BACKGROUND/AIM:The typical insulin deficiency in type 1 diabetes mellitus has general effects on metabolism and also affects bone quality.MATERIALS AND METHODS:Two diabetic rat lines (BB/OK; BB.6KWR) and two non-diabetic rat strains (KWR and BB.14+18KWR), as control group, were included in the study. Bone mineral density, bone mineral content and body structure measurements were performed. The measurements took place before the onset of diabetes mellitus Results: A comparison of the groups showed increased bone density values of the diabetic rats in relation to the control groups. A new finding of increased bone density in the diabetic rats occurs.CONCLUSION:Diabetic rats showed no osteoporotic bone metabolism before the onset of clinically relevant type 1 diabetes mellitus, but rather increased bone metabolic activity.

The genetic architecture of obesity is multifactorial. We have previously identified a quantitative trait locus (QTL) on rat chromosome 10 in a F2 cross of Wistar Ottawa Karlsburg (WOKW) and Dark Agouti (DA) rats responsible for obesity-related traits. The QTL was confirmed in congenic DA.WOKW10 rats. To pinpoint the region carrying causal genes, we established two new subcongenic lines, L1 and L2, with smaller refined segments of chromosome 10 to identify novel candidate genes. All lines were extensively characterized under different diet conditions. We employed transcriptome analysis in visceral adipose tissue (VAT) by RNA-Seq technology to identify potential underlying genes in the segregating regions. Three candidate genes were measured in human paired samples of VAT and subcutaneous (SC) AT (SAT) (N=304) individuals with a wide range of body weight and glucose homeostasis parameters. DA.WOKW and L1 subcongenic lines were protected against body fat gain under high-fat diet (HFD), whereas L2 and DA had significantly more body fat after high-fat feeding. Interestingly, adipocyte size distribution in SAT and epigonadal AT of L1 subcongenic rats did not undergo typical ballooning under HFD and the number of preadipocytes in AT was significantly elevated in L2 compared with L1 and parental rats. Transcriptome analysis identified three candidate genes in VAT on rat chromosome 10. In humans, these candidate genes were differentially expressed between SAT and VAT. Moreover, HID1 mRNA significantly correlates with parameters of obesity and glucose metabolism. Our data suggest novel candidate genes for obesity that map on rat chromosome 10 in an interval 102.2–104.7 Mb and are strongly associated with body fat mass regulation, preadipocyte number and adipocyte size in rats. Among those genes, AT head involution defective (HID1) mRNA expression may be relevant for human fat distribution and glucose homeostasis.

BACKGROUND:Metabolic syndrome is a complex disorder characterized by the presence of insulin resistance (IR), type 2 diabetes mellitus (T2DM), impaired glucose tolerance (IGT), or impaired fasting glucose (IFG), plus at least two of the following conditions--hypertension, hyperlipidemia, obesity, and microalbuminuria. Metabolic syndrome exposes patients to a greater risk of developing cardiovascular disease (CVD) and is often associated with elevated levels of homocysteine (Hcy). In the current work, we analyzed the expression of nicotinamide N-methyltransferase (NNMT). Because NNMT is involved in Hcy metabolism and participates in the regulation of the cellular and plasma levels of this compound, we explored the role played by the enzyme in metabolic syndrome.METHODS:Real-time PCR, immunohistochemistry, western blot analysis, and catalytic activity assay were performed to evaluate NNMT expression levels in adipose tissue from 10 Wistar Ottawa Karlsburg W (WOKW) rats, which are an animal model for metabolic syndrome, and from 10 Dark Agouti (DA) rats as the disease-resistant control strain.RESULTS:NNMT mRNA, protein, and activity levels were significantly higher in adipose tissue obtained from WOKW rats compared with those observed in adipose tissue of DA rats.CONCLUSION:Data reported in this study represent the first evidence supporting the hypothesis that NNMT could play an important role in the pathogenesis of metabolic syndrome and could have a potential for the development of a targeted therapy.

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(-/-)) 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 whole-body insulin sensitivity in LRep1(-/-) 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, Pparγ, 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.

BB, bio breeding; BB/OK, bio breeding/ ottawa karlsburg; BB.6S, BB.4S and BB.1K, congenic derivates of bb/ok

L-arginine is the source of all forms of nitric oxide (NO), which can be an extremely relevant factor in the treatment and reversal of important diseases. This observation prompted us to use L-arginine in BB/OK (Bio Breeding/Ottawa Karlsburg) rats developing insulin-dependent type 1 diabetes to evaluate the effect of L-arginine on the prevention of this disease. BB/OK rats were given L-arginine in drinking water (2%) during pregnancy and to the progeny (group 1), to newborn (group 2) and not given (group 3) up to an age of 30 weeks. Diabetes frequency and age at onset of diabetes were recorded in all BB/ OK rats. The mRNA expression of genes (Nfkb2, Il10, Il1b, Rarres 2, Pparg, Adipoq, Lep and Slc2a4) was measured in subcutaneous and visceral adipose tissue in BB/OK rats which did not develop diabetes up to an age of 30 weeks. Diabetes frequency was reduced in the L-arginine supplemented BB/OK compared to the untreated BB/OK rats (group 1: p<0.001 and group 2: p<0.05 vs. control group 3). Group 2 showed gender specific differences, because more females than males developed diabetes (94/59%; p<0.05). Gene expression in subcutaneous and visceral adipose tissue was reduced in the L-arginine drinking BB/OK rats compared to the control group. L-arginine in drinking water can protect from type 1 diabetes development in a sex specific manner. Because L-arginine is a precursor of NO, it may be concluded that this manipulation normalized NO activity in s cells, partially preventing type 1 diabetes development.

The aim of this study was to evaluate the effects of a high-fat diet (HFD) on oxidative indexes in WistarOttawaKarlsburg W (WOKW) rats used as a model of metabolic syndrome in comparison with Dark Agouti (DA) rats used as a control strain. This syndrome is defined by the occurrence of two or more risk factors including obesity, hypertension, dyslipidemia, and insulin resistance. Forty rats were used in the study and the effect of HFD was evaluated in terms of body weight and both hemoglobin and CoQ oxidative status. Moreover, 16 rats (8 of each strain) were supplemented with 3 mg/100 g b.w. of CoQ10 for 1 month in view of its beneficial properties in cardiovascular disease due to its antioxidant activity in the lipid environment. HFD promoted an increase in body weight, in particular in WOKW males, and in the methemoglobin (met-Hb) index in both strains. Moreover, HFD promoted endogenous CoQ10 oxidation. CoQ10 supplementation was able to efficiently counteract the HFD pro-oxidant effects, preventing met-Hb formation and CoQ oxidation.

Background Recent studies revealed that autophagy is up-regulated in obese individuals, as evidenced by increased expression of autophagy related genes. As argued elsewhere, it is possible that initially insulin resistance functions as an adaptive mechanism to increase autophagy in order to protect cells against death. We have shown that Wistar Ottawa Karlsburg W (RT1 u ) rats (WOKW) develop a metabolic syndrome with insulin resistance in adipose tissue, closely resembling the human disease. Therefore, the aim of this study was to characterize the autophagy phenotype in WOKW rats to clarify the interrelation between insulin resistance and autophagy in adipose tissue. Methods Subcutaneous and epidydimal adipose tissue samples of 5-months-old WOKW and healthy LEW.1 W male rats were investigated and protein levels (Western blot and immunhistochemistry) of key autophagy genes, including Atg5, Atg7, LC3-II/LC3-I and apoptosis marker cleaved caspase-3 were analyzed. Results WOKW rats displayed a significant increase of autophagy related proteins (Atg5, Atg7) in adipose tissue compared with LEW.1 W. This increase was predominantly found in epididymal adipose tissue. Furthermore, the LC3-II/LC3-I ratio as a marker of autophagosomes was significantly up-regulated in subcutaneous adipose tissue of WOKW rats. Cleaved caspase-3 was just slightly detectable in visceral adipose tissue and not detected in subcutaneous fat. Conclusion Insulin resistance in adipose tissue of obese WOKW rats is associated with up-regulation of differing autophagy markers in visceral and subcutaneous fat depots. This fact not only qualifies the WOKW rat for further detailed analysis of genetic determinants of metabolic syndrome but also highlights its suitability for autophagy research.

STUDY DESIGN:An experimental animal study.OBJECTIVE:To investigate histomorphometric and radiographical changes in the BB.4S rat model after PEEK (polyetheretherketone) nonfusion interspinous device implantation.SUMMARY OF BACKGROUND DATA:Clinical effectiveness of the PEEK nonfusion spine implant Wallis (Abbott, Bordeaux, France; now Zimmer, Warsaw, IN) is well documented. However, there is a lack of evidence on the long-term effects of this implant on bone, in particular its influence on structural changes of bone elements of the lumbar spine.METHODS:Twenty-four male BB.4S rats aged 11 weeks underwent surgery for implantation of a PEEK nonfusion interspinous device or for a sham procedure in 3 groups of 8 animals each: (1) implantation at level L4-L5; (2) implantation at level L5-L6; and (3) sham surgery. Eleven weeks postoperatively osteolyses at the implant-bone interface were measured via radiograph, bone mineral density of vertebral bodies was analyzed using osteodensitometry, and bone mineral content as well as resorption of the spinous processes were examined by histomorphometry. RESULTS.: Resorption of the spinous processes at the site of the interspinous implant was found in all treated segments. There was no significant difference in either bone density of vertebral bodies or histomorphometric structure of the spinous processes between adjacent vertebral bodies, between treated and untreated segments and between groups.CONCLUSION:These findings indicate that resorption of spinous processes because of a result of implant loosening, inhibit the targeted load redistribution through the PEEK nonfusion interspinous device in the lumbar spinal segment of the rat. This leads to reduced long-term stability of the implant in the animal model. These results suggest that PEEK nonfusion interspinous devices like the Wallis implants may have time-limited effects and should only be used for specified indications.

It has previously been shown that high-calorie diet alters the function of the mammalian circadian clock and that obesity has an influence on circadian organization of hormone secretion. That prompted us to test whether inbred Wistar Ottawa Karlsburg W (RT1(u)) (WOKW) rats developing facets of the metabolic syndrome show changes in their metabolic profiles under different feeding conditions (high-fat, high-sugar versus control diet) and under two different 12 h:12 h light-dark (LD) cycles. At the age of four weeks, these rats were divided into four groups. Groups 1 and 2 were kept under initial LD cycle (lights on at 05:00 h). Group 1 was fed with a normal rat diet while group 2 received a high-fat, high-sugar diet from 10 up to the age of 21 weeks. Groups 3 and 4 were kept under a shifted LD cycle (lights on at 11:00 h). Group 3 was given a normal diet while group 4 received a high-fat, high-sugar diet from an age like groups 1 and 2. Several metabolic traits were studied during the observation period of 21 weeks. The blood samples were obtained 2 h before lights off. Body weight gain (P < 0.001), leptin (P < 0.001), triglycerides (P < 0.001) and cholesterol (P < 0.05) were significantly reduced in group 4 versus group 2, but comparable between control groups (1 versus 3). The insulin concentrations were reduced in groups 3 and 4 versus groups 1 and 2 without effect of diet. In conclusion, the results provide evidence that light conditions influence diet induced changes in phenotypic traits like body weight gain, lipids as well as hormone levels (insulin and leptin) in WOKW rats.

Introduction Clinical effectiveness of the PEEK nonfusion interspinous device is well documented. However, there is a lack of evidence on the long-term behavior of this implant in regard to bone in general and especially its influence on structural changes of bone elements of the lumbar spine. Aim of our study was to investigate histomorphometric and radiographic changes in the BB.4Sd1 rat model after implantation of an interspinous PEEK implant. Materials and Methods Total 24 male BB.4Sd1 rats 11-week old underwent surgery for implantation of an interspinous PEEK device or for a sham procedure in three groups of eight animals each viz: (1) implantation at level L4-5, (2) implantation at level L5-6, and (3) sham surgery. Total 11 weeks postoperatively osteolyses at the implant-bone interface were measured via X-ray, bone mineral density of vertebral bodies was analyzed using osteodensitometry, and bone mineral content as well as osteolyses of the spinous processes examined by histomorphometry. Results Osteolyses of the spinous processes at the site of the interspinous implant were found in all treated segments. There was no significant difference in either bone density of vertebral bodies or histomorphometric structure of the spinous processes. Conclusion Osteolyses of spinous processes as a result of the implant becoming unsecure inhibit the targeted load redistribution through the interspinous PEEK device in the lumbar spinal segment of the rat. This leads to reduced long-term benefits of the implant in the animal model. These results suggest that the interspinous PEEK implants should only be used under strict indication and in consideration of their time-limited effects. I confirm having declared any potential conflict of interest for all authors listed on this abstract Yes Disclosure of Interest None declared Holinka J, Krepler P, Matzner M, et al. Stabilising effect of dynamic interspinous spacers in degenerative low-grade lumbar instability. International Orthopaedics 2011;35:395–400

Background: Because inbred rat strains are widely used as laboratory models, knowledge of phenotypic and genetic variations between strains will be useful to obtain insight into the relationship between different strains.Methods and Results: We studied phenotypic traits: of each strain - BN/K, DA/K and WOKW -10 male rats were studied for body weight and serum constituents at an age of 10 and 30 weeks. In addition, a total of 95 rats were studied for life expectancy. At an age of 30 weeks, these male rats were killed by an overdose of anesthetic (Sevofluran, Abbott), and the subcutaneous and visceral adipose tissue as well as bone tissue were removed to study the expression of 20 genes. There were significant differences in body weight, serum lipids and leptin at an age of 30 weeks between strains. Regarding life expectancy, BN rats lived longest (1072 +/- 228d). The highest gene expression was found in bone of BN rats. In adipose tissues, Nfkb1 is only expressed in subcutaneous adipocytes, and 5 genes, Col2a1, Mmp9, Tnfa, Ins1 and Cyp24a1, are not expressed in adipocytes. The ranking BN = DA>WOKW was observed in only one gene in subcutaneous (Fto) and visceral adipocytes (Col6a1). There were no significant differences in gene expression of one gene in subcutaneous adipocytes and of 3 genes in visceral adipocytes. Comparing the gene expression in visceral and subcutaneous adipocytes, only one gene showed a comparable behavior (Bmp1).Conclusion: From these results, it can be concluded that obvious phenotypic differences are caused by genetic differences between three rat strains, BN, DA and WOKW, as supported by gene expression studies in bone and adipose tissues. Especially BN rats can be used to study the genetic basis of long life.

A high-fat diet (HFD) has been recognized as a risk factor for diseases such as dyslipidemia, atherosclerosis, obesity, and osteoporosis. However, studies analyzing gene expression after HFD in bone are rare. That prompted us to analyze the expression of selected genes in bone of 4-week-old diabetes-prone B(io)B(reeding) rats. Two breeding pairs were fed a HFD (+10 % tallow) or were fed a normal diet (ND; Ssniff R-Z) before mating and afterward during pregnancy. After the birth of progeny, parents continued to be given HFD or ND until the progeny was weaned (3 weeks). Thereafter, offspring were weaned and were fed the same food as their parents up to an age of 4 weeks. Body weight was measured at an age of 4 weeks, and subsequently 13 HFD rats and 13 ND rats were killed and the tibial bone was harvested to analyze the expression of 53 genes in bone. All rats fed HFD were significantly heavier than rats fed ND after 3 and 4 weeks. The diet also influenced the expression of genes in bone. There were significant differences in 20 out of 53 genes studied between rats fed HFD compared with rats fed ND. Four out of 20 had a lower and 17 out of 20 genes a higher expression in HFD rats, but differences in gene expression showed obvious differences between males and females. There were only two genes that were similarly different between males and females: Bmp4 and Atf4. Two genes, Foxg1 and Npy , were inversely expressed in males and females. It seems that the gene expression is differently regulated by diet during pregnancy and later in life between males and females. Nevertheless, it cannot be excluded that HFD also acts as an epigenetic factor in the development of offspring in utero.

Objectives/Hypothesis: It is generally assumed that glycemic control in diabetic patients is important in optimizing wound healing. The goal of this study was to examine tympanic membrane (TM) wound healing in spontaneously diabetic rats depending on the diabetic metabolic state compared to nondiabetic control animals.Study Design: Prospective controlled study in experimental animals.Methods: Right-sided myringotomy was performed in 20 normoglycemic rats, 17 well-compensated, and 23 poorly compensated diabetic rats. TMs were observed for a total of 3 weeks. Effect of diabetic metabolic state on the healing of the TMs was evaluated by closure rates and histology.Results: Diabetic rats showed a significant delay in TM wound healing compared to the control group, but there were no significant differences between both diabetes groups.Conclusions: Glycemic control does not influence TM wound repair in an animal model of type 1 diabetes.

B(io) B(reedding)/O(ttawa) K(alsburg) rats spontaneously develop insulin-dependent type 1 diabetes. Days before BB/OK rats become diabetic, their body seems to be flabby which may be attributed to loss of subcutaneous fat. However, the rats are normoglycemic and manifest 3-4 days later. This observation prompted us to search for possibilities to avoid the loss of adipose tissue. BB/OK rats were subcutaneously grafted with visceral adipose tissue.In total, 34 (71%) out of 48 male and 23 (49%) out of 47 female BB/OK rats grafted with adipose tissue developed type 1 diabetes so that significantly more females than males were protected from diabetes development (p=0.03). In the control group, 17 (85%) out of 20 male and 20 (95%) out of 21 female BB/OK rats were diabetic.Adipose tissue transplantation can protect BB/OK rats from type 1 diabetes development in a sex specific manner. One could conclude that the manipulations have influenced fat accumulation and/or fat metabolism which prevent type 1 diabetes development in about 50% of BB/OK rats. This idea is supported by the finding that a mutation in the leptin receptor of NOD mice suppresses type 1 diabetes progression. (C) 2011 Elsevier B.V. All rights reserved.

Congenic BB rat strains carrying a SHR segment (D4Got41-Tacr1; 60.5-122.8 Mb; BB.4S) or a WOKW segment (D4Got41-Fabp1; 60.5-104.6 Mb; BB.4W) of chromosome 4 within the BB/OK background develop facets of the metabolic syndrome when compared with their parental BB/OK rats. To narrow down potential genes involved in the pathophysiology of metabolic syndrome, gene expression studies in adipose tissues of BB/OK, BB.4S, and BB.4W rats were initiated. Total RNA of subcutaneous and epididymal adipose tissue of BB/OK (n=10), congenic BB.4S (n=8), and BB.4W (n=9) males at an age of 4 weeks was isolated. The mRNA expression of 92 genes involved in obesity, insulin resistance and other metabolic traits was measured by RT-PCR. Significant differences in gene expression were only found in Repin1 in both adipose tissues. Congenic BB.4W showed significantly lower gene expression than did BB.4S and BB/OK. Our findings and newly published findings of Repin1 in 3T3-L1 adipocytes support the hypothesis that Repin1 may affect the development of facets of the metabolic syndrome.

Background It is well known that lipid metabolism plays an important role in the early stages of type 1 diabetes (T1D). For that reason, we examined factors that influence lipid metabolism of BioBreeding/Ottawa Kalsburg (BB/OK) rats that spontaneously develop an insulin-dependent T1D.Methods BB/OK female rats were fed a high-fat diet during pregnancy (Ssniff R-Z + 10% tallow) and their progeny were also given this diet up to an age of 30 weeks (n = 55) or 4 weeks (n = 14) to study gene expression of Pparg, Fasn, Lep, Adipoq, Repin1, Rarres 2, and Glut4 in adipose tissue. Forty-two BB/OK rats fed the normal diet (Ssniff R-Z) during pregnancy and the observation period served as controls.Results The high-fat diet significantly decreased diabetes frequency in BB/OK rats when compared with control rats (71 versus 95%, p = 0.002). Although this difference was also reflected in the male rats (68 versus 100%, p = 0.003), no significant variation was observed in female rats (73 versus 90%, p = 0.23). The high-fat diet resulted in significantly reduced mRNA expression of examined genes in subcutaneous adipose tissue, but not in visceral adipose tissue, except for Fasn and Repin1 expression.Conclusions A high-fat diet seems to protect BB/OK rats from T1D in a sex-specific manner. The data suggest that a high-fat diet might influence fat accumulation and/or fat metabolism and prevent T1D development in male rats, which is supported by changes in adipose tissue gene expression. Copyright. (C) 2011 John Wiley & Sons, Ltd.

Our previous study found that when injected with Nociceptin/Orphanin FQ (N/OFQ) into the brain, male Dark Agouti (DA) rats, which are resistant to metabolic syndrome, have greater hyperphagia than male Wistar Ottawa Karlsburg W (WOKW) animals, which are prone to this disease. We attributed this difference to the fact that these two strains have different cocaine-amphetamine regulated transcript peptide (Cart) gene sequences and expression. In order to address this hypothesis, the present work focused on sex differences and analyzed not only male but also female N/OFQ-induced (0.25 and 0.5 nmol/rat) food intake in terms of their Cart and N/OFQ receptor gene expression in the hypothalamic area. In N/OFQ-naive WOKW females, cart gene expression is extremely elevated compared to N/OFQ-naive WOKW males. When male and female WOKW littermates are stimulated with N/OFQ, the food intake of females is significantly lower than that of the males. Granted, the N/OFQ feeding behavior experiments were not performed on the animals measured for Cart gene expression, but nonetheless, the responses observed in littermates point to an interesting avenue for further inquiry.

Background and aims: In the last years the intensive research showed that nitric oxide (NO) is not only a very much meaning bioactive signaling messenger in the heart circulation system, but it plays also a role in the case of the fight against infections and in the case of the controlling of the nervous system. As known L-arginine is the substrate for the synthesis of NO. Some years ago it has been reported that supplementation of L-arginine reduces atherosclerosis in rabbits and reduces the arterial pressure in hypertensive rats. Other authors suggested that exogenously administered L-arginine might improve the clinical manifestation of diabetes mellitus and decrease the oxidative stress in the gastrointestinal tract. In addition, the improvement of metabolic and cardiovascular health are mediated by enhancing the flux of muscle- and kidney-derived amino acids to pancreatic and vascular endothelial cells leading to the intracellular production of NO. That resulted in normalization of insulin secretion, vascular tone and insulin sensitivity. This observation prompted us to search for possibilities to prevent diabetes development. BB rats which are an animal model of type 1 diabetes sharing many common features with human insulin-dependent type 1 diabetes were treated with L-arginine.