RESUMEN
Quantitative trait locus (QTL) analyses of intercross populations between widely used mouse inbred strains provide a powerful approach for uncovering genetic factors that influence susceptibility to atherosclerosis. Epistatic interactions are common in complex phenotypes and depend on genetic backgrounds. To dissect genetic architecture of atherosclerosis, we analyzed F2 progeny from a cross between apolipoprotein E-null mice on DBA/2J (DBA-apoE) and C57BL/6J (B6-apoE) genetic backgrounds and compared the results with those from two previous F2 crosses of apolipoprotein E-null mice on 129S6/SvEvTac (129-apoE) and DBA-apoE backgrounds, and B6-apoE and 129-apoE backgrounds. In these round-robin crosses, in which each parental strain was crossed with two others, large-effect QTLs are expected to be detectable at least in two crosses. On the other hand, observation of QTLs in one cross only may indicate epistasis and/or absence of statistical power. For atherosclerosis at the aortic arch, Aath4 on chromosome (Chr)2:66 cM follows the first pattern, with significant QTL peaks in (DBAx129)F2 and (B6xDBA)F2 mice but not in (B6x129)F2 mice. We conclude that genetic variants unique to DBA/2J at Aath4 confer susceptibility to atherosclerosis at the aortic arch. A similar pattern was observed for Aath5 on chr10:35 cM, verifying that the variants unique to DBA/2J at this locus protect against arch plaque development. However, multiple loci, including Aath1 (Chr1:49 cM), and Aath2 (Chr1:70 cM) follow the second type of pattern, showing significant peaks in only one of the three crosses (B6-apoE x 129-apoE). As for atherosclerosis at aortic root, the majority of QTLs, including Ath29 (Chr9:33 cM), Ath44 (Chr1:68 cM) and Ath45 (Chr2:83 cM), was also inconsistent, being significant in only one of the three crosses. Only the QTL on Chr7:37 cM was consistently suggestive in two of the three crosses. Thus QTL analysis of round-robin crosses revealed the genetic architecture of atherosclerosis.
Asunto(s)
Apolipoproteínas E/genética , Aterosclerosis/genética , Sitios de Carácter Cuantitativo , Animales , Aorta/patología , Aterosclerosis/patología , Mapeo Cromosómico , Cruzamientos Genéticos , Lípidos/sangre , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Ratones Transgénicos , Placa Aterosclerótica/patologíaRESUMEN
OBJECTIVE: Arch atherosclerosis 4 (Aath4) is a quantitative trait locus for atherosclerotic plaque formation in the inner curve of the aortic arch previously identified in an F2 cross of Apoe-/- mice on DBA/2J and 129S6 backgrounds. C-mer proto-oncogene tyrosine kinase (Mertk), coding for a ligand-activated transmembrane tyrosine kinase, is a candidate gene within the same chromosomal region. Our objective was to determine whether strain differences in Mertk influence plaque formation. APPROACH AND RESULTS: To dissect the strain effects of Mertk on atherosclerosis, we first established a congenic mouse line (Aath4aDBA/DBA ) in which a 5' region of Aath4 of DBA/2J, including Mertk, was backcrossed onto a 129S6-Apoe-/- background. The resulting Aath4aDBA/DBA male mice developed significantly larger plaques compared with control mice (Aath4a129/129 ), proving that the DBA/2J allele of Aath4a is proatherogenic. Thioglycollate-elicited peritoneal macrophages from Aath4aDBA/DBA mice express less than 50% of Mertk mRNA and cell-surface MERTK protein compared with those from the control mice. Moreover, both large and small peritoneal Aath4aDBA/DBA macrophages showed reduced phagocytosis of apoptotic cells. When Mertk cDNAs from 129S6 and DBA/2J mice were overexpressed in HEK293T (human embryonic kidney 293T) cells, phagocytosis of apoptotic cells was equally enhanced in direct proportion to Mertk levels, indicating that phagocytosis is modulated by the amount of MERTK, but that it is not affected by MERTK amino acid differences between 129S6 and DBA/2J. CONCLUSIONS: Reduced transcription of Mertk, rather than differences in MERTK protein structure, determines the reduced efficiency of apoptotic cell clearance in the Aath4aDBA/DBA mice, which, in turn, contributes to their increased susceptibility to atherosclerosis.
Asunto(s)
Aorta Torácica/enzimología , Enfermedades de la Aorta/genética , Aterosclerosis/genética , Cromosomas de los Mamíferos , Haplotipos , Macrófagos Peritoneales/enzimología , Fagocitosis , Placa Aterosclerótica , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Tirosina Quinasas Receptoras/metabolismo , Animales , Aorta Torácica/patología , Enfermedades de la Aorta/enzimología , Enfermedades de la Aorta/patología , Apolipoproteínas E/deficiencia , Apolipoproteínas E/genética , Apoptosis , Aterosclerosis/enzimología , Aterosclerosis/patología , Modelos Animales de Enfermedad , Regulación hacia Abajo , Regulación Enzimológica de la Expresión Génica , Predisposición Genética a la Enfermedad , Células HEK293 , Humanos , Células Jurkat , Macrófagos Peritoneales/patología , Ratones de la Cepa 129 , Ratones Congénicos , Ratones Endogámicos DBA , Ratones Noqueados , Fenotipo , Regiones Promotoras Genéticas , Proto-Oncogenes Mas , Proteínas Proto-Oncogénicas/deficiencia , Proteínas Proto-Oncogénicas/genética , Proteínas Tirosina Quinasas Receptoras/deficiencia , Proteínas Tirosina Quinasas Receptoras/genética , Factores de Riesgo , Especificidad de la Especie , Transcripción Genética , Transfección , Calcificación Vascular/enzimología , Calcificación Vascular/genética , Calcificación Vascular/patología , Tirosina Quinasa c-MerRESUMEN
The genetic background of apolipoprotein E (apoE) deficient mice influences atherosclerotic plaque development. We previously reported three quantitative trait loci (QTL), Aath1-Aath3, that affect aortic arch atherosclerosis independently of those in the aortic root in a cross between C57BL6 apoEKO mice (B6-apoE) and 129S6 apoEKO mice (129-apoE). To gain further insight into genetic factors that influence atherosclerosis at different vascular locations, we analyzed 335 F2 mice from an intercross between 129-apoE and apoEKO mice on a DBA/2J genetic background (DBA-apoE). The extent of atherosclerosis in the aortic arch was very similar in the two parental strains. Nevertheless, a genome-wide scan identified two significant QTL for plaque size in the aortic arch: Aath4 on Chromosome (Chr) 2 at 137 Mb and Aath5 on Chr 10 at 51 Mb. The DBA alleles of Aath4 and Aath5 respectively confer susceptibility and resistance to aortic arch atherosclerosis over 129 alleles. Both QTL are also independent of those affecting plaque size at the aortic root. Genome analysis suggests that athero-susceptibility of Aath4 in DBA may be contributed by multiple genes, including Mertk and Cd93, that play roles in phagocytosis of apoptotic cells and modulate inflammation. A candidate gene for Aath5 is Stab2, the DBA allele of which is associated with 10 times higher plasma hyaluronan than the 129 allele. Overall, our identification of two new QTL that affect atherosclerosis in an aortic arch-specific manner further supports the involvement of distinct pathological processes at different vascular locations.
Asunto(s)
Aorta Torácica/patología , Enfermedades de la Aorta/genética , Apolipoproteínas E/genética , Aterosclerosis/genética , Sitios de Carácter Cuantitativo , Alelos , Animales , Enfermedades de la Aorta/patología , Aterosclerosis/patología , Cruzamientos Genéticos , Predisposición Genética a la Enfermedad , Haplotipos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Ratones Noqueados , FenotipoRESUMEN
Aldosterone-independent mechanisms may contribute to K(+) homeostasis. We studied aldosterone synthase knockout (AS(-/-)) mice to define renal control mechanisms of K(+) homeostasis in complete aldosterone deficiency. AS(-/-) mice were normokalemic and tolerated a physiologic dietary K(+) load (2% K(+), 2 days) without signs of illness, except some degree of polyuria. With supraphysiologic K(+) intake (5% K(+)), AS(-/-) mice decompensated and became hyperkalemic. High-K(+) diets induced upregulation of the renal outer medullary K(+) channel in AS(-/-) mice, whereas upregulation of the epithelial sodium channel (ENaC) sufficient to increase the electrochemical driving force for K(+) excretion was detected only with a 2% K(+) diet. Phosphorylation of the thiazide-sensitive NaCl cotransporter was consistently lower in AS(-/-) mice than in AS(+/+) mice and was downregulated in mice of both genotypes in response to increased K(+) intake. Inhibition of the angiotensin II type 1 receptor reduced renal creatinine clearance and apical ENaC localization, and caused severe hyperkalemia in AS(-/-) mice. In contrast with the kidney, the distal colon of AS(-/-) mice did not respond to dietary K(+) loading, as indicated by Ussing-type chamber experiments. Thus, renal adaptation to a physiologic, but not supraphysiologic, K(+) load can be achieved in aldosterone deficiency by aldosterone-independent activation of the renal outer medullary K(+) channel and ENaC, to which angiotensin II may contribute. Enhanced urinary flow and reduced activity of the thiazide-sensitive NaCl cotransporter may support renal adaptation by activation of flow-dependent K(+) secretion and increased intratubular availability of Na(+) that can be reabsorbed in exchange for K(+) secreted.
Asunto(s)
Citocromo P-450 CYP11B2/deficiencia , Homeostasis/fisiología , Hipoaldosteronismo/metabolismo , Riñón/metabolismo , Potasio/metabolismo , Angiotensina II/metabolismo , Animales , Citocromo P-450 CYP11B2/genética , Citocromo P-450 CYP11B2/metabolismo , Modelos Animales de Enfermedad , Canales Epiteliales de Sodio/efectos de los fármacos , Canales Epiteliales de Sodio/metabolismo , Hipoaldosteronismo/fisiopatología , Masculino , Ratones , Ratones Noqueados , Canales de Potasio/efectos de los fármacos , Canales de Potasio/metabolismo , Potasio en la Dieta/farmacologíaRESUMEN
The prostanoid thromboxane A2 has been implicated to contribute to the pathogenesis of many cardiovascular diseases, including hypertension. To study the role of vascular thromboxane-prostanoid (TP) receptors in blood pressure regulation, we generated mice with cell-specific deletion of TP receptors in smooth muscle using Cre/Loxp technology. We crossed the KISM22α-Cre transgenic mouse line expressing Cre recombinase in smooth muscle cells with a mouse line bearing a conditional allele of the Tbxa2r gene (Tp(flox)). In KISM22α-Cre(+)Tp(flox/flox) (TP-SMKO) mice, TP receptors were efficiently deleted from vascular smooth muscle cells. In TP-SMKOs, acute vasoconstrictor responses to the TP agonist U46619 were attenuated to a similar extent in both the peripheral and renal circulations. Yet, acute vascular responses to angiotensin II were unaffected at baseline and after chronic angiotensin II administration. Infusion of high-dose U46619 caused circulatory collapse and death in a majority of control mice but had negligible hemodynamic effects in TP-SMKOs, which were completely protected from U46619-induced sudden death. Baseline blood pressures were normal in TP-SMKOs. However, the absence of TP receptors in vascular smooth muscle cells was associated with significant attenuation of angiotensin II-induced hypertension and diminished vascular remodeling. This was also associated with reduced urinary thromboxane production after chronic angiotensin II. Thus, TP receptors in vascular smooth muscle cells play a major role in mediating the actions of thromboxane A(2) in TP agonist-induced shock, hypertension, and vascular remodeling of the aorta.
Asunto(s)
Aorta/metabolismo , Hipertensión/genética , Músculo Liso Vascular/metabolismo , Miocitos del Músculo Liso/metabolismo , Receptores de Tromboxanos/genética , Ácido 15-Hidroxi-11 alfa,9 alfa-(epoximetano)prosta-5,13-dienoico/farmacología , Animales , Aorta/efectos de los fármacos , Presión Sanguínea/efectos de los fármacos , Muerte Súbita , Hipertensión/metabolismo , Ratones , Ratones Noqueados , Músculo Liso Vascular/efectos de los fármacos , Miocitos del Músculo Liso/efectos de los fármacos , Receptores de Tromboxanos/metabolismo , Tromboxano A2/farmacología , Vasoconstricción/efectos de los fármacos , Vasoconstrictores/farmacologíaRESUMEN
To characterize gene expression networks linked to AT(1) angiotensin receptors in the kidney, we carried out genome-wide transcriptional analysis of RNA from kidneys of wild-type (WT) and AT(1A) receptor-deficient mice (KOs) at baseline and after 2 days of angiotensin II infusion (1,000 ng·kg(-1)·min(-1)). At baseline, 405 genes were differentially expressed (>1.5×) between WT and KO kidneys. Of these, >80% were upregulated in the KO group including genes involved in inflammation, oxidative stress, and cell proliferation. After 2 days of angiotensin II infusion in WT mice, expression of ≈805 genes was altered (18% upregulated, 82% repressed). Genes in metabolism and ion transport pathways were upregulated while there was attenuated expression of genes protective against oxidative stress including glutathione synthetase and mitochondrial superoxide dismutase 2. Angiotensin II infusion had little effect on blood pressure in KOs. Nonetheless, expression of >250 genes was altered in kidneys from KO mice during angiotensin II infusion; 14% were upregulated, while 86% were repressed including genes involved in immune responses, angiogenesis, and glutathione metabolism. Between WT and KO kidneys during angiotensin II infusion, 728 genes were differentially expressed; 10% were increased and 90% were decreased in the WT group. Differentially regulated pathways included those involved in ion transport, immune responses, metabolism, apoptosis, cell proliferation, and oxidative stress. This genome-wide assessment should facilitate identification of critical distal pathways linked to blood pressure regulation.
Asunto(s)
Perfilación de la Expresión Génica , Riñón/metabolismo , Receptor de Angiotensina Tipo 1/genética , Angiotensina II/administración & dosificación , Animales , Análisis por Conglomerados , Femenino , Expresión Génica/efectos de los fármacos , Redes Reguladoras de Genes , Hipertensión/genética , Hipertensión/fisiopatología , Proteínas Inmediatas-Precoces/genética , Proteínas Inmediatas-Precoces/metabolismo , Immunoblotting , Infusiones Subcutáneas , Masculino , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Análisis de Secuencia por Matrices de Oligonucleótidos , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Receptor de Angiotensina Tipo 1/deficiencia , Reproducibilidad de los Resultados , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Vasoconstrictores/administración & dosificaciónRESUMEN
To test the hypothesis that angiotensin (Ang) II induces profibrotic gene expression through endogenous aldosterone, we measured the effect of 4 h infusion (600 ng/kg x min) of Ang II on tissue mRNA expression of plasminogen activator inhibitor 1 (PAI-1), preproendothelin-1 (ppET-1), TGF-beta, and osteopontin in wild-type (WT), aldosterone synthase-deficient (AS(-/-)), and AS(-/-) mice treated with aldosterone (either 500 ng/d for 7 d or 250 ng as a concurrent 4 h infusion). Ang II increased aldosterone in WT (P < 0.001) but not in AS(-/-) mice. Aldosterone (7 d) normalized basal aldosterone concentrations in AS(-/-) mice; however, there was no further effect of Ang II on aldosterone (P = NS). Basal cardiac and aortic PAI-1 and ppET-1 expression were similar in WT and AS(-/-) mice. Ang II-stimulated PAI-1 (P < 0.001) and ppET-1 expression (P = 0.01) was diminished in the heart of AS(-/-) mice; treatment with aldosterone for 4 h or 7 d restored PAI-1 and ppET-1 mRNA responsiveness to Ang II in the heart. Ang II increased PAI-1 (P = 0.01) expression in the aorta of AS(-/-) as well as WT mice. In the kidney, basal PAI-1, ppET-1, and TGF-beta mRNA expression was increased in AS(-/-) compared with WT mice and correlated with plasma renin activity. Ang II did not stimulate osteopontin or TGF-beta expression in the heart or kidney. Endogenous aldosterone contributes to the acute stimulatory effect of Ang II on PAI-1 and ppET-1 mRNA expression in the heart; renin activity correlates with basal profibrotic gene expression in the kidney.
Asunto(s)
Aldosterona/fisiología , Angiotensina II/farmacología , Aorta/efectos de los fármacos , Endotelina-1/genética , Corazón/efectos de los fármacos , Inhibidor 1 de Activador Plasminogénico/genética , Aldosterona/sangre , Aldosterona/farmacología , Animales , Aorta/metabolismo , Presión Sanguínea/efectos de los fármacos , Citocromo P-450 CYP11B2/genética , Interacciones Farmacológicas , Endotelina-1/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Miocardio/metabolismo , Inhibidor 1 de Activador Plasminogénico/metabolismo , Receptor de Angiotensina Tipo 1/metabolismo , Factores de Tiempo , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Actions of the lipid mediator thromboxane (Tx) A2 acting through the TP receptor contribute to the pathogenesis of cardiovascular disease. To further explore the role of TxA2 in hypertension, we examined the consequences of deficiency of the TP receptor on the course of hypertension associated with endothelial dysfunction and salt sensitivity. To this end, the nitric oxide synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME) was administered to TP-deficient (Tp-/-) and wild-type (Tp+/+) control mice in drinking water for 21 wk along with a high-salt (HS; 6% NaCl) diet. Administration of L-NAME increased urinary excretion of TxB2 to a similar extent in both Tp+/+ and Tp-/- animals. L-NAME also caused significant and sustained elevations in blood pressure that reached a maximum between weeks 3 and 6. However, the severity of hypertension was attenuated in the Tp-/- mice throughout the study period (P<0.001). At the end of the study, the wild-type mice developed significant cardiac hypertrophy (23.6+/-2% increase in heart-to-body weight ratio). The severity of cardiac hypertrophy was attenuated in the TP-deficient group (11.1+/-2.6%; P<0.05). In contrast, kidney hypertrophy was exaggerated in the Tp-/- mice compared with controls (37.1+/-5.4 vs. 12.3+/-2.3%; P<0.01). Moreover, the severity of glomerulosclerosis, tubule vacuolization, and interstitial chronic inflammation was also enhanced in the Tp-/- group (P<0.01). Thus, in L-NAME hypertension, TP receptors contribute to elevated blood pressure and cardiac hypertrophy. In this model, TP receptors also provided unexpected protection against kidney injury.
Asunto(s)
Inhibidores Enzimáticos/toxicidad , Hipertensión Renal/inducido químicamente , Hipertensión Renal/metabolismo , NG-Nitroarginina Metil Éster/toxicidad , Receptores de Tromboxano A2 y Prostaglandina H2/metabolismo , Albuminuria/inducido químicamente , Albuminuria/complicaciones , Albuminuria/metabolismo , Animales , Presión Sanguínea/fisiología , Cardiomegalia/complicaciones , Modelos Animales de Enfermedad , Ingestión de Líquidos , Ingestión de Alimentos , Hipertensión Renal/complicaciones , Isoprostanos/orina , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Óxido Nítrico/antagonistas & inhibidores , Óxido Nítrico/metabolismo , Sistema Renina-Angiotensina/fisiología , Sodio en la Dieta/farmacología , Tromboxano B2/orinaRESUMEN
Activation of the renin-angiotensin system contributes to the progression of chronic kidney disease. Based on the known cellular effects of ANG II to promote inflammation, we posited that stimulation of lymphocyte responses by ANG II might contribute to the pathogenesis of hypertensive kidney injury. We therefore examined the effects of the immunosuppressive agent mycophenolate mofetil (MMF) on the course of hypertension and kidney disease induced by chronic infusion of ANG II in 129/SvEv mice. Although it had no effect on the severity of hypertension or cardiac hypertrophy, treatment with MMF significantly reduced albuminuria and ameliorated kidney injury, decreasing glomerulosclerosis and reducing lymphocyte infiltration into the renal interstitium. Attenuation of renal pathology with MMF was associated with reduced expression of mRNAs for the proinflammatory cytokines interferon-gamma and tumor necrosis factor-alpha and the profibrotic cytokine transforming growth factor-beta. As infiltration of the kidney by T lymphocytes was a prominent feature of ANG II-dependent renal injury, we carried out experiments examining the effects of ANG II on lymphocytes in vitro. We find that exposure of splenic lymphocytes to ANG II causes prominent rearrangements of the actin cytoskeleton. These actions require the activity of Rho kinase. Thus, ANG II exaggerates hypertensive kidney injury by stimulating lymphocyte responses. These proinflammatory actions of ANG II seem to have a proclivity for inducing kidney injury while having negligible actions in the pathogenesis of cardiac hypertrophy.
Asunto(s)
Angiotensina II/efectos adversos , Angiotensina II/farmacología , Hipertensión/complicaciones , Enfermedades Renales/etiología , Linfocitos T/efectos de los fármacos , Vasoconstrictores/farmacología , Albuminuria/tratamiento farmacológico , Albuminuria/prevención & control , Animales , Cardiomegalia/etiología , Proliferación Celular/efectos de los fármacos , Citoesqueleto/efectos de los fármacos , Modelos Animales de Enfermedad , Inmunosupresores/farmacología , Inmunosupresores/uso terapéutico , Interferón gamma/metabolismo , Enfermedades Renales/metabolismo , Enfermedades Renales/patología , Masculino , Ratones , Ratones Noqueados , Ácido Micofenólico/análogos & derivados , Ácido Micofenólico/farmacología , Ácido Micofenólico/uso terapéutico , Cloruro de Sodio Dietético/farmacología , Linfocitos T/patología , Factor de Crecimiento Transformador beta/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Vasoconstrictores/efectos adversosRESUMEN
To study the effects of modestly increased expression of aldosterone synthase (AS), we generated mice (AS(hi/hi)) by replacing the 3' untranslated region of AS mRNA with that from a stable mRNA. AS(hi/hi) mice on a normal-salt diet had 1.5 times the wild-type AS mRNA in adrenals, although their blood pressure and plasma aldosterone did not differ from wild-type mice. Changes in dietary salt did not affect the blood pressure of wild-type mice, but AS(hi/hi) mice had approximately 10-mm Hg higher blood pressure on a high-salt diet than on a low-salt diet and than wild-type mice on either diet. The AS(hi/hi) mice on a high-salt diet also had higher plasma aldosterone, lower plasma potassium, and greater renal expression of the alpha subunit of epithelial sodium channel compared with wild-type mice. The AS(hi/hi) mice on a high-salt diet also had more water intake and urine volume and less urine osmolality than wild-type mice. On a low-salt diet, AS(hi/hi) mice maintained normal blood pressure with less activation of the renin-angiotensin-aldosterone system than wild-type mice. The AS(hi/hi) mice also had less water intake and urine volume and higher urine osmolality than wild-type mice. On a medium high-salt diet, AS(hi/hi) mice were more susceptible than wild-type mice to infusion of angiotensin II, having a higher blood pressure, greater cardiac hypertrophy, and increased oxidative stress. Thus, a modest increase in AS expression makes blood pressure more sensitive to salt, suggesting that genetically increased AS expression in humans may contribute to hypertension and cardiovascular complications in societies with high-salt diets.
Asunto(s)
Presión Sanguínea/efectos de los fármacos , Citocromo P-450 CYP11B2/metabolismo , Hiperaldosteronismo/complicaciones , Hipertensión/enzimología , Sodio en la Dieta/farmacología , Aldosterona/metabolismo , Angiotensina II/farmacología , Animales , Presión Sanguínea/fisiología , Citocromo P-450 CYP11B2/genética , Femenino , Hiperaldosteronismo/genética , Hiperaldosteronismo/metabolismo , Hipertensión/etiología , Masculino , Ratones , Ratones Mutantes , Óxido Nítrico/metabolismo , Estrés Oxidativo/efectos de los fármacos , ARN Mensajero/metabolismo , Renina/metabolismo , Sistema Renina-Angiotensina/efectos de los fármacos , Sistema Renina-Angiotensina/fisiología , Sodio/metabolismo , Vasoconstrictores/farmacologíaRESUMEN
We have determined that differences in expression of aldosterone synthase (AS) affect responses to a low-salt diet. In AS-null mice (AS(-/-)), but not in wild-type, low salt significantly decreased plasma sodium and increased potassium. The increased urine volume (1.5xwild-type) and decreased urine osmolality (0.7xwild-type), present in AS(-/-) mice on normal salt, became more severe (2.3xwild-type and 0.5xwild-type) on low salt, but neither changed in wild-type. In both genotypes, plasma vasopressin was similar on normal and low salt, and desmopressin injection significantly increased urine osmolality. Renal mRNA levels for aquaporin 1 and 3 were unchanged by genotype or diet and epithelial sodium channel and Na(+)-K(+)-2Cl(-)-cotransporter by genotype. In AS(-/-) mice, aquaporin 2 mRNA increased on normal salt, whereas Na(+)Cl(-)-cotransporter and cortex K(+) channel mRNAs decreased on both diets. The low blood pressure of AS(-/-) mice was decreased further by low salt, despite additional increases in renin, intrarenal arterial wall thickness, and macula densa cyclogenase-2 mRNA. In AS(+/-) mice on normal salt, adrenal AS mRNA was slightly decreased (0.7xwild-type), but blood pressure was normal. On low salt, their blood pressure was less than wild-type (101+/-2 mm Hg versus 106+/-2 mm Hg), even though renin mRNA increased to 2xwild-type. We conclude that aldosterone is critical for urine concentration and maintenance of blood pressure and even a mild reduction of AS expression makes blood pressure sensitive to low salt, suggesting that genetic differences of AS levels in humans may influence how blood pressure responds to dietary salt.
Asunto(s)
Aldosterona/genética , Presión Sanguínea/genética , Citocromo P-450 CYP11B2/genética , Homeostasis/genética , Glándulas Suprarrenales/fisiología , Animales , Fármacos Antidiuréticos/administración & dosificación , Desamino Arginina Vasopresina/administración & dosificación , Dieta Hiposódica , Expresión Génica , Riñón/fisiología , Ratones , Orina/fisiología , Vasopresinas/agonistas , Vasopresinas/sangre , Vasopresinas/fisiologíaRESUMEN
To explore the effects of decreased amounts or absence of aldosterone, we have disrupted the gene coding for aldosterone synthase (AS) in mice and investigated blood pressure and kidney function in AS+/+, AS+/-, and AS-/- mice. AS+/- mice have normal blood pressures and show no abnormalities in electrolytes or kidney gene expression, but they have significantly higher than normal urine volume and lower urine osmolality. In contrast, the AS-/- mice have low blood pressure, abnormal electrolyte homeostasis (increased plasma concentrations of K+, Ca2+, and Mg2+ and decreased concentrations of HCO3(-) and Cl- but no difference in the plasma Na+ level), and disturbances in water metabolism (higher urine output, decreased urine osmolality, and impaired urine concentrating and diluting ability). Absence of aldosterone in the AS-/- mice induced several compensatory changes: an increased food intake-to-body weight ratio, an elevated plasma concentration of glucocorticoids, and strong activation of the renin-angiotensin system. Parallel with the markedly increased synthesis and release of renin, the AS-/- mice showed increased expression of cyclooxygenase-2 (COX-2) in macula densa. On salt supplementation, plasma electrolyte concentrations and kidney renin and COX-2 levels became similar to those of wild-type mice, but the lower blood pressure of the AS-/- mice was not corrected. Thus absence of aldosterone in AS-/- mice results in impairment of Na+ reabsorption in the distal nephron, decreased blood pressure, and strong renin-angiotensin system activation. Our data show the substantial correction of these abnormalities, except the low blood pressure, by high dietary salt does not depend on aldosterone.
Asunto(s)
Aldosterona/metabolismo , Hipotensión/fisiopatología , Riñón/fisiología , Equilibrio Ácido-Base , Animales , Presión Sanguínea , Peso Corporal , Citocromo P-450 CYP11B2/genética , Citocromo P-450 CYP11B2/metabolismo , Electrólitos/sangre , Electrólitos/orina , Homeostasis , Riñón/patología , Riñón/fisiopatología , Ratones , Ratones Mutantes , Ratones Transgénicos , Tamaño de los Órganos , Renina/metabolismoRESUMEN
To study the effects of decreased amounts or absence of aldosterone on development and endocrine function, we have disrupted the mouse gene, Cyp11b2, coding for aldosterone synthase (AS) by replacing its first two exons with sequences coding for enhanced green fluorescent protein. The null pups fail to thrive postnatally, and about 30% die between d 7 and 28. Aldosterone in plasma and AS mRNA in adrenal glands are undetectable in the null mice. Adult AS-null mice are small, weigh 75% of wild type, are hypotensive, have increased concentrations of plasma K(+) and corticosterone, and a decreased concentration of plasma Cl(-). Their plasma renin and angiotensin II concentrations are 45x and 4x wild type. The adrenal cortex is disorganized and has cells that contain marked accumulations of lipid. The zona glomerulosa is widened and includes easily detectable renin-containing cells, not seen in the wild-type adrenal gland. In the AS-/- adrenals, the level of mRNA for Cyp11b1, coding for 11beta-hydroxylase, is 150% wild type. The adrenal glands of the null mice consequently show evidence of a greatly activated renin-angiotensin system and up-regulation of glucocorticoid production. In the AS-null mice enhanced green fluorescent protein fluorescence is mainly at the boundary between the cortex and medulla, where apoptotic cells are numerous. These data are consistent with the absence of aldosterone in the AS-null mice inducing an increased cell-turnover of cells in the adrenals that normally become AS expressing and their migration to the medullary boundary where they apoptose.
Asunto(s)
Corteza Suprarrenal/fisiopatología , Aldosterona/metabolismo , Homeostasis/fisiología , Hipotensión/fisiopatología , Corteza Suprarrenal/metabolismo , Corteza Suprarrenal/patología , Animales , Peso Corporal , Cloruros/sangre , Corticosterona/sangre , Citocromo P-450 CYP11B2/genética , Citocromo P-450 CYP11B2/metabolismo , Expresión Génica , Hipotensión/sangre , Hipotensión/patología , Ratones , Ratones Endogámicos , Ratones Mutantes , Tamaño de los Órganos , Fenotipo , Potasio/sangre , Renina/metabolismoRESUMEN
The urethrogenital (UG) reflex is a spinal sexual reflex that consists of autonomic and somatic nerve activity and vaginal, uterine, and anal sphincter contractions. The UG reflex is under tonic descending inhibition by neurons in the region of the nucleus paragigantocellularis (nPGi). The location of spinal neurons activated by the UG reflex was examined in the female rat using the immediate early gene, c-fos. In addition, the descending inputs from the nPGi onto fos-activated neurons was examined using the anterograde tracer biotin dextran amine injected into the nPGi. The UG reflex resulted in a significant increase in fos-positive nuclei in segments T12-S1, compared with experimental controls in which the UG reflex was not activated. Spinal circuits involved in the UG reflex include neurons relaying afferent information from the pudendal sensory nerve, in the dorsal horn and medial cord of L5-S1. Efferent output includes preganglionic neurons located in the lateral gray of L5-S1 and lateral and medial gray of T13-L2. Spinal interneurons involved in the UG reflex were found close to the preganglionic neurons and in the dorsal horn and intermediate and medial gray of T12-S1. NPGi inputs were found primarily on the autonomic efferents and interneurons in the medial and intermediate gray. These studies demonstrate multisegmental spinal circuits activated with the UG reflex and demonstrate that the descending inhibition from the nPGi is by means of preganglionic and somatic efferents and spinal interneurons closely associated with the efferent output.