272.Decreased expression of oestrogen receptor β in the reproductive tract of pregnant relaxin-deficient (Rlx - / - ) mice

The peptide hormone relaxin (RLX) is reported to directly affect uterine oestrogen receptors (ERs) in the rat (1). Treatment of immature ovariectomised rats with porcine RLX causes a decrease in uterine ERβ mRNA levels within 6 h. However, RLX has no effect on ERα expression. As both ERβ1 and ERβ2 inhibit ER-mediated transcriptional activity, this RLX-induced downregulation in ERβ could be a prerequisite for oestrogen to exert its effects on target tissues. The aim of the current study was to use relaxin-deficient (Rlx–/–) pregnant mice to investigate if relaxin deficiency results in alterations in either ERβ or ERα mRNA expression in reproductive tissues. Cervix and vagina tissues were obtained from adult C57/Blk6J wild-type mice at five stages of gestation (Days 7.5, 10.5, 14.5, 17.5, 18.5 pc) and Rlx–/– littermates on Days 7.5, 14.5 and 18.5 pc. Q-PCR with TaqMan probes in the Opticon 2 thermal cycler (MJ Research, GeneWorks) was used to quantify ERα and ERα gene expression. ERα mRNA levels were significantly (P < 0.05; ANOVA) increased in the cervix/vagina on Days 17.5 and 18.5 pc in Rlx+/+ mice. The increase in ERα in Rlx+/+ mice was negatively correlated with a significant decrease in ERβ expression from Day 14.5 pc. In contrast, there was no decrease in ERβ expression in the cervix/vagina of Rlx–/– mice; ERβ mRNA levels were significantly (P < 0.05) higher compared to Rlx+/+ mice on Days 14.5 or 18.5 pc. However, there was no corresponding reduction in ERα expression in the cervix/vagina of the Rlx–/– mice, so that ERα mRNA levels were still elevated at term despite the maintenance of high ERα expression. In summary, these data show changes in ERα expression in the cervix/vagina of relaxin-deficient mice, which may subsequently affect ERα-mediated transcriptional activity. (1) Pillai et al. (2002) Biol. Reprod. 67, 1919–1926.

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201. Effects of relaxin deficiency on matrix metalloproteinase expression in the cervix and vagina of pregnant mice

Reproduction Fertility and Development ◽

10.1071/srb05abs201 ◽

2005 ◽

Vol 17 (9) ◽

pp. 76

Author(s):

J. T. McGuane ◽

H. M. Gehring ◽

L. J. Parry

Keyword(s):

Reproductive Tract ◽

Late Gestation ◽

Mrna Levels ◽

Biochemical Processes ◽

Fluorogenic Probes ◽

Pregnant Mice ◽

Ecm Degradation ◽

Stromal Collagen

The major functions of relaxin are associated with female reproductive physiology, especially the regulation of biochemical processes involved in the remodelling of the reproductive tract at term. Studies in relaxin deficient mice (Rlx–/–) demonstrate that although females give birth to live young without apparent dystocia, they have abnormal cervices and vaginae. This phenotype is attributed to an increase in stromal collagen, but the mechanism(s) by which relaxin regulates extracellular matrix (ECM) production in reproductive tissues is poorly understood. In this study, we assessed the expression of matrix metalloproteinases (MMPs) in the cervix and vagina of pregnant wild-type (Rlx+/+) and Rlx–/– mice. Tissues were obtained from adult C57/Blk6J Rlx+/+ mice on days 7.5, 14.5, 17.5, 18.5 pc and Rlx–/– littermates on days 7.5, 14.5 and 18.5 pc. Real-time PCR using dual-labelled fluorogenic probes was performed in an Opticon 2 cycler (MJ Research) to quantify MMP-2, -3, -7, -9 and -13 gene expression. In the cervix and vagina of Rlx+/+ mice, only MMP-2 mRNA levels were significantly higher at term compared with earlier stages of gestation. There were significant decreases in MMP-7 and -13 expression at term, but no change in MMP-3 and -9. In contrast, MMP-3, -7, -9 and -13 mRNA levels were significantly higher in the cervix and vagina of late pregnant Rlx–/– mice. The expression of MMP-2 did not differ between Rlx+/+ and Rlx–/– mice at term. Despite the higher expression of the majority of MMPs we examined in Rlx–/– mice, there was no histological evidence of increased ECM degradation in the cervix and vagina in late gestation. Although previous in vitro studies suggest that relaxin positively regulates MMP activity, our data demonstrate that relaxin deficiency does not result in decreased MMP expression in the mouse cervix and vagina in vivo.

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Differential response to myocardial reperfusion injury in eNOS-deficient mice

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00855.2001 ◽

2002 ◽

Vol 282 (6) ◽

pp. H2422-H2426 ◽

Cited By ~ 55

Author(s):

Brent R. Sharp ◽

Steven P. Jones ◽

David M. Rimmer ◽

David J. Lefer

Keyword(s):

Ischemia Reperfusion ◽

Myocardial Necrosis ◽

In Vivo Model ◽

Pcr Analysis ◽

Inos Mrna ◽

Mrna Levels ◽

Wild Type ◽

Significant Difference ◽

Deficient Mice

Two strains of endothelial nitric oxide synthase (eNOS)-deficient (−/−) mice have been developed that respond differently to myocardial ischemia-reperfusion (MI/R). We evaluated both strains of eNOS−/− mice in an in vivo model of MI/R. Harvard (Har) eNOS−/− mice ( n = 12) experienced an 84% increase in myocardial necrosis compared with wild-type controls ( P < 0.05). University of North Carolina (UNC) eNOS−/−( n = 10) exhibited a 52% reduction in myocardial injury versus wild-type controls ( P < 0.05). PCR analysis of myocardial inducible NO synthase (iNOS) mRNA levels revealed a significant ( P < 0.05) increase in the UNC eNOS−/− mice compared with wild-type mice, and there was no significant difference between the Har eNOS−/− and wild-type mice. UNC eNOS−/− mice treated with an iNOS inhibitor (1400W) exacerbated the extent of myocardial necrosis. When treated with 1400W, Har eNOS−/− did not exhibit a significant increase in myocardial necrosis. These data demonstrate that two distinct strains of eNOS−/− mice display opposite responses to MI/R. Although the protection seen in the UNC eNOS−/− mouse may result from compensatory increases in iNOS, other genes may be involved.

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Disassociation of insulin action and Akt/FOXO signaling in skeletal muscle of older Akt-deficient mice

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.00358.2012 ◽

2012 ◽

Vol 303 (11) ◽

pp. R1186-R1194 ◽

Cited By ~ 7

Author(s):

Thomas H. Reynolds ◽

Erin Merrell ◽

Nicholas Cinquino ◽

Megan Gaugler ◽

Lily Ng

Keyword(s):

High Fat Diet ◽

Insulin Action ◽

Mrna Levels ◽

Wild Type ◽

Akt Phosphorylation ◽

Protein Levels ◽

Akt Isoforms ◽

Forkhead Box ◽

Gene Ablation ◽

Deficient Mice

The purpose of the present study was to determine the effect of Akt gene ablation on Akt/Forkhead Box O (FOXO) signaling and atrogene expression. This was accomplished by studying wild-type (WT) and isoform-specific Akt knockout (Akt1−/− and Akt2−/−) mice. The ability of insulin to promote Akt phosphorylation on Ser473 was significantly lower in extensor digitorum longus (EDL) and soleus muscles from Akt1−/− and Akt2−/− mice compared with WT mice. Total Akt1 protein levels were significantly lower in EDL muscles of Akt2−/− mice compared with WT mice, a process that appears to be posttranscriptionally regulated as Akt1 mRNA levels were unchanged. The loss of Akt1 protein in EDL muscles of Akt2−/− mice does not appear to be due to insulin resistance because 4 mo of a high-fat diet failed to reduce Akt1 protein levels in muscles of WT mice. Although FOXO3a phosphorylation and atrogin-1 expression were unaltered in muscles of Akt1−/− and Akt2−/− mice, the expression of the atrogenes Bnip3 and gabarapl were significantly elevated in muscles of both Akt1 and Akt2 knockout mice. Finally, the expression of striated activator of Rho signaling was significantly increased in muscles of Akt2−/− mice compared with Akt1−/− and WT mice. Our results demonstrate that the ablation of Akt isoforms disassociates insulin action and Akt/FOXO signaling to atrogenes.

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Decreased Npas4 and Arc mRNA Levels in the Hippocampus of Aged Memory-Impaired Wild-Type But Not Memory Preserved 11β-HSD1 Deficient Mice

Journal of Neuroendocrinology ◽

10.1111/jne.12339 ◽

2016 ◽

Vol 28 (1) ◽

Cited By ~ 12

Author(s):

J. Qiu ◽

D. R. Dunbar ◽

J. Noble ◽

C. Cairns ◽

R. Carter ◽

...

Keyword(s):

Mrna Levels ◽

Wild Type ◽

Deficient Mice

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Palmitoleic acid (n-7) increases white adipocyte lipolysis and lipase content in a PPARα-dependent manner

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00082.2013 ◽

2013 ◽

Vol 305 (9) ◽

pp. E1093-E1102 ◽

Cited By ~ 43

Author(s):

Andressa Bolsoni-Lopes ◽

William T. Festuccia ◽

Talita S. M. Farias ◽

Patricia Chimin ◽

Francisco L. Torres-Leal ◽

...

Keyword(s):

Fatty Acid ◽

Palmitoleic Acid ◽

Whole Body ◽

Mrna Levels ◽

Dependent Manner ◽

Wild Type ◽

Acid Incorporation ◽

Gene And Protein Expression ◽

Fatty Acid Incorporation ◽

Deficient Mice

We investigated whether palmitoleic acid, a fatty acid that enhances whole body glucose disposal and suppresses hepatic steatosis, modulates triacylglycerol (TAG) metabolism in adipocytes. For this, both differentiated 3T3-L1 cells treated with either palmitoleic acid (16:1n7, 200 μM) or palmitic acid (16:0, 200 μM) for 24 h and primary adipocytes from wild-type or PPARα-deficient mice treated with 16:1n7 (300 mg·kg−1·day−1) or oleic acid (18:1n9, 300 mg·kg−1·day−1) by gavage for 10 days were evaluated for lipolysis, TAG, and glycerol 3-phosphate synthesis and gene and protein expression profile. Treatment of differentiated 3T3-L1 cells with 16:1n7, but not 16:0, increased basal and isoproterenol-stimulated lipolysis, mRNA levels of adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) and protein content of ATGL and pSer660-HSL. Such increase in lipolysis induced by 16:1n7, which can be prevented by pharmacological inhibition of PPARα, was associated with higher rates of PPARα binding to DNA. In contrast to lipolysis, both 16:1n7 and 16:0 increased fatty acid incorporation into TAG and glycerol 3-phosphate synthesis from glucose without affecting glyceroneogenesis and glycerokinase expression. Corroborating in vitro findings, treatment of wild-type but not PPARα-deficient mice with 16:1n7 increased primary adipocyte basal and stimulated lipolysis and ATGL and HSL mRNA levels. In contrast to lipolysis, however, 16:1n7 treatment increased fatty acid incorporation into TAG and glycerol 3-phosphate synthesis from glucose in both wild-type and PPARα-deficient mice. In conclusion, palmitoleic acid increases adipocyte lipolysis and lipases by a mechanism that requires a functional PPARα.

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Hypoxia induces B-type natriuretic peptide release in cell lines derived from human cardiomyocytes

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00250.2009 ◽

2009 ◽

Vol 297 (2) ◽

pp. H550-H555 ◽

Cited By ~ 44

Author(s):

Gregori Casals ◽

Josefa Ros ◽

Alessandro Sionis ◽

Mercy M. Davidson ◽

Manuel Morales-Ruiz ◽

...

Keyword(s):

Natriuretic Peptide ◽

Transcriptional Activity ◽

Molecular Mechanisms ◽

Ventricular Myocytes ◽

Hypoxia Inducible Factor ◽

Peptide Hormone ◽

Mrna Levels ◽

Human Cardiomyocytes ◽

Hypoxic Conditions

B-type natriuretic peptide (BNP) is a peptide hormone of myocardial origin with significant cardioprotective properties. Patients with myocardial ischemia present with high levels of BNP in plasma and elevated expression in the myocardium. However, the molecular mechanisms of BNP induction in the ischemic myocardium are not well understood. The aim of the investigation was to assess whether myocardial hypoxia induces the production of BNP in human ventricular myocytes. To test the hypothesis that reduced oxygen tension can directly stimulate BNP gene expression and release in the absence of hemodynamic or neurohormonal stimuli, we used an in vitro model system of cultured human ventricular myocytes (AC16 cells). Cells were cultured under normoxic (21% O2) or hypoxic (5% O2) conditions for up to 48 h. The accumulation of BNP, atrial natriuretic peptide (ANP), and vascular endothelial growth factor (VEGF) was then measured. Hypoxia stimulated the protein release of BNP and VEGF but not ANP. In concordance, the increased mRNA levels of BNP and VEGF but not ANP were found on culturing AC16 cells under hypoxic conditions. The analysis of the transcriptional activity of the hypoxia-inducible factor 1 (HIF-1) in nuclear extracts showed that HIF-1 activity was induced under hypoxic conditions. Finally, the treatment of AC16 cells with the HIF-1 inhibitor rotenone in hypoxia inhibited BNP and VEGF release. In conclusion, these data indicate that hypoxia induces the synthesis and secretion of BNP in human ventricular myocytes, likely through HIF-1-enhanced transcriptional activity.

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Bach1Deficiency and Accompanying Overexpression of Heme Oxygenase-1 Do Not Influence Aging or Tumorigenesis in Mice

Oxidative Medicine and Cellular Longevity ◽

10.1155/2014/757901 ◽

2014 ◽

Vol 2014 ◽

pp. 1-12 ◽

Cited By ~ 12

Author(s):

Kazushige Ota ◽

Andrey Brydun ◽

Ari Itoh-Nakadai ◽

Jiying Sun ◽

Kazuhiko Igarashi

Keyword(s):

Oxidative Stress ◽

Stress Response ◽

Heme Oxygenase ◽

Transcriptional Activity ◽

Oxidative Stress Response ◽

Acute Injury ◽

Heme Oxygenase 1 ◽

Wild Type ◽

Environmental Perturbations ◽

Deficient Mice

Oxidative stress contributes to both aging and tumorigenesis. The transcription factor Bach1, a regulator of oxidative stress response, augments oxidative stress by repressing the expression of heme oxygenase-1 (HO-1) gene (Hmox1) and suppresses oxidative stress-induced cellular senescence by restricting the p53 transcriptional activity. Here we investigated the lifelong effects ofBach1deficiency on mice.Bach1-deficient mice showed longevity similar to wild-type mice. Although HO-1 was upregulated in the cells ofBach1-deficient animals, the levels of ROS inBach1-deficient HSCs were comparable to those in wild-type cells.Bach1−/−;p53−/−mice succumbed to spontaneous cancers as frequently asp53-deficient mice.Bach1deficiency significantly altered transcriptome in the liver of the young mice, which surprisingly became similar to that of wild-type mice during the course of aging. The transcriptome adaptation toBach1deficiency may reflect how oxidative stress response is tuned upon genetic and environmental perturbations. We concluded thatBach1deficiency and accompanying overexpression of HO-1 did not influence aging or p53 deficiency-driven tumorigenesis. Our results suggest that it is useful to target Bach1 for acute injury responses without inducing any apparent deteriorative effect.

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Genetic Targeting of Relaxin and Insulin-Like Factor 3 Receptors in Mice

Endocrinology ◽

10.1210/en.2004-0515 ◽

2004 ◽

Vol 145 (10) ◽

pp. 4712-4720 ◽

Cited By ~ 97

Author(s):

Aparna A. Kamat ◽

Shu Feng ◽

Natalia V. Bogatcheva ◽

Anne Truong ◽

Colin E. Bishop ◽

...

Keyword(s):

Knockout Mice ◽

Peptide Hormone ◽

Vascular Bundles ◽

Biological Processes ◽

Wild Type ◽

Putative Receptor ◽

Collagen Accumulation ◽

Genetic Targeting ◽

Deficient Mice

Abstract Relaxin (RLN) is a small peptide hormone that affects a variety of biological processes. Rln1 knockout mice exhibit abnormal nipple development, prolonged parturition, agerelated pulmonary fibrosis, and abnormalities in the testes and prostate. We describe here RLN receptor Lgr7-deficient mice. Mutant females have grossly underdeveloped nipples and are unable to feed their progeny. Some Lgr7−/− females were unable to deliver their pups. Histological analysis of Lgr7 mutant lung tissues demonstrates increased collagen accumulation and fibrosis surrounding the bronchioles and the vascular bundles, absent in wild-type animals. However, Lgr7-deficient males do not exhibit abnormalities in the testes or prostate as seen in Rln1 knockout mice. Lgr7-deficient females with additional deletion of Lgr8 (Great), another putative receptor for RLN, are fertile and have normal-sized litters. Double mutant males have normal-sized prostate and testes, suggesting that Lgr8 does not account for differences in Rln1−/− and Lgr7−/− phenotypes. Transgenic overexpression of Insl3, the cognate ligand for Lgr8, does not rescue the mutant phenotype of Lgr7-deficient female mice indicating nonoverlapping functions of the two receptors. Our data indicate that neither Insl3 nor Lgr8 contribute to the RLN signaling pathway. We conclude that the Insl3/Lgr8 and Rln1/Lgr7 actions do not overlap in vivo.

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Nighttime Administration of Nicotine Improves Hepatic Glucose Metabolism via the Hypothalamic Orexin System in Mice

Endocrinology ◽

10.1210/en.2015-1488 ◽

2016 ◽

Vol 157 (1) ◽

pp. 195-206 ◽

Cited By ~ 10

Author(s):

Hiroshi Tsuneki ◽

Takashi Nagata ◽

Mikio Fujita ◽

Kanta Kon ◽

Naizhen Wu ◽

...

Keyword(s):

Blood Glucose ◽

Glucose Metabolism ◽

Oral Intake ◽

Mrna Levels ◽

Active Period ◽

Wild Type ◽

Glucose Levels ◽

Blood Glucose Levels ◽

Deficient Mice

Abstract Nicotine is known to affect the metabolism of glucose; however, the underlying mechanism remains unclear. Therefore, we here investigated whether nicotine promoted the central regulation of glucose metabolism, which is closely linked to the circadian system. The oral intake of nicotine in drinking water, which mainly occurred during the nighttime active period, enhanced daily hypothalamic prepro-orexin gene expression and reduced hyperglycemia in type 2 diabetic db/db mice without affecting body weight, body fat content, and serum levels of insulin. Nicotine administered at the active period appears to be responsible for the effect on blood glucose, because nighttime but not daytime injections of nicotine lowered blood glucose levels in db/db mice. The chronic oral treatment with nicotine suppressed the mRNA levels of glucose-6-phosphatase, the rate-limiting enzyme of gluconeogenesis, in the liver of db/db and wild-type control mice. In the pyruvate tolerance test to evaluate hepatic gluconeogenic activity, the oral nicotine treatment moderately suppressed glucose elevations in normal mice and mice lacking dopamine receptors, whereas this effect was abolished in orexin-deficient mice and hepatic parasympathectomized mice. Under high-fat diet conditions, the oral intake of nicotine lowered blood glucose levels at the daytime resting period in wild-type, but not orexin-deficient, mice. These results indicated that the chronic daily administration of nicotine suppressed hepatic gluconeogenesis via the hypothalamic orexin-parasympathetic nervous system. Thus, the results of the present study may provide an insight into novel chronotherapy for type 2 diabetes that targets the central cholinergic and orexinergic systems.

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