RESUMEN

The present experiments in mice were performed to determine the steady-state effects of exogenous adenosine on the vascular resistance of the whole kidney, of superficial blood vessels, and of afferent arterioles. The steady-state effect of an intravenous infusion of adenosine (5, 10, and 20 microg/min) in wild-type mice was vasodilatation as evidenced by significant reductions of renal and superficial vascular resistance. Resistance decreases were augmented in adenosine 1 receptor (A1AR) -/- mice. Renal vasodilatation by the A2aAR agonist CGS 21680A [2-p-(2-carboxyethyl)phenethyl-amino-5'-N-ethylcarboxamido-adenosine hydrochloride] (0.25, 0.5, and 1 microg/kg/min) and inhibition of adenosine-induced relaxation by the A2aAR antagonist ZM-241385 [4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-yl-amino]ethyl)phenol] (20 mg/kg) suggests that the reduction of renovascular resistance was largely mediated by A2aAR. After treatment with Nomega-nitro-L-arginine methyl ester (L-NAME) adenosine was unable to alter superficial blood flow and resistance significantly indicating that adenosine-induced dilatation is NO-dependent. Absence of a dilatory effect in endothelial nitric-oxide synthase (NOS) -/- mice suggests endothelial NOS as the source of NO. When infused into the subcapsular interstitium, adenosine reduced superficial blood flow through A1AR activation. Adenosine (10(-7) M) constricted isolated perfused afferent arterioles when added to the bath but not when added to the luminal perfusate. Luminal adenosine caused vasoconstriction in the presence of L-NAME or the A2AR antagonist 3,7-dimethyl-1-(2-propynyl)xanthine. Our data show that global elevation of renal adenosine causes steady-state vasorelaxation resulting from adenosine 2 receptor (A2AR)-mediated generation of NO. In contrast, selective augmentation of adenosine around afferent arterioles causes persistent vasoconstriction, indicating A1AR dominance. Thus, adenosine is a renal constrictor only when it can interact with afferent arteriolar A1AR without affecting the bulk of renal A2AR at the same time.

Asunto(s)

Adenosina/farmacología , Riñón/efectos de los fármacos , Receptores Purinérgicos P1/fisiología , Vasoconstrictores/farmacología , Vasodilatadores/farmacología , Adenosina/análogos & derivados , Animales , Arteriolas/efectos de los fármacos , Arteriolas/fisiología , Riñón/irrigación sanguínea , Riñón/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos , Ratones Noqueados

RESUMEN

CONTEXT: Incidence of end-stage renal disease due to hypertension has increased in recent decades, but the optimal strategy for treatment of hypertension to prevent renal failure is unknown, especially among African Americans. OBJECTIVE: To compare the effects of an angiotensin-converting enzyme (ACE) inhibitor (ramipril), a dihydropyridine calcium channel blocker (amlodipine), and a beta-blocker (metoprolol) on hypertensive renal disease progression. DESIGN, SETTING, AND PARTICIPANTS: Interim analysis of a randomized, double-blind, 3 x 2 factorial trial conducted in 1094 African Americans aged 18 to 70 years with hypertensive renal disease (glomerular filtration rate [GFR] of 20-65 mL/min per 1.73 m(2)) enrolled between February 1995 and September 1998. This report compares the ramipril and amlodipine groups following discontinuation of the amlodipine intervention in September 2000. INTERVENTIONS: Participants were randomly assigned to receive amlodipine, 5 to 10 mg/d (n = 217), ramipril, 2.5 to 10 mg/d (n = 436), or metoprolol, 50 to 200 mg/d (n = 441), with other agents added to achieve 1 of 2 blood pressure goals. MAIN OUTCOME MEASURES: The primary outcome measure was the rate of change in GFR; the main secondary outcome was a composite index of the clinical end points of reduction in GFR of more than 50% or 25 mL/min per 1.73 m(2), end-stage renal disease, or death. RESULTS: Among participants with a urinary protein to creatinine ratio of >0.22 (corresponding approximately to proteinuria of more than 300 mg/d), the ramipril group had a 36% (2.02 [SE, 0.74] mL/min per 1.73 m(2)/y) slower mean decline in GFR over 3 years (P =.006) and a 48% reduced risk of the clinical end points vs the amlodipine group (95% confidence interval [CI], 20%-66%). In the entire cohort, there was no significant difference in mean GFR decline from baseline to 3 years between treatment groups (P =.38). However, compared with the amlodipine group, after adjustment for baseline covariates the ramipril group had a 38% reduced risk of clinical end points (95% CI, 13%-56%), a 36% slower mean decline in GFR after 3 months (P =.002), and less proteinuria (P<.001). CONCLUSION: Ramipril, compared with amlodipine, retards renal disease progression in patients with hypertensive renal disease and proteinuria and may offer benefit to patients without proteinuria.

Asunto(s)

Amlodipino/uso terapéutico , Inhibidores de la Enzima Convertidora de Angiotensina/uso terapéutico , Antihipertensivos/uso terapéutico , Bloqueadores de los Canales de Calcio/uso terapéutico , Hipertensión/complicaciones , Hipertensión/tratamiento farmacológico , Fallo Renal Crónico/prevención & control , Nefroesclerosis/complicaciones , Nefroesclerosis/tratamiento farmacológico , Ramipril/uso terapéutico , Antagonistas Adrenérgicos beta/uso terapéutico , Adulto , Negro o Afroamericano , Anciano , Método Doble Ciego , Femenino , Tasa de Filtración Glomerular , Humanos , Fallo Renal Crónico/etiología , Masculino , Metoprolol/uso terapéutico , Persona de Mediana Edad , Modelos de Riesgos Proporcionales , Proteinuria/etiología

RESUMEN

Experiments were performed in mice to investigate whether cyclooxygenase-2 (COX-2) in epithelial cells near the tubulovascular contact point (macula densa and TAL cells) may regulate renin gene expression in juxtaglomerular granular cells. Renin activity, afferent arteriolar granularity, and renin mRNA were determined in wild-type mice and in COX-2-knockout mice on control and low-NaCl diets. Renin activity in microdissected glomeruli assessed as angiotensin I formation in the presence of excess substrate and afferent arteriolar granularity determined by direct visualization and immunostaining were significantly reduced in COX-2 -/- compared with wild-type animals. Similarly, renal cortical mRNA levels were lower in COX-2 -/- than in wild-type mice. Maintaining mice on a low-salt diet for 14 days induced an increase in renin mRNA, afferent arteriolar granularity, and renin activity in wild-type mice. In contrast, renin mRNA and renin granularity did not significantly increase in low-salt-treated COX-2 -/- mice, whereas the increase in juxtaglomerular renin enzyme activity was markedly attenuated, but not fully blocked. In additional experiments we found that COX-2 mRNA was increased in angiotensin type 1A receptor-knockout mice compared with wild-type mice. We conclude that COX-2 in the tubulovascular contact region is a critical determinant of renin synthesis in granular cells under resting conditions and that it participates in the stimulation of renin expression caused by a low-NaCl intake.

Asunto(s)

Dieta Hiposódica , Isoenzimas/genética , Prostaglandina-Endoperóxido Sintasas/genética , Renina/biosíntesis , Animales , Arteriolas/citología , Arteriolas/metabolismo , Ciclooxigenasa 2 , Células Epiteliales/metabolismo , Femenino , Tasa de Filtración Glomerular , Isoenzimas/deficiencia , Aparato Yuxtaglomerular/irrigación sanguínea , Aparato Yuxtaglomerular/citología , Aparato Yuxtaglomerular/metabolismo , Corteza Renal/irrigación sanguínea , Corteza Renal/metabolismo , Túbulos Renales/irrigación sanguínea , Túbulos Renales/citología , Túbulos Renales/metabolismo , Ratones , Ratones Noqueados , Prostaglandina-Endoperóxido Sintasas/deficiencia , ARN Mensajero/biosíntesis , Receptor de Angiotensina Tipo 1 , Receptores de Angiotensina/deficiencia , Receptores de Angiotensina/genética , Renina/genética

RESUMEN

Reducing luminal NaCl concentration in the macula densa region of the nephron stimulates renin secretion, and this response is blocked by a specific inhibitor of cyclooxygenase-2 (COX-2) (Traynor, T. R., Smart, A., Briggs, J. P., and Schnermann, J. (1999) Am. J. Physiol. Renal Physiol. 277, F706-710). To study whether low NaCl activates COX-2 activity or expression we clonally derived a macula densa cell line (MMDD1 cells) from SV-40 transgenic mice using fluorescence-activated cell sorting of renal tubular cells labeled with segment-specific fluorescent lectins. MMDD1 cells express COX-2, bNOS, NKCC2, and ROMK, but not Tamm-Horsfall protein, and showed rapid (86)Rb(+) uptake that was inhibited by a reduction in NaCl concentration and by bumetanide or furosemide. Isosmotic exposure of MMDD1 cells to low NaCl (60 mm) caused a prompt and time-dependent stimulation of prostaglandin E(2) (PGE(2)) release that was prevented by the COX-2 specific inhibitor NS-398 (10 microm). Reducing NaCl to 60 and 6 mm for 16 h increased COX-2 expression in a chloride-dependent fashion. Low NaCl phosphorylated p38 kinase within 30 min and ERK1/2 kinases within 15 min without changing total MAP kinase levels. Low NaCl-stimulated PGE(2) release and COX-2 expression was inhibited by SB 203580 and PD 98059 (10 microm), inhibitors of p38 and ERK kinase pathways. We conclude that low chloride stimulates PGE(2) release and COX-2 expression in MMDD1 cells through activation of MAP kinases.

Asunto(s)

Dinoprostona/metabolismo , Isoenzimas/metabolismo , Corteza Renal/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Prostaglandina-Endoperóxido Sintasas/metabolismo , Cloruro de Sodio/farmacología , Animales , Animales Modificados Genéticamente , Línea Celular , Ciclooxigenasa 2 , Corteza Renal/citología , Corteza Renal/enzimología , Ratones , Fosforilación , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa

RESUMEN

We have previously shown that hypertonicity stimulates cyclooxygenase-2 (COX-2) expression in cultured medullary epithelial cells. The aims of the present study were (i) to examine the role of cytoplasmic signaling through MAPK pathways in tonicity regulation of COX-2 expression in collecting duct cells and (ii) to assess the possible contribution of COX-2 to the survival of inner medullary collecting duct (IMCD) cells under hypertonic conditions. In mIMCD-K2 cells, a cell line derived from mouse IMCDs, hypertonicity induced a marked increase in COX-2 protein expression. The stimulation was reduced significantly by inhibition of MEK1 (PD-98059, 5-50 microm) and p38 (SB-203580, 5-100 microm) and was almost abolished by the combination of the two compounds. To study the role of JNK in tonicity-stimulated COX-2 expression, IMCD-3 cell lines stably transfected with dominant-negative mutants of three JNKs (JNK-1, -2, and -3) were used. Hypertonicity-stimulated COX-2 protein expression was significantly reduced in dominant-negative JNK-2-expressing cells and was unchanged in dominant-negative JNK-1- and JNK-3-expressing cells compared with controls. The reduction of COX-2 expression was associated with greatly reduced viability of dominant-negative JNK-2-expressing cells during hypertonicity treatment. 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2) (2-8 microm), an inhibitor of Src kinases, reduced the tonicity-stimulated COX-2 expression in a dose-dependent manner, whereas PP3, an inactive analog of PP2, had no effect. Inhibition of COX-2 activity by NS-398 (30-90 microm) and SC-58236 (10-20 microm) significantly reduced viability of mIMCD-K2 cells subjected to prolonged hypertonic treatment. We conclude that 1) all three members of the MAPK family (ERK, JNK-2, and p38) as well as Src kinases are required for tonicity-stimulated COX-2 expression in mouse collecting duct cells and that 2) COX-2 may play a role in cell survival of medullary cells under hypertonic conditions.

Asunto(s)

Isoenzimas/metabolismo , Túbulos Renales Colectores/enzimología , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Prostaglandina-Endoperóxido Sintasas/metabolismo , Animales , Línea Celular , Ciclooxigenasa 2 , Dinoprostona/metabolismo , Activación Enzimática , Inhibidores Enzimáticos/farmacología , Isoenzimas/efectos de los fármacos , Proteínas Quinasas JNK Activadas por Mitógenos , Túbulos Renales Colectores/citología , Ratones , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidores , Proteínas Quinasas Activadas por Mitógenos/genética , Nitrobencenos/farmacología , Concentración Osmolar , Prostaglandina-Endoperóxido Sintasas/efectos de los fármacos , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Sulfonamidas/farmacología

RESUMEN

The role of thromboxane (TP) in the vasoconstriction induced by tubuloglomerular feedback or 18-h ureteral obstruction was studied in wild type mice (TP +/+), and in heterozygous (TP +/-) and homozygous TP receptor knockout mice (TPR -/-). TGF function was assessed from the response of stop flow pressure (PSF) to a maximum increase in loop of Henle flow rate (0-30 nL min-1). PSF fell by 6.4 +/- 0.4 mmHg in wild-type mice, by 6.1 +/- 0.6 mmHg in TP +/-, and by 7.9 +/- 0.7 mmHg in TP -/- mice. In the presence of the TP receptor agonist U46,619 (10-5 M) the PSF reduction increased to 10. 4 +/- 0.8 mmHg in TP +/+, and to 10.6 +/- 2.8 mmHg in TP +/-, but was unchanged at 7.7 +/- 0.7 mmHg in TP -/-. Mean arterial blood pressures were comparable between groups (103 +/- 3 mmHg in TP +/+, 113 +/- 4.6 in TP +/- and 113 +/- 2.4 mmHg in TP -/- mice). Intratubular pressure following unilateral ureteral obstruction was significantly higher in TP -/- than in TP +/+ mice both in the early phase (0-3 h) and late phase (18 h) of obstruction. These results indicate that chronic TP receptor deficiency does not significantly alter maximum TGF responses in mice, and that it is accompanied by exaggerated vasodilatation during short-term unilateral ureteral obstruction and attenuated vasoconstriction during longer lasting obstruction. We conclude that thromboxane is primarily a regulator of renal vascular tone under pathophysiological conditions.

Asunto(s)

Glomérulos Renales/fisiología , Túbulos Renales/fisiología , Receptores de Tromboxanos/deficiencia , Obstrucción Ureteral/fisiopatología , Vasoconstricción/fisiología , Animales , Retroalimentación , Ratones , Ratones Noqueados/genética , Receptores de Tromboxanos/genética

RESUMEN

OBJECTIVE: To determine mortality, morbidity, and costs attributable to surgical-site infections (SSIs) in the 1990s. DESIGN: A matched follow-up study of a cohort of patients with SSI, matched one-to-one with patients without SSI. SETTING: A 415-bed community hospital. STUDY POPULATION: 255 pairs of patients with and without SSI were matched on age, procedure, National Nosocomial Infection Surveillance System risk index, date of surgery, and surgeon. OUTCOME MEASURES: Mortality, excess length of hospitalization, and extra direct costs attributable to SSI; relative risk for intensive care unit (ICU) admission and for readmission to the hospital. RESULTS: Of the 255 pairs, 20 infected patients (7.8%) and 9 uninfected patients (3.5%) died during the postoperative hospitalization (relative risk [RR], 2.2; 95% confidence interval [CI95], 1.1-4.5). Seventy-four infected patients (29%) and 46 uninfected patients (18%) required ICU admission (RR, 1.6; CI95, 1.3-2.0). The median length of hospitalization was 11 days for infected patients and 6 days for uninfected patients. The extra hospital stay attributable to SSI was 6.5 days (CI95, 5-8 days). The median direct costs of hospitalization were $7,531 for infected patients and $3,844 for uninfected patients. The excess direct costs attributable to SSI were $3,089 (CI95, $2,139-$4,163). Among the 229 pairs who survived the initial hospitalization, 94 infected patients (41%) and 17 uninfected patients (7%) required readmission to the hospital within 30 days of discharge (RR, 5.5; CI95, 4.0-7.7). When the second hospitalization was included, the total excess hospitalization and direct costs attributable to SSI were 12 days and $5,038, respectively. CONCLUSIONS: In the 1990s, patients who develop SSI have longer and costlier hospitalizations than patients who do not develop such infections. They are twice as likely to die, 60% more likely to spend time in an ICU, and more than five times more likely to be readmitted to the hospital. Programs that reduce the incidence of SSI can substantially decrease morbidity and mortality and reduce the economic burden for patients and hospitals.

Asunto(s)

Infección Hospitalaria/economía , Costos de Hospital/estadística & datos numéricos , Tiempo de Internación/economía , Infección de la Herida Quirúrgica/economía , Adulto , Anciano , Anciano de 80 o más Años , Estudios de Casos y Controles , Infección Hospitalaria/complicaciones , Infección Hospitalaria/mortalidad , Estudios de Seguimiento , Costos de la Atención en Salud , Hospitales Comunitarios , Humanos , Persona de Mediana Edad , North Carolina , Readmisión del Paciente/economía , Infección de la Herida Quirúrgica/complicaciones , Infección de la Herida Quirúrgica/mortalidad

Asunto(s)

Isoenzimas/metabolismo , Prostaglandina-Endoperóxido Sintasas/metabolismo , Prostaglandinas/biosíntesis , Vejiga Urinaria/fisiología , Animales , Ciclooxigenasa 1 , Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa/uso terapéutico , Homeostasis , Humanos , Proteínas de la Membrana , Vejiga Urinaria/enzimología

RESUMEN

The discovery that 15-deoxy-Delta12,14-prostaglandin J2 (15d-PGJ2) is a ligand for the gamma-isoform of peroxisome proliferator-activated receptor (PPAR) suggests nuclear signaling by prostaglandins. Studies were undertaken to determine the nephron localization of PPAR isoforms and their heterodimer partners, retinoid X receptors (RXR), and to evaluate the function of this system in the kidney. PPARalpha mRNA, determined by RT-PCR, was found predominately in cortex and further localized to proximal convoluted tubule (PCT); PPARgamma was abundant in renal inner medulla, localized to inner medullary collecting duct (IMCD) and renal medullary interstitial cells (RMIC); PPARbeta, the ubiquitous form of PPAR, was abundant in all nephron segments examined. RXRalpha was localized to PCT and IMCD, whereas RXRbeta was expressed in almost all nephron segments examined. mRNA expression of acyl-CoA synthase (ACS), a known PPAR target gene, was stimulated in renal cortex of rats fed with fenofibrate, but the expression was not significantly altered in either cortex or inner medulla of rats fed with troglitazone. In cultured RMIC cells, both troglitazone and 15d-PGJ2 significantly inhibited cell proliferation and dramatically altered cell shape by induction of cell process formation. We conclude that PPAR and RXR isoforms are expressed in a nephron segment-specific manner, suggesting distinct functions, with PPARalpha being involved in energy metabolism through regulating ACS in PCT and with PPARgamma being involved in modulating RMIC growth and differentiation.

Asunto(s)

Riñón/metabolismo , Nefronas/metabolismo , Receptores Citoplasmáticos y Nucleares/genética , Receptores de Ácido Retinoico/genética , Factores de Transcripción/genética , Transcripción Genética , Tejido Adiposo/metabolismo , Empalme Alternativo , Animales , Animales Recién Nacidos , Células Cultivadas , Coenzima A Ligasas/genética , Cartilla de ADN , Desarrollo Embrionario y Fetal , Regulación del Desarrollo de la Expresión Génica , Riñón/embriología , Corteza Renal/metabolismo , Médula Renal/metabolismo , Túbulos Renales Colectores/metabolismo , Masculino , Especificidad de Órganos , Isoformas de Proteínas/genética , ARN Mensajero/genética , Ratas , Ratas Sprague-Dawley , Receptores X Retinoide , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa

RESUMEN

Previous results from our laboratory have shown that in the isolated perfused juxtaglomerular apparatus, nonselective inhibitors of cyclooxygenase (COX) activity prevent the stimulation of renin secretion by a reduction in luminal NaCl concentration at the macula densa. The present studies were performed to examine which COX isoform is involved in NaCl-dependent renin secretion. In the absence of COX inhibitors, a reduction in luminal NaCl (from Na 141/Cl 120 mM to Na 26/Cl 7 mM) caused an increase in renin secretion rate from 4.5 +/- 1.8 to 26.1 +/- 7.4 nGU/min (P < 0.01, n = 19). The presence of the COX-1 inhibitor valerylsalicylate (500 microM) in lumen and bath did not affect the stimulation of renin secretion by a reduction in luminal NaCl concentration (5 +/- 1.8 nGU/min at high NaCl, and 30.5 +/- 9.4 nGU/min at low NaCl; P < 0.01, n = 8). In contrast, the specific COX-2 inhibitor NS-398 (50 microM) in lumen and bath abolished the stimulating effect of low luminal NaCl (12.8 +/- 3.9 nGU/min at high NaCl, and 10.7 +/- 3.1 nGU/min at low NaCl; NS, n = 15). The finding that COX-2 is critically involved in macula densa control of renin secretion indicates that the COX-2-expressing epithelial cells in the tubuloglomerular contact area are a likely source of prostaglandins participating in the signaling pathway between the macula densa and renin-producing granular cells.

Asunto(s)

Inhibidores de la Ciclooxigenasa/farmacología , Isoenzimas/efectos de los fármacos , Aparato Yuxtaglomerular/fisiología , Túbulos Renales Distales/fisiología , Prostaglandina-Endoperóxido Sintasas/efectos de los fármacos , Renina/antagonistas & inhibidores , Renina/metabolismo , Animales , Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa 2 , Femenino , Técnicas In Vitro , Nitrobencenos/farmacología , Concentración Osmolar , Conejos , Salicilatos/farmacología , Cloruro de Sodio/farmacología , Sulfonamidas/farmacología

Asunto(s)

Túbulos Renales Distales/metabolismo , Cloruro de Sodio/metabolismo , Animales , Retroalimentación , Tasa de Filtración Glomerular , Humanos , Glomérulos Renales/metabolismo , Óxido Nítrico/fisiología

RESUMEN

Renal medullary prostaglandins are believed to exert an important functional role in antagonizing vasopressin effects in dehydration. Studies were undertaken to determine the effect of hyperosmolality on cyclooxygenase (COX) isoform expression in the renal medulla. COX-1 and COX-2 mRNA and protein levels were determined by RT-PCR or Western blotting in Sprague-Dawley rats on varying water intakes, in Brattleboro rats and in Long-Evans controls. Over a wide range of urinary tonicity, COX-2 expression correlated closely with urine osmolality levels (R = 0.872). COX-1 levels did not vary. Immunolocalization showed that the stimulation of COX-2 expression by dehydration occurred predominantly in the collecting duct. Hypertonicity caused by addition of NaCl produced a dose- and time-dependent stimulation of COX-2 expression in mIMCD-K2 cells as well as in MDCK cells. COX-1 was unaffected. In the same cell lines, mannitol, sucrose, and raffinose also had a stimulatory effect. The tonicity-stimulated COX-2 expression in mIMCD-K2 cells was almost completely blocked by a tyrosine kinase inhibitor, genistein at 100 microM. In MDCK cells transfected with a 2.7-kb COX-2 promoter and lacZ reporter construct, NaCl induced a twofold increase in beta-galactosidase activity. Using mIMCD-K2 cells, hypertonic NaCl (600 mosmol/kgH(2)O for 24 h) induced a 33-fold increase in PGE(2) release determined by enzyme immunoassay, an effect completely blocked by 3 microM indomethacin or the COX-2-specific blocker N-(2-cyclohexy-4-nitrophenyl)methanesulfonamide (NS-398). We conclude that in inner medulla, COX-2 but not COX-1 is upregulated by hyperosmolality.

Asunto(s)

Isoenzimas/biosíntesis , Médula Renal/metabolismo , Peroxidasas/biosíntesis , Prostaglandina-Endoperóxido Sintasas/biosíntesis , Animales , Células Cultivadas , Ciclooxigenasa 1 , Ciclooxigenasa 2 , Deshidratación , Técnicas In Vitro , Isoenzimas/genética , Médula Renal/citología , Proteínas de la Membrana , Prostaglandina-Endoperóxido Sintasas/genética , ARN Mensajero/biosíntesis , Ratas , Ratas Brattleboro , Ratas Long-Evans , Ratas Sprague-Dawley , Cloruro de Sodio/farmacología

RESUMEN

Induction of the inducible cyclooxygenase isoform COX-2 is likely to be an important mechanism for increased prostaglandin production in renal inflammation. We examined the effect of lipopolysaccharide (LPS) on regional renal COX-2 expression in the rat. In the inner medulla, LPS injection (4 mg/kg ip) induced a twofold and 2.5-fold increase in the levels of COX-2 mRNA and COX-2 protein, respectively. In contrast, COX-2 expression in the renal cortex was not significantly altered. COX-2 promoter transgenic mice were created using the 2.7-kb flanking region of the rat COX-2 gene. In these animals, LPS injection induced reporter gene expression predominately in the inner medulla. The LPS receptor CD14, usually regarded as a monocyte/macrophage-specific marker, was found to be abundantly expressed in the inner medulla and in dissected inner medullary collecting duct (IMCD) cells, suggesting that it may mediate medullary COX-2 induction. CD14 was present only at low levels in cortex and cortical segments, including glomeruli. In cultured cells, it was abundant in mouse IMCD (mIMCD-K2) cells and renal medullary interstitial cells, but largely undetectable in mesangial cells and M1 cells, a cell line derived from mouse cortical collecting ducts. In the mIMCD-K2 cell line, LPS significantly induced COX-2 mRNA expression, with concomitant induction of CD14. LPS-stimulated COX-2 expression was reduced by the addition of an anti-CD14 monoclonal antibody to the culture medium. These results demonstrate that LPS selectively stimulates COX-2 expression in the renal inner medulla through a CD14-dependent mechanism.

Asunto(s)

Isoenzimas/biosíntesis , Riñón/metabolismo , Receptores de Lipopolisacáridos/fisiología , Lipopolisacáridos/farmacología , Prostaglandina-Endoperóxido Sintasas/biosíntesis , Animales , Ciclooxigenasa 2 , Isoenzimas/genética , Riñón/citología , Riñón/efectos de los fármacos , Receptores de Lipopolisacáridos/genética , Masculino , Prostaglandina-Endoperóxido Sintasas/genética , ARN Mensajero/biosíntesis , Ratas , Ratas Sprague-Dawley

RESUMEN

In these experiments, we used a strain of angiotensin converting enzyme (ACE) germline null mutant mice, generated by J. H. Krege and co-workers (J. H. Krege, S. W. M. John, L. L. Langenbach, J. B. Hodgin, J. R. Hagaman, E. S. Bachman, J. C. Jennette, D. A. O'Brien, and O. Smithies. Nature 375: 146-148, 1995), to examine the effect of chronic ACE deficiency on the magnitude of tubuloglomerular feedback (TGF) responses. The genotype was determined by PCR on DNA extracted from the tail and was verified after each experiment by assessment of the blood pressure response to an injection of ANG I. To assess TGF responsiveness, we determined the change in stop-flow pressure (PSF) caused by increasing NaCl concentration at the macula densa by using micropuncture techniques. When loop of Henle flow rate was increased from 0 to 40 nl/min, PSF fell from a mean of 42.3 +/- 1.95 to 33.6 +/- 2.09 mmHg (n = 6, P = 0.005) in wild-type mice (+/+), fell from 40.6 +/- 2.35 to 38.6 +/- 1.93 mmHg in heterozygous (+/-) mice (n = 7, P = 0.014), and did not change in homozygous ACE (-/-) mice [36.7 +/- 2.02 mmHg vs. 36.4 +/- 2.01 mmHg; n = 4, P = not significant (NS)]. During an infusion of ANG II at a dose that did not significantly elevate blood pressure (70 ng. kg-1. min-1), TGF response magnitude (PSF 0 - PSF 40) increased from 6.5 +/- 1.4 to 9.8 +/- 1.19 mmHg in +/+ (P = 0.006), from 1.14 +/- 0.42 to 4.6 +/- 1.3 mmHg in +/- (P = 0.016), and from 0.42 +/- 0.25 to 4.02 +/- 1.06 in -/- mice (P = 0.05). Absence of TGF responses in ACE null mutant mice and restoration of near-normal responses during an acute infusion of ANG II supports previous conclusions that ANG II is an essential component in the signal transmission pathway that links the macula densa with the glomerular vascular pole.

Asunto(s)

Retroalimentación/fisiología , Aparato Yuxtaglomerular/fisiología , Glomérulos Renales/metabolismo , Túbulos Renales/metabolismo , Peptidil-Dipeptidasa A/genética , Angiotensina II/farmacología , Animales , Presión Sanguínea/fisiología , Cartilla de ADN , Retroalimentación/efectos de los fármacos , Femenino , Genotipo , Masculino , Ratones , Ratones Noqueados , Circulación Renal/efectos de los fármacos , Circulación Renal/fisiología , Sistema Renina-Angiotensina/fisiología , Cloruro de Sodio/farmacología , Vasoconstricción/fisiología

RESUMEN

The nephron is derived from the ureteric bud and metanephric mesenchyme and develops into a complex epithelial structure with a wide variety of phenotypes along its length. This segmental variation in expression of molecules provides an approach to understand the lineage of unique segments. The present study evaluated the expression of four relatively well-localized molecules--renin, Tamm-Horsfall protein (THP), oxytocin receptor (OTR), and the vasopressin type 2 receptor (V2R)--in cultured mouse-rat chimeric metanephric kidneys using reverse transcription-polymerase chain reaction (RT-PCR). Chimeric kidneys were formed by 1) separating the ureteric bud (U) from the metanephric mesenchyme (M) of mouse (m) at E11 and rat (r) at E13 days of gestation and 2) recombining the ureteric bud of one species with the metanephric mesenchyme of the other species (i.e., UrMm and UmM(r), followed by filter culture until differentiated. Species-specific restriction enzymes for all four genes were chosen to digest the PCR product from either rat or mouse. RT-PCR was performed for each mRNA species and the products digested. The V2R product from the UrMm chimera was cleaved by a restriction enzyme known to digest only rat product, suggesting the PCR product was produced predominantly by cells derived from the ureteric bud. The renin, OTR, and THP products from both chimeras were cleaved equally well by species-specific restriction enzymes, suggesting the products were made by cells originating from both the ureteric bud and the metanephric mesenchyme. These studies demonstrate that the cultured chimeric metanephric model is useful to study segment lineage. The results suggest that the lineage of at least certain portions of the nephron is heterogenous.

Asunto(s)

Quimera , Nefronas/citología , Nefronas/embriología , Animales , Secuencia de Bases , Quimera/genética , Cartilla de ADN/genética , Ratones , Datos de Secuencia Molecular , Mucoproteínas/genética , Nefronas/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Receptores de Oxitocina/genética , Receptores de Vasopresinas/genética , Renina/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Homología de Secuencia de Ácido Nucleico , Especificidad de la Especie , Uromodulina

RESUMEN

The National Institutes of Health (NIH) currently anticipates a period of growth in budgets, making renal community input into research planning and prioritizing particularly important. In considering which women's health issues should be given high priority for research study, the questions of both public health impact and scientific opportunity should be considered. A series of health topics in which there is some evidence for particular impact on women's health are briefly reviewed, and the process the NIH uses for community input and priority setting are discussed.

Asunto(s)

Fallo Renal Crónico/epidemiología , National Institutes of Health (U.S.) , Nefrología/tendencias , Salud de la Mujer , Femenino , Prioridades en Salud , Humanos , Masculino , Investigación/tendencias , Factores Sexuales , Estados Unidos/epidemiología

RESUMEN

Studies were performed to investigate the regulatory mechanism of bladder cyclooxygenase-2 (COX-2) expression after outlet obstruction. In situ hybridization of murine bladder tissues using COX-2-specific riboprobes demonstrated that COX-2 expression was induced predominantly in the bladder smooth muscle cells after outlet obstruction. To study the effect of increased mechanical stretch on COX isoform expression, cultured rat bladder smooth muscle cells were grown on silicone elastomer-bottomed plates coated with collagen type I and were subjected to continuous cycles of stretch/relaxation for variable duration. COX-1 mRNA levels did not change with stretch. COX-2 expression increased in a time-dependent manner after stretch, with maximal mRNA and protein levels occurring after 4 h. PGE2 levels increased more than 40-fold in the culture media after stretch, consistent with increased COX activity, and this was reduced to near completion in the presence of a COX-2 inhibitor, NS-398. Exposure to stretch over a 48-h period induced a 4.7 +/- 0.6-fold increase in tritiated thymidine incorporation rate. This increase in DNA synthesis was markedly suppressed when the cells were stretched in the presence of NS-398. We conclude that in bladder obstruction COX-2 activation occurs predominantly in the smooth muscle cells in response to mechanical stretch. Our findings also suggest that stretch-activated COX-2 expression may participate in bladder smooth muscle cell proliferation and thereby play a role in pathological bladder wall thickening after obstruction.

Asunto(s)

Isoenzimas/metabolismo , Músculo Liso/enzimología , Prostaglandina-Endoperóxido Sintasas/metabolismo , Obstrucción del Cuello de la Vejiga Urinaria/enzimología , Vejiga Urinaria/enzimología , Animales , División Celular/fisiología , Ciclooxigenasa 2 , Femenino , Isoenzimas/genética , Isoenzimas/fisiología , Ratones , Ratones Endogámicos , Músculo Liso/citología , Prostaglandina-Endoperóxido Sintasas/genética , Prostaglandina-Endoperóxido Sintasas/fisiología , ARN Mensajero/metabolismo , Ratas , Ratas Endogámicas Lew , Estrés Mecánico , Vejiga Urinaria/citología

RESUMEN

The effect of the adenosine type 1 receptor agonist N6-cyclohexyladenosine (CHA) on glomerular vascular reactivity was studied in male angiotensin II type 1A (AT1A) receptor knockout mice (9). Vascular reactivity was assessed as the response of stop-flow pressure (PSF) to infusion of CHA into loops of Henle using micropuncture techniques. In AT1A +/+ mice at ambient arterial blood pressure (96.7 +/- 2.8 mmHg), the presence of CHA (10 (-5) M) in the perfusate increased PSF responses from 6.8 +/- 0.6 to 14.3 +/- 0.9 mmHg when the loop of Henle of the index nephron was perfused and from 0.7 +/- 0.3 to 12.3 +/- 1.0 mmHg when the loop of an adjacent nephron was perfused. At reduced arterial blood pressure (82.8 +/- 1. 3 mmHg), index nephron perfusion with CHA increased PSF responses from 4.5 +/- 0.3 to 9.4 +/- 0.4 mmHg. In AT1A -/- mice with a mean arterial blood pressure of 80 +/- 1.9 mmHg, CHA increased PSF responses only from 0.1 +/- 0.3 to 3.6 +/- 0.54 mmHg during index nephron perfusion and from 0.25 +/- 0.2 to 2.7 +/- 0.55 mmHg during adjacent nephron perfusion, significantly less than in wild-type animals (P < 0.001). Responses to CHA were intermediate in AT1A +/- mice. Thus AT1A receptor knockout mice show a markedly reduced constrictor response to CHA both in the presence and absence of simultaneous activation of the tubuloglomerular feedback system. These data support the notion of a functional interaction between adenosine and angiotensin II in the regulation of afferent arteriolar tone.

Asunto(s)

Glomérulos Renales/irrigación sanguínea , Receptores de Angiotensina/deficiencia , Receptores Purinérgicos P1/fisiología , Circulación Renal/fisiología , Vasoconstricción/fisiología , Adenosina/análogos & derivados , Adenosina/farmacología , Animales , Presión Sanguínea/fisiología , Masculino , Ratones , Ratones Noqueados/genética , Perfusión , Presión , ARN Mensajero/metabolismo , Receptores de Angiotensina/genética , Receptores Purinérgicos P1/genética , Sistema Vasomotor/efectos de los fármacos

RESUMEN

Grb7, a signaling protein whose physiological function is unknown, binds receptor tyrosine kinases important for normal kidney development. By investigating and correlating Grb7 gene expression with that reported for Grb7-binding receptors, we provide clues to Grb7 function(s). RT-PCR and immunoblot were used to demonstrate Grb7 gene and protein expression in the mature kidney. Additional RT-PCR studies detected gene expression in all microdissected adult nephron segments examined, except glomeruli, and in the mouse metanephric kidney from embryonic day 11 (E11) through to day 17 (E17). In situ hybridization at E14 demonstrated the following cellular pattern of localization: Grb7 mRNA in metanephric epithelia of mesenchymal and ureteric bud origin; no expression in the undifferentiated mesenchyme; and little expression in podocyte-destined cells or primitive glomeruli. Grb7 mRNA was also present in the epithelia of the lung and gut at E14. Thus Grb7 may have a basic function in growth factor signaling in terminally differentiated epithelia along the nephron and in developing epithelia in the kidney, lung, and gut. It is localized in a pattern permissive for a role in Her2 and Ret receptor signaling.

Asunto(s)

Riñón/embriología , Riñón/fisiología , Proteínas/fisiología , Transducción de Señal/fisiología , Animales , Factor de Crecimiento Epidérmico/fisiología , Receptores ErbB/fisiología , Proteína Adaptadora GRB7 , Regulación del Desarrollo de la Expresión Génica , Hibridación in Situ , Masculino , Ratones , ARN Mensajero/análisis , Ratas , Ratas Sprague-Dawley

RESUMEN

Luminal [NaCl] at the macula densa (MD) has two established effects: regulation of glomerular arteriolar resistance through tubuloglomerular feedback (TGF) and control of renin secretion. TGF acts as a minute-to-minute stabilizer of distal salt delivery, thereby minimizing the impact of random perturbations in filtration and absorption forces on NaCl excretion. During long-lasting perturbations of MD [NaCl], control of renin secretion becomes the dominant function of the MD. The potentially maladaptive effect of TGF under chronic conditions is prevented by TGF adaptations permitting adjustments in glomerular filtration rate to occur. TGF adaptation is mechanistically coupled to the endpoint targeted by chronic deviations in MD [NaCl], the rate of local and systemic angiotensin II generation. Studies of TGF in transgenic mice are expected to provide further insights into the mechanisms mediating between luminal [NaCl] and afferent arterioles. TGF responses are virtually abolished in mice in which either the AT1A gene or the angiotensin converting enzyme gene is rendered nonfunctional by homologous recombination. In contrast, TGF responses are unaltered in nitric oxide synthase I knockout mice. Thus, an intact renin-angiotensin system appears to be critical for the TGF signaling pathway.

Asunto(s)

Retroalimentación/fisiología , Glomérulos Renales/fisiología , Túbulos Renales/fisiología , Animales , Glomérulos Renales/irrigación sanguínea , Túbulos Renales/irrigación sanguínea , Circulación Renal/fisiología