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Biomedical subjects

R A Gomez

Publications and source records attributed to R A Gomez.

At least 37 records · Page 2Linked to original sources
PubMed1997

Biomechanical coupling in renin-releasing cells.

The renin-angiotensin system is a major regulatory system controlling extracellular fluid volume and blood pressure. The rate-limiting enzyme in this hormonal cascade is renin, which is synthesized and secreted into the circulation by renal juxtaglomerular (JG) cells. The renal baroreceptor is a key physiologic regulator of renin secretion, whereby a change in renal perfusion pressure is sensed by these cells and results in a change in renin release. However, the mechanism, direct or indirect, underlying pressure transduction is unknown. We studied the direct application of mechanical stretch to rat JG cells and human renin-expressing (CaLu-6) cells on the release of renin. JG cells released a low level of baseline renin, comprising < 5% of their total renin content. By contrast, renin secretion from CaLu-6 cells comprised approximately 30% of cellular stores, yet was also stimulated twofold by 10 microM forskolin (P </= 0.001). In JG cells, mechanical stretch inhibited basal renin release by 42% (P < 0.01) and forskolin-stimulated renin release by 25% (P < 0.05). In CaLu-6 cells, stretch inhibited basal- and forskolin-stimulated renin release by 30 and 26%, respectively (both P < 0.01). Northern blot analysis demonstrated a stretch-induced reduction in baseline renin mRNA accumulation of 26% (P < 0.05) in JG and 46% (P < 0.05) in CaLu-6 cells. The data demonstrate that mechanical stretch in renin-releasing cells inhibits basal and stimulated renin release accompanied by a decrease in renin mRNA accumulation. Further studies will be necessary to characterize the intracellular events mediating biomechanical coupling in renin-expressing cells and the relationship of this signaling pathway to the in vivo baroreceptor control of renin secretion.

Animals↗
PubMed1997

Oxygen regulates vascular endothelial growth factor-mediated vasculogenesis and tubulogenesis.

To determine whether low oxygen is a stimulus for endothelial cell differentiation and vascular development in the kidney, we examined the effect of low oxygen on rat metanephric organ culture, a model known to recapitulate nephrogenesis in the absence of vessels. After 6 days in culture in standard (20% O2) or low oxygen (1-3% O2) conditions, metanephric kidney growth and morphology were assessed by DNA measurement, and light and electron microscopy. DNA content was higher in 3% O2-treated explants (2.5 +/- 0.17 microgram/kidney, n = 9) than in 20% O2 explants (1.5 +/- 0.09 microgram/kidney, n = 9), P < 0.05. Low oxygen induced proliferation of tubular epithelial cells, resulting in enhanced number of tubules of similar size. Endothelial cells forming capillaries were localized in 3% O2 explants by light and electron microscopy and by immunocytochemistry using endothelial cell markers. Flt-1, Flk-1, and ACE-containing cells were detected in 3% O2-treated explants, whereas 20% O2 explants were virtually negative. VEGF mRNA levels were 10-fold higher in 3% O2-treated explants than in 20% O2-treated explants. Addition of anti-VEGF antibodies to 3% O2-treated explants prevented low oxygen-induced growth and endothelial cell differentiation and proliferation. Our data indicate that low oxygen stimulates growth by cell proliferation and induces tubulogenesis, endothelial cell differentiation, and vasculogenesis in metanephric kidneys in culture. Upregulation of VEGF expression by low oxygen and prevention of low oxygen-induced tubulogenesis and vasculogenesis by anti-VEGF antibodies indicate that these changes were mediated by VEGF. These data suggest that low oxygen is the stimulus to initiate renal vascularization.

Animals↗
PubMed1997

Differential expression of angiotensin II receptors during early renal morphogenesis.

Angiotensin II (ANG II) and its receptors, AT1 and AT2, may modulate kidney development. To define the temporal and spatial distribution of AT1 and AT2 receptors and their mRNAs during nephrogenesis, fetal, newborn, and adult rat kidneys were studied using reverse transcription-polymerase chain reaction and radioligand binding autoradiography. AT1 expression was minimal at embryonic day 14 (E14), highly expressed at E20, and persisted into adulthood. Conversely, AT2 expression was easily detected from E14 through postnatal day 7 but was undetectable by postnatal day 28. At E14, 76% of the receptors were AT2, 24% were AT1, and both were found in the undifferentiated mesenchyme. By E17, AT1 comprised 40% of the receptors and localized to mature nephron segments, whereas AT2 remained within both condensed mesenchyme and differentiating epithelia. The dissociation constants for AT1 and AT2 were 0.45 +/- 0.09 nM and 0.73 +/- 0.15 nM, respectively, at E17, similar to adult values. By E20, AT1 and AT2 colocalized to the outer medullary stripe, deep nephrons, medullary rays, and blood vessels, while AT2 continued to predominate in the actively differentiating cortex. The presence of both subtypes of receptors capable of binding ANG II during early nephrogenesis and the time-dependent and structure-specific regulation of receptor localization confirm a regulated developmental program for receptor expression and suggest important roles for AT1 and AT2 in renal morphogenesis.

Aging↗
PubMed1997

Zis: a developmentally regulated gene expressed in juxtaglomerular cells.

Renal juxtaglomerular (JG) cells are specialized myoepithelioid cells located in the afferent arteriole at the entrance to the glomerulus. Their main function and distinctive feature is the synthesis and release of renin, the key hormone-enzyme of the renin-angiotensin system that regulates arterial blood pressure. Despite their relevance to health and disease, not much is known about factors that confer and/or maintain JG cell identity. To identify genes uniquely expressed in JG cells, we used a cell culture model and RNA differential display. JG cells cultured for 2 days express renin and renin mRNA, but after 10 days in culture they no longer contain or release renin and renin mRNA is reduced 700-fold. We report one cDNA differentially expressed in the 2-day JG cell culture that detects a 2.6-kb mRNA expressed at higher levels in newborn than adult kidney. Screening a 2-day culture JG cell cDNA library yielded clones representing differentially spliced transcripts. These cDNAs encode one unique protein (Zis) containing zinc fingers and domains characteristic of splicing factors and RNA binding proteins. Northern blot analysis confirmed Zis mRNA expression in differentiated JG cells, and identified an additional unique 1.5-kb transcript. The Zis transcripts are developmentally regulated in kidney and a number of other organs. The features of the Zis protein and its organ distribution suggest a possible role in regulation of transcription and/or splicing, both important steps for controlling developmentally expressed genes.

Aging↗
PubMed1997

Aberrant renal vascular morphology and renin expression in mutant mice lacking angiotensin-converting enzyme.

To determine whether angiotensin-converting enzyme plays a role in the development and maintenance of normal renal architecture, the renal morphology of 10- to 12-month-old female mice homozygous for a disruption of the converting enzyme gene was compared with that of age-matched wild-type mice. Tubular obstruction, dilatation, and atrophy were present in all kidneys from the homozygous mutant mice but absent in wild types; two kidneys from 4 mutant mice but none from the wild types were hydronephrotic. The entire arterial vascular tree, microdissected from mice with no converting enzyme, was grossly distorted in comparison to the vasculature of wild-type mice; all intrarenal arterial vessels were widened and thickened, including the terminal (afferent) arterioles. In wild-type mice kidneys, renin-positive cells were detected exclusively in a juxtaglomerular localization. In contrast, abnormal distribution of renin immunostaining was observed in mice without converting enzyme; scattered renin-positive cells were seen along the arterial vessels, often in a perivascular localization, and interstitial renin-positive cells surrounded glomeruli. Kidney renin mRNA was increased more than 32-fold in the mutant mice compared with wild types. Northern blot analysis revealed that this increase included the accumulation of large amounts of smaller renin RNA transcripts. In summary, mice lacking the converting enzyme exhibit abnormal renal vessels and tubules. Renin synthesis is increased, accompanied by the presence of small renin mRNA species, and renin is present mainly in interstitial and perivascular cells. We conclude that angiotensin-converting enzyme is necessary to preserve normal kidney architecture and the normal pattern of renin expression.

Animals↗
PubMed1997

Angiotensin's role in renal development.

All the components of the renin-angiotensin system (RAS) are highly expressed in the developing kidney in a unique spatial and temporal pattern that is associated with nephrogenesis, vascularization, and the proper architectural and functional development of this organ. Pharmacological inhibition of the RAS results in structural and functional developmental abnormalities of the kidneys in several animal species, including humans. Similarly, altered renal morphology and functional abnormalities have been described in mice with targeted inactivation of the angiotensinogen (Agt) and the angiotensin-converting enzyme (ACE) genes. In contrast, inactivation of angiotensin receptors have not resulted in renal morphological abnormalities, suggesting a redundancy at this level of the RAS cascade that prevents the development of renal pathology. More importantly, inactivation of the ACE or Ao genes results in a renal phenotype remarkably similar to that obtained with pharmacological inhibition of the RAS. Taken together, the available information suggests that angiotensin is necessary for normal kidney development and for the maintenance of the functional and structural integrity of the adult kidney.

Adult↗
PubMed1996

Renal interstitial sclerosis in aging: effects of enalapril and nifedipine.

The effects of nifedipine and enalapril on age-associated renal interstitial fibrosis were investigated in 60 CF1 female mice. Mice received 20 mg enalapril (ENAL) per L (N = 20), or 40 mg nifedipine (NIF) per L (N = 20) in their drinking water. Control (CONT) mice received tap water ad libitum. The percentages of both interstitial peritubular sclerosis (IPS) in cortex and interstitial medullary sclerosis (IMS) were determined. Kidney tissue was studied using immunological techniques and optical (OM) and electron microscopy (EM) to analyze the expression of renin. alpha-SM-actin and vimentine expression were also evaluated. The results showed that blood pressure levels in ENAL or NIF animals were not different from those of CONT. Renin expression was observed in arcuate vessels (AV) in ENAL animals, whereas no renin staining in AV was found in either NIF or CONT animals. Renin immunoreactivity in the juxtaglomerular apparatus was more intense in ENAL mice, as compared with NIF or CONT animals. Laboratory testing showed the following values: proteinuria (mg/mL): CONT 6.1 +/- 0.6, NIF 11.2 +/- 2.3, and ENAL 1.0 +/- 0.6 (P < 0.05); creatinine: CONT 1.37 +/- 0.24, NIF 0.87 +/- 0.16, and ENAL 0.63 +/- 0.1 (P < 0.01). The percentages of interstitial sclerosis were: %IPS: CONT 18.12 +/- 1.1, NIF 17.40 +/- 0.9, and ENAL 3.42 +/- 1.3 (P < 0.01); %IMS: CONT 23.41 +/- 1.5, NIF 21.80 +/- 1.9, and ENAL 6.12 +/- 1.2 (P < 0.01). Percentages of alpha-SM-actin expression were: CONT 13.10 +/- 1.9, NIF 13.80 +/- 0.2, and ENAL 1.00 +/- 0.1 (P < 0.01). Vimentine staining showed no differences among the groups. It was concluded that enalapril reduces the peritubular and medullar interstitial fibrosis, whereas nifedipine has no effect.

Actins↗
PubMed1996

Renal apoptosis and clusterin following ureteral obstruction: the role of maturation.

PURPOSE: Unilateral ureteral obstruction (UUO) increases renal apoptosis as well as clusterin, which in turn is modulated by angiotensin II. The present study was designed to investigate the role of maturation in these responses. MATERIALS AND METHODS: Neonatal and adult Sprague-Dawley rats underwent UUO or sham operation. Fourteen days later, renal apoptosis was quantitated by flow cytometry, and renal messenger RNA (mRNA) was quantitated for renin and clusterin. Apoptotic cells, macrophages, clusterin and Dolichos biflorus agglutinin were localized by immunohistochemistry. RESULTS: Following ipsilateral UUO, renal DNA content increased in the adult but decreased in the neonate. Renal macrophage infiltration induced by UUO was 2-fold greater in adults than in neonates, while distal tubular apoptosis was 2-fold greater in neonatal than in adult kidneys. Unilateral ureteral obstruction markedly increased renal renin mRNA expression in the neonate, but not in the adult, while clusterin expression was greater in adults than in neonates. Clusterin was localized to some, but not all, distal tubules containing apoptotic cells. CONCLUSIONS: We conclude that, compared with the adult, UUO in the neonate induces greater apoptosis, which in turn contributes to reduced renal DNA. This may be modulated by relative suppression of clusterin in the obstructed neonatal kidney due to greater activation of the renin-angiotensin system.

Aging↗
PubMed1995

Developmental consequences of the renin-angiotensin system.

Molecular, cellular, and physiological studies indicate that the renin-angiotensin system (RAS) is highly expressed during early kidney development. We propose that a major function of the RAS during early embryonic development is the modulation of growth processes that lead the primitive kidney into a properly differentiated and architecturally organized organ suited for independent extrauterine life. As development progresses, the RAS acquires new and overlapping functions such as the endocrine and paracrine regulation of blood pressure and renal hemodynamics. Disease states in adult mammals often result in expression of RAS genes and phenotypic changes resembling the embryonic pattern, emphasizing the importance of undertaking developmental studies. Because of their importance in health and disease, the immediate challenge is to identify the mechanisms that regulate the unique development of the RAS and its role(s) in normal and abnormal growth processes.

Aging↗
PubMed1995

Regulation of renal growth factors and clusterin by AT1 receptors during neonatal ureteral obstruction.

Unilateral ureteral obstruction (UUO) in the neonate impairs growth of the ipsilateral kidney. Since renal renin expression is increased by UUO, we hypothesized that, by activation of AT1 receptors, angiotensin II (ANG II) regulates expression of transforming growth factor-beta 1 (TGF-beta 1) and epidermal growth factor (EGF) in the obstructed kidney. Sprague-Dawley rats underwent left UUO or sham operation within the first 48 h of life and received losartan, 40 mg.kg-1.day-1, or saline. After 14 days, steady-state renal mRNA was determined for renin, TGF-beta 1, EGF, and clusterin. Losartan reduced the DNA content of the intact kidneys but did not further decrease that of the obstructed kidney. Losartan increased renal renin expression and decreased EGF expression by 80%, regardless of UUO. In contrast, losartan reduced TGF-beta 1 expression by 34% in obstructed kidneys but did not affect TGF-beta 1 in intact kidneys. Losartan increased clusterin expression by 60% in obstructed kidneys and seven-fold in intact kidneys. We conclude that activation of the ANG II AT1 receptor is necessary for normal renal growth and that TGF-beta 1 is regulated by AT1 receptors in the obstructed, but not intact, kidneys. Through AT1 receptors, endogenous ANG II stimulates EGF and inhibits clusterin expression.

Angiotensin II↗
PubMed1995

Angiotensin II regulates nephrogenesis and renal vascular development.

To test the hypothesis that angiotensin II (ANG II) is necessary for normal embryonic and postnatal kidney development, the effect of angiotensin receptor blockade or angiotensin converting enzyme inhibition on nephrovascular development was studied in newborn Sprague-Dawley rats and in Rana catesbeiana tadpoles undergoing prometamorphosis. Blockade of ANG II type 1 receptor (AT1) in newborn rats induced an arrest in nephrovascular maturation and renal growth, resulting in altered kidney architecture, characterized by fewer, thicker, and shorter afferent arterioles, reduced glomerular size and number, and tubular dilatation. Inhibition of ANG II generation in tadpoles induced even more marked developmental renal abnormalities. Blockade of ANG II type 2 receptor (AT2) in newborn rats did not alter renal growth or morphology. Results indicate that ANG II regulates nephrovascular development, a role that is conserved across species.

Angiotensin II↗
PubMed1995

Characterization of a synthetic peptide antibody recognizing rat kidney renin and prorenin.

A fourteen amino acid peptide from the cDNA predicted rat renin amino acid sequence was synthesized and used to produce a polyclonal antibody. The ability of this antibody to recognize rat renin was characterized and compared to that of a previously tested polyclonal anti-rat renin antibody raised against purified rat renin to verify its usefulness as a reagent in renin processing. The anti-renin peptide antibody and anti-rat renin antibody were evaluated for specificity of recognition of rat renin by immunohistochemical staining and immunoblotting. The ability of the anti-renin peptide antibody to inhibit renin enzymatic activity and to immunoprecipitate rat recombinant protein was also tested. Anti-renin peptide antibody and anti-rat renin antibody had identical recognition of rat renin and prorenin in immunohistochemical stains of rat kidney section and immunoblots. However the anti-renin peptide antibody does not inhibit renin enzymatic activity nor does it immunoprecipitate recombinant rat prorenin. We have produced and characterized an antirenin peptide antibody which recognizes denaturated rat renin and prorenin in a way that is identical to that of an antibody raised against native rat renin. The anti-renin peptide antibody does not recognize nondenatured renin because it did not inhibit renin enzymatic activity nor was it able to immunoprecipitate renin or prorenin. The technique of using synthetic peptides to produce antibody recognizing at renin is useful because low yields of native renin purified from rat kidney are generally insufficient for antibody production. This limits the widespread availability of antibodies that recognize rat renin.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1994

Neonatal ureteral obstruction stimulates recruitment of renin-secreting renal cortical cells.

Unilateral ureteral obstruction (UUO) in the neonate increases ipsilateral renal renin gene expression, an effect which is mediated by renal nerves. To determine whether neonatal UUO alters the number of renal cortical cells secreting renin and whether this change is modulated by renal nerve activity, newborn Sprague-Dawley rats were subjected to left UUO, right uninephrectomy, or sham operation and studied four weeks thereafter. To evaluate the importance of renal nerves in this response, an additional group of animals underwent chemical sympathectomy with guanethidine. Ureteral obstruction was associated with marked reduction in renal mass in the obstructed kidney and contralateral compensatory hypertrophy, changes which were not altered by sympathectomy. Renin messenger RNA and renal renin content were elevated in the obstructed kidney. The number of cells secreting renin, measured by the reverse hemolytic plaque assay, was markedly increased in the obstructed kidney (45 +/- 18 plaques/slide vs. 11 +/- 1 plaques/slide in sham animals), but not in the opposite kidney or following uninephrectomy. This effect was not significantly altered by sympathectomy. There was no change in the amount of renin secreted per cell or in the secretory response to Ca++. These results show that UUO results in recruitment of cells not previously secreting renin by a mechanism independent of renal nerve activity. This recruitment occurs without alteration of the quantity of renin secreted per cell or in the normal regulatory effect of Ca++ on renin secretion. An increase in the number of renin-secreting cells may contribute to the activation of the renin-angiotensin system, and thus to the vasoconstriction observed following ureteral obstruction.

Animals↗
PubMed1994

Neonatal renal function and physiology.

Birth rapidly changes the demands placed on the kidneys with respect to infant homeostasis. Conceptional age (gestational plus postnatal), general health, and medical management may independently, or, in concert, give rise to important metabolic abnormalities marked by apparent renal functional inadequacies. The chronology of the renal functional changes occurring with maturation in infants born before or at term is now well described. The confounding effects of treatment on the development of renal function in very low birth weight infants are also becoming more apparent. However, the mechanisms responsible for these changes are just becoming to be understood with the use of molecular biologic techniques.

Age Factors↗
PubMed1994

Bridging the gap between physiology and molecular biology: new approaches to perpetual questions.

Living organisms are the result of precise and complex associations of regulatory systems in which active biomolecules interact with one another and respond to the challenges of growth and development, alterations in the environment, and disease. Understanding of body homeostasis may be accomplished at various levels of scientific endeavor. Physiological research has brought about an enormous understanding of the fundamental principles that sustain life in health and disease. The field of molecular biology has provided new tools and strategies with which to examine physiological processes as viewed from the level of fundamental biomolecules. The integration of both fields as "molecular physiology" has provided the opportunity for another level of scientific understanding and the opening of new avenues of research. Renin is one such molecule that participates in the control of several diverse physiological responses including changes in blood pressure, fluid and electrolyte homeostasis, renal function, and perhaps some elements of growth and differentiation. Because of the authors' bias, this review article will use renin to introduce many of the techniques of molecular biology and illustrate the areas of ongoing and potential interdependent activities resulting in the emerging field of molecular physiology.

Animals↗
PubMed1994

Angiotensin II type 1 receptor: role in renal growth and gene expression during normal development.

To determine whether angiotensin II (ANG II) modulates renal growth and renin and angiotensin type 1 (AT1) gene expression via AT1 during development, weanling rats were given ANG II antagonist losartan (DuP 753) for 3 wk. Body weight (g), kidney weight (g), and kidney weight-to-body weight ratio were lower in losartan-treated rats (162 +/- 7, 1.6 +/- 0.06, and 9.5 +/- 0.1 x 10(-3)) than in control rats (184 +/- 5, 1.8 +/- 0.07, and 10.1 +/- 0.1 x 10(-3); P < 0.05). Renal DNA content (mg/kidney) was lower in losartan-treated (2.4 +/- 0.17) than in control rats (3.3 +/- 0.31; P < 0.05), whereas protein-to-DNA and RNA-to-DNA ratios were similar in losartan-treated and control rats. Renin mRNA levels were sevenfold higher in losartan-treated than in control rats, as determined by quantitative standardized dot blot analysis. In addition, blockade of AT1 with losartan induced recruitment of renin-synthesizing and renin-containing cells in the renal vasculature, as determined by immunocytochemistry and in situ hybridization. To establish whether AT1 blockade has a direct effect on renin gene expression, freshly isolated renin-producing cells were exposed in vitro to losartan (10(-6) M) or culture media (control). Losartan induced a twofold increase in steady-state renin mRNA levels above control (P < 0.05). Intrarenal AT1 mRNA levels were not altered by losartan given either in vivo or in vitro to freshly dispersed cells. To define whether immature renin-secreting cells are responsive to ANG II, renin release was determined by reverse hemolytic plaque assay.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗