bluuebunny/crossref_metadata_2025 huggingface dataset:
ATP is released from guinea pig ureter epithelium on distension
Authors: Knight G. E. , Bodin P. , De Groat W. C. , Burnstock G.
Published: March 2015

Distension of the perfused guinea pig ureter at pressures from 20 to 700 cmH2O increased the amount of ATP released from the epithelium in a pressure-dependent manner. During basal perfusion (40 μl/min), the perfusate contained 10 pmol/ml ATP; this increased 10- to 50-fold at various distending pressures. ATP was released from epithelial cells during distension as mechanical removal of the urothelium blocked release. No lactate dehydrogenase was detected in the perfusate, and scanning electron microscopy confirmed an intact urothelium after distension. ATP was not released due to the activation of stretch-activated channels, as gadolinium (10 μM) failed to affect ATP release. Glibenclamide (10 μM), known to inhibit two members of the ATP-binding cassette (ABC) protein family, did not affect ATP release after distension; nor did verapamil (10 μM). In contrast, both monensin (100 μM) and brefeldin A (10 μM), which interfere with vesicular formation or trafficking, inhibited distension-evoked ATP release, which was Ca2+-dependent. This suggests that ATP release from the ureter epithelium might be mediated by vesicular exocytosis. The role of ATP released by distension of hollow visceral organs is discussed in relation to the concept of purinergic mechanosensory transductions, with special reference to nociception and the activation of P2X3receptors on the subepithelial sensory nerves.
Diabetic kidney disease in thedb/dbmouse
Authors: Sharma Kumar , McCue Peter , Dunn Stephen R.
Published: March 2015

Diabetic nephropathy is increasing in incidence and is now the number one cause of end-stage renal disease in the industrialized world. To gain insight into the genetic susceptibility and pathophysiology of diabetic nephropathy, an appropriate mouse model of diabetic nephropathy would be critical. A large number of mouse models of diabetes have been identified and their kidney disease characterized to various degrees. Perhaps the best characterized and most intensively investigated model is the db/ db mouse. Because this model appears to exhibit the most consistent and robust increase in albuminuria and mesangial matrix expansion, it has been used as a model of progressive diabetic renal disease. In this review, we present the findings from various studies on the renal pathology of the db/ db mouse model of diabetes in the context of human diabetic nephropathy. Furthermore, we discuss shortfalls of assessing functional renal disease in mouse models of diabetic kidney disease.
Role of cAMP-PKA-PLC signaling cascade on dopamine-induced PKC-mediated inhibition of renal Na+-K+-ATPase activity
Authors: Gomes Pedro , Soares-da-Silva P.
Published: March 2015

We studied the molecular events set into motion by stimulation of D1-like receptors downstream of Na+-K+-ATPase, while measuring apical-to-basal ouabain-sensitive, amphotericin B-induced increases in short-circuit current in opossum kidney (OK) cells. The D1-like receptor agonist SKF-38393 decreased Na+-K+-ATPase activity (IC50, 130 nM). This effect was prevented by the D1-like receptor antagonist SKF-83566, overnight cholera toxin treatment, the protein kinase A (PKA) antagonist H-89, or the PKC antagonist chelerythrine, but not the mitogen-activated PK inhibitor PD-098059 or phosphatidylinositol 3-kinase inhibitors wortmannin and LY-294002. Dibutyryl cAMP (DBcAMP) and phorbol 12,13-dibutyrate (PDBu) both effectively reduced Na+-K+-ATPase activity. PKA downregulation abolished the inhibitory effects of SKF-38393 and DBcAMP but not those of PDBu. PKC downregulation abolished inhibition by PDBu, SKF-38393, and DBcAMP. The phospholipase C (PLC) inhibitor U-73122 prevented inhibition by SKF-38393 and DBcAMP. However, DBcAMP increased PLC activity. Although OK cells express both Gsα and Gq/11α proteins, D1-like receptors are coupled to Gsα proteins only, as evidenced by studies in cells treated overnight with specific antibodies raised against Gsα and Gq/11α proteins. We conclude that PLC and Na+-K+-ATPase are effector proteins for PKA and PKC, respectively, after stimulation of D1-like receptors coupled to Gsα proteins, in a sequence of events that begins with adenylyl cyclase-PKA system activation followed by PLC-PKC system activation.
Mathematical models of renal fluid and electrolyte transport: acknowledging our uncertainty
Authors: Weinstein Alan M.
Published: March 2015

Mathematical models of renal tubular function, with detail at the cellular level, have been developed for most nephron segments, and these have generally been successful at capturing the overall bookkeeping of solute and water transport. Nevertheless, considerable uncertainty remains about important transport events along the nephron. The examples presented include the role of proximal tubule tight junctions in water transport and in regulation of Na+transport, the mechanism by which axial flow in proximal tubule modulates solute reabsorption, the effect of formate on proximal Cl−transport, the assessment of potassium transport along collecting duct segments inaccessible to micropuncture, the assignment of pathways for peritubular Cl−exit in outer medullary collecting duct, and the interaction of carbonic anhydrase-sensitive and -insensitive pathways for base exit from inner medullary collecting duct. Some of these uncertainties have had intense experimental interest well before they were cast as modeling problems. Indeed, many of the renal tubular models have been developed based on data acquired over two or three decades. Nevertheless, some uncertainties have been delineated as the result of model exploration and represent communications from the modelers back to the experimental community that certain issues should not be considered closed. With respect to model refinement, incorporating more biophysical detail about individual transporters will certainly enhance model reliability, but ultimate confidence in tubular models will still be contingent on experimental development of critical information at the tubular level.
Characterization ofcis-acting element in renal NaPi-2 cotransporter mRNA that determines mRNA stability
Authors: Moz Yulia , Silver Justin , Naveh-Many Tally
Published: March 2015

Hypophosphatemia leads to an increase in Na+-Picotransporter (NaPi-2) mRNA levels. This increase is posttranscriptional and correlates with a more stable transcript mediated by the terminal 698 nt of the NaPi-2 mRNA. A 71-nt binding element was identified with renal proteins from rats fed control and low-Pi(−Pi) diet. The binding of −Pirenal proteins to this transcript was increased compared with control proteins. The functionality of the ciselement was demonstrated by an in vitro degradation assay. −Pirenal proteins stabilized transcripts that included the cis element compared with control renal extracts. The full-length NaPi-2 transcript, but not control transcripts, was stabilized by −Piextracts. Insertion of the binding element into green fluorescent protein (GFP) as a reporter gene decreased chimeric GFP mRNA levels in transfection experiments. Our results suggest that the protein-binding region of the NaPi-2 mRNA functions as a cis-acting instability element. In hypophosphatemia there is increased binding to the cis-acting element and subsequent stabilization of NaPi-2 mRNA.
Rapid microplate method for PAH estimation
Authors: Agarwal Rajiv ,   (With the Technical Assistance of Shawn D. Chase)
Published: March 2015

Evaluation of renal hemodynamics requires estimation of effective renal plasma flow, which is commonly measured by the renal clearance of p-aminohippuric acid (PAH). There are many existing methods for PAH assay that are complicated, expensive, or time consuming. We describe a rapid, precise, and accurate microplate-based assay of PAH using p-dimethylaminocinnamaldehyde, which produces a red color on reaction with PAH, and compare it with a reference HPLC method. Renal PAH clearances were measured in 10 volunteers, and clearances were calculated by using the new and HPLC methods. There was excellent agreement between the HPLC and the microplate method of PAH assay. The average ratio of microplate to HPLC method was nearly 1.0, and the limits of agreement (2 SD) for plasma, urine, and clearances were 17.2, 19.3, and 25.5%, respectively. Intraday coefficient of variation for urine and plasma was <7%; interday coefficient of variation was <6% for urine and plasma samples. The microplate method is a reliable alternative to a reference HPLC method and can be performed for a fraction of the cost, time, and reagents.
CNP gene expression is activated by Wnt signaling and correlates withWnt4expression during renal injury
Authors: Surendran Kameswaran , Simon Theodore C.
Published: March 2015

C-type natriuretic peptide (CNP) regulates salt excretion, vascular tone, and fibroblast proliferation and activation. CNP inhibits fibroblast activation in vitro and fibrosis in vivo, but endogenous CNP gene ( Nppc) expression during tissue fibrosis has not been reported. We determined that Nppc is induced in renal tubular epithelia and then in interstitial myofibroblasts after unilateral ureteral obstruction (UUO). Induction of Nppcoccurred in identical cell populations to those in which Wnt4 is induced after renal injury. In addition, Nppc was activated in Wnt4-expressing cells during nephrogenesis. Wnt signaling components β-catenin and T cell factor/lymphoid enhancer binding factor (TCF/LEF) specifically bound to cognate elements in the Nppc proximal promoter. Wnt-4, β-catenin, and LEF-1 activated an Nppc transgene in cultured cells, and transgene activation by Wnt-4 and LEF-1 was dependent on the presence of intact cognate elements. These findings suggest that Wnt-4 stimulates Nppc in a TCF/LEF-dependent manner after renal injury and thus may contribute to limiting renal fibrosis.
NO dependency of RBF and autoregulation in the spontaneously hypertensive rat
Authors: Racasan Simona , Joles Jaap A. , Boer Peter , Koomans Hein A. , Braam Branko
Published: March 2015

In the spontaneously hypertensive rat (SHR), renal blood flow (RBF) has been reported to be very dependent on nitric oxide (NO); however, autoregulation is normal, albeit shifted to higher perfusion pressures. To test the hypothesis that in the SHR NO dependency of RBF autoregulation is diminished, we investigated RBF autoregulation in anesthetized young male SHR and normotensive Wistar-Kyoto (WKY) rats before and during acute intravenous NO synthase (NOS) inhibition with Nω-nitro-l-arginine (l-NNA) and urinary excretion of nitrate plus nitrite (UNOxV) at different renal perfusion pressures (RPP). Under baseline conditions, SHR had higher mean arterial pressure (147 ± 4 mmHg) and renal vascular resistance (16 ± 1 U) than WKY (105 ± 4 mmHg and 10 ± 0.5 U, respectively, P < 0.05). RBF was similar (9.4 ± 0.5 vs. 10.3 ± 0.1 ml · min-1· g kidney wt-1). Acute NOS blockade increased mean arterial pressure similarly, but there was significantly more reduction in RBF and hence an enhanced increase in renal vascular resistance in SHR (to 36 ± 3 vs. 17 ± 1 U in WKY, P < 0.001). The renal vasculature of SHR is thus strongly dependent on NO in maintaining basal RBF. The lower limit of autoregulation was higher in SHR than WKY in the baseline situation (85 ± 3 vs. 71 ± 2 mmHg, P < 0.05). Acute l-NNA administration did not decrease the lower limit in the SHR (to 81 ± 3 mmHg, not significant) and decreased the lower limit to 63 ± 2 mmHg ( P < 0.05) in the WKY. The degree of compensation as a measure of autoregulatory efficiency attained at spontaneous perfusion pressures was comparable in SHR vs. WKY but with a shift of the curve toward higher perfusion pressures in SHR. Acute NOS blockade only increased the degree of compensation in WKY. Remarkably, UNOxV was significantly lower at spontaneous RPP in SHR. After reduction of RPP, the observed decrease in UNOxV was significantly more pronounced in WKY than in SHR. In conclusion, the renal circulation in SHR is dependent on high levels of NO; however, the capacity to modulate NO in response to RPP-induced changes in shear stress seems to be limited.
Early transcriptional effects of aldosterone in a mouse inner medullary collecting duct cell line
Authors: Gumz Michelle L. , Popp Michael P. , Wingo Charles S. , Cain Brian D.
Published: March 2015

The mineralocorticoid aldosterone is a major regulator of Na+ and acid-base balance and control of blood pressure. Although the long-term effects of aldosterone have been extensively studied, the early aldosterone-responsive genes remain largely unknown. Using DNA array technology, we have characterized changes in gene expression after 1 h of exposure to aldosterone in a mouse inner medullary collecting duct cell line, mIMCD-3. Results from three independent microarray experiments revealed that the expression of many transcripts was affected by aldosterone treatment. Northern blot analysis confirmed the upregulation of four distinct transcripts identified by the microarray analysis, namely, the serum and glucose-regulated kinase sgk, connective tissue growth factor, period homolog, and preproendothelin. Immunoblot analysis for preproendothelin demonstrated increased protein expression. Following the levels of the four transcripts over time showed that each had a unique pattern of expression, suggesting that the cellular response to aldosterone is complex. The results presented here represent a novel list of early aldosterone-responsive transcripts and provide new avenues for elucidating the mechanism of acute aldosterone action in the kidney.
Cyclooxygenase-2 products compensate for inhibition of nitric oxide regulation of renal perfusion
Authors: Beierwaltes William H.
Published: March 2015

Cyclooxygenase (COX)-2 is in the macula densa, cosegregating with neuronal nitric oxide synthase (nNOS). It is hypothesized that in response to acute inhibition of NOS, the influence of COX-2-derived prostanoids is exaggerated, compensating for renal vasoconstriction. Blood pressure (BP) and renal blood flow (RBF) were measured after selective COX-2 inhibition with NS-398 followed by NOS inhibition with l-nitro arginine methyl ester (l-NAME) or after l-NAME followed by NS-398. BP was 106 ± 4 mmHg and was unaffected by NS-398.l-NAME after NS-398 increased BP by 27 ± 2 mmHg, decreased RBF by one-half, and doubled renal vascular resistance (RVR; P < 0.001). Initial l-NAME increased BP by 26 ± 3 mmHg ( P < 0.001) and decreased RBF by 44% ( P < 0.001), doubling RVR. Afterl-NAME, NS-398 induced a further 7 ± 3-mmHg rise in BP ( P < 0.05), decreased RBF by 20% ( P < 0.025), and increased RVR by 23% ( P < 0.01). The constrictor response to COX-2 inhibition after l-NAME could not be duplicated by either selective nNOS inhibition or NOS-independent renal vasoconstriction. Acute NOS inhibition unmasked renal vasoconstriction with COX-2 inhibition, suggesting that the influence of COX-2-derived vasodilator eicosanoids is exaggerated to maintain renal perfusion, compensating for the acute loss of NO.
Comparative analysis of ontogenic changes in renal and intestinal biotin transport in the rat
Authors: Nabokina Svetlana M. , Subramanian Veedamali S. , Said Hamid M.
Published: March 2015

Biotin, an essential water-soluble micronutrient, cannot be synthesized by mammals; rather, it is obtained from exogenous sources via uptake by intestinal epithelia. Renal epithelia reclaim the vitamin that is filtered in the glomeruli. Both epithelia take up biotin via the sodium-dependent multivitamin transporter (SMVT). Little is known about ontogenic regulation of the renal and intestinal biotin transport processes and about the mechanism(s) involved in any such regulation. In this study, we sought to examine and compare ontogenic aspects of the renal and intestinal biotin uptake processes using purified brush-border membrane vesicles (BBMV) isolated from the kidney cortex and jejunum of suckling and adult rats. Clear ontogenic changes were observed in the intestinal biotin uptake process, which were mediated via changes in Vmaxand apparent Km. Parallel changes were also seen in protein, mRNA, and transcription rate of SMVT as indicated by results of Western blotting, RT-PCR, and nuclear run-on assays, respectively. In contrast, biotin uptake by renal BBMV did not show ontogenic changes; i.e., it was similar in suckling and adult rats. Also, the levels of SMVT protein and mRNA were similar in the kidneys of both age groups. These data show that biotin uptake by renal and intestinal epithelial cells responds differently to ontogenic regulation. In addition, the ontogenic changes observed in the intestinal biotin uptake process involve the entry step of the vitamin at the BBM and appear to be mediated via a transcriptional mechanism(s).
Thick ascending limb-specific expression of Cre recombinase
Authors: Stricklett Peter K. , Taylor Deborah , Nelson Raoul D. , Kohan Donald E.
Published: March 2015

Evaluation of thick ascending limb (TAL) function has been hindered by the limited ability to selectively examine the function of this nephron segment in vivo. To address this, a Cre/loxP strategy was employed whereby the Tamm-Horsfall (THP) promoter was used to drive Cre recombinase expression in transgenic mice. The THP gene was cloned from a mouse genomic library, and 3.7 kb of the mouse THP 5′-flanking region containing the first noncoding exon of the THP gene were inserted upstream of an epitope-tagged Cre recombinase (THP-CreTag). THP-CreTag transgenic mice were bred with ROSA26-enhanced yellow fluorescent protein (eYFP) mice (contain a loxP-flanked “STOP” sequence 5′ to eYFP), and doubly heterozygous offspring were analyzed. THP and eYFP were expressed in an identical pattern with predominant localization to the renal outer medulla without expression in nonrenal tissues. eYFP did not colocalize with thiazide-sensitive cotransporter (distal tubule) or neuronal nitric oxide synthase (macula densa) expression. THP mRNA expression was detected only in kidney, whereas CreTag mRNA was also present in testes. These data indicate that THP-CreTag transgenic mice can be used for TAL-specific gene recombination in the kidney.
Carbonic anhydrase XII mRNA encodes a hydratase that is differentially expressed along the rabbit nephron
Authors: Schwartz George J. , Kittelberger Anne M. , Watkins Richard H. , O'Reilly Michael A.
Published: March 2015

Membrane-bound carbonic anhydrase (CA) facilitates acidification in the kidney. Although most hydratase activity is considered due to CA IV, some in the basolateral membranes could be attributed to CA XII. Indeed, CA IV is glycosylphosphatidylinositol anchored, connoting apical polarization, but CA IV immunoreactivity has been detected on basolateral membranes of proximal tubules. Herein, we determined whether CA XII mRNA was expressed in acidifying segments of the rabbit nephron. The open reading frame of CA XII was sequenced from a rabbit kidney cortex cDNA library; it was 83% identical to human CA XII and coded for a 355-amino acid single-pass transmembrane protein. Northern blot analysis revealed an abundant 4.5-kb message in kidney cortex, medulla, and colon. By in situ hybridization, CA XII mRNA was expressed by proximal convoluted and straight tubules, cortical and medullary collecting ducts, and papillary epithelium. By RT-PCR, CA XII mRNA was abundantly expressed in cortical and medullary collecting ducts and thick ascending limb of Henle's loop; it was also expressed in proximal convoluted and straight tubules but not in glomeruli or S3 segments. FLAG-CA XII of ∼40 kDa expressed in Escherichia coli showed hydratase activity that was inhibited by 0.1 mM acetazolamide. Unlike CA IV, expressed CA XII activity was inhibited by 1% SDS, suggesting insufficient disulfide linkages to stabilize the molecule. Western blotting of expressed CA XII with two anti-rabbit CA IV peptide antibodies showed no cross-reactivity. Our findings indicate that CA XII may contribute to the membrane CA activity of proximal tubules and collecting ducts.
A role for angiotensin II AT1receptors in ureteric bud cell branching
Authors: Iosipiv Igor V. , Schroeder Mercedes
Published: March 2015

Gene-targeting studies in mice demonstrate that the renin-angiotensin system is required for the proper development of the renal medulla. In the absence of angiotensin II (ANG II) or the ANG II type 1 (AT1) receptor, mice exhibit poor papillary development and a severe urinary-concentrating defect. These findings imply that the ureteric bud (UB) and its branches are targets for ANG II actions during renal development. However, direct evidence linking ANG II with UB-branching morphogenesis does not exist. Using immunohistochemistry, we demonstrated that UB-derived epithelia express angiotensinogen (Ao) and the AT1receptor during murine metanephrogenesis. Ao and AT1receptors are expressed in the UB branches and to a lesser extent in the stromal mesenchyme. AT1receptor expression in UB-derived epithelia increased from embryo day 12 to day 16 and was observed on both luminal and basolateral membranes. In accord with these findings, cultured murine UB cells express AT1receptor protein and mRNA. Treatment of UB cells cultured in three-dimensional type I collagen gels with ANG II (10–7to 10–5M) elicits a dose-related increase in the number of cells that have primary and secondary branches. These effects of ANG II on UB branching are abrogated by pretreatment with the AT1receptor antagonist candesartan. These data demonstrate a direct and independent role for ANG II acting via AT1receptors on UB cell branching in vitro. The presence of Ao in the stroma and AT1on UB cells supports the notion that cross talk between stroma and epithelial cells is crucial to epithelial branching morphogenesis in the developing kidney.
pH-responsive stabilization of glutamate dehydrogenase mRNA in LLC-PK1-F+cells
Authors: Schroeder Jill M. , Liu Wenlin , Curthoys Norman P.
Published: March 2015

During chronic metabolic acidosis, the adaptive increase in rat renal ammoniagenesis is sustained, in part, by increased expression of mitochondrial glutaminase (GA) and glutamate dehydrogenase (GDH) enzymes. The increase in GA activity results from the pH-responsive stabilization of GA mRNA. The 3′-untranslated region (3′-UTR) of GA mRNA contains a direct repeat of an eight-base AU-rich element (ARE) that binds ζ-crystallin/NADPH:quinone reductase (ζ-crystallin) with high affinity and functions as a pH-response element. RNA EMSAs established that ζ-crystallin also binds to the full-length 3′-UTR of GDH mRNA. This region contains four eight-base sequences that are 88% identical to one of the two GA AREs. Direct binding assays and competition studies indicate that the two individual eight-base AREs from GA mRNA and the four individual GDH sequences bind ζ-crystallin with different affinities. Insertion of the 3′-UTR of GDH cDNA into a β-globin expression vector (pβG) produced a chimeric mRNA that was stabilized when LLC-PK1-F+cells were transferred to acidic medium. A pH-responsive stabilization was also observed using a βG construct that contained only the single GDH4 ARE and a destabilizing element from phospho enolpyruvate carboxykinase mRNA. Therefore, during acidosis, the pH-responsive stabilization of GDH mRNA may be accomplished by the same mechanism that affects an increase in GA mRNA.
Expression of the prostaglandin F receptor (FP) gene along the mouse genitourinary tract
Authors: Saito Osamu , Guan Youfei , Qi Zhonghua , Davis Linda S. , Kömhoff Martin , Sugimoto Yukihiko , Narumiya Shuh , Breyer Richard M. , Breyer Matthew D.
Published: March 2015

PGF2αis one of the major prostanoids produced by the kidney. The cellular effects of PGF2αare mediated by a G protein-coupled transmembrane receptor designated the FP receptor. Both in situ hybridization and β-galactosidase knocked into the endogenous FP locus were used to determine the cellular distribution of the mouse FP receptor. Specific labeling was detected in the kidney, ovary, and uterus. Abundant FP expression in ovarian follicles and uterus is consistent with previous reports of failed parturition in FP−/− mice. In the kidney, coexpression of the mFP mRNA with the thiazide-sensitive cotransporter defined its expression in the distal convoluted tubule (DCT). FP receptor was also present in aquaporin-2-positive cortical collecting ducts (CCD). No FP mRNA was detected in glomeruli, proximal tubules, or thick ascending limbs. Intrarenal expression of the FP receptor in the DCT and CCD suggests an important role for the FP receptor regulating water and solute transport in these segments of the nephron.
Distribution of transcellular calcium and sodium transport pathways along mouse distal nephron
Authors: Loffing Johannes , Loffing-Cueni Dominique , Valderrabano Victor , Kläusli Lea , Hebert Steven C. , Rossier Bernard C. , Hoenderop Joost G. J. , Bindels René J. M. , Kaissling Brigitte
Published: March 2015

First published August 15, 2001; 10.1152/ajprenal. 00085.2001.—The organization of Na+and Ca2+transport pathways along the mouse distal nephron is incompletely known. We revealed by immunohistochemistry a set of Ca2+and Na+transport proteins along the mouse distal convolution. The thiazide-sensitive Na+-Cl−cotransporter (NCC) characterized the distal convoluted tubule (DCT). The amiloride-sensitive epithelial Na+channel (ENaC) colocalized with NCC in late DCT (DCT2) and extended to the downstream connecting tubule (CNT) and collecting duct (CD). In early DCT (DCT1), the basolateral Ca2+-extruding proteins [Na+/Ca2+exchanger (NCX), plasma membrane Ca2+-ATPase (PCMA)] and the cytoplasmic Ca2+-binding protein calbindin D28K(CB) were found at very low levels, whereas the cytoplasmic Ca2+/Mg2+-binding protein parvalbumin was highly abundant. NCX, PMCA, and CB prevailed in DCT2 and CNT, where we located the apical epithelial Ca2+channel (ECaC1). Its subcellular localization changed from apical in DCT2 to exclusively cytoplasmic at the end of CNT. NCX and PMCA decreased in parallel with the fading of ECaC1 in the apical membrane. All three of them were undetectable in CD. These findings disclose DCT2 and CNT as major sites for transcellular Ca2+transport in the mouse distal nephron. Cellular colocalization of Ca2+and Na+transport pathways suggests their mutual interactions in transport regulation.
Urogenital distribution of a mouse membrane-associated prostaglandin E2synthase
Authors: Guan Youfei , Zhang Yahua , Schneider André , Riendeau Denis , Mancini Joseph A. , Davis Linda , Kömhoff Martin , Breyer Richard M. , Breyer Matthew D.
Published: March 2015

First published August 15, 2001; 10.1152/ajprenal.00116.2001.—PGE2plays a critical role in regulating renal function and facilitating reproduction. One of the rate-limiting biosynthetic enzymes in PGE2synthesis is the terminal PGE2synthase (PGES). In the present studies, we report the functional expression of a membrane-associated murine PGES (mPGES) and its expression in urogenital tissues. Two independent cDNA clones sharing an identical open reading frame of 459 bp and encoding a peptide of 153 amino acids, but differing in the 3′-untranslated region, were identified. Assays for enzymatic activity, using microsomes prepared from cells transfected with mPGES cDNA, showed that these cDNA sequences encode a functional protein that catalyzes the conversion of PGH2to PGE2. Constitutive expression of mPGES was highest in the mouse kidney, ovary, and urinary bladder but was also expressed at lower levels in uterus and testis. Renal mPGES expression was predominantly localized to epithelia of distal tubules and medullary collecting ducts. High expression was also seen in transitional epithelial cells of bladder and ureter and in the primary and secondary follicles in the ovary. In conclusion, mPGES is constitutively expressed along the urogenital tract, where it may have important roles in normal physiology and disease.
LPS-sensory peptide communication in experimental cystitis
Authors: Saban M. R. , Saban R. , Hammond T. G. , Haak-Frendscho M. , Steinberg H. , Tengowski M. W. , Bjorling D. E.
Published: March 2015

Stimulation of sensory nerves can lead to release of peptides such as substance P (SP) and consequently to neurogenic inflammation. We studied the role of bacterial lipopolysaccharide (LPS) in regulating SP-induced inflammation. Experimental cystitis was induced in female mice by intravesical instillation of SP, LPS, or fluorescein-labeled LPS. Uptake of fluorescein-labeled LPS was determined by confocal analysis, and bladder inflammation was determined by morphological analysis. SP was infused into the bladders of some mice 24 h after exposure to LPS. In vitro studies determined the capacity of LPS and SP to induce histamine and cytokine release by the bladder. LPS was taken up by urothelial cells and distributed systemically. Twenty-four hours after instillation of LPS or SP, bladder inflammation was characterized by edema and leukocytic infiltration of the bladder wall. LPS pretreatment enhanced neutrophil infiltration induced by SP, increased in vitro release of histamine, tumor necrosis factor-α, and interferon-γ, and significantly reduced transforming growth factor-β1 release. These findings suggest that LPS amplifies neurogenic inflammation, thereby playing a role in the pathogenesis of neurogenic cystitis.
Spontaneous renal blood flow autoregulation curves in conscious sinoaortic baroreceptor-denervated rats
Authors: Pires Silene L. S. , Julien Claude , Chapuis Bruno , Sassard Jean , Barrès Christian
Published: March 2015

These experiments examined whether the conscious sinoaortic baroreceptor-denervated (SAD) rat, owing to its high spontaneous arterial pressure (AP) variability, might represent a model for renal blood flow (RBF) autoregulation studies. In eight SAD and six baroreceptor-intact rats, AP and RBF were recorded (1-h periods) before and after furosemide (10 mg/kg followed by 10 mg · kg−1· h−1iv)administration. In control conditions, AP variability was markedly enhanced in SAD rats (coefficient of variation: 16.0 ± 1.2 vs. 5.4 ± 0.5% in intact rats), whereas RBF variability was only slightly increased (8.7 ± 0.6 vs. 6.1 ± 0.5% in intact rats), suggesting buffering by autoregulatory mechanisms. In SAD rats, but not in intact rats, the AP-RBF relationships could be modeled with a four-parameter sigmoid Weibull equation ( r2= 0.24 ± 0.07, 3,600 data pairs/rat), allowing for estimation of an autoregulatory plateau (10.1 ± 0.7 ml/min) and a lower limit of RBF autoregulation (PLL= 93 ± 6 mmHg, defined as AP at RBF 5% below the plateau). After furosemide treatment, autoregulation curves ( r2= 0.49 ± 0.07) in SAD rats were shifted downward (plateau = 8.6 ± 0.8 ml/min) and rightward (PLL= 102 ± 5 mmHg). In five of six intact rats, PLLbecame measurable (104 ± 1 mmHg), albeit with limited accuracy ( r2= 0.09 ± 0.03). In conclusion, the conscious SAD rat offers the possibility of describing RBF autoregulation curves under dynamic, unforced conditions. The tubuloglomerular feedback and myogenic mechanisms cooperate in setting PLLand thus in stabilizing RBF during spontaneous depressor episodes.
Analysis of NaCl transport in thin ascending limb of Henle's loop in CLC-K1 null mice
Authors: Liu Wen , Morimoto Tetsuji , Kondo Yoshiaki , Iinuma Kazuie , Uchida Shinichi , Sasaki Sei , Marumo Fumiaki , Imai Masashi
Published: March 2015

To characterize the nature of NaCl transport in the thin ascending limb (tAL), we examined the transport properties of Na+and Cl−using in vitro microperfusion of the tAL in CLC-K1 null mice. In the presence of a transmural NaCl concentration gradient (100 mM higher in the lumen), the transepithelial diffusion voltage ( Vd) was 15.5 ± 1.0 and −7.6 ± 1.4 mV in CLC-K1+/+and CLC-K1−/−mice, respectively. Neither Cl−transport inhibitor 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB) nor acidification of the bathing fluid changed the Vdvalues in CLC-K1−/−mice. The addition of 300 μg/ml protamine, a selective blocker of paracellular conductance, to the bath increased the Vdvalues by 5.6 ± 0.7 and 12.6 ± 1.5 mV ( P < 0.001) in CLC-K1+/+and CLC-K1−/−mice, respectively. Although efflux coefficients of36Cl were significantly decreased in CLC-K1−/−mice (188.3 ± 25.6 in 4 tubules vs. 17.2 ± 7.0 × 10−5cm/s in 6 tubules), those of22Na were not different between CLC-K1+/+and CLC-K1−/−mice. These results clearly indicate that the major component of Cl−transport sensitive to NPPB or pH is mediated by CLC-K1 in the tAL.
Reduction of renal immune cell infiltration results in blood pressure control in genetically hypertensive rats
Authors: Rodríguez-Iturbe Bernardo , Quiroz Yasmir , Nava Mayerly , Bonet Lizzette , Chávez Maribel , Herrera-Acosta Jaime , Johnson Richard J. , Pons Héctor A.
Published: March 2015

Immunocompetent cells infiltrate the kidney in several models of experimental hypertension. We have previously shown that reduction of this infiltrate results in prevention of salt-sensitive hypertension induced by short-term angiotensin II infusion and nitric oxide inhibition (Quiroz Y, Pons H, Gordon KI, Rincón J, Chávez M, Parra G, Herrera-Acosta J, Gómez-Garre D, Largo R, Egido J, Johnson RJ, and Rodrı́guez-Iturbe B. Am J Physiol Renal Physiol281: F38–F47, 2001; Rodrı́guez-Iturbe B, Pons H, Quiroz Y, Gordon K, Rincón J, Chávez M, Parra G, Herrera-Acosta J, Gómez-Garre D, Largo R, Egido J, and Johnson RJ. Kidney Int 59: 2222–2232, 2001). We therefore studied whether hypertension could be controlled in genetically hypertensive rats [spontaneously hypertensive rats (SHR)] by the administration of 20 mg · kg−1· day−1of the immunosuppressive drug mycophenolate mofetil (MMF group; n = 35). Other SHR received vehicle ( n= 35), and Wistar-Kyoto rats ( n = 20) were used as controls. MMF or vehicle was given in two separate 4-wk periods, separated by a 3-wk interval. Systemic hypertension was reduced to normal levels in both periods of MMF treatment in association with a reduction in lymphocyte, macrophage, and angiotensin II-positive cells infiltrating the kidney. Oxidative stress was also reduced by MMF, as indicated by a reduction in urinary malondialdehyde (MDA), renal MDA content, and superoxide-positive cells, and was highly correlated with blood pressure levels. We conclude that the renal immune infiltrate plays a major role in the hypertension in SHR.
Apical H+/base transporters mediating bicarbonate absorption and pHi regulation in the OMCD
Authors: Yip Kay-Pong , Tsuruoka Shuichi , Schwartz George J. , Kurtz Ira
Published: March 2015

The outer medullary collecting duct (OMCD) plays an important role in mediating transepithelial HCO[Formula: see text] transport ([Formula: see text]) and urinary acidification. HCO[Formula: see text] absorption by type A intercalated cells in the OMCD inner stripe (OMCDis) segment is thought to by mediated by an apical vacuolar H+-ATPase and H+-K+-ATPase coupled to a basolateral Cl−-HCO[Formula: see text] exchanger (AE1). Besides these Na+-independent transporters, previous studies have shown that OMCDis type A intercalated cells have an apical electroneutral EIPA-sensitive, DIDS-insensitive Na+-HCO[Formula: see text] cotransporter (NBC3); a basolateral Na+/H+ antiporter; and a basolateral Na+-K+-ATPase. In this study, we reexamined the Na+ dependence of transepithelial Na+ transport in the OMCDis and determined the role of apical NBC3 in intracellular (pHi) regulation in OMCDis type A intercalated cells. Control tubules absorbed HCO[Formula: see text] at a rate of ∼13 pmol · min−1 · mm−1. Lowering luminal Na+ from 140 to 40 mM decreased[Formula: see text] by ∼15% without a change in transepithelial potential ( V te). Furthermore, 50 μM EIPA (lumen) also decreased [Formula: see text] by ∼13% without a change in V te. The effect of lowering luminal Na+ and adding EIPA were not additive. These results demonstrate that[Formula: see text] in the OMCDis is in part Na+ dependent. In separate experiments, the pHi recovery rate after an NH[Formula: see text] prepulse was monitored in single type A intercalated cells with confocal fluorescence microscopy. The pHi recovery rate was ∼0.21 pH/min in Na+-containing solutions and decreased to ∼0.16 pH/min with EIPA (50 μM, lumen). In tubules perfused/bathed without Na+, luminal Na+ addition resulted in a pHi recovery rate of ∼0.36 pH/min, whereas the Na+-independent recovery rate was ∼0.16 pH/min. EIPA (50 μM, lumen) decreased the Na+-dependent pHirecovery rate to ∼0.07 pH/min. The Na+-independent recovery rate was decreased to ∼0.06 pH/min by bafilomycin (10 nM, lumen) and to ∼0.10 pH/min using Schering 28080 (10 μM, lumen). These findings indicate that NBC3 contributes to pHiregulation in OMCDis type A intercalated cells and plays only a minor role in mediating[Formula: see text] in the OMCDis.
Hypochlorous acid alters bronchial epithelial cell membrane properties and prevention by extracellular glutathione
Authors: Venglarik Charles J. , Girón-Calle Julio , Wigley Amanda F. , Malle Ernst , Watanabe Nobuo , Forman Henry Jay
Published: March 2015

In chronic inflammatory diseases of the airways, such as cystic fibrosis, hypochlorous acid (HOCl) generated by neutrophils is involved in airway injury. We examined the effects of HOCl on 16HBE14o– bronchial epithelial cells by bolus addition or by generation with glucose oxidase plus myeloperoxidase. HOCl produced both carbonyl formation of a discreet number of proteins and modification of surface targets that were recognized by an antibody raised against HOCl-modified protein. Bolus or enzymatically generated HOCl decreased transepithelial resistance, but surprisingly bolus HOCl also increased short-circuit current. Glutathione in lung epithelial lining fluid is an excellent scavenger of HOCl; however, glutathione content is lower in cystic fibrosis epithelial lining fluid due to deficient glutathione transport to the apical side of bronchial-tracheal epithelial cells (Gao L, Kim KJ, Yankaskas JR, and Forman HJ. Am J Physiol Lung Cell Mol Physiol 277: L113–L118, 1999). We found that alteration of the GSH content of apical fluid above 16HBE14o– cells was protective because all HOCl-induced changes were delayed or eliminated by exogenous glutathione within the physiological range. Extrapolating this to cystic fibrosis suggests that HOCl can alter cell function without destruction but that elevating glutathione could be protective.
Exercise increases prolactin-receptor expression on human lymphocytes
Authors: Dohi Keiichiro , Kraemer William J. , Mastro Andrea M.
Published: March 2015

Plasma prolactin has been shown to increase during stress; the immune system is responsive to prolactin and affected by stress. Therefore, this study was undertaken to investigate the effects of acute graded, maximal treadmill exercise on prolactin-receptor expression by lymphocytes. Eight healthy men underwent one exercise and one nonexercise session. Blood was sampled immediately before and after the exercise. On the day of the nonexercise session, two resting blood samples were obtained at the same times as the exercise session samples to act as baseline data. Plasma prolactin concentrations were significantly elevated in response to exercise and correlated positively with total prolactin-receptor expression per B lymphocyte. An increase in total prolactin-receptor expression per B lymphocyte in response to exercise also was observed. In addition, exercise significantly increased the total number of circulating B lymphocytes expressing prolactin receptor as well as the total number of circulating B lymphocytes. These data support the idea that exercise may enhance the interaction between immune target cells and prolactin, a stress hormone capable of enhancing immune function.
Voluntary muscle activation varies with age and muscle group
Authors: Jakobi Jennifer M. , Rice Charles L.
Published: March 2015

The consistency and the number of attempts required to achieve maximal voluntary muscle activation have not been documented and compared between young and old adults. Furthermore, few studies have contrasted activation between functional pairs of muscle groups, and no study has tested upper limb muscles. The purpose of this study was to measure and compare voluntary muscle activation of the elbow flexors and extensors in young and old men over two separate test sessions. With the method of twitch interpolation to measure activation, six young (24 ± 1 yr) and six old (83 ± 4 yr) men performed five maximal voluntary contractions (MVC) during each session for each muscle group. Elbow flexion and extension MVC was less (43 and 47%, respectively) in the old men, yet the best maximal voluntary muscle activation was similar between age groups. However, when all 10 attempts at MVC were compared, the mean activation scores were slightly less (∼5%) in the elbow extensors but were ∼11% less ( P < 0.001) in the elbow flexors of old men, compared with young men. During the second session, there was a significant improvement of 13% ( P< 0.005) in mean elbow flexor activation in the old men. There were no session differences for either muscle group for the young men. The results indicate that, for aged men, elbow flexor maximal activation is achieved less frequently compared with elbow extensors, and thus mean activation for elbow flexors is less than for elbow extensors. However, if sufficient attempts are provided, the best effort for the old men is not different from that of the young men for either muscle group.
Historical perspectives on lung edema clearance
Authors: Crandall Edward D. , Effros Richard M.
Published: March 2015

Early studies of fluid transport across the pulmonary epithelium were conducted in intact animals or isolated lungs. Although the location and cells responsible for transport cannot be determined with studies in whole mammalian lungs, such preparations remain indispensable for determining the physiological and clinical relevance of in vitro investigations of cells and their transport proteins. Three different approaches have been used to study transport and exchange between the vascular and air space compartments in intact lungs. Some of the advantages and limitations of these methods are briefly reviewed here.
Attenuated hepatosplanchnic uptake of lactate during intense exercise in humans
Authors: Nielsen H. B. , Clemmesen J. O. , Skak C. , Ott P. , Secher N. H.
Published: March 2015

We evaluated whether the increase in blood lactate with intense exercise is influenced by a low hepatosplanchnic blood flow as assessed by indocyanine green dye elimination and blood sampling from an artery and the hepatic vein in eight men. The hepatosplanchnic blood flow decreased from a resting value of 1.6 ± 0.1 to 0.7 ± 0.1 (SE) l/min during exercise. Yet the hepatosplanchnic O2uptake increased from 67 ± 3 to 93 ± 13 ml/min, and the output of glucose increased from 1.1 ± 0.1 to 2.1 ± 0.3 mmol/min ( P < 0.05). Even at the lowest hepatosplanchnic venous hemoglobin O2 saturation during exercise of 6%, the average concentration of glucose in arterial blood was maintained close to the resting level (5.2 ± 0.2 vs. 5.5 ± 0.2 mmol/l), whereas the difference between arterial and hepatic venous blood glucose increased to a maximum of 22 mmol/l. In arterial blood, the concentration of lactate increased from 1.1 ± 0.2 to 6.0 ± 1.0 mmol/l, and the hepatosplanchnic uptake of lactate was elevated from 0.4 ± 0.06 to 1.0 ± 0.05 mmol/min during exercise ( P < 0.05). However, when the hepatosplanchnic venous hemoglobin O2 saturation became low, the arterial and hepatosplanchnic venous blood lactate difference approached zero. Even with a marked reduction in its blood flow, exercise did not challenge the ability of the liver to maintain blood glucose homeostasis. However, it appeared that the contribution of the Cori cycle decreased, and the accumulation of lactate in blood became influenced by the reduced hepatosplanchnic blood flow.
Effect of exercise training at different intensities on fat metabolism of obese men
Authors: Van Aggel-Leijssen Dorien P. C. , Saris Wim H. M. , Wagenmakers Anton J. M. , Senden Joan M. , Van Baak Marleen A.
Published: March 2015

The present study investigated the effect of exercise training at different intensities on fat oxidation in obese men. Twenty-four healthy male obese subjects were randomly divided in either a low- [40% maximal oxygen consumption (V˙o 2 max)] or high-intensity exercise training program (70%V˙o 2 max) for 12 wk, or a nonexercising control group. Before and after the intervention, measurements of fat metabolism at rest and during exercise were performed by using indirect calorimetry, [U-13C]palmitate, and [1,2-13C]acetate. Furthermore, body composition and maximal aerobic capacity were measured. Total fat oxidation did not change at rest in any group. During exercise, after low-intensity exercise training, fat oxidation was increased by 40% ( P < 0.05) because of an increased non-plasma fatty acid oxidation ( P < 0.05). High-intensity exercise training did not affect total fat oxidation during exercise. Changes in fat oxidation were not significantly different among groups. It was concluded that low-intensity exercise training in obese subjects seemed to increase fat oxidation during exercise but not at rest. No effect of high-intensity exercise training on fat oxidation could be shown.
Regulation of TGF-β ligand and receptor expression in neonatal rat lungs exposed to chronic hypoxia
Authors: Vicencio Alfin G. , Eickelberg Oliver , Stankewich Michael C. , Kashgarian Michael , Haddad Gabriel G.
Published: March 2015

Long-term effects of hypoxia are largely due to its modulatory effects on proliferation and differentiation of epithelial and endothelial cells, processes also regulated by the transforming growth factor (TGF)-β system. We investigated the effects of hypoxia on the TGF-β system in rat lungs from different developmental stages. Sprague-Dawley rats were exposed to 9.5% oxygen during either the first 2 wk of life or adulthood. Analysis revealed an arrest of alveolarization in hypoxic postnatal day 14 rats. Bioactive TGF-β levels in bronchoalveolar lavage fluid were increased in these animals, and Western blot analysis revealed upregulation of TGF-β receptor (TβR) I and II. None of these changes was observed in hypoxic adults. Hypoxia did, however, lead to decreased expression of TβRIII in both postnatal day 14 and adult rats. Immunohistochemical analysis localized TβRI-III predominantly to bronchiolar and alveolar epithelium; these patterns did not change with hypoxia. Thus we observed changes in TGF-β activity and TβR isotype expression in rat lung that parallel the arrest in alveolarization seen with chronic hypoxia in early development. These alterations may partly explain the morphological changes observed in hypoxia.