bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of chloride channel blockers on rat renal vascular responses to angiotensin II and norepinephrine
Authors: Steendahl Joen , Holstein-Rathlou Niels-Henrik , Sorensen Charlotte Mehlin , Salomonsson Max
Published: January 2004

The aim of the present study was to investigate the role of Ca2+-activated Cl-channels in the renal vasoconstriction elicited by angiotensin II (ANG II) and norepinephrine (NE). Renal blood flow (RBF) was measured in vivo using electromagnetic flowmetry. Ratiometric photometry of fura 2 fluorescence was used to estimate intracellular free Ca2+concentration ([Ca2+]i) in isolated preglomerular vessels from rat kidneys. Renal arterial injection of ANG II (2-4 ng) and NE (20-40 ng) produced a transient decrease in RBF. Administration of ANG II (10-7M) and NE (5 × 10-6M) to the isolated preglomerular vessels caused a prompt increase in [Ca2+]i. Renal preinfusion of DIDS (0.6 and 1.25 μmol/min) attenuated the ANG II-induced vasoconstriction to ∼35% of the control response, whereas the effects of NE were unaltered. Niflumic acid (0.14 and 0.28 μmol/min) and 2-[(2-cyclopentenyl-6,7-dichloro-2,3-dihydro-2-methyl-1-oxo-1 H-inden-5-yl)oxy]acetic acid (IAA-94; 0.045 and 0.09 μmol/min) did not affect the vasoconstrictive responses of these compounds. Pretreatment with niflumic acid (50 μM) or IAA-94 (30 μM) for 2 min decreased baseline [Ca2+]ibut did not change the magnitude of the [Ca2+]iresponse to ANG II and NE in the isolated vessels. The present results do not support the hypothesis that Ca2+-activated Cl-channels play a crucial role in the hemodynamic effects of ANG II and NE in rat renal vasculature.
Greater tolerance of renal medullary cells for a slow increase in osmolality is associated with enhanced expression of HSP70 and other osmoprotective genes
Authors: Cai Qi , Ferraris Joan D. , Burg Maurice B.
Published: January 2004

In tests of osmotic tolerance of renal inner medullary cells in tissue culture, osmolality has usually been increased in a single step, whereas in vivo the increase occurs gradually over several hours. We previously found that more passage 2 mouse inner medullary epithelial (p2mIME) cells survive a linear increase in NaCl and urea from 640 to 1,640 mosmol/kgH2O over 20 h (which is similar to the change that may occur in vivo) than they do a step increase. The present studies examine accompanying differences in gene expression. Among mRNAs of genes known to be protective, tonicity-responsive enhancer binding protein and aldose reductase increase with a linear but decrease with a step increase; betaine transporter BGT1 decreases with a step but not a linear increase; heat shock protein 70.1 ( HSP70.1) and HSP70.3 increase more with a linear than a step increase; and osmotic stress protein 94 and heme oxygenase-1 increase with a linear but decrease with a step increase. mRNAs for known urea-responsive proteins, GADD153 and Egr-1, increase with both a step and linear increase. A step increase in urea alone reduces mRNAs, similar to the combination of NaCl and urea, but a step increase in NaCl alone does not. HSP70 protein increases substantially with a linear rise in osmolality but does not change significantly with a step rise. We speculate that poorer survival of p2mIME cells with a step than with linear increase in NaCl and urea is accounted for, at least in part, by urea-induced suppression of protective genes, particularly HSP70.
Dietary salt intake modulates progression of antithymocyte serum nephritis through alteration of glomerular angiotensin II receptor expression
Authors: Suzuki Hiroyuki , Yamamoto Tatsuo , Ikegaya Naoki , Hishida Akira
Published: January 2004

Dietary salt intake modulates the renin-angiotensin system (RAS); however, little is known about the effect of salt intake on the progression of glomerulonephritis. We investigated the glomerular expression of TGF-β1type I (TβRI) and II (TβRII) TGF-β receptors and RAS components in rats with antithymocyte serum (ATS) nephritis on normal (NSI)-, low (LSI)-, and high-salt intake (HSI) and on HSI rats receiving candesartan cilexetil (CC) and LSI rats receiving PD-123319. Glomerular lesions were less severe in rats on LSI and aggravated in those on HSI compared with those on NSI. Intrarenal renin and glomerular ANG II levels were significantly higher in LSI and lower in HSI rats. In ATS nephritis, HSI increased glomerular TβRI, TβRII, and ANG II type 1 receptor (AT1R), and decreased glomerular ANG II type 2 receptor (AT2R), whereas LSI decreased glomerular TGF-β1and TβRI and increased glomerular AT2R. CC ameliorated glomerular lesions, reduced glomerular TGF-β1and TβRII, and increased glomerular AT2R. PD-123319 aggravated glomerular lesions and increased glomerular TGF-β1and TβRII. Our results suggest that dietary salt intake influences progression of ATS nephritis by modulating glomerular TGF-β1and TβR expression resulting, at least in part, from altered glomerular AT1R and AT2R expression.
Interaction of the metal chelator DMPS with OAT1 and OAT3 in intact isolated rabbit renal proximal tubules
Authors: Lungkaphin A. , Chatsudthipong V. , Evans K. K. , Groves C. E. , Wright S. H. , Dantzler W. H.
Published: January 2004

2,3-Dimercapto-1-propanesulfonic acid (DMPS) is used clinically to increase urinary excretion of heavy metals, including mercury and arsenic. We used single S2 segments and suspensions of rabbit renal proximal tubules (RPT) to test the interaction of this anionic heavy metal chelator with basolateral transporters OAT1 and OAT3. RTPCR revealed expression of both transporters in single S2 segments. [3H]PAH and3H-labeled estrone sulfate ([3H]ES) were used as specific substrates for rbOAT1 and rbOAT3, respectively. PAH and ES were transported into nonperfused single RPT segments with Ktvalues of 67 ± 20 and 3.4 ± 1.2 μM, respectively, and into tubule suspensions with Ktvalues of 58 ± 17 and 7.7 ± 2.1 μM, respectively. Reduced DMPS (DMPSH) inhibited uptake of both substrates into single tubule segments with Kappvalues of 405 ± 49 μM (for [3H]PAH) and 320 ± 66 μM (for [3H]ES). Oxidized DMPS (DMPSS), the prevalent form in the blood, also inhibited uptakes of [3H]PAH ( Kappof 766 ± 190 μM) and [3H]ES (696 ± 166 μM). Inward gradients of ES, DMPSH, and DMPSS trans-stimulated the 30-s efflux of preloaded [3H]ES across the basolateral membrane of RPT. Similarly, DMPSH, and PAH itself, trans-stimulated the 15-s efflux of [3H]PAH. In contrast, efflux of [3H]PAH was inhibited by the presence of DMPSS in the bathing medium. These data suggest that, whereas both OAT1 and OAT3 probably transport DMPSH, DMPSS transport may be limited to OAT3. This is the first evidence showing that both OAT1 and OAT3 can transport DMPS across the basolateral membrane of RPT.
Vitamin D3upregulates plasma membrane Ca2+-ATPase expression and potentiates apico-basal Ca2+flux in MDCK cells
Authors: Kip Sertac N. , Strehler Emanuel E.
Published: January 2004

Plasma membrane Ca2+-ATPases (PMCAs) are a ubiquitous system for the expulsion of Ca2+from eukaryotic cells. In tight monolayers of polarized Madin-Darby canine kidney (MDCK) cells representing a distal kidney tubule model, PMCAs are responsible for about one-third of the vectorial Ca2+transport under resting conditions, with the remainder being provided by the Na+/Ca2+exchanger. Vitamin D3(VitD) is known to increase PMCA expression and activity in Ca2+-transporting tissues such as the intestine, as well as in osteoblasts and Madin-Darby bovine kidney epithelial cells. We found that VitD upregulated the expression of the PMCAs (mainly PMCA4b) in MDCK cell lysates at the RNA and protein level in a time- and dose-dependent manner. Interestingly, VitD caused a decrease of the PMCAs in the apical plasma membrane fraction and a concomitant increase of the pumps in the basolateral membrane. Functional studies demonstrated that transcellular45Ca2+flux from the apical-to-basolateral compartment was significantly enhanced by VitD. These findings demonstrate that VitD is a positive regulator of the PMCAs in MDCK epithelial cells. The correlation of decreased apical/increased basolateral expression of the PMCAs with an increase in transcellular Ca2+flux from the apical (urine) toward the basolateral (blood) compartment indicates the physiological relevance of VitD function in kidney tubular Ca2+reabsorption.
Chronic nitric oxide synthase inhibition exacerbates renal dysfunction in cirrhotic rats
Authors: Græbe Martin , Brønd Lone , Christensen Sten , Nielsen Søren , Olsen Niels V. , Jonassen Thomas E. N.
Published: January 2004

The present study investigated sodium balance and renal tubular function in cirrhotic rats with chronic blockade of the nitric oxide (NO) system. Rats were treated with the nonselective NO synthase inhibitor NG-nitro-l-arginine methyl ester (l-NAME) starting on the day of common bile duct ligation (CBL). Three weeks of daily sodium balance studies showed that CBL rats developed sodium retention compared with sham-operated rats and that l-NAME treatment dose dependently deteriorated cumulative sodium balance by reducing urinary sodium excretion. Five weeks after CBL, renal clearance studies were performed, followed by Western blotting of the electroneutral type 3 sodium/proton exchanger (NHE3) and the Na-K-ATPase present in proximal tubules. Untreated CBL rats showed a decreased proximal reabsorption with a concomitant reduction of NHE3 and Na-K-ATPase levels, indicating that tubular segments distal to the proximal tubules were responsible for the increased sodium reabsorption. l-NAME-treated CBL rats showed an increased proximal reabsorption measured by the lithium clearance method and showed a marked increase in NHE3 and Na-K-ATPase protein levels. Our results show that chronic l-NAME treatment exacerbates the sodium retention found in CBL rats by a significant increase in proximal tubular reabsorption.
Low endogenous glucocorticoid allows induction of kidney cortical cyclooxygenase-2 during postnatal rat development
Authors: Madsen Kirsten , Stubbe Jane , Yang Tianxin , Skøtt Ole , Bachmann Sebastian , Jensen Boye L.
Published: January 2004

In postnatal weeks 2–4, cyclooxygenase-2 (COX-2) is induced in the rat kidney cortex where it is critically involved in final stages of kidney development. We examined whether changes in circulating gluco- or mineralocorticosteroids or in their renal receptors regulate postnatal COX-2 induction. Plasma corticosterone concentration peaked at birth, decreased to low levels at days 3- 13, and increased to adult levels from day 22. Aldosterone peaked at birth and then stabilized at adult levels. Gluco- and mineralocorticoid receptor (GR and MR) mRNAs were expressed stably in kidney before, during, and after COX-2 induction. 11β-Hydroxysteroid dehydrogenase 2 was induced shortly after birth and was widely distributed in the whole collecting duct system in the suckling period and then returned to an adult pattern. Supplementation with corticosterone (20 mg·kg-1·day-1) or GR-specific dexamethasone (1 mg·kg-1·day-1) during low endogenous corticosterone suppressed renal COX-2 mRNA and protein and led to a restricted distribution of COX-2 immunolabeling. The ability of glucocorticoids to affect COX-2 was reflected in colocalization of GR-α and COX-2 immunoreactivity and mRNAs in thick ascending limb of Henle's loop. The MR antagonist potassium canrenoate (20 mg·kg-1·day-1) enhanced COX-2 expression from days 5 to 10, but low MR-specific concentrations of DOCA (1 mg·kg-1·day-1) had no effect on COX-2. Renomedullary interstitial cells expressed GR-α and COX-2. Dexamethasone suppressed COX-2 in these cells. Thus low plasma concentrations of corticosterone allowed for cortical and medullary COX-2 induction during postnatal kidney development. Increased circulating glucocorticoid in the postnatal period may damage late renal development through inhibition of COX-2.
Local anesthetics worsen renal function after ischemia-reperfusion injury in rats
Authors: Lee H. Thomas , Krichevsky Igor E. , Xu Hua , Ota-Setlik Ayuko , D'Agati Vivette D. , Emala Charles W.
Published: January 2004

Local anesthetics are widely used during the perioperative period, even in patients with preexisting renal disease. However, local anesthestics have been shown to cause cell death in multiple cell lines, including human kidney proximal tubule cells. We questioned whether local anesthetics potentiate renal dysfunction after ischemia-reperfusion (I/R) injury in vivo. Rats were implanted with subcutaneous miniosmotic pumps that continuously delivered lidocaine (2 mg·kg-1·h-1), bupivacaine (0.4 mg·kg-1·h-1), tetracaine (1 mg·kg-1·h-1), or saline vehicle, and 6 h later the rats were subjected to 30 min of renal ischemia or to sham operation. Renal function was assessed by measurement of plasma creatinine at 24 and 48 h after renal I/R injury in the presence or absence of chronic infusions of local anesthetics and correlated to histological changes indicative of necrosis. The degree of renal apoptosis was assessed by three methods: 1) DNA fragmentation detected by terminal deoxynucleotidyl transferase biotin-dUTP nick-end labeling staining, 2) DNA laddering detected after agarose gel electrophoresis, and 3) morphological identification of apoptotic tubules at the corticomedullary junction. We also measured the expression of the proinflammatory markers ICAM-1 and TNF-α. Continuous local anesthetic infusion with renal I/R injury resulted in an increased magnitude and duration of renal dysfunction compared with the saline-infused I/R group. Additionally, both apoptotic and necrotic renal cell death as well as inflammatory changes were significantly potentiated in local anesthetic-treated rat kidneys. Local anesthetic infusion alone without I/R injury had no effect on renal function. We conclude that local anesthetics potentiated renal injury after I/R by increasing necrosis, apoptosis, and inflammation.
Angiotensin II-dependent increased expression of Na+-glucose cotransporter in hypertension
Authors: Bautista Rocío , Manning Rebeca , Martinez Flavio , del Carmen Avila-Casado Maria , Soto Virginia , Medina Armando , Escalante Bruno
Published: January 2004

Glucose uptake is increased in hypertension. Thus we investigated Na+-glucose cotransporter (SGLT2) activity and expression in proximal tubules from renovascular hypertensive rats. Sham-operated rats, aortic coarctation rats, and aortic coarctation rats treated with either ramipril (2.5 mg·kg-1·day-1for 21 days) or losartan (10 mg·kg-1·day-1for 21 days) were used. Na+-dependent glucose uptake was measured in brush-border membrane vesicles (BBMV). Vmaxin BBMV from hypertensive rats was greater compared with those from normotensive rats (3 ± 0.2 vs. 1.5 ± 0.1 nmol·mg protein-1·min-1) without a change in Km. Renal immunostaining was greater, and Western blot analysis and RT-PCR showed a higher expression of SGLT2 in hypertensive rats than in normotensive rats (1,029 ± 71 vs. 5,003 ± 292, 199 ± 15 vs. 95 ± 10, and 1.4 ± 0.2 vs. 0.3 ± 0.1 arbitrary units, respectively). In rats treated with either ramipril or losartan, Vmaxdecreased to 2.1 ± 0.3 and 1.8 ± 0.4 nmol·mg protein-1·min-1, respectively, as well as did the intensity of immunostaining and levels of protein and mRNA. We suggest that in renovascular hypertension, angiotensin II induced SGLT2 via the AT1receptor, which was evidenced at both the functional and expression levels, probably contributing to increased absorption of Na+and thereby to the development or maintenance of hypertension.
Arginine vasopressin stimulates H+-ATPase in MDCK cells via V1(cell Ca2+) and V2(cAMP) receptors
Authors: Oliveira-Souza Maria , Musa-Aziz Raif , Malnic Gerhard , de Mello Aires Margarida
Published: January 2004

The effect of arginine vasopressin (AVP) and/or atrial natriuretic peptide (ANP) on the regulation of intracellular pH (pHi) via H+-ATPase and of cytosolic calcium ([Ca2+]i) was investigated in Madin-Darby canine kidney (MDCK) cells by the fluorescent probes BCECF-AM and fluo-4-AM, respectively. The pHirecovery rate was examined after intracellular acidification following an NH4Cl pulse, in the presence of zero Na+plus Schering 28080 (a specific inhibitor of H+-K+-ATPase). AVP (10-12-10-6M) increased the rate of pHirecovery and [Ca2+]iin a dose-dependent manner. V1- or V2-receptor antagonists impaired the effect of AVP on both processes, and DDAVP (10-12-10-6M; a V2-selective agonist) caused a dose-dependent stimulation of them. [Ca2+]ior cAMP (as increased by 10-5M thapsigargin or 8-BrcAMP, respectively) alone had no effect on H+-ATPase, but their synergic action was necessary to stimulate H+-ATPase. In agreement with these findings, ANP (10-6M) or dimethyl-BAPTA-AM (5 × 10-5M), impairing the increase of [Ca2+]iin response to AVP, blocks the stimulatory effect of AVP on H+-ATPase.
Antidiuretic hormone resistance in the neonatal cortical collecting tubule is mediated in part by elevated phosphodiesterase activity
Authors: Quigley Raymond , Chakravarty Sumana , Baum Michel
Published: January 2004

Neonates cannot concentrate their urine to the same degree as adults. One of the key factors in concentrating the urine is the renal collecting duct osmotic water permeability ( Pf) response to antidiuretic hormone (ADH). Neonatal cortical collecting ducts have a blunted Pf response to ADH compared with adult tubules ( Pf: 119.0 ± 12.5 vs. 260.1 ± 29.5 μm/s, P < 0.05). We found that the phosphodiesterase activity in the neonatal collecting ducts was higher than that in the adult collecting ducts (3,970 ± 510 vs. 2,440 ± 220 cpm·μg tubular protein-1·20 min-1, P < 0.05). After pretreatment of in vitro microperfused tubules with the nonspecific phosphodiesterase inhibitor IBMX (10-6 M in the bath), the Pf response to ADH in neonatal collecting ducts was 271.4 ± 51.7 μm/s, which was identical to that of the adult collecting duct [315.3 ± 31.3 μm/s, P = not significant (NS)]. Rolipram, a specific type IV phosphodiesterase inhibitor, lowered the elevated phosphodiesterase activity in the neonatal tubules to that in the adult tubules (2,460 ± 210 vs. 2,160 ± 230 cpm·μg tubular protein-1·20 min-1, P = NS). Neonatal tubules pretreated with rolipram (10-5 M) in the bath also had a Pf response to ADH that was comparable to that of the adult tubules (258.2 ± 17.0 vs. 271.4 ± 32.6 μm/s, P = NS). Thus the elevated phosphodiesterase activity in the neonatal tubules appears to be due to an increase in type IV phosphodiesterase activity. Hence, one of the key factors in the decreased ability of neonates to concentrate their urine is overactivity of phosphodiesterase in the cortical collecting duct that blunts the neonatal collecting duct Pf response to ADH.
Development of an experimental system for the study of urethral biomechanical function
Authors: Jankowski Ron J. , Prantil Rachelle L. , Fraser Matthew O. , Chancellor Michael B. , de Groat William C. , Huard Johnny , Vorp David A.
Published: January 2004

Despite its principal mechanical function in the storage and release of urine, the biomechanical properties of the urethra have remained largely unexplored. The purpose of this study was to develop and validate an experimental model that can be used for evaluating whole urethral tissue in such a manner. Bladder-urethral specimens were excised from halothane-anesthetized female rats and mounted at in vivo length within the experimental apparatus consisting of a tissue perfusion chamber, an adjustable fluid column, and a laser micrometer. Outer diameter measurements were made at proximal, mid, and distal axial locations in response to increases in intraluminal pressure and after addition of various muscle-responsive agents. Basal smooth muscle tone and regional variations in compliance were detected through pressure-diameter responses. Chemically evoked contractile responses were measured and correspond to regional compositions of intrinsic smooth and striated muscle components. The results presented illustrate the utility of this system, which should permit a more thorough characterization of structure-function relationships and urethral biomechanical function in relation to normal and dysfunctional tissue states.
Stanniocalcin-1, an inhibitor of macrophage chemotaxis and chemokinesis
Authors: Kanellis John , Bick Roger , Garcia Gabriela , Truong Luan , Tsao Chun Chui , Etemadmoghadam Dariush , Poindexter Brian , Feng Lili , Johnson Richard J. , Sheikh-Hamad David
Published: January 2004

In macrophages, changes in intracellular calcium have been associated with activation of cellular processes that regulate cell adhesion and motility and are important for the response of macrophages to antigenic stimuli. The mammalian counterpart of the fish calcium-regulating hormone stanniocalcin-1 (STC1) is expressed in multiple organs including the thymus and spleen, and hence, we hypothesized that it may have a role in modulating the immune/inflammatory response. Using murine macrophage-like (RAW264.7) and human monoblast-like (U937) cells to study chemotaxis in vitro, we found that STC1 attenuated chemokinesis and diminished the chemotactic response to monocyte chemotactic protein-1 (MCP-1) and stromal cell-derived factor-1α. Consistent with these findings, STC1 blunted the rise in intracellular calcium following MCP-1 stimulation in RAW264.7 cells. In vivo studies suggested differential expression of STC1 in obstructed kidney and localization to macrophages. MCP-1 and STC1 transcripts were both upregulated following ureteric obstruction, suggesting a functional association between the two genes. Our data suggest a role for mammalian STC1 in modulating the immune/inflammatory response.
Angiotensin II induces thrombospondin-1 production in human mesangial cells via p38 MAPK and JNK: a mechanism for activation of latent TGF-β1
Authors: Naito Takayuki , Masaki Takao , Nikolic-Paterson David J. , Tanji Chie , Yorioka Noriaki , Kohno Nobuoki
Published: January 2004

ANG II induces secretion and activation of transforming growth factor-β (TGF-β) by glomerular mesangial cells. However, the mechanisms that operate this are unclear. Thrombospondin-1 (TSP-1), which is produced by mesangial cells in damaged glomeruli, is one of several molecules known to activate the latent TGF-β1complex. Therefore, we examined whether the ANG II-induced activation of latent TGF-β1in human mesangial cells (HMC) operates via TSP-1. The addition of ANG II (1-100 nM) to HMC significantly increased TSP-1 mRNA within 6 h, followed by an increase in TSP-1 protein production as shown by Western blot analysis of cells and immunoassay of the culture supernatant. Production of ANG II-induced TSP-1 mRNA and protein was completely inhibited by an ANG II type 1 (AT1)-receptor antagonist but was unaffected by an AT2-receptor antagonist. Use of a TSP-1-specific blocking peptide demonstrated that the ANG II-induced activation of latent TGF-β1operates via TSP-1. Next, we investigated the role of ERK1/2, p38 MAPK, and JNK in ANG II-induced TSP-1 production in HMC. The addition of the upstream ERK1/2 inhibitor PD-98059 did not affect ANG II-induced TSP-1 production, whereas addition of either the p38 MAPK inhibitor SB-203580 or the JNK inhibitor SP-600125 significantly reduced TSP-1 production. In conclusion, this study has demonstrated that ANG II-induced activation of latent TGF-β1in HMC operates via TSP-1. Furthermore, ANG II-induced TSP-1 production is dependent on p38 MAPK and JNK signaling.
Renal expression and activity of the germinal center kinase SK2
Authors: Cybulsky Andrey V. , Takano Tomoko , Papillon Joan , Khadir Abdelkrim , Bijian Krikor , Chien Chu-Chun , Alpers Charles E. , Rabb Hamid
Published: January 2004

Rat fetal kidney mRNA was analyzed by RT-PCR to identify protein kinases. This screening demonstrated expression of a protein kinase consistent with SK2, a group II germinal center kinase and homolog of human Ste20-like kinase (SLK). SK2 mRNA, protein expression, and kinase activity were increased in rat fetal kidney homogenates ( embryonic days 17–21) compared with adult controls. In adult kidneys subjected to cross-clamping of the renal artery, followed by reperfusion, SK2 mRNA, protein expression, and kinase activity were increased compared with untreated contralateral controls. By immunohistochemistry, SK2 expression was evident mainly in the cytoplasm of tubular epithelial cells in fetal and adult kidneys. There was also some expression in developing and mature podocytes, but staining of the interstitium was negative. In cultured renal tubular epithelial cells, SK2 kinase activity was increased after incubation with serum, or after exposure to chemical anoxia plus reexposure to glucose. Stable overexpression of SLK reduced cell proliferation and increased apoptosis and exacerbated apoptosis and necrosis after chemical anoxia plus reexposure to glucose. Thus SK2 is a renal epithelial protein kinase whose expression and activity are increased during development and recovery from acute renal failure, where tubular epithelial regeneration may recapitulate developmental processes. The actions of SK2 appear to be antiproliferative and may facilitate cell injury.
Update of extracellular matrix, its receptors, and cell adhesion molecules in mammalian nephrogenesis
Authors: Kanwar Yashpal S. , Wada Jun , Lin Sun , Danesh Farhad R. , Chugh Sumant S. , Yang Qiwei , Banerjee Tushar , Lomasney Jon W.
Published: January 2004

One of the hallmarks of mammalian nephrogenesis includes a mesenchymal-epithelial transition that is accomplished by intercalation of the ureteric bud, an epithelium-lined tubelike structure, into an undifferentiated mesenchyme, and the latter then undergoes an inductive transformation and differentiates into an epithelial phenotype. At the same time, the differentiating mesenchyme reciprocates by inducing branching morphogenesis of the ureteric bud, which forms a treelike structure with dichotomous iterations. These reciprocal inductive interactions lead to the development of a functioning nephron unit made up of a glomerulus and proximal and distal tubules. The inductive interactions and differentiation events are modulated by a number of transcription factors, protooncogenes, and growth factors and their receptors, which regulate the expression of target morphogenetic modulators including the ECM, integrin receptors, and cell adhesion molecules. These target macromolecules exhibit spatiotemporal and stage-specific developmental regulation in the metanephros. The ECM molecules expressed at the epithelial-mesenchymal interface are perhaps the most relevant and conducive to the paracrine-juxtacrine interactions in a scenario where the ligand is expressed in the mesenchyme while the receptor is located in the ureteric bud epithelium or vice versa. In addition, expression of the target ECM macromolecules is regulated by matrix metalloproteinases and their inhibitors to generate a concentration gradient at the interface to further propel epithelial-mesenchymal interactions so that nephrogenesis can proceed seamlessly. In this review, we discuss and update our current understanding of the role of the ECM and related macromolecules with respect to metanephric development.
Regulation of sodium-dicarboxylate cotransporter-3 from winter flounder kidney by protein kinase C
Authors: Hagos Yohannes , Burckhardt Birgitta C. , Larsen Alexander , Mathys Christian , Gronow Tobias , Bahn Andrew , Wolff Natascha A. , Burckhardt Gerhard , Steffgen Jürgen
Published: January 2004

The sodium dicarboxylate cotransporter located at the basolateral side supplies renal proximal tubule cells with Krebs cycle intermediates and maintains the driving force for the exchange of organic anions like PAH against α-ketoglutarate through the organic anion transporter-1. Recently, we cloned sodium dicarboxylate cotransporter-3 from winter flounder kidney (fNaDC-3). To understand the regulation of fNaDC-3, we preincubated fNaDC-3-expressing oocytes with PMA, a PKC activator. PMA dose and time dependently inhibited fNaDC-3-mediated succinate uptake. Simultaneous preincubation of fNaDC-3-expressing oocytes with 50 nM PMA and either staurosporine or RO 31-8220 for 30 min attenuated PKC-mediated inhibition of succinate uptake. Site-directed mutagenesis of the five putative PKC sites (S7, T167, S174, T188, and S396) resulted in no change in PKC-mediated inhibition of the transporter. In electrophysiological studies performed at -60 mV, the K0.5for succinate was not significantly affected (56 ± 13 vs. 42 ± 19 μM), but Δ Imaxwas reduced from -139 ± 49 to -20 ± 8 nA by PMA (50 nM, 30 min). Immunofluorescence analysis of fNaDC-3-expressing oocytes revealed that PMA leads to an endocytosis of fNaDC-3 protein. In conclusion, fNaDC-3 expressed in oocytes is downregulated by PMA through endocytosis. PKC consensus sites appear not to be important for this process.
Developmental approaches to kidney tissue engineering
Authors: Steer Dylan L. , Nigam Sanjay K.
Published: January 2004

Recent advances in our understanding of the developmental biology of the kidney, as well as the establishment of novel in vitro model systems, have potential implications for kidney tissue engineering. These advances include delineation of the roles of a number of growth factors in the developmental programs of branching morphogenesis and mesenchymal differentiation, a new understanding of the roles of the extracellular matrix, identification of potential “renal” stem cells, the ex vivo propagation and subsequent recombination of isolated components of the kidney, and successful transplantation of renal primordia into adult hosts. This review will examine these advances in the context of approaches to tissue engineering. Finally, novel approaches that synthesize advances in both cell-based and organ-based approaches are proposed.
Decreased amiloride-sensitive Na+absorption in collecting duct principal cells isolated from BPK ARPKD mice
Authors: Veizis Elias I. , Carlin Cathleen R. , Cotton Calvin U.
Published: January 2004

The main feature of polycystic kidney diseases (PKD) is formation and progressive enlargement of renal cysts. Alterations in epithelial cell proliferation, extracellular matrix, and ion transport are thought to contribute to cyst enlargement and loss of renal function. Abnormal Cl-secretion is implicated in cyst enlargement in autosomal dominant PKD (ADPKD), but little is known about transport abnormalities in autosomal recessive PKD (ARPKD). We developed a method to isolate collecting duct (CD) principal cells (site of the lesion in ARPKD) from normal and ARPKD mice. A transgenic mouse (Hoxb7/GFP) in which enhanced green fluorescent protein (GFP) is expressed in CDs was bred with an ARPKD mouse (BPK), and GFP-positive cells from normal and cystic mice were selected by fluorescence-activated cell sorting. GFP-positive CD cells (>95 ± 3%) obtained from either normal or cystic mice formed high-resistance, polarized epithelial monolayers. Expression patterns for marker proteins and the presence of a central cilium confirmed that the monolayers are composed of principal cells. Under basal conditions, the Cl-secretory responses elicited by elevation of cAMP or calcium were not significantly different between normal and cystic monolayers. In contrast, the amiloride-sensitive short-circuit current was significantly reduced in monolayers of cells isolated from cystic mice (12.9 ± 1.6 μA/cm2; n = 10) compared with monolayers of cells isolated from normal mice (27.3 ± 3.4 μA/cm2; n = 12). The results of these studies suggest that epithelial sodium channel-mediated sodium absorption is decreased in principal cells of ARPKD CD cysts and that the reduction in sodium absorption may contribute to the accumulation of luminal fluid.
Inhibition of endocytosis causes phosphorylation (S256)-independent plasma membrane accumulation of AQP2
Authors: Lu Hua , Sun Tian-Xiao , Bouley Richard , Blackburn Karen , McLaughlin Margaret , Brown Dennis
Published: January 2004

Inhibition of clathrin-mediated endocytosis by expression of a GTPase-deficient dynamin mutant (dynamin-2/K44A) for 16 h results in an accumulation of plasma membrane aquaporin-2 (AQP2) in epithelial cells stably transfected with wild-type AQP2. We now show a similar effect of K44A dynamin in LLC-PK1cells transfected with an S256 phosphorylation-deficient AQP2 mutant, AQP2(S256A), and in AQP2-transfected inner medullary collecting duct (IMCD) cells. More acute blockade of endocytosis in these cells with the cholesterol-depleting agent methyl-β-cyclodextrin (mβCD; 10 mM) resulted in a rapid and extensive cell-surface accumulation of both wild-type AQP2 and AQP2 (S256A) within 15 min after treatment. This effect was similar to that induced by treatment of the cells with vasopressin. Blockade of endocytosis by mβCD was confirmed using quantitative analysis of FITC-dextran uptake and AQP2 membrane insertion was verified by cell-surface biotinylation. These data indicate that AQP2 recycles constitutively and rapidly between intracellular stores and the cell surface in LLC-PK1 and IMCD cells. The constitutive trafficking process is not dependent on phosphorylation of the serine-256 residue of AQP2, which is, however, an essential step for regulated vasopressin/cAMP-mediated translocation of AQP2. Our data show that rapid and extensive plasma membrane accumulation of AQP2 can occur in a vasopressin receptor (V2R)- and phosphorylation-independent manner, pointing to a potential means of bypassing the mutated V2R in X-linked nephrogenic diabetes insipidus to achieve cell surface expression of AQP2.
Troglitazone acts by PPARγ and PPARγ-independent pathways on LLC-PK1-F+acid-base metabolism
Authors: Welbourne Tomas , Friday Ellen , Fowler Rocky , Turturro Francesco , Nissim Itzhak
Published: January 2004

Troglitazone was studied in pH-sensitive LLC-PK1-F+cells to determine the effect on pHiand glutamine metabolism as well as the role of peroxisome proliferator-activated receptor (PPARγ)-dependent and PPARγ-independent signaling pathways. Troglitazone induces a dose-dependent cellular acidosis that occurs within 4 min and persists over 18 h as a result of inhibiting Na+/H+exchanger-mediated acid extrusion. Cellular acidosis was associated with glutamine-dependent augmented [15N]ammonium production and decreased [15N]alanine formation from15N-labeled glutamine. The shift in glutamine metabolism from alanine to ammoniagenesis appears within 3 h and is associated after 18 h with both a reduction in assayable alanine aminotransferase (ALT) activity as well as cellular acidosis. The relative contribution of troglitazone-induced cellular acidosis vs. the decrease in assayable ALT activity to alanine production could be demonstrated. The PPARγ antagonist bisphenol A diglycide ether (BADGE) reversed both the troglitazone-induced cellular acidosis and ammoniagenesis but enhanced the troglitazone reduction of assayable ALT activity; BADGE also blocked troglitazone induction of peroxisome proliferator response element-driven firefly luciferase activity. The protein kinase C (PKC) inhibitor chelerythrine mimics troglitazone effects, whereas phorbol ester reverses the effects on ammoniagenesis consistent with troglitazone negatively regulating the DAG/PKC/ERK pathway. Although functional PPARγ signaling occurs in this cell line, the major troglitazone-induced acid-base responses appear to be mediated by pathway(s) involving PKC/ERK.
A1adenosine receptor knockout mice exhibit increased renal injury following ischemia and reperfusion
Authors: Lee H. Thomas , Xu Hua , Nasr Samih H. , Schnermann Jurgen , Emala Charles W.
Published: January 2004

Controversy exists regarding the effect of A1adenosine receptor (AR) activation in the kidney during ischemia and reperfusion (I/R) injury. We sought to further characterize the role of A1ARs in modulating renal function after I/R renal injury using both pharmacological and gene deletion approaches in mice. A1AR knockout mice (A1KO) or their wild-type littermate controls (A1WT) were subjected to 30 min of renal ischemia. Some A1WT mice were subjected to 30 min of renal ischemia with or without pretreatment with 1,3-dipropyl-8-cyclopentylxanthine (DPCPX) or 2-chrolo-cyclopentyladenosine (CCPA), selective A1AR antagonist and agonist, respectively. Plasma creatinine and renal histology were compared 24 h after renal injury. A1KO mice exhibited significantly higher creatinines and worsened renal histology compared with A1WT controls following renal I/R injury. A1WT mice pretreated with the A1AR antagonist or agonist demonstrated significantly worsened or improved renal function, respectively, after I/R injury. In addition, A1WT mice pretreated with DPCPX or CCPA showed significantly increased or reduced markers of renal inflammation, respectively (renal myeloperoxidase activity, renal tubular neutrophil infiltration, ICAM-1, TNF-α, and IL-1β mRNA expression), while demonstrating no differences in indicators of apoptosis. In conclusion, we demonstrate that endogenous or exogenous preischemic activation of A1ARs protects against renal I/R injury in vivo via mechanisms leading to decreased necrosis and inflammation.
Protection of renal cells from cisplatin toxicity by cell cycle inhibitors
Authors: Price Peter M. , Safirstein Robert L. , Megyesi Judit
Published: January 2004

The optimal use of cisplatin as a chemotherapeutic drug has been limited by its nephrotoxicity. Murine models have been used to study cisplatin-induced acute renal failure. After cisplatin administration, cells of the S3 segment in the renal proximal tubule are especially sensitive and undergo extensive necrosis in vivo. Similarly, cultured proximal tubule cells undergo apoptosis in vitro after cisplatin exposure. We have shown in vivo that kidney cells enter the cell cycle after cisplatin administration but that cell cycle-inhibitory proteins p21 and 14-3-3σ are also upregulated. These proteins coordinate the cell cycle, and deletion of either of the genes resulted in increased nephrotoxicity in vivo or increased cell death in vitro after exposure to cisplatin. However, it was not known whether cell cycle inhibition before acute renal failure could protect from cisplatin-induced cell death, especially in cells with functional p21 and 14-3-3σ genes. Using several cell cycle inhibitors, including a p21 adenovirus, and the drugs roscovitine and olomoucine, we have been able to completely protect a mouse kidney proximal tubule cell culture from cisplatin-induced apoptosis. The protection by p21 was independent of an effect on the cell cycle and was likely caused by selective inhibition of caspase-dependent and -independent cell death pathways in the cells.
Suppressions of chronic glomerular injuries and TGF-β1production by HGF in attenuation of murine diabetic nephropathy
Authors: Mizuno Shinya , Nakamura Toshikazu
Published: January 2004

Diabetic nephropathy is now the leading cause of end-stage renal diseases, and glomerular sclerotic injury is an initial event that provokes renal dysfunction during processes of diabetes-linked kidney disease. Growing evidence shows that transforming growth factor-β1(TGF-β1) plays a key role in this process, especially in eliciting hypertrophy and matrix overaccumulation. Thus it is important to find a ligand system to antagonize the TGF-β1-mediated pathogenesis under high-glucose conditions. Herein, we provide evidence that hepatocyte growth factor (HGF) targets mesangial cells, suppresses TGF-β1production, and minimizes glomerular sclerotic changes, using streptozotocin-induced diabetic mice. In our murine model, glomerular sclerogenesis (such as tuft area expansion and collagen deposition) progressed between 6 and 10 wk after the induction of hyperglycemia, during a natural course of diabetic disease. Glomerular HGF expression levels in the diabetic kidney transiently increased but then declined below a basal level, with manifestation of glomerular sclerogenesis. When anti-HGF IgG was injected into mice for 2 wk (i.e., from weeks 4 to 6 after onset of hyperglycemia), these glomerular changes were significantly aggravated. When recombinant HGF was injected into the mice for 4 wk (i.e., between 6 and 10 wk following streptozotocin treatment), the progression of glomerular hypertrophy and sclerosis was almost completely inhibited, even though glucose levels remained unchanged (>500 mg/dl). Even more important, HGF repressed TGF-β1production in glomerular mesangial cells even under hyperglycemic conditions both in vitro and in vivo. Consequently, not only albuminuria but also tubulointerstitial fibrogenesis were attenuated by HGF. Overall, HGF therapy inhibited the onset of renal dysfunction in the diabetic mice. On the basis of these findings, we wish to emphasize that HGF plays physiological and therapeutic roles in blocking renal fibrogenesis during a course of diabetic nephropathy.
Lack of UT-B in vasa recta and red blood cells prevents urea-induced improvement of urinary concentrating ability
Authors: Bankir Lise , Chen Kai , Yang Baoxue
Published: January 2004

Recycling of urea within the renal medulla is known to play an important role in the capacity of the kidney to concentrate urine. This recycling occurs simultaneously through a tubular and a vascular route (i.e., through the loops of Henle and vasa recta, respectively). In the present study, transgenic mice with a selective deficiency in UT-B (the urea transporter protein expressed in descending vasa recta and red blood cells), were used to evaluate the specific contribution of vascular urea recycling to overall urine-concentrating ability (UCA). The renal handling of urea was studied in normal conditions and after acute or chronic alterations in urea excretion (acute urea loading or variations in protein intake, respectively). In normal conditions, UT-B null mice exhibited a 44% elevation in plasma urea (Purea), a normal creatinine clearance, but a 25% decrease in urea clearance, with no change in that of sodium and potassium. Acute urea loading induced a progressive increase in urinary urea concentration (Uurea) in wild-type mice and a subsequent improvement in their UCA in contrast to UT-B null mice, in which urinary osmolality and Uureadid not rise, due to the failure to accumulate urea in the medulla. With increasing protein intake (from 10 to 40% protein in diet, leading to a 5-fold increase in urea excretion), Pureawas further increased in null mice while little change was observed in wild-type mice, and null mice were not able to increase Uureaas did wild-type mice. In conclusion, this study in UT-B-deficient mice reveals that countercurrent exchange of urea in renal medullary vessels and red blood cells accounts for a major part of the kidney's concentrating ability and for the adaptation of renal urea handling during a high-protein intake.
Kidney function in early diabetes: the tubular hypothesis of glomerular filtration
Authors: Thomson Scott C. , Vallon Volker , Blantz Roland C.
Published: January 2004

At the onset of diabetes mellitus, the glomerular filtration rate becomes supranormal. Discovery science has identified many abnormalities in the early diabetic kidney that apparently contribute to this phenotype. A serviceable understanding of the early diabetic kidney requires this information to fit together. It is the purpose of this article to present an archetype that explains multiple nuances of kidney function in early diabetes. We refer to this archetype as the “tubular hypothesis of glomerular filtration.” Its basic tenet is that strange effects of diabetes on glomerular filtration stem from primary effects on the proximal tubule or loop of Henle that impact glomerular filtration by feedback through the macula densa. This theory can explain diabetic hyperfiltration, a paradoxical effect of dietary salt on glomerular filtration rate in diabetes, and the renal response to dietary protein and amino acid infusion in diabetes. The discussion centers on the kidney as an integrated system of parts rather than on the specific cellular mechanisms that comprise those parts.
SLC26A7: a basolateral Cl-/HCO3-exchanger specific to intercalated cells of the outer medullary collecting duct
Authors: Petrovic Snezana , Barone Sharon , Xu Jie , Conforti Laura , Ma Liyun , Kujala Minna , Kere Juha , Soleimani Manoocher
Published: January 2004

The outer medullary collecting duct (OMCD) plays an important role in bicarbonate reabsorption and acid-base regulation. An apical V-type H+-ATPase and a basolateral [Formula: see text] exchanger, located in intercalated cells of OMCD, mediate the bicarbonate reabsorption. Here we report the identification of a new basolateral [Formula: see text] exchanger in OMCD intercalated cells in rat kidney. Northern hybridizations demonstrated the predominant expression of this transporter, also known as SLC26A7, in the outer medulla, with lower expression levels in the inner medulla. SLC26A7 was recognized as a ∼90-kDa band in the outer medulla by immunoblot analysis and was localized on the basolateral membrane of a subset of OMCD cells by immunocytochemical staining. No labeling was detected in the cortex. Double-immunofluorescence labeling with the aquaporin-2 and SLC26A7 antibodies or anion exchanger-1 and SLC26A7 antibodies identified the SLC26A7-expressing cells as α-intercalated cells. Functional studies in oocytes demonstrated that increasing the osmolality of the media (to simulate the physiological milieu in the medulla) increased the [Formula: see text] exchanger activity mediated via SLC26A7 by about threefold ( P < 0.02 vs. normal condition). We propose that SLC26A7 is a basolateral [Formula: see text] exchanger in intercalated cells of the OMCD and may play an important role in bicarbonate reabsorption in medullary collecting duct.
Application of difference gel electrophoresis to the identification of inner medullary collecting duct proteins
Authors: Hoffert Jason D. , van Balkom Bas W. M. , Chou Chung-Lin , Knepper Mark A.
Published: January 2004

In this study, we present a standardized approach to purification of native inner medullary collecting duct (IMCD) cells from rat kidney for proteomic analysis and apply the approach to identification of abundant proteins utilizing two-dimensional difference gel electrophoresis (DIGE) coupled with matrix-assisted laser desorption-ionization-time of flight mass spectrometry. Fractionation of inner medullary cell suspensions by low-speed centrifugation gave a highly purified IMCD cell fraction in which aquaporin-2 was enriched 10-fold. When DIGE was initially applied to rat inner medullas fractionated into IMCD cells (labeled with Cy3) and non-IMCD cells (labeled with Cy5), we identified 50 highly abundant proteins expressed in the IMCD cells. These proteins, identifiable without subcellular fractionation, included chiefly enzymes, structural proteins, and signaling intermediates. An additional 35 proteins were found predominantly in the non-IMCD cell types. Proteins that were highly enriched in the IMCD fraction included cytokeratin 8, cytokeratin 18, transglutaminase II, aminopeptidase B, T-plastin, heat shock protein (HSP) 27, HSP70, and lactate dehydrogenase A. Semiquantitative immunoblotting and immunohistochemistry confirmed relative expression levels and distribution of selected proteins. An additional 40 IMCD proteins were identified in separate experiments aimed at further enrichment of proteins through optimization of sample loading. These studies document the applicability of a standardized approach to purification of IMCD cells for proteomic analysis of IMCD proteins and demonstrate the feasibility of largescale identification of proteins in the native IMCD cell.
Lipoxins: endogenous regulators of inflammation
Authors: McMahon Blaithin , Godson Catherine
Published: January 2004

Over the past decade, compelling in vivo and in vitro studies have highlighted lipoxins (LXs) and aspirin-triggered LXs (ATLs) as endogenously produced anti-inflammatory eicosanoids. LXs and ATLs elicit distinct anti-inflammatory and proresolution bioactions that include inhibition of leukocyte-mediated injury, stimulation of macrophage clearance of apoptotic neutrophils, repression of proinflammatory cytokine production, modulation of cytokine-stimulated metalloproteinase activity, and inhibition of cell proliferation and migration. An overview of recent advances in LX physiology is provided, with particular emphasis on the cellular and molecular processes involved. These data coupled with in vivo models of inflammatory diseases suggest that LX bioactions may be amenable to pharmacological mimicry for therapeutic gain.