H+-ATPase activity on unilateral ureteral obstruction: interaction of endogenous nitric oxide and angiotensin II.
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BACKGROUND: A number of cytokines, vasoactive compounds, chemoattractant molecules, and growth factors are up-regulated in obstruction. Following the onset of ureteral obstruction, angiotensin II production is rapidly stimulated. Cytokine-induced expression of inducible nitric oxide synthase (iNOS) has been reported in primary cultures of inner medullary collecting duct (IMCD) cells. We found that the defective urinary acidification in unilateral ureteral obstruction (UUO) includes an intensive decrease in bafilomycin-sensitive H+-ATPase activity in microdissected IMCD segments. METHODS: To investigate the interaction between endogenous nitric oxide and angiotensin II on H+-ATPase activity, we used microdissected IMCD segments of unilaterally obstructed, contralateral, and control kidneys to measure the bafilomycin-sensitive ATPase activity and nitric oxide synthase (NOS) activity. The generated NO was also evaluated. RESULTS: Preincubation of obstructed IMCD segments in the presence of a competitive inhibitor of NOS, NG-nitro-L-arginine methyl ester (L-NAME) 1 mmol/L, and in the presence of a specific inhibitor of calcium/calmodulin-independent NOS (iNOS), aminoguanidine 1 mmol/L, each for 60 minutes, significantly increased bafilomycin-sensitive H+-ATPase. A greater increase on iNOS activity (fmol [3H] citrulline/min/microg protein) and a lesser increase in calcium/calmodulin-dependent NOS activity (cNOS) were observed in the obstructed renal medulla. This inhibitory effect of obstruction was abolished when IMCDs were incubated with 10-5 to 10-8 mol/L losartan. Decreasing doses of the angiotensin II type 1 (AT1) receptor inhibitor caused an increase in bafilomycin-sensitive H+-ATPase, with a maximum increase at 10-8 mol/L losartan. A decrease on iNOS activity was demonstrated in the obstructed renal medulla incubated with losartan in concentrations of 10-5 to 10-8 mol/L, the same losartan concentrations that showed recovery of vacuolar H+-ATPase activity. Similarly, a decrease on the generation of NO after incubation with losartan 10-5 to 10-8 mol/L was shown. CONCLUSION: From these results, we suggest that endogenous NO increased by iNOS is involved in the inhibition of H+-ATPase activity in obstructed IMCD segments. The recovery of H+-ATPase activity in IMCD of obstructed kidneys induced by losartan may be related to a decrease of inducible NOS activity. (+info)
BACKGROUND: Interstitial expression of transforming growth factor-beta1 (TGF-beta1) is important in tubulointerstitial fibrosis, a common process in most progressive renal diseases. However, no effective therapy for progressive interstitial fibrosis is known. Recently, we developed an artificial viral envelope (AVE)-type hemagglutinating virus of Japan (HVJ) liposome-mediated retrograde ureteral gene transfer method, which allowed us to introduce the genetic material selectively into renal interstitial fibroblasts. METHOD: We introduced antisense or scrambled oligodeoxynucleotides (ODNs) for TGF-beta 1 into interstitial fibroblasts in rats with unilateral ureteral obstruction, a model of interstitial fibrosis, to block interstitial fibrosis by retrograde ureteral injection of AVE-type HVJ liposomes. RESULTS: TGF-beta 1 and type I collagen mRNA increased markedly in the interstitium of untreated obstructed kidneys, and those were not affected by scrambled ODN transfection. Northern analysis and in situ hybridization revealed that the levels of TGF-beta 1 and type I collagen mRNA were dramatically decreased in antisense ODN-transfected obstructed kidneys. Consequently, the interstitial fibrotic area of the obstructed kidneys treated with antisense ODN was significantly less than that of the obstructed kidneys untreated or treated with scrambled ODN. CONCLUSION: The introduction of TGF-beta 1 antisense ODN into interstitial fibroblasts may be a potential therapeutic maneuver for interstitial fibrosis. (+info)
Unilateral ureteral obstruction in neonatal rats leads to renal insufficiency in adulthood.
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BACKGROUND: Although unilateral ureteropelvic junction obstruction is the most common cause of congenital obstructive nephropathy in infants and children, management remains controversial, and follow-up after pyeloplasty is generally limited to the pediatric ages. We have developed a model of temporary unilateral ureteral obstruction (UUO) in the neonatal rat: One month following the relief of five-day UUO, the glomerular filtration rate (GFR) of the postobstructed kidney was normal despite a 40% reduction in the number of glomeruli and residual vascular, glomerular, tubular, and interstitial injury. METHODS: To determine whether hyperfiltration and residual injury of remaining nephrons leads to progression of renal insufficiency in later life, 31 rats were sham operated or subjected to left UUO at one day of age, with relief of UUO five days later, and were studied at one year of age. GFR was measured by inulin clearance, and the number of glomeruli, tubular atrophy, glomerular sclerosis, and interstitial fibrosis were measured by histomorphometry in sham, obstructed (UUO), and intact opposite kidneys. Intrarenal macrophages and alpha-smooth muscle actin were identified by immunohistochemistry. RESULTS: Despite relief of UUO, ultimate growth of the postobstructed kidney was impaired. The number of glomeruli was reduced by 40%, and GFR was decreased by 80%. However, despite significant compensatory growth of the opposite kidney, there was no compensatory increase in GFR, and proteinuria was increased. Moreover, glomerular sclerosis, tubular atrophy, macrophage infiltration, and interstitial fibrosis were significantly increased not only in the postobstructed kidney, but also in the opposite kidney. CONCLUSIONS: Although GFR is initially maintained following relief of five-day UUO in the neonatal rat, there is eventual profound loss of function of the postobstructed and opposite kidneys because of progressive tubulointerstitial and glomerular damage. These findings suggest that despite normal postoperative GFR in infancy, children undergoing pyeloplasty for ureteropelvic junction obstruction should be followed into adulthood. Elucidation of the cellular response to temporary UUO may lead to improved methods to assess renal growth, injury, and functional reserve in patients with congenital obstructive nephropathy. (+info)
Salutary role for angiotensin in partial urinary tract obstruction.
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BACKGROUND: In the neonatal period, angiotensin II (Ang II) is up-regulated and induces a timely development of the renal pelvis and ureteral peristalsis, thereby protecting the kidney from hydronephrosis. We tested the possibility that in adulthood, Ang II may act salutarily on the kidney structure during partial urinary tract obstruction by inducing adaptive changes in the peristaltic machinery. METHODS: Adult male Sprague-Dawley rats were subjected to partial unilateral ureteral obstruction (UUO) and divided into two groups, that is, those treated with (group L, N = 21) and those without (group C, N = 21) an angiotensin type 1 (AT1) receptor antagonist (losartan). Control animals were sham operated (N = 10). Rats were sacrificed either at day 7 or day 14. RESULTS: The degree of hydronephrosis determined morphometrically was significantly more severe in group L than group C at both day 7 and day 14, indicating that Ang II inhibition accentuated hydronephrosis. The measurement of upstream pressure within the partially ligated ureter in vivo revealed that losartan significantly attenuates the frequency of ureteral peristaltic activities. In in vitro studies using ureteral strips harvested from normal adult Sprague-Dawley rats (N = 10), Ang II (10(-8) mol/L) was shown to augment contraction, which was completely inhibited by losartan (10(-6) mol/L). CONCLUSIONS: Ang II has a salutary effect of protecting kidneys from hydronephrosis during partial ureteral obstruction through its ability to augment ureteral peristalsis. (+info)
MAG3-F0 scintigraphy in decision making for emergency intervention in renal colic after helical CT positive for a urolith.
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Patients with renal colic are evaluated with clinical, laboratory, and imaging methods for stratification for emergency decompression, medical treatment, or discharge and follow up. The current standard practice is heavily based on unenhanced helical CT for detecting uroliths. However, the presence of a urolith does not necessarily mean that the kidney is obstructed and requires emergency decompression. In this study, technetium-mercaptoacetyltriglycine (MAG3) diuretic scintirenography was used to detect obstruction in patients with renal colic. The contribution of this test to patient management after positive findings from helical CT was also studied. METHODS: Diagnostic criteria were established on the basis of previous experience with 60 patients who had renal colic and had undergone radiography of the kidneys, ureters, and urinary bladder (KUB) and diuretic Tc-MAG3 scintirenography and were followed up to correlate scintigraphic findings with clinical outcome. Subsequently, 80 patients with renal colic underwent scintigraphy within 12 h of presentation in the emergency room, after abdominal helical CT showed findings positive for calculus and suggestive of obstruction. After therapeutic oral or intravenous hydration and analgesics, diuretic dynamic renal scintigraphy (flow, function, delayed imaging) was performed after intravenous injections of 10 mCi (370 MBq) 99mTc-MAG3 and 40 mg furosemide (at zero time, or F0). Results were available soon after completion of the study and were considered in patient management. Four characteristic patterns of scintirenography, essential in patient stratification and treatment, had been standardized and were used for interpretation of the studies: the unobstructed kidney; the partially obstructed kidney, proximally or distally obstructed, with mild to severe obstruction and impairment of function; the totally obstructed kidney, with arrested renal function; and the unobstructed but dysfunctioning kidney after decompression, or stunned kidney. RESULTS: Among the 80 patients with positive helical CT findings, 56.5% were found to have obstruction by scintigraphy (32.5% partially, 24% completely); the remaining 43.5% did not have obstruction (21% without an indication of recent obstruction and 22.5% with stunned kidneys after spontaneous decompression). Occasionally, findings of preexistent urine extravasation or infection were present. Patients who, by scintigraphy, never had obstruction or had experienced spontaneous decompression did not require admission or emergency intervention; those with complete or severe obstruction required admission and decompression for relief of pain or restoration of function, whereas those with mild obstruction were treated variably with forced fluids, analgesics, or, less frequently, elective surgery. Outcome information from clinical examination, imaging, and interventional findings indicated that this stratification was successful. The test caused no side effects. CONCLUSION: For renal colic, clinical selection, KUB radiography, and even positive helical CT findings were all found to have a low positive predictive value for obstruction (in this study, 35%, 32%, and 56% respectively). Anatomic studies, including helical CT, should be followed by diuretic MAG3-F0 scintirenography to diagnose and quantify or exclude obstruction, detect spontaneous decompression, and appropriately stratify patients for emergency intervention, observation and medical therapy, or further work-up and discharge with referral to the clinic. (+info)
Antibody to transforming growth factor-beta ameliorates tubular apoptosis in unilateral ureteral obstruction.
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BACKGROUND: Unilateral ureteral obstruction (UUO) is characterized by progressive renal atrophy, renal interstitial fibrosis, an increase in renal transforming growth factor-beta (TGF-beta), and renal tubular apoptosis. The present study was undertaken to determine the effect of a monoclonal antibody to TGF-beta (1D11) in UUO. METHODS: Mechanical stretch was applied to tubular epithelial cells (NRK-52E) by a computer-assisted system. Three doses of 1D11 (either 0.5, 2, or 4 mg/rat) were administered to rats one day prior to UUO and every two days thereafter, and kidneys were harvested at day 13. Fibrosis was assessed by measuring tissue hydroxyproline and mRNA for collagen and fibronectin. Apoptosis was assessed with the terminal deoxy transferase uridine triphosphate nick end-labeling assay. TGF-beta levels were determined by bioassay. Western blot and immunostaining were used to identify proliferating cell nuclear antigen (PCNA), p53, bcl-2, and inducible nitric oxide synthase (iNOS). RESULTS: Stretch significantly induced apoptosis in NRK-52E cells, which was accompanied by an increased release of TGF-beta; 1D11 (10 microg/mL) totally inhibited stretch-induced apoptosis. Control obstructed kidney contained 20-fold higher TGF-beta as compared with its unobstructed kidney; 1D11 neutralized tissue TGF-beta of the obstructed kidney. Control obstructed kidney exhibited significantly more fibrosis and tubular apoptosis than its unobstructed counterpart, which was blunted by 1D11. In contrast, 1D11 significantly increased tubular proliferation. p53 immunostaining was localized to renal tubular nuclei of control obstructed kidney and was diminished by 1D11. In contrast, bcl-2 was up-regulated in the 1D11-treated obstructed kidney. Total NOS activity and iNOS activity of the obstructed kidney were increased by 1D11 treatment. CONCLUSION: The present study strongly suggests that an antibody to TGF-beta is a promising agent to prevent renal tubular fibrosis and apoptosis in UUO. (+info)
Genetically modified bone marrow-derived vehicle cells site specifically deliver an anti-inflammatory cytokine to inflamed interstitium of obstructive nephropathy.
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In this study, we used genetically modified bone marrow-derived CD11b(+)CD18(+) vehicle cells to deliver IL-1 receptor antagonist (IL-1ra) for treatment of inflamed renal interstitium in an animal model of unilateral ureteral obstruction (UUO). Vehicle cells that expressed the ICAM-1 ligands, CD11b and CD18, were obtained from bone marrow cells of DBA/2j mice and adenovirally transduced with the IL-1ra gene or glucocerebrosidase (GC) gene ex vivo. In kidneys treated to develop UUO, levels of ICAM-1, IL-1 beta, and IL-1R expression increased within 3 days compared with contralateral untreated kidneys in the same mice. Similarly, the macrophage infiltration in the cortical interstitium increased after 3 days in UUO kidneys, but not untreated kidneys. After UUO developed, DBA/2j mice were injected i.v. with either IL-1ra(+) vehicle cells (IL-1ra-treated mice) or GC(+) vehicle cells (GC-treated mice) at 24 h after UUO. Six days after the injection of these vehicle cells, marked increase of CD11b(+) IL-1ra(+) vehicle cells was observed in the ICAM-1-positive interstitium of UUO kidneys from IL-1ra-treated mice. In contrast, no CD11b(+) IL-1ra(+) cells appeared in ICAM-1-negative contralateral kidneys from these mice. Furthermore, the infiltration of macrophages (p < 0.001), expression of ICAM-1 (p < 0.005), and presence of alpha-smooth muscle actin (p = 0.005) in the interstitium of UUO kidneys were significantly decreased in IL-1ra-treated mice compared with GC-treated mice. These findings suggest that IL-1 may contribute to the development of renal interstitial injury and that our method can deliver a functioning gene encoding an antiinflammatory cytokine gene specifically at that site by interacting with local adhesion molecules. (+info)
Renal interstitial fibrosis is reduced in angiotensin II type 1a receptor-deficient mice.
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Unilateral ureteral obstruction (UUO) results in tubulointerstitial fibrosis of the affected kidney by stimulating the renin-angiotensin system. This study established a UUO model in angiotensin type 1a receptor (AT1a) deficient (mutant) mice to elucidate the role of angiotensin II through AT1a on the fibrosis of the obstructed kidney (OBK). The relative volume of the tubulointerstitium was measured by an image analyzer; deposition of collagen types III and IV and monocyte/macrophage infiltration were histologically examined using specific antibodies. Also determined were the mRNA levels of transforming growth factor-beta by Northern blot analysis. Nuclear factor-kappaB activity was assessed by gel shift assay. UUO in wild mice resulted in a marked expansion of relative volume of the tubulointerstitium, together with increased deposition of collagen types III and IV and number of infiltrated monocytes/macrophages in the interstitium, relative to sham-operated mice. In comparison, these changes were significantly lower in mutant mice with UUO. The mRNA level of transforming growth factor-beta was significantly higher in the OBK of wild mice with UUO compared with sham-operated mice. In contrast, the increase in mRNA level in the OBK of mutant mice was significantly less than in wild mice. Finally, UUO resulted in activation of nuclear factor-kappaB in wild mice but was inhibited in the OBK of mutant mice. The results provide direct evidence that angiotensin II acting via the AT1a plays a pivotal role in the development of tubulointerstitial fibrosis in UUO. (+info)