Endothelin-1 transgenic mice develop glomerulosclerosis, interstitial fibrosis, and renal cysts but not hypertension.

The human endothelin-1 (ET-1) gene under the control of its natural promoter was transferred into the germline of mice. The transgene was expressed predominantly in the brain, lung, and kidney. Transgene expression was associated with a pathological phenotype manifested by signs such as age-dependent development of renal cysts, interstitial fibrosis of the kidneys, and glomerulosclerosis leading to a progressive decrease in glomerular filtration rate. This pathology developed in spite of only slightly elevated plasma and tissue ET-1 concentrations. Blood pressure was not affected even after the development of an impaired glomerular filtration rate. Therefore, these transgenic lines provide a new blood pressure-independent animal model of ET-1-induced renal pathology leading to renal fibrosis and fatal kidney disease.     

Muscle-specific PPARgamma-deficient mice develop increased adiposity and insulin resistance but respond to thiazolidinediones

Activation of peroxisome proliferator-activated receptor gamma (PPARgamma) by thiazolidinediones (TZDs) improves insulin resistance by increasing insulin-stimulated glucose disposal in skeletal muscle. It remains debatable whether the effect of TZDs on muscle is direct or indirect via adipose tissue. We therefore generated mice with muscle-specific PPARgamma knockout (MuPPARgammaKO) using Cre/loxP recombination. Interestingly, MuPPARgammaKO mice developed excess adiposity despite reduced dietary intake. Although insulin-stimulated glucose uptake in muscle was not impaired, MuPPARgammaKO mice had whole-body insulin resistance with a 36% reduction (P < 0.05) in the glucose infusion rate required to maintain euglycemia during hyperinsulinemic clamp, primarily due to dramatic impairment in hepatic insulin action. When placed on a high-fat diet, MuPPARgammaKO mice developed hyperinsulinemia and impaired glucose homeostasis identical to controls. Simultaneous treatment with TZD ameliorated these high fat-induced defects in MuPPARgammaKO mice to a degree identical to controls. There was also altered expression of several lipid metabolism genes in the muscle of MuPPARgammaKO mice. Thus, muscle PPARgamma is not required for the antidiabetic effects of TZDs, but has a hitherto unsuspected role for maintenance of normal adiposity, whole-body insulin sensitivity, and hepatic insulin action. The tissue crosstalk mediating these effects is perhaps due to altered lipid metabolism in muscle.     

Interstitial insulin concentrations determine glucose uptake rates but not insulin resistance in lean and obese men.

Insulin action and obesity are both correlated with the density of muscle capillary supply in humans. Since the altered muscle anatomy in the obese might affect interstitial insulin concentrations and reduce insulin action, we have cannulated peripheral lymphatic vessels in lean and obese males, and compared peripheral lymph insulin concentrations with whole body glucose uptake during a euglycemic, hyperinsulinemic clamp. Lymph insulin concentrations in the lower limb averaged only 34% of arterial insulin concentrations during 150 min of insulin infusion. Obese subjects had the highest arterial (P < or = 0.0001) and lymph insulin (P < 0.005) concentrations, but the lowest glucose uptake rates (P < 0.002). In contrast to the initial steep rise then plateau of arterial insulins, both lymph insulin and whole body glucose uptake rates rose slowly and did not consistently reach a plateau. In each individual, the glucose uptake closely correlated with peripheral lymphatic insulin concentrations (mean r2 = 0.95). The coupling between glucose uptake and lymph insulin (glucose uptake/pmol insulin) was much steeper in lean subjects than in the obese (P < or = 0.0001). These results indicate that even if insulin diffusion into tissues is rate limiting for insulin action, a tissue defect rather than an insulin diffusion defect causes insulin resistance in obese subjects.     

Recombinant Eimeria protozoan protein elicits resistance to acute phlebovirus infection in mice but not hamsters

A protein antigen from an Eimeria protozoan has recently been reported to induce antitumor activity in mice. This activity most likely results from the strong induction of interkeukin-12 (IL-12) and gamma interferon (IFN-gamma), which are also essential factors in the establishment of protective immunity against viral infection. We evaluated recombinant Eimeria antigen (rEA) as a potential immunotherapeutic agent in mouse and hamster models of acute phleboviral disease. Punta Toro virus (PTV) was highly sensitive to a single dose of nanogram quantities of rEA in the mouse infection model. Intraperitoneal treatment with rEA also reduced virus load and liver damage associated with PTV infection. IL-12 was elicited following exposure of uninfected mice to quantities of rEA of 10 ng or greater, and the levels peaked at between 3 and 8 h postexposure. IFN-gamma release was induced more slowly and required less rEA (1 ng) to produce a significant rise in systemic levels. The induction of IL-12 and IFN-gamma involved in the coordination of innate and adaptive immune responses to microbial pathogens required myeloid differentiation factor 88, a signaling adaptor shared by most members of the Toll-like receptor (TLR) family. Despite encouraging results in the murine system, rEA failed to protect hamsters challenged with PTV. Our findings suggest that hamsters may lack functional TLR11, which has recently been shown to recognize a profilin-like protein homologous to rEA from the protozoan Toxoplasma gondii. Further investigation into the immunostimulatory capacity of rEA in other mammalian systems is necessary.     

Evidence for a central but not a peripheral analgesic effect of clomipramine in rats

The effect of clomipramine (CMI), a tricyclic antidepressant, was studied on an acute inflammatory pain model in an attempt to understand its potential antinociceptive activity, the involvement of a central and/or peripheral component and its influence on the inflammatory process. When administered (i.v.) before the inflammatory agent, carrageenan (CAR), CMI (0.125, 0.25 and 0.5 mg/kg) completely prevented the development of the hyperalgesia for 70-120 min according to the doses. This antinociceptive effect was suppressed by naloxone (100 micrograms/kg i.v.) for 65 min. Neither higher doses (1, 2 and 20 mg/kg, i.v.) nor CMI injected into the inflamed paw (15 min before CAR) modified pain thresholds. Moreover, CMI (0.5 and 2 mg/kg, i.v.) administered 15 min before CAR markedly increased the volume of the CAR-induced oedema. These results (1) demonstrate an opioid-dependent antinociceptive effect of CMI on this model, the doses used being lower than those active in thermal or electrical tests, and (2) tend to exclude a peripheral mechanism and an NSAID-like anti-inflammatory activity suggested by previous in vitro studies.     

Pregnancy and amyloidogenesis: I. Offspring of amyloidotic mice are not predisposed to develop amyloidosis.

Amyloid enhancing factor (AEF) is a substance formed during amyloidogenesis that accelerates amyloid deposition in tissues. The administration of AEF followed by AgNO3 (inflammatory stimulus) to mice results in amyloidosis within 6 days. The purpose of the study was to determine whether the offspring of amyloidotic mice are exposed to maternal AEF during pregnancy and therefore become predisposed to the development of amyloidosis on inflammatory stimulus. To that end female mice were made amyloidotic by the administration of AEF and AgNO3, made pre-amyloidotic (a condition associated with self-generation of AEF) with a short course of casein, or treated with exogenous AEF without AgNO3; then mating and conception took place. The possible priming of offspring with maternal AEF was studied by the administration of AgNO3 alone (without AEF) to the offspring and the determination of the presence of amyloid deposits in their spleens. Despite the long-term effect of AEF and its high activity, amyloidosis did not develop in any of the studied offspring, implying that the newborn mice were not primed by maternal AEF. These findings suggest that amyloidotic mothers do not predispose their offspring to the risk of developing amyloidosis, probably because maternal AEF does not cross the placenta.     

瘦素在肥胖哮喘小鼠气道重构中的作用

目的探讨脂肪因子瘦素在支气管哮喘气道重构中的作用。方法高脂饮食制造肥胖模型,经腹腔注射与雾化吸入卵蛋白(OVA)制作慢性哮喘模型。45只雌性C57/6J小鼠随机分为对照组,单纯哮喘组,肥胖哮喘组,每组15只,于末次激发48 h内处死小鼠;计数支气管肺泡灌洗液(BALF)中细胞总数及分类,肺组织HE观察各组小鼠肺组织病理变化及ELISA法测定血清中瘦素水平。结果肥胖哮喘组小鼠BALF中白细胞总数及嗜酸性粒细胞比例以及气道管壁厚度和气道平滑肌厚度均较单纯哮喘组和对照组显著增加(P0.05)。对照组血清瘦素为(1 438.54±208.14)pg/mL,明显低于单纯哮喘和肥胖哮喘组[(1 677.75±214.36)pg/mL,(2 114.20±189.15)pg/mL,P0.05)],血清瘦素水平与气管壁厚度和气道平滑肌厚度呈正相关(P0.05)。结论肥胖通过瘦素参与了哮喘气道重构过程,纠正瘦素失衡有望成为肥胖哮喘患者治疗的新方向。     

Indomethacin, alone and combined with prochlorperazine and caffeine, but not sumatriptan, abolishes peripheral and central sensitization in in vivo models of migraine.

Recently it has been proposed that the throbbing pain of migraine is mediated by sensitization of peripheral trigeminovascular neurons, and that cutaneous allodynia of migraine is mediated by sensitization of central trigeminovascular neurons, and, moreover, that the triptans are less effective in aborting a migraine attack if the central sensitization is already established. The combination of indomethacin, prochlorperazine, and caffeine (IndoProCaf) is a drug of well-established use in the acute treatment of migraine. The aim of this study was to investigate whether the 3 active principles of IndoProCaf, alone and combined, compared to sumatriptan, were able to abolish the peripheral sensitization induced by kainic acid and the central sensitization induced by N-methyl-D-aspartate (NMDA) in in vivo models of hyperalgesia. The study showed that indomethacin or IndoProCaf is able to abolish both the kainic acid-induced and the NMDA-induced hyperalgesia. If administered at different times, IndoProCaf was always effective in reversing the kainic acid-induced hyperalgesia. Sumatriptan was not able to reverse either the kainic acid-induced or the NMDA-induced hyperalgesia. The efficacy of indomethacin, alone and combined with prochlorperazine and caffeine, in abolishing peripheral and central sensitization in in vivo models of hyperalgesia is a further explanation of the clinical efficacy of IndoProCaf in the treatment of migraine. PERSPECTIVE: This study suggests that, although triptans were shown to be able to abort migraine attacks only if given before the establishment of cutaneous allodynia and central sensitization, IndoProCaf should be able to abort migraine attacks independently from the time of administration, because it is able to abolish an already established peripheral and central sensitization.     

Syntaxin 4 heterozygous knockout mice develop muscle insulin resistance

To investigate the physiological function of syntaxin 4 in the regulation of GLUT4 vesicle trafficking, we used homologous recombination to generate syntaxin 4-knockout mice. Homozygotic disruption of the syntaxin 4 gene results in early embryonic lethality, whereas heterozygous knockout mice, Syn4(+/-), had normal viability with no significant impairment in growth, development, or reproduction. However, the Syn4(+/-) mice manifested impaired glucose tolerance with a 50% reduction in whole-body glucose uptake. This defect was attributed to a 50% reduction in skeletal muscle glucose transport determined by 2-deoxyglucose uptake during hyperinsulinemic-euglycemic clamp procedures. In parallel, insulin-stimulated GLUT4 translocation in skeletal muscle was also significantly reduced in these mice. In contrast, Syn4(+/-) mice displayed normal insulin-stimulated glucose uptake and metabolism in adipose tissue and liver. Together, these data demonstrate that syntaxin 4 plays a critical physiological role in insulin-stimulated glucose uptake in skeletal muscle. Furthermore, reduction in syntaxin 4 protein levels in this tissue can account for the impairment in whole-body insulin-stimulated glucose metabolism in this animal model.     

CXCL12 mediates CCR7-independent homing of central memory cells, but not naive T cells, in peripheral lymph nodes

Central memory CD8(+) T cells (T(CM)) confer superior protective immunity against infections compared with other T cell subsets. T(CM) recirculate mainly through secondary lymphoid organs, including peripheral lymph nodes (PLNs). Here, we report that T(CM), unlike naive T cells, can home to PLNs in both a CCR7-dependent and -independent manner. Homing experiments in paucity of lymph node T cells (plt/plt) mice, which do not express CCR7 ligands in secondary lymphoid organs, revealed that T(CM) migrate to PLNs at approximately 20% of wild-type (WT) levels, whereas homing of naive T cells was reduced by 95%. Accordingly, a large fraction of endogenous CD8(+) T cells in plt/plt PLNs displayed a T(CM) phenotype. Intravital microscopy of plt/plt subiliac lymph nodes showed that T(CM) rolled and firmly adhered (sticking) in high endothelial venules (HEVs), whereas naive T cells were incapable of sticking. Sticking of T(CM) in plt/plt HEVs was pertussis toxin sensitive and was blocked by anti-CXCL12 (SDF-1alpha). Anti-CXCL12 also reduced homing of T(CM) to PLNs in WT animals by 20%, indicating a nonredundant role for this chemokine in the presence of physiologic CCR7 agonists. Together, these data distinguish naive T cells from T(CM), whereby only the latter display greater migratory flexibility by virtue of their increased responsiveness to both CCR7 ligands and CXCL12 during homing to PLN.     

Insulin resistance, role of leptin and leptin receptor

Leptin, the obese gene product, is an adipocyte-derived satiety factor which is involved in the regulation of food intake and energy expenditure. Obesity often accompanies insulin resistance and high levels of leptin. In in vitro studies, leptin has been reported to increase fatty acid oxidation and decrease fatty acid synthesis in adipocytes and hepatocytes. The direct effects of leptin on glucose metabolism and insulin signaling have not been clarified yet. In in vivo studies, however, leptin has been reported to improve insulin sensitivity and glucose metabolism in normal and obese rodents acting mainly through hypothalamus. Moreover leptin has been reported to have antidiabetic effects in insulin-deficient diabetes rats and lipoatrophic diabetes mice. It is suggested that leptin modulates insulin sensitivity and glucose disposal and that leptin may have a pathophysiological and therapeutic implications in diabetes.     

既要发展现代技术 也要继承经典方法

科学技术和临床医学的飞速发展，极大地促进了检验医学技术的现代化进程，使检验医学在疾病的诊断、治疗、预防和康复中发挥着不可替代的重要作用。     

Novel SHV-derived extended-spectrum beta-lactamase, SHV-57, that confers resistance to ceftazidime but not cefazolin

A new SHV-derived extended-spectrum beta-lactamase, SHV-57, that confers high-level resistance to ceftazidime but not cefotaxime or cefazolin was identified from a national surveillance study conducted in Taiwan in 1998. An Escherichia coli isolate resistant to ampicillin, cephalothin, and ceftazidime but sensitive to cefoxitin, ceftriaxone, cefotaxime, imipenem, and a narrow-spectrum cephem (cefazolin) was isolated from the urine of a patient treated with beta-lactam antibiotics. Resistance to beta-lactams was conjugatively transferred with a plasmid of about 50 kbp. The pI of this enzyme was 8.3. The sequence of the gene was determined, and the open reading frame of the gene was found to consist of 861 bases (GenBank accession number AY223863). Kinetic parameters showed that SHV-57 had a poor affinity to cefazolin. The K(m) value toward cefazolin (5.57 x 10(3) muM) was extremely high in comparison to those toward ceftazidime (30.9 muM) and penicillin G (67 muM), indicating its low affinity to cefazolin. Although the K(m) value of the beta-lactamase inhibitor was too high for the study of catalytic activity (k(cat)), indicating the low k(cat) of SHV-57, the SHV-57 carrier was highly susceptible to a beta-lactam-beta-lactamase inhibitor combination. Comparison of the three-dimensional molecular model of SHV-57 with that of the SHV-1 beta-lactamase suggests that the substitution of arginine for leucine-169 in the Omega loop is important for the substrate specificity.     

Acute insulin administration does not affect plasma leptin levels in lean or obese subjects

Abstract. Whether leptin levels are related to insulin sensitivity or subject to acute regulation by insulin is not known. In 12 obese [body mass index (BMI) = 34.0 ±1.5 kg m^-2] and 12 lean (BMI = 22.2 ±0.6 kg m^-2) non-diabetic subjects, plasma leptin concentrations were measured in the fasting state and during 2 hours of euglycaemic hyperinsulinaemia (˜600 pmol L^-2). Fasting plasma leptin was significantly higher in obese (26.6 ±3.2) than in lean subjects (6.4 ±1.2 ng mL^-1, P = 0.0001), and in women (21.1 ±3.3) than in men (7.3 = 2.3 ng mL^-1, P = 0.01). In univariate analysis, fasting plasma leptin was strongly related to all anthropometric measures (body weight, fat mass, percent fat mass, waist and hip circumferences). In multiple regression, per cent adiposity, hip circumference and duration of obesity explained 90% of the variability in fasting leptin concentrations. Fasting and stimulated (OGTT) insulin levels, insulin sensitivity (22.6 ±1.9 vs 36.7 ±2.0 μmol min^-1 kg^-1 in lean and obese subjects, respectively, P < 0.0001), glucose area, and serum triglycerides were positively related to fasting plasma leptin concentrations; none of these associations, however, was statistically significant after adjusting for BMI. During the clamp, plasma leptin concentrations remained constant in both lean and obese subjects. We conclude that neither insulin levels nor sensitivity relate to leptin levels independently of fat mass, and that leptin is not subject to acute (2 hours) regulation by insulin in lean or obese humans.     

Nocturnal rise of leptin in lean, obese, and non-insulin-dependent diabetes mellitus subjects.

We studied 24-h profiles of circulating leptin levels using a sensitive and specific RIA in lean controls and obese subjects with or without non-insulin-dependent diabetes mellitus (NIDDM) during normal routine activity. Serum leptin levels were significantly higher in obese (41.7 +/- 9.0 ng/ml; n = 11) and obese NIDDM (30.8 +/- 6.7; n = 9) subjects compared with those in lean controls (12.0 +/- 4.4, n = 6). In all the three groups, serum leptin levels were highest between midnight and early morning hours and lowest around noon to midafternoon. The nocturnal rise in leptin levels was significant when data were analyzed by ANOVA (lean: F = 3.17, P < 0.0001, n = 4; obese: F = 2.02, P < 0.005, n = 11; and obese NIDDM: F = 4.9, P < 0.0001, n = 5). The average circadian amplitude between acrophase and nadir was 75.6% in lean, 51.7%, in obese and 60.7% in obese NIDDM groups, respectively. No significant correlations (P > 0.05) were observed between circulating levels of leptin and either insulin or glucose levels in any of the 20 subjects studied for 24-h profiles. The nocturnal rise in leptin observed in the present study resembles those reported for prolactin, thyroid-stimulating hormone, and free fatty acids. We speculate that the nocturnal rise in leptin could have an effect in suppressing appetite during the night while sleeping.