scholarly journals Expression and Distribution of Heme Oxygenase-2 mRNA and Protein in Rat Kidney

1998 ◽  
Vol 46 (2) ◽  
pp. 249-256 ◽  
Author(s):  
Ying Hu ◽  
Ning Ma ◽  
Miao Yang ◽  
Reiji Semba
Keyword(s):  
Epithelial Cells ◽  
Heme Oxygenase ◽  
Cellular Localization ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Constitutive Expression ◽  
Renal Tubules ◽  
Thick Ascending Limb ◽  
Protection Assay ◽  
Rna Protection

Recent studies suggest that carbon monoxide (CO), which is formed by the enzyme heme oxygenase (HO) during the conversion of heme to biliverdin, shares some of the chemical and biological properties of nitric oxide (NO) and may play roles similar to those of NO. Heme oxygenase activity in the kidney has been reported for many years, and there are some reports on the expression of mRNA for two HO isozymes (HO-1 and HO-2) and cellular localization of HO-1 protein. However, cellular localization of HO-2 protein in the kidney under normal conditions has not been reported. In the present study we examined the expression and distribution of HO-2 mRNA and HO-2 protein in rat kidney using RNA protection assay and light and electron immunocytochemistry. RNA protection assay confirmed constitutive expression of HO-2 transcript in rat kidney. HO-2 immunoreactivity was selectively found in epithelial cells of the thick ascending limb and distal convoluted tubule, connecting tubule cells, and principal cells of the collecting duct. These results suggest that HO-2 is synthesized in the kidney and that HO-2 in the epithelial cells of renal tubules may serve as a source for CO generation under normal conditions.

Immunolocalization of AE2 anion exchanger in rat kidney

1997 ◽  
Vol 273 (4) ◽  
pp. F601-F614 ◽  
Author(s):  
Seth L. Alper ◽  
Alan K. Stuart-Tilley ◽  
Daniel Biemesderfer ◽  
Boris E. Shmukler ◽  
Dennis Brown
Keyword(s):  
Epithelial Cells ◽  
Anion Exchanger ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Sodium Dodecyl ◽  
Colchicine Treatment ◽  
Cell Types ◽  
Staining Intensity ◽  
Thick Ascending Limb ◽  
Renal Epithelial Cell

The cellular and subcellular localizations of the AE2 anion exchanger in rat kidney have remained elusive despite detection of moderately abundant AE2 mRNA and AE2 polypeptide in all kidney regions. In this report a simple epitope unmasking technique has allowed the immunolocalization of AE2 antigenic sites in basolateral membranes of several rat kidney tubular epithelial cells. AE2 immunostaining was faint or absent in the glomerulus and proximal tubule, present in descending and ascending thin limbs, and stronger in the medullary thick ascending limb (MTAL). A lower staining intensity was found in cortical thick ascending limbs and even less in the distal convoluted tubule. In contrast, there was an enhanced staining in the macula densa. In principal cells (PC) of the connecting segment, AE2 was undetectable but gradually increased in intensity along the collecting duct, with strongest staining in inner medullary collecting duct (IMCD) PC. A sodium dodecyl sulfate-sensitive AE2-related Golgi epitope was also detected in some interstitial and endothelial cells of the inner medulla and in epithelial cells of IMCD and MTAL. Colchicine treatment of the intact animal altered the distribution of this Golgi-associated epitope but left plasmalemmal AE2 undisturbed. Reverse transcription-polymerase chain reaction detected AE2a, AE2b, and AE2c2 but not AE2c1 transcripts in rat kidney mRNA. The results suggest a widespread occurrence of the AE2 protein in several renal epithelial cell types.

Monoaminergic innervation of the rat kidney: a quantitative study

1990 ◽  
Vol 259 (3) ◽  
pp. F503-F511 ◽  
Author(s):  
L. Barajas ◽  
K. Powers
Keyword(s):  
Collecting Duct ◽  
Rat Kidney ◽  
Sympathetic Innervation ◽  
Renal Tubules ◽  
Vascular Bundles ◽  
Thick Ascending Limb ◽  
High Concentration ◽  
Renal Vasculature ◽  
Indirect Measure ◽  
Afferent Arterioles

The sympathetic innervation of the renal tubules and vasculature was characterized by measuring the overlap of accumulations of autoradiographic grains (AAGs) on these structures in autoradiograms of kidney sections from rats injected with tritiated norepinephrine. AAG overlap was used as an indirect measure of the innervation of those structures. The renal vasculature showed x 4.5 more AAG overlap than observed on renal tubules. The greatest amount of AAG overlap occurred on afferent arterioles, followed by efferent arterioles, interlobular arteries, cortical capillaries, arcuate arteries, and renal veins. High concentration of AAGs occurred along the vascular bundles of the outer stripe. In the tubular nephron the proximal tubule had the greatest amount of AAG overlap, followed by the cortical thick ascending limb of Henle, the connecting tubule, the distal convoluted tubule, and the collecting duct. It was found that afferent arterioles had significantly higher mean density of AAG overlap than efferent arterioles for the superficial, midcortical, and juxtamedullary (vascular bundles excluded) renal cortex. There was consistently more AAG perimeter facing the interstitium than overlapping the vasculature. These observations, together with the ultrastructural distribution of synaptic vesicles in varicosities, suggest that the interstitium might be an additional pathway of neurotransmitter access to the effector structures.

scholarly journals The sodium-activated sodium channel is expressed in the rat kidney thick ascending limb and collecting duct cells and is upregulated during high salt intake

2012 ◽  
Vol 303 (1) ◽  
pp. F105-F109 ◽  
Author(s):  
Lucienne S. Lara ◽  
Ryousuke Satou ◽  
Camille R. T. Bourgeois ◽  
Alexis A. Gonzalez ◽  
Andrea Zsombok ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Salt Intake ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Extracellular Fluid ◽  
High Salt ◽  
Thick Ascending Limb ◽  
Principal Cells ◽  
High Salt Intake

Increased dietary salt triggers oxidative stress and kidney injury in salt-sensitive hypertension; however, the mechanism for sensing increased extracellular Na+ concentration ([Na+]) remains unclear. A Na+-activated Na+ channel (Na sensor) described in the brain operates as a sensor of extracellular fluid [Na+]; nonetheless, its presence in the kidney has not been established. In the present study, we demonstrated the gene expression of the Na sensor by RT-PCR and Western blotting in the Sprague-Dawley rat kidney. Using immunofluorescence, the Na sensor was localized to the luminal side in tubular epithelial cells of collecting ducts colocalizing with aquaporin-2, a marker of principal cells, and in thick ascending limb, colocalizing with the glycoprotein Tamm-Horsfall. To determine the effect of a high-salt diet (HSD) on Na sensor gene expression, we quantified its transcript and protein levels primarily in renal medullas from control rats and rats subjected to 8% NaCl for 7 days ( n = 5). HSD increased Na sensor expression levels (mRNA: from 1.2 ± 0.2 to 5.1 ± 1.3 au; protein: from 0.98 ± 0.15 to 1.74 ± 0.28 au P < 0.05) in the kidney medulla, but not in the cortex. These data indicate that rat kidney epithelial cells of the thick ascending limb and principal cells of the collecting duct possess a Na sensor that is upregulated by HSD, suggesting an important role in monitoring changes in tubular fluid [Na+].

RT-PCR microlocalization of mRNA for guanylyl cyclase-coupled ANF receptor in rat kidney

1991 ◽  
Vol 261 (6) ◽  
pp. F1080-F1087 ◽  
Author(s):  
Y. Terada ◽  
T. Moriyama ◽  
B. M. Martin ◽  
M. A. Knepper ◽  
A. Garcia-Perez
Keyword(s):  
Guanylyl Cyclase ◽  
Renal Tubule ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Renal Tubules ◽  
Thick Ascending Limb ◽  
Rt Pcr ◽  
Vasa Recta ◽  
Vascular Elements ◽  
Medullary Collecting Duct

Microlocalization of mRNA coding for the guanylyl cyclase-coupled atrial natriuretic factor (ANF) receptor was carried out in the rat kidney. We used a combination of reverse transcription and polymerase chain reaction (RT-PCR) in individual microdissected renal tubule segments, glomeruli, and vasa recta bundles. Relative quantitation of the resulting amplified cDNA utilized densitometry of autoradiograms from Southern blots probed with a specific 32P-labeled probe. Among renal tubule segments, the largest signal was found in the terminal inner medullary collecting duct (IMCD). Slightly smaller signals were found in the initial IMCD and in loop of Henle segments from the inner medulla. Readily detectable signals were also seen in the following segments (in descending order): cortical collecting duct, proximal convoluted tubule, medullary thick ascending limb, cortical thick ascending limb, distal convoluted tubule, and outer medullary collecting duct. Large signals were also detected in glomeruli and in vasa recta bundles from the inner stripe of the outer medulla. Based on these results, we conclude that 1) renal microlocalization of specific mRNAs coding for hormone receptors is feasible through application of the RT-PCR procedure in microdissected renal tubules and vascular elements, and 2) the gene for the guanylyl cyclase-coupled ANF receptor is broadly expressed along the nephron, raising the possibility that multiple sites of ANF action are present.

Expression and cellular localization of mRNA encoding the "gastric" isoform of H(+)-K(+)-ATPase alpha-subunit in rat kidney

1995 ◽  
Vol 268 (1) ◽  
pp. F99-F109 ◽  
Author(s):  
K. Y. Ahn ◽  
B. C. Kone
Keyword(s):  
Cellular Localization ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Molecular Probes ◽  
Alpha Subunit ◽  
Thick Ascending Limb ◽  
Rt Pcr ◽  
Direct Demonstration ◽  
Specific Hybridization ◽  
Pcr Products

The distribution of transcripts encoding the gastric H(+)-K(+)-adenosinetriphosphatase (ATPase) alpha-subunit in the normal rat kidney was studied by reverse transcription-polymerase chain reaction (RT-PCR), combined with DNA sequence analysis and renal microdissection, and by nonradioactive in situ hybridization of fixed kidney sections using highly specific molecular probes. RT-PCR products corresponding to the gastric H(+)-K(+)-ATPase alpha-subunit were detected in the cortex, outer and inner medulla, and in isolated cortical (CCD) and inner medullary collecting ducts (IMCD). With digoxigenin-labeled cRNAs derived from the 5' and 3' ends of the gastric H(+)-K(+)-ATPase alpha-subunit cDNA, specific hybridization signal was detected prominently in all the cells of the connecting segment and CCD, the intercalated cells of the outer medullary collecting duct, the IMCD, and the renal pelvic epithelium lining the secondary pouches. Weak labeling was noted in the S3 segment of the proximal tubule, the distal convoluted tubule, and the cortical thick ascending limb of Henle. Hybridization with the sense probes produced no cellular labeling. These data provide the first direct demonstration for the expression and cellular distribution of mRNA encoding the gastric H(+)-K(+)-ATPase alpha-subunit in the normal, potassium-replete kidney, and they provide essential tools for the molecular analysis of renal acid base and potassium transport under physiological and pathophysiological conditions.

Prostanoid signaling, localization, and expression of IP receptors in rat thick ascending limb cells

1998 ◽  
Vol 275 (6) ◽  
pp. F904-F914 ◽  
Author(s):  
Richard L. Hébert ◽  
Tim O’Connor ◽  
Chris Neville ◽  
Kevin D. Burns ◽  
Odette Laneuville ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Rat Kidney ◽  
Receptor Expression ◽  
Molecular Evidence ◽  
Thick Ascending Limb ◽  
Renal Epithelial Cells ◽  
Receptor Mrna ◽  
Ip Receptor ◽  
Receptor Cdna ◽  
Nucleotide Binding Proteins

It is widely held that only one prostacyclin (IP) receptor exists that can couple to guanine stimulatory nucleotide binding proteins (Gs) leading to activation of adenyl cyclase. Although IP receptor mRNA is expressed in vascular arterial smooth muscle cells and platelets, with lower level expression in mature thymocytes, splenic lymphocytes, and megakaryocytes, there is no molecular evidence for IP receptor expression in renal epithelial cells. The purpose of the present study was to obtain molecular evidence for the expression and localization of the IP receptor and to study the signaling pathways of IP receptor in rat medullary thick ascending limb (MTAL). Biochemical studies showed that IP prostanoids do not increase cAMP in rat MTAL. However, in the presence of vasopressin, inhibition of cAMP formation by prostacyclin (PGI2) analogs is pertussis toxin sensitive and does not activate protein kinase C. In situ hybridization studies localized IP receptor mRNA expression to MTAL in the rat kidney outer medulla. The results of RT-PCR of freshly isolated RNA from MTAL, with primers specific for the mouse IP receptor cDNA, produced an amplification product of the correct predicted size that contained an expected Nco I endonuclease restriction site. We conclude that rat renal epithelial cells express the IP receptor, coupled to inhibition of cAMP production.

Intrarenal and Cellular Localization of CLC-K2 Protein in the Mouse Kidney

2001 ◽  
Vol 12 (7) ◽  
pp. 1327-1334 ◽  
Author(s):  
KATSUKI KOBAYASHI ◽  
SHINICHI UCHIDA ◽  
SHUKI MIZUTANI ◽  
SEI SASAKI ◽  
FUMIAKI MARUMO
Keyword(s):  
Cellular Localization ◽  
Collecting Duct ◽  
Type A ◽  
Mouse Kidney ◽  
Thick Ascending Limb ◽  
Intercalated Cells ◽  
Basolateral Membranes ◽  
Nephron Segments ◽  
Henle's Loop ◽  
Distal Tubules

Abstract. CLC-K2, a kidney-specific member of the CLC chloride channel family, is thought to play an important role in the transepithelial Cl- transport in the kidney. This consensus was first reached shortly after it was demonstrated that the mutations of the human CLCNKB gene resulted in Bartter's syndrome type III. To clarify the pathogenesis, the exact intrarenal and cellular localization of CLC-K2 by immunohistochemistry of the Clcnk1-/- mouse kidney were investigated by use of an anti-CLC-K antibody that recognized both CLC-K1 and CLC-K2. CLC-K2 is expressed in the thick ascending limb of Henle's loop and distal tubules, where it is localized to the basolateral membranes. The localization of CLC-K2 to these nephron segments strongly implies that CLC-K2 confers the basolateral chloride conductance in the thick ascending limb of Henle's loop and distal tubules, where Cl- is taken up by the bumetanide-sensitive Na-K-2Cl cotransporter or the thiazide-sensitive Na-Cl cotransporter at the apical membranes. CLC-K2 expression was also shown to extend into the connecting tubule in the basolateral membrane. CLC-K2 was found in basolateral membranes of the type A intercalated cells residing along the collecting duct. This localization strongly suggests that CLC-K2 confers the basolateral conductance in the type A intercalated cells where Cl- is taken up by the anion exchanger in exchange for HCO3- at the basolateral membranes. These aspects of CLC-K2 localization suggest that CLC-K2 is important in Cl- transport in the distal nephron segments.

scholarly journals Localization of phosphatidylinositol phosphate kinase IIγ in kidney to a membrane trafficking compartment within specialized cells of the nephron

2008 ◽  
Vol 295 (5) ◽  
pp. F1422-F1430 ◽  
Author(s):  
Jonathan H. Clarke ◽  
Piers C. Emson ◽  
Robin F. Irvine
Keyword(s):  
Epithelial Cells ◽  
Membrane Trafficking ◽  
Kidney Cell ◽  
Collecting Duct ◽  
Thick Ascending Limb ◽  
Phosphatidylinositol Phosphate ◽  
The Matrix ◽  
Phosphatidylinositol 4

PIP4Ks (type II phosphatidylinositol 4-phosphate kinases) are phosphatidylinositol 5-phosphate (PtdIns5P) 4-kinases, believed primarily to regulate cellular PtdIns5P levels. In this study, we investigated the expression, localization, and associated biological activity of the least-studied PIP4K isoform, PIP4Kγ. Quantitative RT-PCR and in situ hybridization revealed that compared with PIP4Kα and PIP4Kβ, PIP4Kγ is expressed at exceptionally high levels in the kidney, especially the cortex and outer medulla. A specific antibody was raised to PIP4Kγ, and immunohistochemistry with this and with antibodies to specific kidney cell markers showed a restricted expression, primarily distributed in epithelial cells in the thick ascending limb and in the intercalated cells of the collecting duct. In these cells, PIP4Kγ had a vesicular appearance, and transfection of kidney cell lines revealed a partial Golgi localization (primarily the matrix of the cis-Golgi) with an additional presence in an unidentified vesicular compartment. In contrast to PIP4Kα, bacterially expressed recombinant PIP4Kγ was completely inactive but did have the ability to associate with active PIP4Kα in vitro. Overall our data suggest that PIP4Kγ may have a function in the regulation of vesicular transport in specialized kidney epithelial cells.

Regional and segmental localization of AE2 anion exchanger mRNA and protein in rat kidney

1995 ◽  
Vol 269 (4) ◽  
pp. F461-F468 ◽  
Author(s):  
F. C. Brosius ◽  
K. Nguyen ◽  
A. K. Stuart-Tilley ◽  
C. Haller ◽  
J. P. Briggs ◽  
...  
Keyword(s):  
Collecting Duct ◽  
Rat Kidney ◽  
Chloride Concentration ◽  
Transepithelial Transport ◽  
Thick Ascending Limb ◽  
Cortical Collecting Duct ◽  
Base Exchange ◽  
Medullary Thick Ascending Limb ◽  
Nephron Segment ◽  
Medullary Collecting Duct

Chloride/base exchange activity has been detected in every mammalian nephron segment in which it has been sought. However, in contrast to the Cl-/HCO3- exchanger AE1 in type A intercalated cells, localization of AE2 within the kidney has not been reported. We therefore studied AE2 expression in rat kidney. AE2 mRNA was present in cortex, outer medulla, and inner medulla. Semiquantitative polymerase chain reaction of cDNA from microdissected tubules revealed AE2 cDNA levels as follows [copies of cDNA derived per mm tubule (+/- SE)]: proximal convoluted tubule, 688 +/- 161; proximal straight tubule, 652 +/- 189; medullary thick ascending limb, 1,378 +/- 226; cortical thick ascending limb, 741 +/- 24; cortical collecting duct, 909 +/- 71; and outer medullary collecting duct, 579 +/- 132. AE2 cDNA was also amplified in thin limbs and in inner medullary collecting duct. AE2 polypeptide was detected in all kidney regions. AE2 mRNA and protein were also detected in several renal cell lines. The data are compatible with the postulated roles of AE2 in maintenance of intracellular pH and chloride concentration and with its possible participation in transepithelial transport.

Functional Expression of Human Heme Oxygenase-1 Gene in Renal Structure of Spontaneously Hypertensive Rats

2003 ◽  
Vol 228 (5) ◽  
pp. 454-458 ◽  
Author(s):  
Alvin I. Goodman ◽  
Shou Quan ◽  
Liming Yang ◽  
Arika Synghal ◽  
Nader G. Abraham
Keyword(s):  
Blood Pressure ◽  
Heme Oxygenase ◽  
Spontaneously Hypertensive Rats ◽  
Rat Kidney ◽  
Functional Expression ◽  
Heme Oxygenase 1 ◽  
Thick Ascending Limb ◽  
Bile Pigments ◽  
Hypertensive Rats ◽  
Spontaneously Hypertensive

Heme oxygenase (HO), by catabolizing heme to bile pigments, regulates the levels and activity of cellular hemoprotein and HO activity. We examined the effect of delivery of the human HO-1 gene on cellular heme in renal tissue using a retroviral vector. We used a single intracardiac injection of the concentrated infectious viral particles in 5-day-old spontaneously hypertensive rats; 25 were transduced with empty vector and 25 were transduced with the human HO-1 gene. Functional expression of human and rat HO-1 was measured after 2 and 4 weeks. Reverse transcription polymerase chain reaction showed that human HO-1 mRNA was expressed as early as 2 weeks, with the highest levels in the kidney. Western blot analysis showed distribution of human HO-1 protein in rat kidney structures, predominantly in the thick ascending limb of the loop of Henle as well as in proximal tubules and preglomerular arterioles. These areas also demonstrated higher HO activity as measured by increased conversion of heme to bilirubin and carbon monoxide. Functional expression of the human HO-1 gene was associated with a decrease in blood pressure in 4- and 8-week-old spontaneously hypertensive rats. Compared with nontransduced rats, human HO-1 gene overexpression in transduced rats was associated with a 35% decrease in urinary 20-hydroxyeicosatetraenoic acid, a potent vasoconstrictor and an inhibitor of tubular Na+ transport, which may be related to the decrease in blood pressure.

Sign in / Sign up

Export Citation Format

Share Document