Effect of a novel thermogenic beta-adrenoceptor agonist (BRL 26830) on insulin resistance in soleus muscle from obese Zucker rats

Young lean (Fa/?) and obese (fa/fa) rats were treated with the thermogenic beta-adrenoceptor agonist, BRL 26830, for 3 weeks. In lean rats this treatment had no effect on body weight but there was a marked increase in the insulin sensitivity of soleus muscle strips with respect to glycolytic rate. Treatment of obese rats with BRL 26830 produced a small but not significant decrease in body weight but the sensitivity of both glycolysis and glycogen synthesis to insulin was increased so that muscles of treated obese rats showed similar insulin sensitivity to untreated lean rats. It is suggested that such changes are unlikely to be merely a secondary consequence of an anti-obesity action.     

Increased insulin mediated glucose metabolism in fat cells from I versus C57BL mice

The purpose of this study was to determine whether adipocytes from I strain mice, which are characterized by a greater in vivo glucose tolerance than most other strains, had a higher capacity to utilize glucose in response to physiological concentrations of insulin. Using C57BL mice as a control strain, we examined the effect of insulin on glucose metabolism in epididymal and inguinal adipocytes from 2-month-old male mice. Body weight was only slightly less (7%) for the I mice than for the C57BL mice, but fat pad sizes were 60 and 20% less for epididymal and inguinal depots, respectively, in the I mice. Fat cell size was also smaller in epididymal adipocytes from the I mice than from the C57BL mice. Fat cell size of inguinal adipocytes was similar in the two strains. Without insulin the rates of [U-14C]glucose incorporation into CO2 or lipids were twofold higher in cells from the I mice than in those from the C57BL mice. Maximal insulin concentration (2.5 nM) increased glucose metabolism by 140 and 500% in epididymal and inguinal adipose cells, respectively, in the I mice versus 30 and 50% in the C57BL mice. The maximal effect of insulin was reached at a much higher insulin concentration in the I mice than in the C57BL mice. The activity of fatty acid synthetase was four- to sixfold higher in fat cells from I than in those from C57BL mice. These results demonstrate an increased insulin responsiveness of glucose metabolism in fat cells from the I mice related to an increased lipogenic capacity. Furthermore, they show that adipose tissue in mice exhibits significant regional differences in terms of insulin responsiveness of glucose metabolism.     

Increased membrane immunoglobulin capping of B cells from C57Bl/6 lpr/lpr and C57Bl/6 nu/nu mice

When the capping of membrane immunoglobulin on spleen B cells from normal C57Bl/6 mice (B6) is taken as reference, a faster capping rate is found for cells of age-matched B6 mice which are congenic at the lymphoproliferation (lpr) or nude (nu) loci. Though both congenic strains can be characterized by an abnormal T-lineage cell content, the nature of the abnormality itself is very different since B6 nudes lack thymus-processed/influenced lymphocytes whereas B6 mice with the lpr phenotype suffer from an invasion of all lymphoid organs with cells of a particular T-cell subset. Moreover, the more "normal" capping rate of B cells from the double congenic B6 mice (nu/nu, lpr/lpr) is intriguing. Since other mice homozygous at the lpr locus (MRL-1) or at the nu locus (BALB/c nude) also cap faster than their congenic controls (MRL-n and BALB/c, respectively), the observed effects do not appear to depend on a peculiarity of the B6 genetic background. If the faster capping of B cells of nu congenic and of lpr congenic mice had a common origin, it might be that T cells would control in some way the mobility of B-cell membrane immunoglobulins: both congenic mice have in their spleen a very low proportion of mature T cells together with a very high proportion of prethymic/thymic immature T-cell types, either of which might affect B-cell behavioral responses to membrane immunoglobulin clustering.     

D-LEU-4]-OB3, a synthetic leptin agonist, improves hyperglycemic control in C57BL/6J ob/ob mice

We have recently shown that the activity of a synthetic peptide corresponding to amino acid residues 116-130 of secreted mouse leptin is contained in a restricted sequence at the amino terminus of the peptide, between residues 116-122 (Ser-Cys-Ser-Leu-Pro-Gln-Thr, OB3). Substitution of the Leu residue at position 4 of OB3 with its D-isomer ([D-Leu-4]-OB3) enhanced the ability of OB3 (1 mg/day, ip, 7 days) to reduce body weight gain, food and water intake, and serum glucose in female C57BL/6J ob/ob mice. In the present study, we have utilized a pair-feeding approach to demonstrate that the antihyperglycemic action of [D-Leu-4]-OB3 is not solely due to its effects on caloric intake. One group of female C57BL/6J ob/ob mice (n=6) was fed ad libitum, and two additional groups (n=6 per group) were allowed 3.0 g food/mouse daily, an amount previously determined to satisfy [D-Leu-4]-OB3-treated mice. At the end of the 7-day test period, vehicle-injected mice fed ad libitum were approximately 10% heavier than their initial body weights, while pair-fed mice injected with vehicle and [D-Leu-4]-OB3-treated mice lost 5% of their initial body weights. After 1 day of treatment, blood glucose was reduced by 20% in pair-fed vehicle-injected mice, and by 40% in mice given [D-Leu-4]-OB3. Food restriction reduced blood glucose throughout the 7-day study, but not to levels seen in wild-type nonobese C57BL/6J mice of the same sex and age. After 2 days of treatment with [D-Leu-4]-OB3, however, blood glucose was reduced to levels comparable to those seen in wild-type nonobese mice. [D-Leu-4]-OB3 also lowered serum insulin levels by 53% when compared to mice fed ad libitum. Neither pair-feeding nor [D-Leu-4]-OB3 treatment had any apparent effect on thermogenesis. These results suggest that [D-Leu-4]-OB3 exerts its effects on serum glucose not only by suppressing caloric intake, but also through a separate effect on glucose metabolism which may involve increased tissue sensitivity to insulin.     

Pancreatic lesions of small-obese mice (C57BL/6 J-ob/ob) with hyperglucagonemia

Male Small-obese mice (Small-ob) which derived from a C57BL/6 J-ob/ob mouse colony were examined histopathologically at 13-, 39-, and over 52-week-old. C57BL/6 J-ob mice (?/+: Non-ob, ob/ob: Ob) were studied as controls. In Small-ob mice, plasma glucagon concentration was higher than that of the Ob mice (this difference was highly significant), and serum levels for insulin was within normal limits. Microscopically, hypertrophy and hyperplasia of the islets of Langerhans were found only in the pancreas of Ob mice. The increase in the number of A-cells and the decrease in the number of B-cells were revealed immunohistochemically in the islets of Small-ob mice. These changes were more severe with advance of age. In the aged Small-ob mice, perivascular and periductular cell infiltration were found, but inflammatory change of islet tissue was not confirmed in any animals examined. Diabetic symptoms in Small-ob mice seems to stem from the disparity in insulin/glucagon (I/G) ratio associated with hyperglucagonemia which result from increased number of A-cells of pancreatic islets.     

Acute effects of a beta-adrenoceptor agonist (BRL 26830A) on rat brown-adipose-tissue mitochondria. Increased GDP binding and GDP-sensitive proton conductance without changes in the concentration of uncoupling protein.

GDP binding, proton conductance and the specific concentration of uncoupling protein were measured in brown-adipose-tissue mitochondria of rats treated acutely with the novel beta-agonist, BRL 26830A. At 1 h after dosing with BRL 26830A, mitochondrial GDP binding was increased more than 2-fold. The increase in binding resulted from an increase in the number of binding sites. An iterative analysis of Scatchard binding data suggested that there is only one high-affinity GDP-binding site (Kd 0.3 microM) in brown-adipose-tissue mitochondria. The acute increase in GDP binding produced by treatment with BRL 26830A occurred without any alteration in the specific mitochondrial concentration of uncoupling protein, as determined by radioimmunoassay. Treatment with the beta-agonist did, however, lead to a small increase in the GDP-sensitive component of mitochondrial proton conductance. These results indicate that GDP-binding sites on uncoupling protein can be rapidly unmasked after treatment with a brown-fat-specific beta-agonist, and that the increase in binding reflects an increase in the activity of the mitochondrial proton-conductance pathway.     

Glucose oxidation,glucose transport and insulin binding in isolated rat epididymal adipocytes in the presence of anesthetics

Insulin binding and glucose oxidation were measured in isolated rat adipocytes in the presence of several anesthetics; ethanol, n-octanol, pentobarbital, chlorpromazine and tetracaine. Ethanol and chlorpromazine, at anesthetic and pentobarbitol at sub-anesthetic concentrations are inhibitory to both basal and insulin stimulated rates of glucose oxidation. At all concentrations of ethanol, pentobarbital or chlorpromazine tested binding of insulin is not affected. Since anesthetics may alter membrane fluidity, it is suggested that an anesthetic-induced increase in membrane fluidity beyond that which occurs at 37°C is detrimental to glucose oxidation. Of the 5 anesthetics examined, only chlorpromazine (10 μM or less) and tetracaine (500 μM) stimulate glucose oxidation. These two agents are known to bind to a cell's cytoskeletal system; the binding of chlorpromazine to microtubules is entropy driven. The temperature and concentration dependence of chlorpromazine stimulation of glucose oxidation (transport) are consistent with this form of binding. It is proposed that chlorpromazine binds to the cytoskeletal system of the adipocyte and that this system is normally restrictive to the motion of membrane proteins. Disruption of the cytoskeletal system by chlorpromazine or tetracaine would increase the frequency of insulin-receptor and glucose-carrier contact. Activation of glucose transport could ensue.     

Spontaneous, asymmetrical microphthalmia in C57B1/6J mice

An investigation was undertaken to evaluate ocular size and shape by objective and subjective criteria to establish the incidence and laterality of spontaneous microphthalmia in the C57B1/6J mouse strain. The purpose was to provide useful information of normal and abnormal ocular development in a laboratory strain frequently used for teratogenic studies. Ten-week-old C57B1/6J mice were time mated and sacrificed at gestational day 14. Eye normality was evaluated subjectively using veterinary ophthalmology methods, and objectively by a computer derived method. A distinct laterality was noted in the right eye when compared to the left eye. Degree of laterality varied with assessment methodology (19.0% vs. 6.9% and 8.9% vs. 4.0% for the subjective and objective methods respectively). Mean eye area and histogram distributions for the right eye were consistently smaller than the left eye for all anatomical rating groups, thereby suggesting that asymmetry found in the C57B1/6J may be due to an inherent discrepancy in eye size in the strain. The data from this study suggest that when C57B1/6J mouse eyes are used as end points for teratological testing each eye should be treated as an independent unit of measure.     

Adrenal function and the effect of a high-fat diet on C57BL/6J and C57BL/6J-ob/ob mice.

In C57BL/6J mice and the ob/+ and ob/ob mutants total plasma corticosterone levels were found to be statistically different. In C57BL/6J mice the level was 1.9 +/- 0.2 mug/100 ml plasma, in ob/+ mice 8.6 +/- 1.6 mug/100 ml and in ob/ob mice 13.7 +/- 1.5 mug/100 ml. The percentage of protein-bound corticosterone as well as the free endogenous corticosterone levels were also different. Feeding a high-fat diet to young C57BL/6J and C57BL/6J-ob/ob mice for a period of 4 weeks had no effect upon blood glucose, plasma insulin and plasma corticosterone levels. The significantly higher increase in body weight of the high-fat diet groups of both lines of mice was mainly due to fat cell hypertrophy.     

MPTP诱导小鼠黑质区铁摄取和DMT1表达增加

铁在帕金森病(Parkinson‘s disease,PD)的发病机制中起着非常关键的作用,为了探讨PD中铁升高的机制,本实验观察了1-甲基4-苯基—1,2,3,6-四氢吡啶(MPTP)处理小鼠黑质(substantia nigra,SN)内铁摄取及新的铁转运蛋白二价金属离子转运蛋白1(DMT1)的表达变化。结果表明：(1)MPTP处理组小鼠SN内铁染色增高,注射MPTP7d组明显高于3d组。(2)MPTP处理组小鼠,酪氨酸羟化酶(TH)免疫阳性细胞数目显著减少。(3)MPTP处理组小鼠,“-IRE”型DMT1表达在各组中均增加,而“ IRE”型DMT1仅在MPTP处理后7d才出现变化。上述结果提示,这种新发现的哺乳动物跨膜铁转运蛋白表达增加可能是引起MPTP处理小鼠SN中铁升高的关键因素,铁的升高进一步导致DA神经元的死亡。     

The adipose organ of obesity-prone C57BL/6J mice is composed of mixed white and brown adipocytes

White and brown adipocytes are believed to occupy different sites in the body. We studied the anatomical features and quantitative histology of the fat depots in obesity and type 2 diabetes-prone C57BL/6J mice acclimated to warm or cold temperatures. Most of the fat tissue was contained in depots with discrete anatomical features, and most depots contained both white and brown adipocytes. Quantitative analysis showed that cold acclimation induced an increase in brown adipocytes and an almost equal reduction in white adipocytes; however, there were no significant differences in total adipocyte count or any signs of apoptosis or mitosis, in line with the hypothesis of the direct transformation of white into brown adipocytes. The brown adipocyte increase was accompanied by enhanced density of noradrenergic parenchymal nerve fibers, with a significant correlation between the density of these fibers and the number of brown adipocytes. Comparison with data from obesity-resistant Sv129 mice disclosed a significantly different brown adipocyte content in C57BL/6J mice, suggesting that this feature could underpin the propensity of the latter strain to develop obesity. However, the greater C57BL/6J browning capacity can hopefully be harnessed to curb obesity and type 2 diabetes in patients with constitutively low amounts of brown adipose tissue.     

Alterations in mRNA levels, expression, and function of GTP-binding regulatory proteins in adipocytes from obese mice (C57BL/6J-ob/ob)

Messenger RNA levels for the alpha subunit of G-proteins expressed in adipocytes of lean and obese (ob/ob) mice were compared with relative levels of the encoded proteins. Using both toxin labeling and Western blots, expression of Gs alpha, Gi alpha-1, and Gi alpha-3 was decreased by approximately 2-fold in adipocytes of obese mice, while levels of Gi alpha-2 did not differ between the phenotypes. The decreases in Gi alpha-1 and Gs alpha in the obese mouse were attributed to decreased mRNA levels for these proteins. Similar mRNA levels for Gi alpha-3 were noted in both phenotypes, but Gi alpha-2 message was increased 2-fold in the obese mouse. Inhibitory regulation of adipocyte adenylylcyclase through G-proteins was evaluated by comparing the ability of R-PIA to inhibit isoproterenol-stimulated responses between the phenotypes. In spite of the decrease in Gi alpha-1 and Gi alpha-3 in adipocytes from obese mice, R-PIA inhibited adenylylcyclase, cAMP-dependent protein kinase, and lipolysis in similar fashion in both phenotypes. The GTP analog, Gpp(NH)p also inhibited forskolin-stimulated adenylylcyclase in a comparable manner, but the magnitude of the inhibition was slightly less in adipocyte membranes from obese mice. In contrast, the decrease in expression of Gs alpha was translated into substantially poorer activation of isoproterenol-stimulated responses in the obese mouse. The concentration of isoproterenol producing half-maximal activation of adenylylcyclase, protein kinase, and lipolysis did not differ between the phenotypes, but the maximal responses were much lower in cells from obese mice. Similar lipolytic potential in isolated adipocytes from each phenotype and similar total forskolin-stimulated cyclase activity in adipocyte membranes from each phenotype suggest that decreased expression of Gs alpha may contribute to the characteristic alteration in mobilization of triglycerides noted in adipocytes from obese mice.     

Beta 3-adrenoceptor knockout in C57BL/6J mice depresses the occurrence of brown adipocytes in white fat.

White and brown adipocytes are usually located in distinct depots; however, in response to cold, brown adipocytes appear in white fat. This response is mediated by beta-adrenoceptors but there is a controversy about the subtype(s) involved. In the present study, we exposed to cold beta 3-adrenoceptor knockout mice (beta 3KO) on a C57BL/6J genetic background and measured in white adipose tissue the density of multilocular cells and the expression of the brown adipocyte marker uncoupling protein-1 (UCP1). In brown fat of beta 3KO mice, UCP1 expression levels were normal at 24 degrees C as well as after a 10-day cold exposure. Strikingly, under both conditions, in the white fat of beta 3KO mice the levels of UCP1 mRNA and protein as well as the density of multilocular cells were decreased. These results indicate that beta 3-adrenoceptors play a major role in the appearance of brown adipocytes in white fat and suggest that the brown adipocytes present in white fat differ from those in brown fat.     

Identification of Staphylococcus xylosus isolated from C57BL/6J-Nos2(tm1Lau) mice with dermatitis

Coagulase negative staphylococci (CNS) are significant pathogens, particularly in medical device related infections and in immunocompromised patients. Five CNS strains were isolated from 5 NOS2 knockout mice with dermatitis. Histologically, granulomatous dermatitis was found in the skin around the ear with epidermal ulceration. Dermal lesions included pustules, necrosis, and accumulations of neutrophils and macrophages. Isolates of the bacterial strains were identified to be Staphylococcus xylosus by the API STAPH kit and 16S-23S intergenic transcribed spacer-PCR. These results demonstrate the potential of this organism to be an opportunistic pathogen in immunocompromised mice.     

Immune responses to the pertussis toxin in Balb/c and C57B1/6 mice

Abstract Immunization of Balb/c and C57B1/6 mice with the pertussis toxin (Ptx), purified from the culture supernatant of Bordetella pertussis (the whooping cough bacillus) resulted in different immune reactions in these genetically different strains of mice. Antibody responses to Ptx were detected only in Balb/c, whereas both Balb/c and C57B1/6 produced anti-Ptx antibodies when immunized with detoxified Ptx. Also, delayed-type hypersensitivity reactions differ strongly according to the use of Ptx or detoxified Ptx as eliciting antigen.     

6-h advances alter circadian activity patterns,fasting glucose,and insulin levels in C57BL6/J mice

Chronobiological disruptions, including shift work, have been linked to a number of disorders such as fatigue and diabetes. Additionally, there is evidence to support that exercise cannot only counteract fatigue and the onset of diabetes, but also alleviate the other negative symptoms associated with shift work. Therefore, the present study investigated the effects of wheel running and monthly 6-h phase advances on the circadian locomotor activity patterns and glucose and insulin levels in C57BL6/J mice. 6-h phase advances produced decreases in fasting glucose and increases in insulin, and wheel-running was able to alleviate the spike in insulin secretion. Additionally, mice experiencing the shift increased their food intake, despite having no change in body mass. Circadian wheel-running activity was also altered in phase-advanced mice. These results provide further evidence that chronobiological disruptions can lead to alterations in physiology and behavior, and that exercise can alleviate some of those symptoms.     

Lactobacillus plantarum LRCC 5273 isolated from Kimchi ameliorates diet-induced hypercholesterolemia in C57BL/6 mice

ABSTRACT

This study was designed to select potent cholesterol-lowering probiotic strains on HepG2 cell and investigate the effect of selected strain, Lactobacillus plantarum LRCC 5273 and LRCC 5279 in hypercholesterolemic mice. In the results, LP5273 group showed significantly reduced total and LDL cholesterol compared to HCD group. In addition to significantly up-regulated hepatic mRNA expression of LXR-α and CYP7A1, intestinal LXR-α and ABCG5 were significantly up-regulated in LP5273 group. With activation of hepatic and intestinal LXR-α and its target genes, fecal cholesterol and bile acid excretion were increased in LP5273 fed mice. These results suggest that LP5273 ameliorates hypercholesterolemia in mice through the activation of hepatic and intestinal LXR-α, resulting in enhancement of fecal cholesterol and bile acids excretion in the small intestine. The results of present study suggest mechanistic evidences for hypocholesterolemic effects of L. plantarum spp., and may contribute to future researches for prevention of hypercholesterolemia and cardiovascular disease.     

Increased norepinephrine by medium-chain triglyceride attributable to lipolysis in white and brown adipose tissue of C57BL/6J mice

A further investigation of the lipolysis induced by medium-chain triglyceride (MCT) was conducted on C57BL/6J mice fed with a diet containing 2% MCT or 2% long-chain triglyceride (LCT). Blood norepinephrine, body fat and blood lipid variables, and the protein or mRNA expression of the genes relevant to lipolysis were measured and analyzed in the white and brown adipose tissue (WAT, BAT). Decreased body fat and improved blood lipid profiles attributable to MCT were confirmed. A higher level of blood norepinephrine was observed with the MCT diet. The adipose triglyceride lipase (ATGL) activity and its mRNA expression, the expression of protein and mRNA of the beta 3 adrenergic receptor (β3-AR) in both WAT and BAT, and the hormone-sensitive lipase (HSL) activity and its mRNA expression in BAT were significantly increased in the mice with MCT feeding. The lipolysis induced by MCT might be partially mediated by increasing norepinephrine, thereafter signaling the up-regulation of β3-AR, ATGL, and HSL in WAT and BAT.