A pathophysiologic role for T lymphocytes in murine acute cisplatin nephrotoxicity

Recent evidence supports a role for an inflammatory pathogenesis of cisplatin nephrotoxicity, but immune cell-mediated mechanisms in this disease are still largely unknown. The role for T lymphocytes on cisplatin-induced acute kidney injury was examined with C57BL/6 T cell-deficient (nu/nu) mice and CD4- or CD8-deficient mice and their wild-type (WT) littermates. All mice received a single dose of cisplatin at 40 mg/kg (intraperitoneally) and were followed up for 72 h. At 72 h after cisplatin administration, T cell-deficient mice had a marked attenuation in renal dysfunction (serum creatinine 3.2+/-0.5 versus 0.8+/-0.1 mg/dl; P=0.007), kidney tubular injury (scores 1.44+/-0.15 versus 0.22+/-0.08; P<0.0001), and survival. Adoptive transfer of T cells into nu/nu mice followed by cisplatin enhanced renal dysfunction and tubular injury. The increase in renal myeloperoxidase activity after cisplatin administration was blunted in nu/nu mice. Renal TNF-alpha, IL-1beta, and keratinocyte-derived chemokine protein expression was increased in WT mice but not in nu/nu mice after cisplatin administration. T cell levels significantly increased in kidneys of WT mice after cisplatin administration as early as at 1 h, peaked at 12 h, and declined by 24 h. CD4- and, to a lesser degree, CD8-deficient mice were relatively protected from cisplatin-induced mortality and renal dysfunction compared with WT mice. These data demonstrate that T lymphocytes are direct mediators of experimental cisplatin nephrotoxicity. Targeting T lymphocytes could lead to improved ways to administer cisplatin safely to cancer patients.     

Tranilast ameliorates renal tubular damage in unilateral ureteral obstruction

PURPOSE: We determined whether tranilast, the anti-allergic agent N-(3, 4-dimethoxyciannamoyl)-anthranilic acid, would diminish renal transforming growth factor-beta (TGF-beta) levels in unilateral ureteral obstruction and concomitantly affect renal tubular apoptosis and proliferation in that condition. MATERIALS AND METHODS: Tranilast (150 mg./kg.) was administered to rats 1 day before unilateral ureteral obstruction and each day thereafter. Kidneys were harvested day 14 after unilateral ureteral obstruction. Tissue TGF-beta was measured by bioassay using mink lung epithelial cells. Renal tubular proliferation and apoptosis were detected by immunostaining proliferating cell nuclear antigen and the terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate nick end labeling assay, respectively. Fibrosis was assessed by measuring collagen deposition with trichrome stained slides. RESULTS: TGF-beta bioassay showed that obstructed kidneys in controls contained significantly higher mean TGF-beta plus or minus standard deviation than unobstructed kidneys in controls (73.7 +/- 13.6 versus 14.1 +/- 5.5 pg./mg. tissue) and tranilast significantly decreased tissue TGF-beta in obstructed kidneys (15.9 +/- 4.8 pg./mg. tissue). The terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate nick end labeling assay demonstrated that obstructed kidneys in controls had significantly more mean tubular apoptosis than the unobstructed counterparts (36.6 +/- 6.7 versus 5.8 +/- 5.5 nuclei per high power field) and tranilast significantly decreased mean renal tubular apoptosis in obstructed kidneys (16.2 +/- 1.7 nuclei per high power field). In addition, immunostaining proliferating cell nuclear antigen showed that obstructed kidneys in controls had significantly more mean renal tubular proliferation than unobstructed kidneys (20.7 +/- 3.4 versus 6.2 +/- 2.1 per high power field) and tranilast significantly increased proliferating renal tubules in obstructed and unobstructed kidneys (26.5 +/- 8.3 and 14.5 +/- 3.4 per high power field, respectively). Control obstructed kidneys exhibited significantly more fibrosis, which was also blunted by tranilast. CONCLUSIONS: Tranilast significantly decreases tissue TGF-beta, resulting in a reduction in tubular apoptosis and an increase in tubular proliferation. This finding suggests that tranilast is a promising agent for preventing renal tubular damage in unilateral ureteral obstruction.     

Does cyclooxygenase-2 inhibitor prevent renal tissue damage in unilateral ureteral obstruction?

PURPOSE: We determined whether the cyclooxygenase-2 inhibitor etodolac affects renal tubular damage and interstitial fibrosis in unilateral ureteral obstruction. MATERIALS AND METHODS: Etodolac (10 mg./kg.) was administered to rats 1 day before unilateral ureteral obstruction and every day thereafter. Kidneys were harvested at day 14 after unilateral ureteral obstruction. Tissue transforming growth factor-beta and prostaglandin E2 were measured by bioassay using mink lung epithelial cells and enzyme linked immunosorbent-sandwich assay. Renal tubular proliferation and apoptosis were detected by immunostaining with proliferating cellular nuclear antigen and by terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate nick end labeling, respectively. Cyclooxygenase-2 expression was detected by immunohistochemistry. Fibrosis was assessed by measuring collagen deposition in trichrome stained slides. RESULTS: Bioassay showed that in the control group obstructed kidneys contained significantly higher mean transforming growth factor-beta1 than unobstructed kidneys (79.1 +/- 8.3 versus 33.6 +/- 4.2 ng./gm. tissue) and etodolac significantly decrease the mean value in obstructed kidneys (46.2 +/- 10.0 ng./gm. tissue). Assay demonstrated that obstructed control kidneys had significantly more mean tubular apoptosis than their unobstructed counterparts (26.6 +/- 5.4 versus 2.2 +/- 1.4 nuclei per high power field) and etodolac significantly decreased mean renal tubular apoptosis in the obstructed kidneys (16.2 +/- 1.9 nuclei per high power field). In addition, immunostaining with proliferating cellular nuclear antigen showed that obstructed kidneys in the control group had significantly more mean renal tubular proliferation than unobstructed kidneys (9.8 +/- 3.4 versus 3.9 +/- 0.1 per high power field) and etodolac significantly increased mean proliferating renal tubule in the obstructed kidneys (24.9 +/- 4.3 per high power field). Control obstructed kidneys had significantly more fibrosis and prostaglandin E2 production, which were also significantly blunted by etodolac. CONCLUSIONS: The cyclooxygenase-2 inhibitor etodolac significantly reduces tissue transforming growth factor-beta, resulting in decreased tubular damage and interstitial fibrosis. This finding suggests that etodolac is a promising agent for preventing renal tissue damage in unilateral ureteral obstruction.     

Protein kinase C in mouse kidney: effect of the Hyp mutation and phosphate deprivation

To test whether protein kinase C plays a role in the regulation of renal brush border membrane phosphate transport and mitochondrial vitamin D metabolism, we examined the activity, distribution and endogenous substrates of protein kinase C in renal subcellular fractions derived from two mouse models exhibiting perturbations in both renal functions. The X-linked Hyp mouse is characterized by reduced phosphate transport and 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) synthesis relative to normal, whereas the phosphate-deprived mouse exhibits elevated phosphate transport and vitamin D hormone synthesis. Protein kinase C activity was higher in renal cytosol of Hyp mice, when compared to normal littermates (358 +/- 11 vs. 244 +/- 31 pmol 32P/mg prot/min, P less than 0.02), whereas genotype differences in brush border membrane and mitochondrial kinase were not apparent. Phosphate deprivation of normal mice elicited a 50% reduction in brush border membrane protein kinase C (from 819 +/- 56 to 460 +/- 48 pmol 32P/mg prot/min, P less than 0.03), an increase in mitochondrial kinase (from 57 +/- 7 to 87 +/- 10 pmol 32P/mg prot/min, P less than 0.03), and no change in cytosolic kinase activity. Phosphate deprivation of Hyp mice led to an increase in mitochondrial protein kinase C (from 72 +/- 7 to 98 +/- 9 pmol 32P/mg prot/min, P less than 0.03) and no change in either brush border membrane or cytosolic kinase activity.(ABSTRACT TRUNCATED AT 250 WORDS)     

Mouse podocyte complement factor H: the functional analog to human complement receptor 1

Complement factor H (Cfh) is a key plasma protein in humans and animals that serves to limit alternative pathway complement activation in plasma, as well as in local sites such as capillaries of the glomerulus and eye. It was shown that rodent Cfh on platelets is the functional analogue to human erythrocyte complement receptor 1 with a role that is distinct from plasma Cfh and that Cfh is also on cultured rodent podocytes. For investigation of the role of Cfh in the kidney, renal transplants were performed between wild-type (WT) and Cfh(-/-) C57BL/6 mice. For these studies, bilateral native nephrectomies were done so that renal function was dependent solely on the transplanted kidney. Chronic serum sickness was induced by active immunization with apoferritin. Diffuse proliferative glomerulonephritis (GN) occurred in WT kidneys that were transplanted into Cfh(-/-) recipients (n = 8) but not into WT recipients (n = 14), consistent with the importance of plasma Cfh to dictate outcome in this disease model. Relative to the WT recipients of WT kidneys, WT mice with Cfh(-/-) kidneys (n = 12) developed glomerular disease features, including increased albuminuria (82.8 +/- 7.0 versus 45.1 +/- 3.6 microg/mg creatinine; P < 0.001) and blood urea nitrogen levels (54.4 +/- 6.1 versus 44.2 +/- 3.7 mg/dl; P < 0.01). In addition, they had substantial glomerular capillary wall deposits of IgG and C3, which by electron microscopy were present in subendothelial and subepithelial immune deposits, whereas WT kidneys in WT hosts had almost exclusive mesangial deposits. The IgG deposits in Cfh(-/-) kidneys were adjacent to Cfh-deficient podocytes, whereas WT kidneys in a Cfh(-/-) host had podocyte-associated Cfh with absent IgG deposits. These data suggest that locally produced podocyte Cfh is important to process immune complexes in the subepithelial space, where it also limits complement activation. Just as in platelets, rodent podocytes seem to use Cfh as the functional surrogate for human complement receptor 1.     

Late consequences of acute ischemic injury to a solitary kidney

The sequelae of acute ischemic injury to a solitary kidney were assessed in rats subjected to right nephrectomy and transient occlusion of the left renal artery; control rats underwent right nephrectomy alone. Incomplete recovery from ischemic injury at 2 wk (serum creatinine levels of 1.1 +/- 0.2 versus 0.5 +/- 0.1 mg/dl, P < 0.05 for ischemia versus control) was followed by deterioration of renal function at 20 wk (serum creatinine levels of 1.7 +/- 0.4 versus 0.7 +/- 0.1 mg/dl, P < 0.05 for ischemia versus control). Morphologic studies showed that impairment of function after ischemic injury was associated with widespread tubulointerstitial disease. Some tubule segments were atrophic and others exhibited cystic dilation, so that the tubular cell volume fraction was reduced (37 +/- 4 versus 53 +/- 2%, P < 0.05), while the tubular lumen and interstitial volume fractions were increased (31 +/- 4 versus 23 +/- 2% and 29 +/- 2 versus 20 +/- 1%, respectively, both P < 0.05). Many glomeruli retained open capillary loops but were no longer connected to normal tubule segments (63 +/- 8 versus 15 +/- 7% of glomeruli, P < 0.05). There was a strong inverse correlation between the prevalence of such glomeruli and the GFR at 20 wk after ischemia (r2 = 0.79, P < 0.001). Tubulointerstitial disease at that time was accompanied by proteinuria and widespread segmental glomerular tuft injury. The occurrence of similar processes in human patients could contribute to the loss of graft kidneys that suffer ischemic injury during transplantation.     

Increased transforming growth factor-β1 and tubulointerstitial fibrosis in rats with congenital hydronephrosis

OBJECTIVES: Most of our knowledge concerning renal obstruction has been derived from experimental animal models, and it is not yet well defined in spontaneous hydronephrosis. The aim of our study is to evaluate the roles of transforming growth factor-beta1 (TGF-beta1) and apoptosis in congenital hydronephrotic kidneys in comparison with experimental models. METHODS: We made histological studies on kidneys from 6-week-old Wistar-Imamichi rats with congenital unilateral hydronephrosis as well as surgical models of complete or partial unilateral ureteral obstruction. The severity of hydronephrotic kidneys was evaluated on routine hematoxylin and eosin (H&E) stained sections, and the tubulointerstitial fibrosis analyzed morphometrically on Masson's trichrome stained sections. Renal tubular atrophy was assessed on periodic acid Schiff (PAS) stained sections, and tubular cell apoptosis assessed with TUNEL technique. The renal TGF-beta1 level was determined by a sandwich enzyme-linked immunosorbent assay (ELISA). RESULTS: We observed a significant loss of kidney weight with profound compensatory growth of the contralateral kidney in rats with congenital hydronephrosis. Most of the hydronephrotic kidneys were markedly enlarged with dilatation of the collecting system, renal parenchymal thinning, tubular atrophy, interstitial infiltration and fibrosis. The renal TGF-beta1 level was markedly elevated in hydronephrotic kidneys as compared with normal controls (326.01 +/- 30.64 pg/mg protein vs 227.81 +/- 11.07 pg/mg protein, P < 0.01). The tubular apoptotic score in hydronephrotic kidneys was also significantly higher than normal controls (2.17 +/- 0.50/HPF [high power field]vs 0.14 +/- 0.04/HPF, P < 0.01). The increased TGF-beta1 and apoptotic status paralleled the histological changes of tubulointerstitial fibrosis and tubular atrophy. Similar findings were also obtained in experimental obstructive models. CONCLUSION: In comparison with surgical models of partial and complete ureteral obstruction, our data provide solid morphological and molecular evidences of renal obstruction in rats with congenital hydronephrosis.     

The role of heme oxygenase 1 in rapamycin-induced renal dysfunction after ischemia and reperfusion injury

Ischemia and reperfusion injury (IRI) is the main etiology of acute renal failure in native and transplanted kidneys. In the transplantation field, immunosuppressive drugs may play an additional role in acute graft dysfunction. Rapamycin may impair renal regeneration post IRI. Heme oxygenase 1 (HO-1) is a protective gene with anti-inflammatory and anti-apoptotic actions. We investigated whether HO-1 played a role in rapamycin-induced renal dysfunction in an established model of IRI. Rapamycin (3 mg/kg) was administered to mice before being subjected to 45 min of ischemia. Animals subjected to IRI presented with impaired renal function that peaked at 24 h (2.05+/-0.23 mg/dl), decreasing thereafter. Treatment with rapamycin caused even more renal dysfunctions (2.30+/-0.33 mg/dl), sustained up to 120 h after reperfusion (1.54+/-0.4 mg/dl), when compared to the control (0.63+/-0.09 mg/dl, P<0.05). Rapamycin delayed tubular regeneration that was normally higher in the control group at day 5 (68.53+/-2.30 vs 43.63+/-3.11%, P<0.05). HO-1 was markedly upregulated after IRI and its expression was even enhanced by rapamycin (1.32-fold). However, prior induction of HO-1 by cobalt protoporphyrin improved the renal dysfunction imposed by rapamycin, mostly at later time points. These results demonstrated that rapamycin used in ischemic-injured organs could also negatively affect post-transplantation recovery. Modulation of HO-1 expression may represent a feasible approach to limit rapamycin acute toxicity.     

Ghrelin improves renal function in mice with ischemic acute renal failure

Growth hormone and IGF-1 have been suggested to have tissue-protective effects. Ghrelin is a stomach-derived growth hormone secretagogue. The effects of ghrelin on ischemia/reperfusion-induced renal failure in mice were examined. Ischemic acute renal failure was induced by bilateral renal artery clamping for 45 min and reperfusion for 24 h. Ghrelin (100 microg/kg mouse) or vehicle was injected subcutaneously six times before surgery and three times after surgery every 8 h. Twenty-four hours after reperfusion, the right kidney was isolated and perfused. Acetylcholine (ACh)- and adrenomedullin-induced endothelium-dependent vasorelaxation of renal vessels significantly improved in ghrelin-pretreated mice (%Delta renal perfusion pressure by 10(-7) M ACh -63.5 +/- 3.7 versus -41.2 +/- 5.5%; P < 0.05). This change was associated with significant increases of nitric oxide release in the kidneys of ghrelin-treated mice (10(-7) M ACh 35.5 +/- 5.8 versus 16.9 +/- 3.5 fmol/g kidney per min; P < 0.05). Serum concentration of urea nitrogen (53 +/- 7 versus 87 +/- 15 mg/dl; P < 0.05) and renal injury score were significantly lower in the ghrelin group (2.5 +/- 0.8 versus 5.3 +/- 1.5; P < 0.01). Tubular apoptotic index was significantly lower in the ghrelin group (5 +/- 5 versus 28 +/- 4; P < 0.05). Furthermore, the survival rate after the 60-min ischemic period was higher in the ghrelin group (80 versus 20%; P < 0.05). Ghrelin treatment significantly increased the serum level of IGF-1. However, such renal protective effects of ghrelin on ischemia/reperfusion injury were not observed in insulin receptor substrate-2 knockout mice. These results suggest that ghrelin may protect the kidneys from ischemia/reperfusion injury and that this effect is related to an improvement of endothelial function through an IGF-1-mediated pathway.     

The use of the endothelin receptor antagonist, tezosentan, before or after renal ischemia protects renal function

BACKGROUND: Utilization of organs subjected to ischemia/reperfusion (I/R) injury could expand the donor pool. Endothelin (ET) is implicated in renal I/R injury. Therefore, our study compared the effectiveness of pre- and postischemic administration of the ET receptor antagonist, Tezosentan, in preserving renal function. METHODS: In a rat model, a kidney was subjected to 45 min of ischemia along with a contralateral nephrectomy. After 24 hr of reperfusion, renal function was assessed by serum creatinine (Scr), inulin clearance (glomerular filtration rate; GFR), and histology. ET-1 peptide expression was localized using immunohistochemistry. Three groups were studied: I/R untreated (n=17), I/R pretreated (n=11), and I/R posttreated (n=13) with Tezosentan (15 mg/kg, i.v.). RESULTS: Tezosentan significantly decreased (P<0.05) the rise in Scr from I/R injury (2.0+/-0.4 mg/dl, before and 2.9+/-0.4 mg/dl, after treatment) compared with untreated animals (4.2+/-0.4 mg/dl). GFR was significantly increased (P<0.05) from 0.13+/-0.03 ml/min (untreated animals) to 0.74+/-0.16 and 0.47+/-0.14 ml/min (pre- and posttreated animals). Untreated animals had significant cortical acute tubular necrosis, which was almost completely prevented by pretreatment with Tezosentan and markedly reduced by posttreatment. Increased ET-1 peptide expression was noted in the renal vasculature and in the cortical tubular epithelium of kidneys exposed to I/R. CONCLUSIONS: The purpose of this study was to optimize the function of kidneys exposed to I/R injury. Pretreatment as well as posttreatment with Tezosentan successfully decreased Scr, increased GFR, and maintained renal architecture in kidneys after ischemia. Therefore, ET receptor antagonists may be useful to preserve renal function in the transplantation setting.     

Dual response of animals with chronic cyclosporine nephrotoxicity to an acute ischemic injury

Normal kidneys regenerate after acute injury with little development of chronic fibrosis. However, the long-term effects of an acute injury in kidneys with established chronic toxicity induced by cyclosporine (CsA) are not entirely clear. To study the consequences of an ischemia and reperfusion (IR) injury in long-term CsA-treated rats, male Wistar rats (250-300 g) were treated daily with CsA (10 mg/kg) or vehicle (olive oil 1 mL/kg) for 28 days. On day 21, ischemia was performed by clamping the renal vessel for 1 hour. Blood samples were collected on days 0 and 21 (before IR) as well as days 22 and 28. On day 28, the kidneys were collected to examine the mRNA expression of MCP-1 by real-time PCR. For renal function, serum creatinine levels were measured. Twenty-four hours after reperfusion, long-term CsA-treated animals showed better renal function compared with the control group, as demonstrated by serum creatinine levels: 2.2 +/- 0.13 mg/dL vs 2.9 +/- 0.18 mg/dL, respectively (P < .05). However, 1 week after IR, the renal function was worse among the long-term CsA-treated group than the controls: 1.16 +/- 0.08 mg/dL vs 0.8 +/- 0.09 mg/dL, respectively (P < .05). Interestingly, CsA treatment was associated with lower MCP-1 mRNA expression than that in the control group: mean MCP-1 mRNA expression 0.58 +/- 0.13 vs 1.02 +/- 0.12, respectively (P < .05). In conclusion, animals with chronic CsA nephrotoxicity were protected from an acute renal injury, possibly through decreased chemokine production, although at later time points, renal function was clearly impaired, probably by the acceleration of vasculopathy caused by nephrotoxicity.     

Accumulation of indoxyl sulfate in OAT1/3-positive tubular cells in kidneys of patients with chronic renal failure.

Indoxyl sulfate shows nephrotoxicity and is a stimulating factor for progression of chronic renal failure (CRF). Indoxyl sulfate is taken up by renal proximal tubular cells through organic anion transporters 1 and 3 (OAT1/3), and is accumulated in the renal proximal tubular cells of uremic rats. To determine whether indoxyl sulfate is accumulated in human OAT1/3 (hOAT1/3)-positive renal proximal tubular cells, localization of indoxyl sulfate and hOAT1/3 in the kidneys of CRF patients was determined by immunohistochemistry. Kidney samples were obtained by autopsy from 9 CRF patients (mean serum creatinine 4.7 mg/dL, ranging from 2.0 to 14.5 mg/dL) and 9 patients with non-kidney disease (mean serum creatinine 0.6 mg/dL, ranging from 0.4 to 0.9 mg/dL). Immunohistochemistry was performed using antibodies against indoxyl sulfate, hOAT1, and hOAT3. Indoxyl sulfate was localized in the hOAT1- and hOAT3-positive renal tubular cells in the kidneys of CRF patients. The indoxyl sulfate-positive area in the kidneys was markedly increased in the kidneys of CRF patients compared with patients with non-kidney disease. The indoxyl sulfate-positive area was positively correlated with serum creatinine. In conclusion, in CRF patients, indoxyl sulfate is accumulated in the tubular cells with hOAT1 and/or hOAT3 localized at the basolateral membrane. The extent of indoxyl sulfate accumulation in the kidneys is more prominent in those patients with more severe CRF.     

Course of renal injury in the Mpv17-deficient transgenic mouse

The mutant Mpv17 mouse is a transgenic strain that fails to express a protein that is normally expressed in the kidney and that is associated with peroxisomes. The present studies provide a quantitative examination of renal function and structure in this strain compared to its control CFW strain. By 52 wk of age, the mutant strain developed proteinuria (urinary protein to creatinine ratio: 25 +/- 14 versus 3 +/- 1, mutant versus control), albuminuria (urinary albumin to creatinine ratio: 23 +/- 15 versus 0.1 +/- 0.1, mutant versus control), and hypoalbuminemia (2.1 +/- 0.4 versus 2.5 +/- 0.2 G/dl, mutant versus control), but without arterial hypertension or major reduction in filtration (serum creatinine 0.14 +/- 0.04 versus 0.18 +/- 0.12 mg/dl, mutant versus control). The Mpv17 glomeruli were enlarged (0.98 +/- 0.12 versus 0.52 +/- 0.02 micrometer(3) x 10(6), mutant versus control). Glomerular sclerosis became widespread (95 +/- 3 versus 23 +/- 32%, mutant versus control) and was preceded by mesangiolysis and microaneurysms. Tubulointerstitial disease was conspicuous by its absence. The intrarenal vasculature was normal in the mutant mice. Electron microscopy demonstrated focal foot process fusion and mesangiolysis. Thus, this mutant strain of mouse develops proteinuria and a distinct glomerulopathy including mesangiolysis but little interstitial injury all due to the loss of expression of a single gene.     

Report of a patient with a high level of parathyroid hormone-related peptide associated with malignant lymphoma involving multiple bony lesions.

Hypercalcemia in malignant lymphoma is not common. Our case of malignant lymphoma with multiple bony lesions showed hypercalcemia (13 mg/dl) at the time of bone marrow relapse. The serum level of parathyroid hormone-related peptide increased to 142 pmol/l, which may be secreted by malignant lymphoma cells. The course of the patient was aggressive and she died from bone marrow relapse after 6 months of treatment including high-dose methotrexate, which caused acute nonoliguric renal failure. At autopsy there was extensive calcium deposition in the lungs, kidneys and pancreas and prominent tubular damage of kidneys.     

Extracellular signal-regulated kinase inhibition slows disease progression in mice with polycystic kidney disease

The expression of mitogen-activated protein kinases (MAPK) in DBA/2-pcy/pcy (pcy) mice, a murine model of polycystic kidney disease was investigated. Proliferating cell nuclear antigen-positive cells were recognized in cyst epithelium from embryonic day 14.5 to 25 wk of age. Extracellular signal-regulated kinase (ERK) was expressed in the renal tubules of control and pcy mice, but stronger immunostaining was observed in cyst epithelium. Phosphorylated ERK was detected only in pcy mice and was localized predominantly in the cysts. p38 MAPK (p38) was no longer expressed after birth in controls but was detected in the cyst epithelium and in occasional tubular cells of pcy mice at all stages examined. c-Jun N-terminal kinase (JNK) was expressed in all tubular segments of controls after neonatal day 7, whereas in pcy kidneys, tubules became positive for JNK after 8 wk, and the cysts expressed little JNK. Administration of an oral MAP/ERK kinase inhibitor, PD184352, 400 mg/kg per d, to 10-wk-old pcy mice daily for the first week and then every third day for 6 additional weeks significantly decreased BP, kidney weight, serum creatinine level, and water intake and significantly increased urine osmolality. The cystic index and expression of phosphorylated ERK and ERK were significantly lower in PD184352-treated pcy mice. These results demonstrate that the expression of MAPK is dysregulated in cyst epithelium and that inhibition of ERK slowed the progression of renal disease in pcy mice.     

Gene therapy expressing amino-terminal truncated monocyte chemoattractant protein-1 prevents renal ischemia-reperfusion injury

Ischemia-reperfusion is closely associated with tissue damage in various organs, including kidney. Despite clinical investigations, useful therapy for renal ischemia-reperfusion injury is not available so far. This study evaluated therapeutic effects of gene therapy expressing an amino-terminal deletion mutant of MCP-1 called 7ND to inhibit monocyte chemoattractant protein (MCP)-1/CCR2 signaling in vivo on renal ischemia-reperfusion injury. 7ND gene was transferred into the femoral muscle of Balb/c mice. Renal artery and vein of the left kidney were occluded with a vascular clamp for 60 min. A large number of infiltrated cells were observed, as was marked acute tubular necrosis in outer medulla after renal ischemia-reperfusion injury in control mice, while these lesions were significantly decreased in 7ND gene-transfected mice. Macrophages in the interstitial region, most of which were CCR2-positive, were markedly decreased in 7ND gene-transfected mice after reperfusion. Although macrophages infiltrated around MCP-1-positive cells in control mice, the smaller number of F4/80-positive cells could infiltrate into the neighbor of MCP-1-positive cells in 7ND-treated mice. These results provide evidence that gene therapy by 7ND is potentially a powerful therapeutic approach to inhibit MCP-1/CCR2 signaling, resulting in rescue from renal ischemia-reperfusion injury.     

The kinase Pyk2 is involved in renal fibrosis by means of mechanical stretch-induced growth factor expression in renal tubules

Unilateral ureteral obstruction is a well-established experimental model of progressive renal fibrosis. We tested whether mechanical stretch and subsequent renal tubular distension might lead to renal fibrosis by first studying renal tubular epithelial cells in culture. We found that mechanical stretch induced reactive oxygen species that in turn activated the cytoplasmic proline-rich tyrosine kinase-2 (Pyk2). This kinase is abundantly expressed in tubular epithelial cells where it is activated by several stimuli. Using mice with deletion of Pyk2 we found that the expression of transforming growth factor-β1 induced by mechanical stretch in renal tubular epithelial cells was significantly reduced. The expression of connective tissue growth factor was also reduced in the Pyk2(-/-) mice. We also found that expression of connective tissue growth factor was independent of transforming growth factor-β1, but dependent on the Rho-associated coiled-coil forming protein kinase pathway. Thus, Pyk2 may be an important initiating factor in renal fibrosis and might be a new therapeutic target for ameliorating renal fibrosis.     

Novel nuclear factor kappa B activation inhibitor prevents inflammatory injury in unilateral ureteral obstruction

PURPOSE: We determined whether the novel nuclear factor kappa B activation inhibitor dehydroxymethylepoxyquinomicin (DHMEQ), which is derived from epoxyquinomicin C, affects renal inflammatory responses in unilateral ureteral obstruction. MATERIALS AND METHODS: DHMEQ (8 mg./kg.) was administered to rats 1 day after unilateral ureteral obstruction and every day thereafter. Kidneys were harvested at day 7 after unilateral ureteral obstruction. Tissue nuclear factor kappa B activity and transforming growth factor-beta were determined by electrophoretic mobility shift assay and bioassay using mink lung epithelial cells, respectively. Renal tubular proliferation and apoptosis were detected by immunostaining proliferating cellular nuclear antigen and the TUNEL (Intergen, Purchase, New York) assay, respectively. Leukocyte infiltration was detected by immunostaining for CD45. Fibrosis was assessed by measuring tissue hydroxyproline content. RESULTS: Unilateral ureteral obstruction for 7 days significantly activated nuclear factor kappa B, induced tubular apoptosis, proliferation and interstitial fibrosis in the obstructed kidney of the control group compared with their unobstructed counterparts (30.3 +/- 4.5 nuclei per high power field versus 1.7 +/- 0.4, 25.7 +/- 3.3 nuclei per high power field versus 3.2 +/- 0.4 and 6.2 +/- 0.3 micromol. hydroxyproline per gm. tissue versus 3.4 +/- 0.1, respectively). Conversely daily administration of DHMEQ (8 mg./kg.) significantly inhibited nuclear factor kappa B activation and decreased mean tubular apoptosis (9.5 +/- 2.1 nuclei per high power field), proliferation (10.2 +/- 2.4 nuclei per high power field) and interstitial fibrosis (4.9 +/- 0.4 micromol. hydroxyproline per gm. tissue) in the obstructed kidney. CONCLUSIONS: Specific inhibition of nuclear factor kappa B can prevent inflammatory renal responses, suggesting that targeting nuclear factor kappa B activation may be feasible for preventing inflammatory kidney diseases.     

p38 Mitogen-activated protein kinase contributes to autoimmune renal injury in MRL-Fas lpr mice

The phosphorylation of p38 mitogen-activated protein kinase (MAPK) is responsible for the production and signal transduction of cytokines and chemokines. This study hypothesized that p38 MAPK activation is required for spontaneous autoimmune renal injury in MRL-Fas(lpr) mice, resembling human lupus erythematosus. FR167653, a specific inhibitor of p38 MAPK, is orally administrated from 3 or 4 mo of age in MRL-Fas(lpr) mice (at doses of 10 or 32mg/kg per day) until 6 mo of age. The phosphorylated p38 MAPK in kidneys of MRL-Fas(lpr) mice was evaluated. The number of phosphorylated p38 MAPK-positive cells was increased in diseased kidneys. The daily oral administration of FR167653 decreased p38 MAPK phosphorylation in kidneys, especially in a group of mice administered FR167653 (32 mg/kg per day) daily from 3 to 6 mo of age. FR167653 reduced the accumulation of macrophages and T cell and prevented kidney pathology, resulting in prolonged survival. In addition, FR167653 reduced expression of MCP-1 and TNF-alpha in the diseased kidneys and cultured tubular epithelial cells. Furthermore, FR167653 decreased IgG levels in the diseased kidneys and circulation. These results suggest that the phosphorylation of p38 MAPK is required for the pathogenesis of renal injury in MRL-Fas(lpr) mice followed by subsequent expression of renal cytokine/chemokine and IgG production. This study provides evidence that the regulation of p38 MAPK is a novel target for the therapy of renal injury in systemic lupus erythematosus.