexamethasone (DEX) is known to induce diabetes and dyslipidemia. We have compared fasting triacylglycerol and cholesterol concentrations across 20 lipoprotein fractions and glucose tolerance in control (standard diet) and DEX-treated 7-month-old males of two rat strains, Brown Norway (BN) and congenic BN.SHR-(Il6-Cd36)/Cub (BN.SHR4). These two inbred strains differ in a defined segment of chromosome 4, originally transferred from the spontaneously hypertensive rat (SHR) including the mutant Cd36 gene, a known target of DEX. Compared to BN, the standard-diet-fed BN.SHR4 showed higher cholesterol and triacylglycerol concentrations across many lipoprotein fractions, particularly in small VLDL and LDL particles. Total cholesterol was decreased by DEX by more than 21% in BN.SHR4 contrasting with the tendency to increase in BN (strain*DEX interaction p = 0.0017). Similar pattern was observed for triacylglycerol concentrations in LDL. The LDL particle size was significantly reduced by DEX in both strains. Also, while control BN and BN.SHR4 displayed comparable glycaemic profiles during oral glucose tolerance test, we observed a markedly blunted DEX induction of glucose intolerance in BN.SHR4 compared to BN. In summary, we report a pharmacogenetic interaction between limited genomic segment with mutated Cd36 gene and dexamethasone-induced glucose intolerance and triacylglycerol and cholesterol redistribution into lipoprotein fractions.

Endogenous retroviruses (ERVs) contribute to a range of germline, as well as somatic mutations in mammals. However, autonomous retrotransposition of potentially active elements has not been demonstrated in the rat genome. We cloned an insertion that disrupted the normal splicing of the Cntrob gene that was subsequently identified as a nonautonomous, novel endogenous retrovirus of the RnERV-K8e family. The RnERV-K8e family is closely related to the recently reported MmERV-K10c elements, but differs from the autonomous mouse MusD or IAP families. In addition, we identified a novel, unexpectedly close relative of RnERV-K8e in the mouse, suggesting ERV-K cross-species transmission between mice and rats. We cloned a potentially autonomous RnERV-K8e element identified by in silico analysis and, using an in vitro retrotransposition assay, demonstrated that it is capable of retrotransposition. This particular element (named Rat-rho, pronounced "retro") encodes a retroviral envelope gene (env); however, env is not required for de novo retrotransposition events. Significant levels of RnERV-K8e-associated genetic polymorphisms were detected among inbred rat strains, suggesting ongoing retrotransposition in the rat genome. This study identifies an ERV-K-type family in rats that shows obvious signs of recent activity. Ongoing retrotranspositional activity may significantly add to genomic variability among inbred rat strains.

Rat hypodactyly (hd) is an autosomal recessive mutation manifesting in homozygotes as reduction or loss of digits II and III. We mapped the hd allele to a short segment of chromosome 10, containing 16 genes. None of these genes has been shown to influence limb development yet. In situ hybridization showed no changes in several important patterning genes (Shh, Fgf8, Bmp2, 4, 7). However, we found that expression of cartilage condensation marker Sox9, and Bmp receptor Bmpr1b (acting as an upstream activator of Sox9 expression) is absent from the subepithelial mesenchyme of the digit condensations II and III. The failure of the chondrogenic condensations to extend towards the subepithelial mesenchyme may reduce the size of digit primordia and underlie the subsequent loss of phalanges and reduction of metacarpals/metatarsals in hd rats.

Aims: Therapeutic administration of retinoids is often accompanied with undesirable side effects, including an increase in lipid levels in up to 45% of treated patients. We tested the hypothesis of whether spontaneously hypertensive rat (SHR) and congenic SHR.PD-(D8Rat42-D8Arb23)/Cub (SHR-Lx) strains, differing only in a 14-gene region of chromosome 8 and previously shown to display differential sensitivity to the teratogenic effects of retinoic acid, could serve as a pharmacogenetic model set of the metabolic side effects of retinoid therapy., Materials & Methods: Male, 15-week old rats (n = 12/strain) of SHR and SHR-Lx strains were fed a high-sucrose diet for 2 weeks and subsequently treated either with all-trans retinoic acid (15 mg/kg) or only with a vehicle for 16 days (n = 6/strain/treatment), while still on the high-sucrose diet. We assessed the morphometric and metabolic profiles of all groups, including glucose tolerance tests, levels of insulin, adiponectin, free fatty acids, concentrations of triglycerides and cholesterol in 20 lipoprotein fractions under conditions of both high-sucrose diet and high-sucrose diet plus all-trans retinoic acid administration., Results & Conclusion: SHR-Lx displayed substantially greater sensitivity to a number of all-trans retinoic acid-induced metabolic dysregulations compared with SHR, resulting in impairment of glucose tolerance, increased visceral adiposity, and substantially greater increase of circulating triglyceride concentrations, accompanied by a shift towards their less favorable distribution into the lipoprotein fractions. These observations closely mimic the common side effects of retinoid therapy in humans, rendering SHR-Lx an experimental pharmacogenetic model of atRA-induced dyslipidemia.

The hypodactylous (hd) locus impairs limb development and spermatogenesis, leading to male infertility in rats. We show that the hd mutation is caused by an insertion of an endogenous retrovirus into intron 10 of the Cntrob gene. The retroviral insertion in hd mutant rats disrupts the normal splicing of Cntrob transcripts and results in the expression of a truncated protein. During the final phase of spermiogenesis, centrobin localizes to the manchette, centrosome, and the marginal ring of the spermatid acroplaxome, where it interacts with keratin 5-containing intermediate filaments. Mutant spermatids show a defective acroplaxome marginal ring and separation of the centrosome from its normal attachment site of the nucleus. This separation correlates with a disruption of head-tail coupling apparatus, leading to spermatid decapitation during the final step of spermiogenesis and the absence of sperm in the epididymis. Cntrob may represent a novel candidate gene for presently unexplained hereditary forms of teratozoospermia and the "easily decapitated sperm syndrome" in humans.

Lx mutation in SHR.Lx rat manifests in homozygotes as hindlimb preaxial polydactyly. It was previously mapped to a chromosome 8 segment containing the Plzf gene. Plzf (promyelocytic leukemia zinc finger protein) influences limb development as a direct repressor of posterior HoxD genes. However, the Plzf coding sequence is intact in the Lx mutants. Using linkage mapping in F2 hybrids, we downsized the segment containing Lx to 155 kb and sequenced conserved noncoding elements (CNEs) inside. A 2,964-bp deletion in Plzf intron 2, never detected in control animals, is the only candidate for Lx. The deletion removes the most deeply conserved CNE in the 155-kb segment, suggesting a regulatory influence on Plzf expression. Correspondingly, using in situ hybridization and quantitative real-time polymerase chain reaction, we found a decrease of Plzf expression in Lx/Lx limb buds with concomitant anterior expansion of expression domains of its targets, Hoxd10-13 genes, in the absence of ectopic Sonic hedgehog expression. Upstream regulation of Plzf in limb buds is currently unknown. We present here the first candidate Plzf cis-regulatory sequence., ((c) 2009 Wiley-Liss, Inc.)

Introduction: Thiazolidinediones are increasingly used drugs for the treatment of Type 2 diabetes. The individual response to thiazolidinedione therapy, ranging from the variable degree of metabolic improvement to harmful side-effects, is empirical, yet the underlying mechanisms remain elusive. In order to assess the pharmacogenomic component of thiazolidinediones' metabolic action, we compared the effect of rosiglitazone in two genetically defined models of metabolic syndrome, polydactylous (PD) and BN.SHR4 inbred rat strains, with their insulin-sensitive, normolipidemic counterpart, the Brown Norway (BN) rat., Materials & Methods: 5-month-old male rats were fed a high-fat diet for 4 weeks, and the experimental groups received rosiglitazone (0.4 mg/100 g body weight) during the last 2 weeks of high-fat diet feeding. We assessed metabolic and morphometric profiles, oxidative stress parameters and gene expression in white adipose tissue., Results: In many followed parameters, we observed genetic background-specific effects of rosiglitazone administration. The mass and the sensitivity of visceral adipose tissue to insulin-stimulated lipogenesis increased with rosiglitazone treatment only in PD, correlating with a PD-specific significant increase in expression of prostaglandin D2 synthase. The glucose tolerance was enhanced in all strains, although fasting plasma glucose was increased by rosiglitazone in BN and BN.SHR4. Among the markers of lipid peroxidation, we observed the rosiglitazone-driven increase of plasma-conjugated dienes only in BN.SHR4. The genes with genotype-specific expression change included ADAM metallopeptidase domain 7, aquaporin 9, carnitine palmitoyltransferase 1B, caveolin 1, catechol-O-methyl transferase, leptin and prostaglandin D2 synthase 2., Conclusion: Rosiglitazone's effects on lipid deposition and insulin sensitivity of peripheral tissues are largely dependent on the genetic background it acts upon.

The importance of early environment, including maternal diet during pregnancy, is suspected to play a major role in pathogenesis of metabolic syndrome and related conditions. One of the proposed mechanisms is a mismatch between the prenatal and postnatal environments, leading to misprogramming of the metabolic and signaling pathways of the developing fetus. We assessed whether the exposure to high-sucrose diet (HSD) alleviates the detrimental effects of sucrose feeding in later life (predictive adaptive hypothesis) in a highly inbred model of metabolic syndrome, the PD/Cub rat. Rat dams were continuously fed either standard or HSD (70% calories as sucrose) starting 1 wk before breeding, throughout pregnancy, at birth, and until weaning of the offspring. After weaning, all male offspring were fed HSD until the age of 20 wk, when detailed metabolic and morphometric profiles were ascertained. The early life exposure to a sucrose-rich diet resulted in distinct responses to longtime postnatal HSD feeding. Offspring of the sucrose-fed mothers displayed higher adiposity and substantial increases in triglyceride liver content together with unfavorable distribution of cholesterol into lipoprotein subfractions. On the other hand, their adiponectin concentrations were significantly higher, and insulin sensitivity of skeletal muscle was enhanced compared with the offspring of standard diet-fed mothers. Triglycerides, free fatty acids, overall glucose tolerance, and the insulin sensitivity of adipose tissue were comparable in both groups. In the genetically identical animals, maternal HSD feeding elicited a variety of subtle effects but did not lead to predictive adaptive protection from most HSD-induced metabolic derangements.

We have developed a new, double-congenic rat strain BN-Lx.SHR2, which carries two distinct segments of chromosome 2 introgressed from the spontaneously hypertensive rat strain (SHR) into the genetic background of congenic strain BN-Lx, which was previously shown to express variety of metabolic syndrome features. In 16-wk-old male rats of BN-Lx and BN-Lx.SHR2 strains, we compared their glucose tolerance and triacylglycerol and cholesterol concentrations in 20 lipoprotein subfractions and the lipoprotein particle sizes under conditions of feeding standard and high-sucrose diets. Introgression of two distinct SHR-derived chromosome 2 segments resulted in decreased adiposity together with aggravation of glucose intolerance in the double-congenic strain. The BN-Lx.SHR2 rats were more sensitive to sucrose-induced rise in triacylglycerolemia. Although the total cholesterol concentrations of the two strains were comparable after the standard diet and even lower in BN-Lx.SHR2 after sucrose feeding, detailed analysis revealed that under both dietary conditions, the double-congenic strain had significantly higher cholesterol concentrations in low-density lipoprotein fractions and lower high-density lipoprotein fractions. We established a new inbred model showing dyslipidemia and mild glucose intolerance without obesity, attributable to specific genomic regions. For the first time, the chromosome 2 segments of SHR origin are shown to influence other than blood pressure-related features of metabolic syndrome or to be involved in relevant nutrigenomic interactions.

We have previously established a congenic strain SHR-Lx that carries a differential segment of rat chromosome 8 introgressed from a model of metabolic syndrome--the polydactylous rat strain--on the genomic background of spontaneously hypertensive rat (SHR). We compared the glucose tolerance and lipid profile of adult SHR and SHR-Lx males under conditions of standard diet and diets enriched in sucrose and cholesterol, respectively. While there was no evident difference between the SHR and SHR-Lx on standard diet, the one-week sucrose administration revealed the congenic strain sensitivity to carbohydrate-induced dyslipidemia conferred by the differential segment with only mild derangement of glucose tolerance. On the other hand, the high-cholesterol diet administration for three-weeks resulted in a contrasting pattern as the congenic strain displayed significantly lower concentrations of free fatty acids and improved glucose tolerance compared to SHR. After one-month washout period, the SHR-Lx showed higher insulin, triglyceride and cholesterol concentrations together with diminished insulin sensitivity of visceral adipose tissue. In summary, we have identified a genomic region syntenic to human chromosome 11q23, which determines complex nutrigenetic interactions under conditions of sucrose- and cholesterol-enriched diets.

The SHR and the PD/Cub are two established rodent models of human metabolic syndrome. Introgression of a ca 30 cM region of rat chromosome 8 from PD/Cub onto the genetic background of SHR was previously shown to influence several of the metabolic syndrome-related traits along with causing the PLS in the SHR-Lx congenic strain. In the process of identification of the causative alleles, we have produced several congenic sublines. The differential segment of SHR-Lx PD5 congenic substrain [SHR.PD(D8Rat42-D8Arb23)/Cub] spans approximately 1.4 Mb encompassing only 14 genes. When comparing the metabolic, morphometric and gene expression profiles of the SHR-Lx PD5 vs. SHR, the polydactyly and several distinct metabolic features observed in the original SHR-Lx congenic were still manifested, suggesting that the responsible genes were "trapped" within the relatively short differential segment of PD/Cub origin in SHR-Lx PD5. Particularly, the SHR-Lx PD5 displayed substantial reduction of insulin sensitivity confined to skeletal muscle. Among the candidate genes, the promyelocytic leukaemia zinc-finger Plzf (Zbtb16) transcription repressor is most likely responsible for the Lx mutation resulting in PLS and could also be involved in the alteration of metabolic pathways. The sequence analysis of the Plzf gene revealed a SNP leading to a threonine to serine substitution in SHR at aminoacid position 208 (T208S). In summary, we have isolated a 1.4 Mb genomic region syntenic to human chromosome 11q23, which, apart from causing polydactyly-luxate syndrome (PLS), affects total body weight, adiposity, lipid profile, insulin sensitivity of skeletal muscle and related gene expression as shown in the SHR-Lx PD5 congenic substrain.

Germinal epithelium of seminiferous tubules in adult, infertile hypodactylous males displays significant reduction in the number of germ-line cells. Detection of apoptosis in the germ-line cells during postnatal differentiation was performed to elucidate the mechanism of the decreased number of germ cells in the testes of adult rats. Evaluation of DNA fragmentation and expression of activated caspase-3 in germ cells did not confirm marked germ cell death during the onset of spermatogenesis as a main cause of significant reduction of germ cells in Hd/Hd testes of adult males. The primary cause of spermatogenesis defect seems to be rather associated with a disorder in the cell cycle regulation and interrelation of germ-line cells with Sertoli cells.

In the current review, we summarize results of genetic analyses of "metabolic syndrome" in the spontaneously hypertensive rat (SHR). These results include (1) linkage analyses in the HXB/BXH recombinant inbred (RI) strains derived from SHR and Brown Norway (BN-Lx) strains which revealed quantitative trait loci (QTL) for hemodynamic and metabolic traits on several chromosomes, (2) genetic isolation of these putative QTL within differential chromosome segments of SHR.BN congenic strains, (3) detailed mapping of these QTL within limited chromosome segments of SHR.BN congenic sublines, (4) sequencing of selected positional candidate genes which revealed important mutations in the Cd36 and Srebp1 SHR genes, (5) functional tests of these candidate genes in SHR transgenic lines, and (6) integrated gene expression profiling and linkage mapping in RI strains which will be used to identify co-regulated genes and to determine co-segregation of transcriptional profiles with physiological and pathophysiological phenotypes.