Calcineurin-inhibition Results in Upregulation of Local Renin and Subsequent Vascular Endothelial Growth Factor Production in Renal Collecting Ducts.

BACKGROUND: Tacrolimus (Tac) and Cyclosporine A (CyA) calcineurin inhibitors (CNIs) are 2 effective immunosuppressants which are essential to prevent allograft rejection. Calcineurin inhibitors are known to be nephrotoxic. However, the precise mechanism of nephrotoxicity is not fully understood. In this study, we investigated the in vivo effects of CNIs on the local renal renin-angiotensin system in the collecting duct (CD). METHODS: Three-week-old mice were treated with either vehicle, CyA (2 mg/kg per day), Tac (0.075 mg/kg per day), CyA + Aliskiren (25 mg/kg per day), or Tac + Aliskiren for 3 weeks. Serum creatinine was measured. Renin and vascular endothelial growth factor (VEGF) contents in CD were evaluated with flow cytometry and multiphoton microscopy. The diameter of vessels was assessed with multiphoton microscopy, and the amount of renal collagen was determined by real-time polymerase chain reaction and Masson staining. RESULTS: The elevated level of serum creatinine in CNI groups was abolished by Aliskiren. Flow cytometric analysis found elevated renin content in principal cells, which was prevented by Aliskiren. This result was further confirmed with multiphoton microscopy. The VEGF content in CD correlated with reduced capillary diameter and with the formation of fibrotic islands. CONCLUSIONS: Calcineurin inhibitors induce production of renin in the CD that may contribute to decreased renal blood flow. In turn, CD responds with increased VEGF production, resulting in disproportional vessel growth, further worsening the local hypoxia and striped fibrosis surrounding the CDs. Aliskiren, a direct renin inhibitor blocks these effects and improves CNI-induced nephropathy by decreasing renin production in the CDs. Our data suggest that Aliskiren may be used for the prevention of CNI nephrotoxicity.

Architecture of vasa recta in the renal inner medulla of the desert rodent Dipodomys merriami: potential impact on the urine concentrating mechanism.

We hypothesize that the inner medulla of the kangaroo rat Dipodomys merriami, a desert rodent that concentrates its urine to over 6,000 mosmol/kg H(2)O, provides unique examples of architectural features necessary for production of highly concentrated urine. To investigate this architecture, inner medullary vascular segments in the outer inner medulla were assessed with immunofluorescence and digital reconstructions from tissue sections. Descending vasa recta (DVR) expressing the urea transporter UT-B and the water channel aquaporin 1 lie at the periphery of groups of collecting ducts (CDs) that coalesce in their descent through the inner medulla. Ascending vasa recta (AVR) lie inside and outside groups of CDs. DVR peel away from vascular bundles at a uniform rate as they descend the inner medulla, and feed into networks of AVR that are associated with organized clusters of CDs. These AVR form interstitial nodal spaces, with each space composed of a single CD, two AVR, and one or more ascending thin limbs or prebend segments, an architecture that may lead to solute compartmentation and fluid fluxes essential to the urine concentrating mechanism. Although we have identified several apparent differences, the tubulovascular architecture of the kangaroo rat inner medulla is remarkably similar to that of the Munich Wistar rat at the level of our analyses. More detailed studies are required for identifying interspecies functional differences.

Connecting tubule glomerular feedback mediates acute tubuloglomerular feedback resetting.

Tubuloglomerular feedback (TGF) and connecting tubule glomerular feedback (CTGF) are mechanisms that control afferent arteriole (Af-Art) tone. TGF, initiated by increased NaCl at the macula densa, causes Af-Art constriction. Prolonged activation of TGF leads to an attenuation or "resetting" of its constrictor effect. The mechanisms of TGF resetting remain incompletely understood. CTGF is initiated by increased NaCl in the connecting tubule and Na(+) entry via epithelial sodium channels (ENaC). Contrary to TGF, CTGF dilates the Af-Art. Here, we hypothesize that CTGF, in part, mediates TGF resetting. We performed micropuncture of individual rat nephrons while measuring stop-flow pressure (P(SF)), an index of glomerular filtration pressure and Af-Art tone. Increases in Af-Art tone cause P(SF) to decrease. TGF responses, measured as the decrease in P(SF) induced by switching late proximal tubule perfusion from 5 to 40 nl/min, were elicited before and after a 30-min period of sustained perfusion of the late proximal tubule at a rate of 40 nl/min designed to induce TGF resetting. TGF responses were 7.3 ± 0.3 and 4.9 ± 0.2 mmHg before and after resetting was induced (P < 0.001, n = 6). When CTGF was inhibited with the ENaC blocker benzamil (1 µM), TGF responses were 9.5 ± 0.3 and 8.8 ± 0.6 mmHg (NS, n = 6), thus resetting was abolished. In the presence of the carbonic anhydrase inhibitor acetazolamide (10 mM), TGF responses were 8.8 ± 0.6 and 3.3 ± 0.4 mmHg before and after resetting (P < 0.001, n = 6). With both acetazolamide and benzamil, TGF responses were 10.4 ± 0.2 and 8.4 ± 0.5 mmHg (P < 0.01, n = 6), thus resetting was attenuated. We conclude that CTGF, in part, mediates acutely induced TGF resetting.

Effect of reduced renal mass on renal ammonia transporter family, Rh C glycoprotein and Rh B glycoprotein, expression.

Kidneys can maintain acid-base homeostasis, despite reduced renal mass, through adaptive changes in net acid excretion, of which ammonia excretion is the predominant component. The present study examines whether these adaptations are associated with changes in the ammonia transporter family members, Rh B glycoprotein (Rhbg) and Rh C glycoprotein (Rhcg). We used normal Sprague-Dawley rats and a 5/6 ablation-infarction model of reduced renal mass; control rats underwent sham operation. After 1 wk, glomerular filtration rate, assessed as creatinine clearance, was decreased, serum bicarbonate was slightly increased, and Na(+) and K(+) were unchanged. Total urinary ammonia excretion was unchanged, but urinary ammonia adjusted for creatinine clearance, an index of per nephron ammonia metabolism, increased significantly. Although reduced renal mass did not alter total Rhcg protein expression, both light microscopy and immunohistochemistry with quantitative morphometric analysis demonstrated hypertrophy of both intercalated cells and principal cells in the cortical and outer medullary collecting duct that was associated with increased apical and basolateral Rhcg polarization. Rhbg expression, analyzed using immunoblot analysis, immunohistochemistry, and measurement of cell-specific expression, was unchanged. We conclude that altered subcellular localization of Rhcg contributes to adaptive changes in single-nephron ammonia metabolism and maintenance of acid-base homeostasis in response to reduced renal mass.

Computer analysis of the significance of the effective osmolality for urea across the inner medullary collecting duct in the operation of a single effect for the counter-current multiplication system.

BACKGROUND: Although urea and water are transported across separate pathways in the apical membrane of the inner medullary collecting duct (IMCD), the existence of a cellular diffusion barrier as an unstirred layer makes it possible to use coefficients of effective osmotic force (sigma*) as equivalent to reflection coefficients. The difference in effective osmolality between urea and NaCl across the IMCD becomes a driving force for water if the compositions of solutes are different between tubular lumen and interstitium. Since reported values for sigma*(urea) are discrepant, we compared the efficiency of a single effect in the counter-current system between an ascending thin limb (ATL) and the IMCD, with the interposition of capillary networks (CNW), between two models with sigma(urea)* = 0.7 (model 1) and sigma(urea)* = 1.0 (model 2). METHODS: The time courses (within 3 s) of solute and the water transport profiles among ATL, CNW, and IMCD were simulated with a computer in the absence of flow in each compartment. RESULTS: In spite of small differences in the profiles of urea and NaCl concentrations between the two models, model 1 displayed a larger volume flux in the IMCD than model 2, resulting in an increase of osmolality in the IMCD and a decrease of osmolality in the ATL. These findings are vital for the operation of the counter-current multiplication system. CONCLUSIONS: The concept of coefficients for effective osmotic force can be applied to the counter-current model between the IMCD and the ATL with the interposition of CNW. The model of sigma(urea)* = 0.7 is more efficient than that of sigma(urea)* = 1.0.

Pig kidney: anatomical relationships between the intrarenal arteries and the kidney collecting system. Applied study for urological research and surgical training.

PURPOSE: We present a systematic study of the anatomical relationships between intrarenal arteries and the kidney collecting system in pigs. MATERIALS AND METHODS: The intrarenal anatomy (collecting system and arteries) was studied in 91, 3-dimensional endocasts of the kidney collecting system together with the intrarenal arteries. RESULTS: Some anatomical details that have importance to help research and surgical training in urology when using the pig as an animal model were observed and described. It was found that there was only 1 artery per kidney. This artery divided into cranial and caudal branches in 85 cases (93.4%). In 6 cases (6.6%) the primary division of the renal artery was in a dorsal and in a ventral branch. In all cases 2 arteries (1 ventral and 1 dorsal) involved the cranial caliceal group. In the dorsal mid zone a dorsal artery originated from the cranial division of the renal artery, which was obliquely positioned in 47.25% of cases. The arterial supply related to the ventral mid zone of the kidney consisted of branches that coursed horizontally in the ventral surface of the renal pelvis in 81.32% of cases. The caudal division of the renal artery supplied the ventral and dorsal surfaces of the caudal caliceal group in 84.62% of cases, while in 15.38% a dorsal artery supplied its dorsal surface. CONCLUSIONS: Although the results of renal and intrarenal anatomy in pigs could not be completely transposed to humans, many similarities in the pig and human intrarenal arteries support its use as the best animal model for urological procedures.

Distribution of cytochrome P-450 4A and 4F isoforms along the nephron in mice.

The production of 20-hydroxyeicosatetraenoic acid (20-HETE) in the kidney is thought to be involved in the control of renal vascular tone and tubular sodium and chloride reabsorption. 20-HETE production in the kidney has been extensively studied in rats and humans and occurs primarily via the actions of P-450 enzymes of the CYP4A and -4F families. Recent advancements in molecular genetics of the mouse have made it possible to disrupt genes in a cell-type-specific fashion. These advances could help in the creation of models that could distinguish between the vascular and tubular actions of 20-HETE. However, isoforms of the CYP4A and -4F families that may be responsible for the production of 20-HETE in the vascular and tubular segments in the kidney of the mouse are presently unknown. The goal of this study was to identify the isoforms of the CYP4A and -4F families along the nephron by RT-PCR of RNA isolated from microdissected renal blood vessels and nephron segments from 16- to 24-wk-old male and female C57BL/6J mice. CYP4A and -4F isoforms were detected in every segment analyzed, with sex differences only observed in the proximal tubule and glomeruli. In the proximal tubular segments from male mice, the 4A10 and -12 isoforms were present, whereas the 4A10 and -14 isoforms were detected in segments from female mice. In glomeruli, sex differences in the expression pattern of CYP4F isoforms were also observed, with male mice expressing the 4F13, -14, and -15 isoforms, whereas female mice expressed the 4F13, -16, and -18 isoforms. These results demonstrate that isolated nephron and renal vessel segments express multiple isoforms of the CYP4A and -4F families; therefore, elimination of a single CYP4A or -4F isoform may not decrease 20-HETE production in all nephron segments or the renal vasculature of male and female mice. However, the importance of CYP4A vs. -4F isoforms to the production of 20-HETE in each of these renal tubular and vascular segments of the mouse remains to be determined.

Tubulovascular nitric oxide crosstalk: buffering of angiotensin II-induced medullary vasoconstriction.

Studies were designed to determine the source of NO responsible for buffering of the angiotensin II (Ang II)-mediated decrease of blood flow in the renal medulla. Intracellular Ca2+ concentration ([Ca2+]i) and NO production ([NO]i) of pericytes and endothelium of the vasa recta were independently measured with the use of fura 2-AM and 4,5-diaminofluorescein diacetate (DAF-2DA), respectively, in microtissue strips of the vascular bundles of the outer medullary vasa recta. Disruption of the endothelium of the vasa recta by perfusion with latex microspheres enabled imaging of the pericytes. Ang II (1 micromol/L) produced an increase of [NO]i of 19+/-6 U in pericytes of the vasa recta when the vessels were adjacent to medullary thick ascending limbs (mTALs). Pericytes of isolated vasa recta without surrounding mTALs showed a rapid peak increase in [Ca2+]i of 248+/-107 nmol/L, with a sustained elevation of 107+/-75 nmol/L, but did not show an increase in [NO]i to either Ang II (1 micromol/L) or the Ca2+ ionophore 4-bromo-A23187 (5 micromol/L). These observations indicated the lack of Ang II and Ca2+-sensitive NO production in pericytes of the vasa recta. In isolated vasa recta with intact endothelium, Ang II reduced [Ca2+]i from 128+/-28 to 62+/-13 nmol/L and failed to increase [NO]i. However, the Ca2+ ionophore did increase [NO]i in the endothelium (47+/-8 U), indicating the presence of Ca2+-sensitive NO production. Significant increases of [NO]i were observed in single isolated mTALs in response to both Ang II (33+/-6 U) and the Ca2+ ionophore (51+/-18 U). We conclude that Ang II increases [Ca2+]i in pericytes of the descending vasa recta as part of its constrictor action and that this vasoconstriction is buffered by the NO from the surrounding tubular elements, such as mTALs.

Histological features of hypovascular or avascular renal cell carcinoma: the experience at four university hospitals.

BACKGROUND: We aimed to evaluate the pathological features of hypovascular or avascular renal cell carcinoma (RCC), by the retrospective review of the histological features that coincide with hypovascular or avascular RCC. METHODS: Seven hundred and ninety-one patients who underwent both preoperative angiography and nephrectomy were examined. Of these patients, the 126 patients (15.9%) who showed hypovascular or avascular angiographic features were selected. Patients with hemorrhage or cyst(s), or with at least 50% necrosis or more in the tumor, and those with renal tumor metastatic to the kidney were excluded. The criteria proposed by the World Health Organization (1998) were adopted for the histological classification. RESULTS: Papillary RCC was the most frequently observed hypovascular or avascular renal tumor (44 cases; 34.9%). The vascularity differed among the variants, i.e., some cases had a basophilic and solid variant with avascular features, while the remaining cases had wide stromal organization showing hypovascular features. The second most frequently observed hyporascular or avascular RCC was chromophobe cell carcinoma (35 cases; 27.8%). No difference in vascularity was detected between variants, except for 2 cases with sarcomatoid changes (avascular features). The third most frequently observed hypovascular or avascular RCC was cyst-associated RCC (29 cases; 23%). All of the 7 RCCs originating in a cyst showed avascular features, and the remaining 22 cystic RCCs showed hypovascular features. The remaining hypovascular or avascular RCCs were cases of clear cell carcinoma accompanied by sarcomatoid changes (8 cases; 6.3%), spindle cell carcinoma (5 cases; 4.0%), and collecting-duct carcinoma (5 cases; 4.0%). CONCLUSION: Hypovascular or avascular RCC can be categorized as non-clear cell carcinoma and some clear cell carcinoma accompanied by sarcomatoid changes.

Interstitial water and solute recovery by inner medullary vasa recta.

A recent model of volume and solute microvascular exchange in the renal medulla was extended by simulating the deposition of NaCl, urea, and water into the medullary interstitium from the loops of Henle and collecting ducts with generation rates that undergo spatial variation within the inner medullary interstitium. To build an exponential osmolality gradient in the inner medulla, as suggested by Koepsell et al. (H. Koepsell, W. E. A. P. Nicholson, W. Kriz, and H. J. Höhling. Pflügers Arch. 350: 167-184, 1974), the ratio of the interstitial area-weighted generation rate of small solutes to that of water must increase along the corticomedullary axis. We satisfied this condition either by holding the area-weighted generation rate of water constant while increasing that of NaCl and urea or by reducing the input rate of water with medullary depth. The latter case, in particular, yielded higher solute concentrations at the papillary tip. Assuming that the fraction of the filtered load recovered by inner medullary vasa recta for water, NaCl, and urea is 1%, 1%, and 40%, respectively, papillary tip osmolality is 1,470 mosmol/kgH(2)O when urea generation and NaCl generation per unit volume of interstitium increase exponentially and linearly, respectively. The inner medullary osmolar gradient also increases further when 1) medullary blood flow is reduced, 2) hydraulic conductivity of descending vasa recta (DVR) is lowered, and 3) vasa recta permeability to NaCl and urea is maximized. The coupling between water and small solute transport, resulting from aquaporin-1-mediated transcellular flux in DVR, also enhances tip osmolality.

Utility of the resistance index ratio in differentiating obstructive from nonobstructive hydronephrosis in children.

PURPOSE: We assessed the utility of the resistance index ratio (RIR) in distinguishing between obstructive and nonobstructive upper urinary tract dilatation in children. METHODS: Twenty-three children (7 days-14 years old) with unilateral dilated collecting systems and a contralateral normal kidney were prospectively evaluated by duplex Doppler sonography. We measured the resistance index (RI) of the intrarenal arteries and calculated the RIR. Ninety-six normal kidneys in 48 controls were also evaluated. RESULTS: Twelve kidneys were proved to be obstructed at the ureteropelvic junction, and 11 were found by renal scintigraphy with furosemide and/or by surgery to have nonobstructive dilatation. The mean RIR differed significantly between the obstructed and dilated nonobstructed kidneys (1.16+/-0.04 versus 1.04+/-0.04, respectively; p < 0.01). After surgical correction of ureteropelvic junction obstruction, the obstructed kidneys showed significant drops in the RIR (1.18+/-0.03 to 1.07+/-0.03, p < 0.01). The RIR showed no statistically significant relationship with age (r = -0.268, p > 0.01); however, the RI declined with increasing age (r= -0.414, p < 0.01). An RIR cut-off value of > or = 1.10 provided a good discriminatory level, resulting in a sensitivity of 92%, a specificity of 97%, a positive predictive value of 85%, and a negative predictive value of 97%. CONCLUSIONS: An RIR cut-off value of > or = 1.10 appears to be an effective parameter for evaluation and follow-up of unilateral obstructive hydronephrosis in children.

Acute obstruction of the renal collecting system: the intrarenal resistive index is a useful yet time-dependent parameter for diagnosis.

The aim of this study was to determine whether the intrarenal resistive index (RI) can be used for the diagnosis of acute obstruction in patients with renal colic and to determine whether the index is time-related. Seventy patients referred to the Emergency Department with acute renal colic and without known associated renal disease underwent duplex Doppler ultrasonography to determine the intrarenal RI at the symptomatic and asymptomatic side. The age range of the patients was 18-72 years. An RI greater than 0.68 and/or an interrenal difference in RI greater than 0.06 and/or an increase in RI of more than 11% compared with the normal side proved reliable cut-off values to diagnose acute renal obstruction. In addition, time dependency of the increase in RI was noted. No significant differences were observed within the first 6 h after the onset of symptoms. From 6 to 48 h, however, the mean RI in the affected kidney (0.70 +/- 0.06; mean +/- SD) was significantly different from that in the normal kidney (0.59 +/- 0.04) (P < 0.001). In the same period the mean difference in RI was 0.08-0.13 (P < 0.001). After 48 h the sensitivity of RI dropped substantially. It is concluded that renal duplex Doppler ultrasonography is useful for diagnosing acute renal obstruction between 6 and 48 h after the onset of symptoms.

The central vessel of the renal countercurrent bundles of two marine elasmobranchs--dogfish (Scyliorhinus caniculus) and skate (Raja erinacea)--as revealed by light and electron microscopy with computer-assisted reconstruction.

The renal countercurrent bundles of elasmobranch fish were studied by light and electron microscopy. The kidneys of the lesser spotted dogfish, Scyliorhinus caniculus Blainville, and the little skate, Raja erinacea Mitchill, were investigated. Three-dimensional reconstruction with computer assistance revealed the spatial association of the renal tubular segments and their relationships to each other, as well as to the microvasculature. Regular association between structures was assessed by quantification of contact points on histological sections. The bundles contain a hairpin loop of neck segment and the beginning of the proximal tubule, PIa. The limbs of this loop closely adhere to each other, and a second loop (the early distal tubule) coils around the first loop at the tip of the bundle. The collecting tubule runs between the two loops, and merges with the collecting duct inside the end portion of the bundle. A single lymph capillary-like vessel originates from a few blind-ended rami at the tip of the bundle and runs in close contact with the collecting tubule along the entire bundle. This central vessel merges via several side branches with the venous sinusoid capillaries of the peritubular blood circulation. Thereby the central vessel provides a channel for convective flow of NaCl-rich fluid unidirectionally to the venous portal system of the mesial tissue zone of the kidney. By the close spatial arrangement of the collecting tubule and the central vessel countercurrent exchange of urea from the collecting tubule urine to the fluid in the central vessel is feasible. Thus, the spatial organisation of renal tubular segments and the central vessel is considered to represent the morphological correlate to urea retention by the kidney of Elasmobranchii.

Endothelin production in the rabbit collecting ducts of acute renal failure models. An immunohistochemical study.

Endothelin 1 (ET-1) production was examined in the rabbit nephron in acute renal failure (ARF) induced by uranyl acetate administration or by clamping the renal artery. Uranyl acetate dissolved in saline was injected intravenously at a dose of 0.8 mg/kg (n = 12). In the ischemic kidney experiment, 60 min of left renal artery clamping was carried out 1 week after removing the right kidney (n = 8). Plasma and urine concentrations of ET-1 were measured 0, 24, and 48 h after treatment, and immunohistochemical studies of renal tissues obtained 48 h after the experiments were carried out using ET-1 monoclonal antibody. The fractional excretion of ET-1 increased from 10 to 89% at 48 h in the uranyl acetate treated group and from 6 to 15% at 24 h in the renal artery clamping group, suggesting the existence of ET-1 secretion from the nephron in both types of ARF. By immunohistochemical examination, strong ET-1 expression was noted only in the collecting ducts. No staining was observed in other parts of the nephron in both types of ARF. The present study indicates that the increased expression of ET-1 probably reflects production by the collecting ducts in both rabbit ARF models.

Compensatory hypertrophy and adaptation in the cortical collecting duct.

The cortical collecting duct (CCD) undergoes hypertrophy and functional adaptation following reduction of renal mass. The nature and mechanisms of these changes have been investigated using microperfusion of isolated CCD from rabbit remnant kidneys. By 1 week after reduction of renal mass, tubule hypertrophy and increased sodium transport are fully developed. The transport adaptations are specific or selective, since bicarbonate transport in these CCD is unchanged. Mineralocorticoids may play an important role in the hypertrophy and increased sodium transport, since plasma aldosterone increases early after reduction of renal mass. Also, adrenalectomy abolishes the changes in size and sodium transport, even with supplementation of aldosterone to unstressed physiologic levels. Epidermal growth factor also has immediate effects on CCD sodium transport; however, the direction of the effect is opposite--an inhibition of transport.

Grade I hydronephrosis: pulsed Doppler US evaluation.

This study was undertaken to determine whether the presence of blood vessels could mimic the appearance of grade I hydronephrosis on sonograms and thus cause false-positive readings. One hundred consecutive patients with grade I hydronephrosis were examined. Sample volumes were obtained with pulsed Doppler ultrasonography (US) at the site of the greatest separation of the central renal sinus echoes to determine if the separation was fluid accumulating in the collecting system, as in obstruction, or if the separation was actually caused by vessels that mimic hydronephrosis. Vascular structures accounted for the separation of the sinus echoes in 43% of patients. In patients 12 years of age or younger, this frequency rose to 61%. The simple procedure of evaluating the renal sinus echo separation with pulsed Doppler US should decrease the frequency of false-positive diagnoses of hydronephrosis and thus diminish the need for further confirmatory testing.