Obob Mice Research Articles

Conjugated linoleic acid ameliorates hepatic steatosis by modulating intestinal permeability and gut microbiota in ob/ob mice

BackgroundConjugated linoleic acid (CLA) is an effective supplement for reducing fat mass, but its effect on hepatic steatosis remains controversial.ObjectiveThis study aims to evaluate the effect of CLA on liver fat accumulation, inflammation, gut microbiome, and intestinal barrier integrity.DesignWild-type (WT) mice and ob/ob (OB) mice were randomly divided into four groups according to the treatment with/without 1% CLA: WT, WT mice treated with CLA (WT-CLA), OB, and OB mice treated with CLA (OB-CLA). Lipid metabolism and hepatic fat accumulation were evaluated by changes in histological and biochemical parameters. Gene expressions related to liver inflammation and intestinal barrier integrity were examined. The effect of CLA on the gut microbiota population was investigated.ResultsThe body weight, fatty tissue mass, and serum lipid levels of the WT-CLA group and OB-CLA group were separately lower than those of the WT group and OB group, but the livers of the WT-CLA group had more fatty lipids, higher triglyceride properties, and saturated fatty acid (FA) composition than those of the WT group, which was contrary to the effect of CLA on OB mice. Real time quantitative PCR results showed that CLA increased hepatic inflammation and intestinal permeability in the WT mice, while it significantly decreased the mRNA expression of liver TNF-α, IFN-γ, and IL-1β and markedly ameliorated intestinal tight junction proteins in the OB mice. The gut microbiota testing indicated a higher abundance of beneficial bacteria (e.g., Lachnoclostridium, Roseburia, Dubosiella, Oscillibacter, and Anaerostipes) and a lower abundance of pro-inflammatory bacteria (e.g., Tyzzerella and Alistipes) in the OB-CLA group than those of the OB group. Correlation analysis suggested that gut microbiota correlated with liver inflammation, intestinal permeability, and hepatic FA composition.ConclusionCLA potentially contributed to ameliorating hepatic steatosis in OB mice via modulating liver inflammation, intestinal permeability, and gut microbiota, which suggests CLA is more suitable for people with obesity or overweight.

Abstract MP227: Single Cell RNA Sequencing Analysis Reveals Insulin Signaling Defects In Cardiomyocytes Derived From A Novel Mouse Model Of Heart Failure With Preserved Ejection Fraction

Heart failure with preserved ejection fraction (HFpEF) is an emerging form of heart failure worldwide with no effective therapies in contrast with heart failure with reserved ejection fraction (HFrEF). To simulate multiple risk-factors associated with HFpEF in clinic, we developed a HFpEF mouse model by introducing cardiac hypertrophy with transverse aortic constriction (TAC) in ObOb ( Lep ob/ob ) mice, which has intrinsic systemic metabolic dysfunctions including obesity and insulin resistance. We first validated pathological changes in diastolic but not systolic parameters in the Ob-TAC vs. Ob-sham mice up to 10 weeks post-TAC by echocardiography. To evaluate the global transcriptome change in difference cell types, we conducted single nuclei RNA sequencing (snRNA-seq) from whole hearts of lean mice (c57), ObOb, and Ob-TAC mice (male only). 10x genomic 3’ GEM kit was used to generate the cDNA library and sequencing was done by Novaseq SP platform. A total of 13k nuclei were recovered from QC, nFeature RNA (&lt 2500) and mitochondrial gene (&lt 5%) filtering. By UMAP dimension reduction analysis, we annotated major cardiac cell types in the integrated snRNA-seq dataset, including 3 clusters of Cardiomyocytes (CMs). By pathway analysis of the differentially expressed genes in each CM clusters, we found that insulin resistance and glucagon pathway were enriched among the up regulated genes in CMs in HFpEF vs. lean control, while cell migration, signal transduction including insulin substrates were down regulated. Thus, we hypothesized that the altered crosstalk between glucagon and insulin signaling might contribute to the development of HFpEF in this mouse modal. This hypothesis was validated in a proof-of-concept study showing significant improvement of HFpEF features by inhibiting the glucagon receptors post-TAC with injection of a glucagon receptor antagonist.

Adipokinome Signatures in Obese Mouse Models Reflect Adipose Tissue Health and Are Associated with Serum Lipid Composition.

Adipocyte and hepatic lipid metabolism govern whole-body metabolic homeostasis, whereas a disbalance of de novo lipogenesis (DNL) in fat and liver might lead to obesity, with severe co-morbidities. Nevertheless, some obese people are metabolically healthy, but the “protective” mechanisms are not yet known in detail. Especially, the adipocyte-derived molecular mediators that indicate adipose functionality are poorly understood. We studied transgenic mice (alb-SREBP-1c) with a “healthy” obese phenotype, and obob mice with hyperphagia-induced “sick” obesity to analyze the impact of the tissue-specific DNL on the secreted proteins, i.e., the adipokinome, of the primary adipose cells by label-free proteomics. Compared to the control mice, adipose DNL is reduced in both obese mouse models. In contrast, the hepatic DNL is reduced in obob but elevated in alb-SREBP-1c mice. To investigate the relationship between lipid metabolism and adipokinomes, we formulated the “liver-to-adipose-tissue DNL” ratio. Knowledge-based analyses of these results revealed adipocyte functionality with proteins, which was involved in tissue remodeling or metabolism in the alb-SREBP-1c mice and in the control mice, but mainly in fibrosis in the obob mice. The adipokinome in “healthy” obesity is similar to that in a normal condition, but it differs from that in “sick” obesity, whereas the serum lipid patterns reflect the “liver-to-adipose-tissue DNL” ratio and are associated with the adipokinome signature.

Novel Insights into the Adipokinome of Obese and Obese/Diabetic Mouse Models.

The group of adipokines comprises hundreds of biological active proteins and peptides released from adipose tissue. Alterations of those complex protein signatures are suggested to play a crucial role in the pathophysiology of multifactorial, metabolic diseases. We hypothesized that also the pathophysiology of type-2-diabetes is linked to the dysregulation of the adipocyte secretome. To test this, we investigated mouse models with monogenic defects in leptin signaling which are susceptible to adipositas (C57BL/6 Cg-Lepob (obob)) or adipositas with diabetes (C57BL/KS Cg-Leprdb (dbdb)) according to their genetic background. At the age of 17 weeks, visceral fat was obtained and primary murine adipocytes were isolated to harvest secretomes. Quantitative proteome analyses (LC-ESI-MS/MS) identified more than 800 potential secreted proteins. The secretome patterns revealed significant differences connected to the pathophysiology of obese mice. Pathway analyses indicated that these differences focus on exosome modelling, but failed to provide more precise specifications. To investigate the relationship of secretome data to insulin sensitivity, we examined the content of diabetogenic lipids, i.e., diacylglycerols (DAGs), identified as key players in lipid-induced insulin resistance. In contrast to obob mice, fat tissue of dbdb mice showed elevated DAG content, especially of DAG species with saturated fatty acid C16:0 and C18:0, while unsaturated fatty acid C16:1 were only changed in obob. Furthermore, DAG signatures of the models specifically correlate to secreted regulated adipokines indicating specific pathways. In conclusion, our data further support the concept that the fat tissue is an endocrine organ that releases bioactive factors corresponding to adipose tissue health status.

Depot-specific differences in fat mass expansion in WT and ob/ob mice.

The study was designed to investigate the cellular mechanisms underlying the differential fat expansion in different fat depots in wild type (WT) and ob/ob (OB) mice. At 6 weeks old, no differences in fat mass were found between SAT and VAT in WT mice, while O-SAT showed significantly higher weight than that of O-VAT. The average adipocyte size of SAT (∼ 4133.47 μm2) was smaller than that of VAT (∼ 7438.91 μm2) in OB mice. O-SAT preadipocytes gained higher triglyceride contents and higher levels of PPARγ and C/EBPα than did O-VAT preadipocytes upon in vitro differentiation. W-SAT and W-VAT displayed no significant differences in fatty acid uptake, while 1.36 fold significantly higher fatty acid uptake was found in O-SAT compared to O-VAT. Approximately 52% of the radioactivity recovered in cellular lipids was found in TAG in O-SAT, which was significantly higher than the other three adipocyte types. Significantly more radiolabelled oleic acid was β-oxidized to CO2 in adipocytes from O-VAT than that from O-SAT. ATP production was significantly lower in W-SAT compared with W-VAT, whereas no significantly ATP level was observed between O-SAT and O-VAT. Expression of UCP-1 in SAT from either WT or OB mice was significantly higher than the counterpart of VAT, which demonstrated higher uncoupled respiration and lower oxidative phosphorylation in SAT. Together, a combined increase in adipogenesis and FA uptake, and decreases in β-oxidation and ATP production, contributed to greater expansion of SAT compared to VAT in obese mice.

Abstract 607: Endothelin-1 Increases ß-Actin Expression During Osteochondrogenesis of Aortic Smooth Muscle Cells from Leptin-Deficient ob/ob Mice

β-actin, a highly conserved gene/protein, which is usually not modulated by different stimuli, is routinely used to normalize target gene or protein expression in experimental biology, and so defined endogenous housekeeping gene. However, investigators showed that β-actin expression could change during cellular growth and differentiation. We hypothesized that endothelin 1 (ET-1), a cellular proliferative agonist, increases β-actin expression and stimulates calcification in vascular smooth muscle cells (SMC) from obob mice. Primary aortic SMC passage 4-6 isolated from obese leptin-deficient (obob) and from C57BL/6 (C57) mice were incubated without (Cont) or with ET-1 50nM (ET) for 0-72h. Cell lysates were assessed for β-actin and RUNX2 expression by western blotting and normalized by GAPDH. ObobET SMC increased β-actin expression after 24h, 48h and 72h (1.68±0.06, 1.66±0.05, 1.73±0.06 respectively) vs obobCont (1.01±0.07), p<.05 n=6. However, C57ET did not change β-actin expression (0.91±0.06, 1.04±0.03, 0.99±0.08) after 24h, 48h and 72h respectively vs. C57Cont (0.95 ± 0.04), p=NS n=6. Concomitantly, obobET SMC increased osteochondrogenic differentiation, by modulating RUNX2 (1.23±0.03 vs obobCont 1±0.01, p<.05 n=3) after 48h, which did not occur in C57 (C57ET 1.07±0.07 vs C57Cont 1.02±0.11), p=NS. Furthermore, obobET increased calcification vs obobCont (1.69±0.025 vs 1.03±0.03) after 14 days p<.05, n=3 and C57ET did not calcify (0.97±0.02 vs C57Cont 1±0.05), p=NS. We showed that ET-1 modulates β-actin expression and osteochodrogenenic differentiation only in obob SMC, thus increasing calcification. In conclusion, β-actin is not a consistent endogenous control in ET-1-stimulated SMC from obob mice during osteochondrogenic differentiation.

Mo1767 Genetic Deletion of IL-25 Increases Susceptibility to High-Fat Diet-Induced Metabolic Dysfunction in Mice

unknown. This study therefore evaluated the effect of the DPP-4 inhibitor, sitagliptin (STG), on diabetes-related colon carcinogenesis in mice. Materials and Methods: Six-week-old male wild-type (WT) and leptin-deficient (ob/ob) C57BL/6J mice were used in this study. The mice received 1,2-dimethlhydrazine (DMH) subcutaneously at a dose of 20 mg/kg body weight three times within a week, and chronic colitis was then induced by administration of two cycles of DSS (each cycle: 3% DSS for 7 days followed by distilled water for 14 days). Sitagliptin (3 mg/kg) was administered to the mice by oral gavage every day during the entire experimental period. The mice were divided into the following three groups: WT group; obob group; and obob+STG group. The mice were sacrificed 28 days after the completion of both cycles of DSS. Results: During the DSS administration cycle, mean body weight decreased significantly in the WT group compared with the obob and obob+STG groups. The disease activity index (DAI) was also increased significantly in the WT group. However, administration of STG did not affect the DAI or body weight. The mean number of tumors was 7.8±4.5, 10.6±3.3, and 6.7±2.4 in the WT, obob, and obob+STG groups, respectively. STG significantly suppressed the occurrence of neoplasia in the obob mice. Mucosal DPP-4 activity and the mucosal concentration of GLP-2 were not significantly different in the three groups. Conclusion: Administration of STG exerts a suppressive effect on the development of colon neoplasia in a murine model of type 2 diabetes. However, STG does not affect the mucosal DPP-4 activity. Because DPP-4 mRNA/DPP-4 activity are highly expressed/detected in the small intestine andDPP-8 and 9 are expressed predominantly in the colon, DPP-4 inhibitors may suppress colon carcinogenesis independent of the GLP pathway.

β‐Actin is regulated by endothelin 1 and is not a consistent endogenous control in aortic smooth muscle cells from leptin‐deficient mice (955.1)

β‐actin, a highly conserved gene/protein, which is usually not modulated by different stimuli, is routinely used to normalize target gene or protein expression in experimental biology, and so defined endogenous housekeeping gene. However, investigators showed that β‐actin expression could change during cellular growth and differentiation. We hypothesized that endothelin 1 (ET‐1), a cellular proliferative agonist, increase β‐actin expression in vascular smooth muscle cells (SMC). Primary aortic SMC passage 4‐6 isolated from obese leptin‐deficient (obob) and from C57BL/6 mice were incubated without (Cont) or with ET‐1 50nM (ET) for 0‐72h. Cell lysates were assessed for β‐actin expression by Western blotting. ObobET SMC increased β‐actin expression after 24h, 48h and 72h (1.68 ±0.06, 1.66 ± 0.05, 1.73 ± 0.06 respectively) normalized by GAPDH vs obobCont (1.01± 0.07), p<.05 n=6. However, C57ET did not change β‐actin expression (0.91 ±0.06, 1.04 ± 0.03, 0.99 ± 0.08) after 24h, 48h and 72h respectively vs. C57Cont (0.95 ± 0.04), p=NS n=6. We showed that ET‐1 modulates β‐actin expression only in obob SMC, but not in C57BL/6 SMC. In conclusion, β‐actin is not a consistent endogenous control in ET‐1‐stimulated SMC from obob mice. Grant Funding Source: Supported by FAPESP 2010/07455‐5

Contrasting effects of leptin on food anticipatory and total locomotor activity.

Obese, leptin deficient obob mice have profoundly decreased activity and increased food seeking behavior. The decreased activity has been attributed to obesity. In mice, we tested the hypothesis that leptin increases total locomotor activity but inhibits food anticipatory activity. We also sought to determine if leptin induced increases in total locomotor activity are independent of changes in body weight and obesity. We studied obob mice and also created a novel transgenic mouse where leptin is over-expressed in a tetracycline-off system and can be abruptly and non-invasively suppressed by doxycycline within few hours. The studies were performed using two independent behavioral assays: home cage activity (HCA) and running wheel activity (RWA). Systemic administration of leptin (150 ng/hr) to obob mice produced a 122%±30% (mean ± SEM) increase (p≤0.01) in locomotor activity within 2 days In addition, cerebroventricular administration of leptin (5 ng/hr) also produced an early and progressive increase in total locomotor activity beginning on the 1st day (+28±8%; p≤0.05) and increasing to +69±23% on day 3 without a decrease in body weight during this time. The increase in activity was restricted to the dark phase. Conversely, in a tet-off transgenic obob mouse line, acute leptin suppression reduced spontaneous locomotor activity. To further define activities that are leptin regulated, we assayed food anticipatory activity (FAA) and found that it was markedly augmented in obob mice compared to wild type mice (+38±6.7 in obob vs +20±6.3% in wild type at peak; mean ± SEM; p≤0.001) and abolished by leptin. Although melanocortin-3 receptors (MC3R) reportedly mediate FAA, we found augmented FAA and preserved inhibitory effects of leptin on FAA in MC3R−/−obob mice. In summary, this study demonstrates that total activity and FAA are regulated independently by leptin. Leptin, acting in the central nervous system and at physiologic levels, produces early increases in locomotor activity before substantial weight loss. In contrast, leptin suppresses augmented food anticipatory activity in obob mice.

Re-dicing the pancreatic β-cell: do microRNAs define cellular identity?

Re-dicing the pancreatic β-cell: do microRNAs define cellular identity?

Rapamycin does not improve insulin sensitivity despite elevated mammalian target of rapamycin complex 1 activity in muscles ofob/obmice

Studies of cultured cells have indicated that the mammalian target of rapamycin complex 1 (mTORC1) mediates the development of insulin resistance. Because a role for mTORC1 in the development of skeletal muscle insulin resistance has not been established, we studied mTORC1 activity in skeletal muscles of ob/ob (OB) mice and wild-type (WT) mice. In vivo insulin action was assessed in muscles of mice 15 min following an intraperitoneal injection of insulin or an equivalent volume of saline. In the basal state, the phosphorylation of S6K on Thr(389), mTOR on Ser(2448), and PRAS40 on Thr(246) were increased significantly in muscles from OB mice compared with WT mice. The increase in basal mTORC1 signaling was associated with an increase in basal PKB phosphorylation on Thr(308) and Ser(473). In the insulin-stimulated state, no differences existed in the phosphorylation of S6K on Thr(389), but PKB phosphorylation on Thr(308) and Ser(473) was significantly reduced in muscles of OB compared with WT mice. Despite elevated mTORC1 activity in OB mice, rapamycin treatment did not improve either glucose tolerance or insulin tolerance. These results indicate that the insulin resistance of OB mice is mediated, in part, by factors other than mTORC1.

Constitutive activation of the mammalian target of rapamycin complex 1 (mTORC1) is associated with skeletal muscle insulin resistance

The present study assessed mTORC1 activity in isolated rat skeletal muscles and in skeletal muscles of Ob/Ob (OB) mice. Epitrochlearis muscles isolated from rats were incubated with insulin (750 uU/ml) and/or amino acids (2.5X MEM). Amino acids produced a significant reduction in the ability of insulin to promote protein kinase B (PKB) phosphorylation, a process that was reversed by rapamycin (100nM). Amino acids did not inhibit the ability of insulin to promote the phosphorylation of PRAS40 on Thr‐246, a site whose phosphorylation enhances mTORC1 activity. In vivo insulin action was assessed in muscles of OB and wild type (WT) mice by intraperitoneal injections of insulin (0.25 U/Kg) or saline. In the basal state, phosphorylation of S6K on Thr‐389 and PRAS40 on Thr‐246 were increased significantly in muscles of OB compared to WT mice. In the insulin‐stimulated state, phosphorylation of S6K on Thr‐389 and PRAS40 on Thr‐246 were similar in muscles of OB compared to WT mice. We also assessed whether or not 5 days of rapamycin treatment (0.75 mg/Kg/day) could improve the insulin resistance of OB mice. Rapamycin treatment exacerbated the insulin resistance of Ob mice, indicating that some mTORC1 activity is necessary for normal in vivo glucose disposal. These results indicate that mTORC1 signaling is associated with insulin resistance, but inhibiting mTORC1 activity with rapamycin does not improve in vivo insulin action.

Impaired Expression of Cardiac Adiponectin in Leptin-Deficient Mice With Viral Myocarditis

A mouse model of encephalomyocarditis (EMC) virus-induced myocarditis was used to investigate the expression of adiponectin in damaged cardiomyocytes. We intraperitoneally injected EMC virus into leptin-deficient ob/ob (OB) mice and wild-type (WT) mice. OB mice were divided into two subgroups consisting of mice with no intervention and mice receiving leptin replacement starting simultaneously with viral inoculation. We determined differences in heart weight, cardiac histological score, numbers of infiltrating and apoptotic cells in the myocardium, expression levels of adiponectin and TNF-alpha mRNA in the heart, adiponectin immunoreactivity in myocytes, adiponectin and TNF-alpha concentrations in the heart, and immunoreactivity of adiponectin receptors in myocytes between OB mice and WT mice. There was significantly decreased adiponectin mRNA expression, immunoreactivity, and protein level in the heart, and reduced immunoreactivity of adiponectin receptor 1 in myocytes from OB mice on days 4 and 8 after viral inoculation as compared with those in WT mice, together with increased cardiac weight, severe inflammatory myocardial damage, and increased levels of cardiac TNF-alpha mRNA and protein. Replacement of leptin in OB mice inhibited the development of severe myocarditis through augmentation of adiponectin mRNA, immunoreactivity, and protein level, increased adiponectin receptor 1 immunoreactivity in myocytes, and suppressed levels of TNF-alpha mRNA and protein. These results suggest that impaired expression of cardiac adiponectin may contribute to the progression of viral myocarditis through enhanced expression of TNF-alpha under a leptin-deficient condition.

Overexpression of APOC1 in obob mice leads to hepatic steatosis and severe hepatic insulin resistance

Obese obob mice with strong overexpression of the human apolipoprotein C1 (APOC1) exhibit excessive free fatty acid (FFA) and triglyceride (TG) levels and severely reduced body weight (due to the absence of subcutaneous adipose tissue) and skin abnormalities. To evaluate the effects of APOC1 overexpression on hepatic and peripheral insulin sensitivity in a less-extreme model, we generated obob mice with mild overexpression of APOC1 (obob/APOC1(+/-)) and performed hyperinsulinemic clamp analysis. Compared with obob littermates, obob/APOC1(+/-) mice showed reduced body weight (-25%) and increased plasma levels of TG (+632%), total cholesterol (+134%), FFA (+65%), glucose (+73%), and insulin (+49%). Hyperinsulinemic clamp analysis revealed severe whole-body and hepatic insulin resistance in obob/APOC1(+/-) mice and, in addition, increased hepatic uptake of FFA and hepatic TG content. Treatment of obob/APOC1(+/-) mice with rosiglitazone strongly improved whole-body insulin sensitivity as well as hepatic insulin sensitivity, despite a further increase of hepatic fatty acid (FA) uptake and a panlobular increase of hepatic TG accumulation. We conclude that overexpression of APOC1 prevents rosiglitazone-induced peripheral FA uptake leading to severe hepatic steatosis. Interestingly, despite rosiglitazone-induced hepatic steatosis, hepatic insulin sensitivity improves dramatically. We hypothesize that the different hepatic fat accumulation and/or decrease in FA intermediates has a major effect on the insulin sensitivity of the liver.

Functional proteomics of nonalcoholic steatohepatitis: mitochondrial proteins as targets of S-adenosylmethionine.

Recent work shows that S-adenosylmethionine (AdoMet) helps maintain normal liver function as chronic hepatic deficiency results in spontaneous development of steatohepatitis and hepatocellular carcinoma. The mechanisms by which these nontraditional functions of AdoMet occur are unknown. Here, we use knockout mice deficient in hepatic AdoMet synthesis (MAT1A(-/-)) to study the proteome of the liver during the development of steatohepatitis. One hundred and seventeen protein spots, differentially expressed during the development of steatohepatitis, were selected and identified by peptide mass fingerprinting. Among them, 12 proteins were found to be affected from birth, when MAT1A(-/-) expression is switched on in WT mouse liver, to the rise of histological lesions, which occurs at approximately 8 months. Of the 12 proteins, 4 [prohibitin 1 (PHB1), cytochrome c oxidase I and II, and ATPase beta-subunit] have known roles in mitochondrial function. We show that the alteration in expression of PHB1 correlates with a loss of mitochondrial function. Experiments in isolated rat hepatocytes indicate that AdoMet regulates PHB1 content, thus suggesting ways by which steatohepatitis may be induced. Importantly, we found the expression of these mitochondrial proteins was abnormal in obob mice and obese patients who are at risk for nonalcoholic steatohepatitis.

Effect of δ 22-5β-taurocholenic acid on hepatic cholesterol and fatty acid in gold thioglucose obese mice fed low- or high-fat diets

We previously demonstrated that hyperglycemic-obese (obob) mice fed a 1% corn oil diet accumulated 10 times as much hepatic cholesterol as did their non-obese (+/?) littermates fed this diet because of difficulty in removal of cholesterol from the liver rather than from increased synthesis. Furthermore, feeding the bile acid analog Δ 22-5β-taurocholenic acid completely prevented the accumulation of hepatic cholesterol in obob mice fed the 1% corn oil diet. The hypothesis to be tested in the current study is that these aspects of cholesterol metabolism in the obob mouse do not occur in the hyperinsulinemic and insulin-resistant gold thioglucose obese mouse. Gold thioglucose obese (gtgo) and non-obese (ngtgo) mice were fed diets containing either 1% corn oil or 40% lard each with or without added taurocholenic acid for 6 weeks and then given a 250 mg meal of [U- 14C]-glucose with incorporation of label into hepatic cholesterol and fatty acid measured 2 hours later. Consistent with earlier results in the obob model, incorporation of labeled glucose was significantly increased in obese compared with non-obese mice fed 1% corn oil and significantly reduced either by feeding 40% lard or by adding taurocholenic acid to the diet. In addition, taurocholenic acid greatly increased incorporation of labeled glucose into hepatic cholesterol in obese or non-obese mice fed either diet. In contrast to obob mice, the percentage of fat in the liver of gtgo mice was increased only 50% compared with ngtgo mice. The comparable increase in obob mice was 480%. Hepatic cholesterol did not increase significantly in the liver of gtgo mice fed 1% corn oil when compared with the ngtgo controls. The comparable increase in obob mice fed 1% corn oil was 350%. Also in marked contrast to obob mice, feeding taurocholenic acid increased hepatic cholesterol compared with non-obese controls fed either diet. The results are discussed in the light of the presence of circulating leptin in gtgo but not in obob mice.

Leptin

A highly conserved protein called ‘leptin’ was recently discovered to play a role in regulation of the balance in humans and rodents. This 167-amino-acid-containing protein is only produced and secreted by mature adipocytes Absence of the protein in mutant obob mice and resistance to its effects in dbdb mice lead to extreme obesity, and gape 11 diabetes mellitus. No mutation of the ob-gene encoding for leptin has been found in obese humans so far. ob mRNA in adipocytes and serum leptin levels are positively related to adipose tissue mass. Receptors for leptin have been found in the choroid plexus and hypothalamus. A feedback inhibition of leptin on hypothalamic neuropeptide Y (NY) production is 1 ostulated as hypothalamic NY concentrations are increased in obob mice and NY induces food intake, insuhn secretion and autonomic nervous system activity. Insulin increases triglyceride stores in fat cells and could thereby stimulate leptin secretion. The ultimate intracellular pathway within the adipocyte that stimulates or shuts off ob mRNA expression and consequent leptin production and secretion remains to be elucidated. Whether leptin will ever come to play a role in the treat ment of human obesity remains an unanswered question at the present time.

Effect of Δ 22-5β-taurocholenic acid and dietary fat on hepatic cholesterol and fatty acid in hyperglycemic-obese mice

Previous work demonstrated that hyperglycemic obese mice ( obob) fed a 1% corn oil diet accumulated cholesterol and fat in the liver when compared with nonobese littermates. The present experiment was carried out to test the hypothesis that feeding the bile acid analog Δ 22-5β-taurocholenic acid would correct this metabolic defect. Liver total fatty acid was greatly elevated in obob mice fed 1% corn oil compared with nonobese controls (21.04 ± 1.23% versus 4.41 ± 0.27%). Incorporation of [U- 14C]glucose into liver fatty acid was inhibited by taurocholenic acid in both obese and nonobese mice fed 1% corn oil. Taurocholenic acid reduced liver fatty acid in obob mice fed 1% corn oil to the level seen in nonobese mice (3.07 ± 0.22% versus 4.41 ± 0.27%). Cholesterol greatly accumulated in the livers of obob mice fed 1% corn oil compared with nonobese mice (0.97 ± 0.08% versus 0.28 ± 0.02%) and was reduced to normal by feeding taurocholenic acid (0.27 ± 0.05% versus 0.28 ± 0.01%) despite greatly elevating rates of incorporation of [U- 14C]glucose into liver cholesterol. Taurocholenic acid significantly reduced liver fat in obese mice fed 40% lard (11.62 ± 1.41% versus 5.87 ± 1.03%) but had no significant effect on the percentage of liver cholesterol (0.68 ± 0.05% versus 0.56 ± 0.04%). It is suggested that the elevation of liver cholesterol in obob mice fed 1% corn oil comes about through a defect in cholesterol removal from the liver, a defect corrected by taurocholenic acid via increasing the excretion of cholesterol and bile acids in the bile by an as yet unknown mechanism. The cholesterol accumulating in obob mice fed 40% lard occurs by a different mechanism than when these mice are fed 1% corn oil since it is unaffected by taurocholenic acid.

Yohimbine and rauwolscine reduce food intake of genetically obese (obob) and lean mice

Multiple behavioral and neurochemical abnormalities are found in the genetically obese mouse, obob, including hyperphagia, elevated hypothalamic norepinephrine (NE) levels, and increases α-1 receptor density. The obese mutant also responds abnormally to neuropharmacological agents. In the current study the α-2 receptor blockers yohimbine and rauwolscine were administered to food-restricted (6-hour food access) obob and lean mice. Yohimbine and rauwolscine significantly reduced the 3- and 6-hour food intake of both obob and lean mice. The obob mice were, however, more sensitive to this anorectic effect than lean mice. Effective anorectic doses of yohimbine did not affect water intake in water-deprived lean mice, suggesting a specific effect of the drug upon food intake. Low doses (50 and 100 μg) of the α-2 agonist clonidine increased the 1-hour food intake of obob mice, but did not affect the food intake of lean mice. No differences were found between obob and lean mice in the number of α-receptors in the hypothalamus. The results suggest that modification of NE release by manipulation of α-2 receptor can alter food intake, and that the obob mutant is particularly sensitive to this effect.

Amphetamine Anorexia and Hypothalamic catecholamines in genetically obese mice ( obob)

Genetically obese mice ( obob) and their lean littermates were acclimated to a restricted food-access schedule of six hours and then treated with various doses of amphetamine (0, 3, 5 or 10 mg/kg). Saline-treated obob mice maintained on this schedule retained the primary characteristics of obob mice fed ad lib, i.e., hyperphagia, hyperglycemia, elevated hypothalamic norepinephrine (NE) levels. Both lean and obese mice treated with amphetamine showed a dose-dependent decrease in food intake and hypothalamic NE levels. In obob mice amphetamine treatment reduced food intake and hypothalamic NE levels to values which were not significantly different from those of similarly treated lean mice. When the drug dose was administered on a body weight basis, however, brain amphetamine levels were twice as high in obob as in lean mice. When the amphetamine dose was adjusted to produce approximately equivalent brain levels of amphetamine in obob and lean mice, the obob mice ate significantly more than lean mice. The results indicate that amphetamine is an effective anorectic agent capable of reducing food intake, body weight, and hypothalamic NE levels in obob mice.