Browsing by Person "Brownstein, D. G."

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Glucocorticoid receptor haploinsufficiency causes hypertension and attenuates hypothalamic-pituitary-adrenal axis and blood pressure adaptions to high-fat diet
(2008-11) Michailidou, Z.; Carter, R. N.; Marshall, E.; Sutherland, H. G.; Brownstein, D. G.; Owen, E.; Cockett, K.; Kelly, V.; Ramage, L.; Al-Dujaili, Emad A. S.; Ross, M.; Maraki, I.; Newton, K.; Holmes, M. C.; Seckl, J.; Morton, N. M.; Kenyon, C. J.; Chapman, K. E.
Glucocorticoid hormones are critical to respond and adapt to stress. Genetic variations in the glucocorticoid receptor (GR) gene alter hypothalamic-pituitary-adrenal (HPA) axis activity and associate with hypertension and susceptibility to metabolic disease. Here we test the hypothesis that reduced GR density alters blood pressure and glucose and lipid homeostasis and limits adaption to obesogenic diet. Heterozygous GR _geo/+ mice were generated from embryonic stem (ES) cells with a gene trap integration of a _-galactosidase-neomycin phosphotransferase (_geo) cassette into the GR gene creating a transcriptionally inactive GR fusion protein. Although GR_geo/+ mice have 50% less functional GR, they have normal lipid and glucose homeostasis due to compensatory HPA axis activation but are hypertensive due to activation of the renin-angiotensin- aldosterone system (RAAS). When challenged with a high-fat diet, weight gain, adiposity, and glucose intolerance were similarly increased in control and GR_geo/+ mice, suggesting preserved control of intermediary metabolism and energy balance. However, whereas a high-fat diet caused HPA activation and increased blood pressure in control mice, these adaptions were attenuated or abolished in GR_geo/+ mice. Thus, reduced GR density balanced by HPA activation leaves glucocorticoid functions unaffected but mineralocorticoid functions increased, causing hypertension. Importantly, reduced GR limits HPA and blood pressure adaptions to obesogenic diet. FASEB.
Targeting Cyp11b1 expression in mice to model sequelae of congenital adrenal hyperplasia
(2008) Mullins, L. J.; Peter, A.; Wrobel, N.; Al-Dujaili, Emad A. S.; McNeilly, J. R.; Brownstein, D. G.; McNeilly, J. R.; Mullins, J. J.; Kenyon, C. J.
We have created transgenic mice in which Cyp11b1, the gene encoding 11_-hydroxylase, has been knocked out. Since mice do not secrete adrenal androgens, this knockout line allows a more detailed investigation of phenotypes associated with congenital adrenal hyperplasia (CAH) without the overwhelming virilisation that characterises patients with CAH. Starting with a BAC containing the mouse Cyp11b1/b2 locus and including flanking up- and downstream sequences, a construct was engineered in which exons 3-7 of Cyp11b1 were substituted with DNA encoding the fluorescent reporter protein ECFP. An IRES site preceding and a farnesylation signal following the ECFP gene were added to the construct which was then homologously recombined in ES cells before injection into blastocysts. Successful targeting of the Cyp11b1 gene was confirmed in ES cells by FISH analysis and in homozygous transgenic mice by the absence of adrenal Cyp11b1 mRNA (RT-PCR) and 11beta-hydroxylase (immunocytochemistry) expression. The expected increase in adrenal mass (3-fold, P<0.001) was observed with changes due to cell hypertrophy rather than hyperplasia. Urinary steroid profiles showed marked reductions in corticosterone (9.5-fold, P<0.001) and concomitant increases of earlier intermediates in the glucocorticoid biosynthetic pathway (deoxycorticosterone 25-fold, P<0.001; progesterone 3-fold, P<0.001). Urinary DHEA and testosterone were higher in both males and females although not to the extent seen in patients with CAH. Estradiol in females was unaffected. Several unexpected phenotypes relating to reproduction were observed. In initial crosses of heterozgotes, offspring carrying the Cyp11b1 null allele were underrepresented. Also female homozygote mice were infertile with poorly defined corpora lutea, endometrial hyperplasia, and late-onset adenomyosis. We conclude that Cyp11b1 null mice exhibit signs of glucocorticoid deficiency, mineralocorticoid and progesterone excess and mild hyperandrogenism. Infertility which is known to be a problem of CAH patients, appears to be caused by abnormalities in uterine and ovarian tissues.