Angiotensin II induces renal oxidant stress in vivo and heme oxygenase-1 in vivo and in vitro

BACKGROUND: Angiotensin II is strongly incriminated in progressive renal injury. There is recent evidence that angiotensin II induces oxidative stress in vitro. We examined the capacity of angiotensin II to induce oxidative stress in vivo and the functional significance of such stress. The capacity of angiotensin II to induce the oxidant-sensitive gene heme oxygenase (HO) in vivo and in vitro was also examined. METHODS: Angiotensin II was administered via mini-osmotic pumps to rats maintained on standard diets. Indices of oxidative stress, including thiobarbituric acid reactive substance, carbonyl protein content, and HO activity, were determined. Indices of oxidative stress and functional markers were also determined in the DOCA salt model. The effect of angiotensin II was studied in rats maintained on antioxidant-deficient diets so as to examine the functional significance of oxidative stress induced by angiotensin II. We also explored the inductive effect of angiotensin II on HO in vivo and whether such actions occur in vitro. RESULTS: Angiotensin II administered in vivo increased kidney content of thiobarbituric acid reactive substances protein carbonyl content, and HO activity. These indices were not present in the kidney of rats treated with DOCA salt for three weeks. Such oxidative stress was functionally significant, since the administration of angiotensin II to rats maintained on a prooxidant diet demonstrated increased proteinuria and decreased creatinine clearance. The stimulatory effect on HO activity was due to induction of HO-1 mRNA, with HO-2 mRNA remaining unchanged. Expression of HO-1 was localized to the renal proximal tubules in vivo. We also demonstrate that angiotensin II at concentrations of 10-8 and 10-7 mol/L induces expression of HO-1 mRNA in LLC-PK1 cells. CONCLUSIONS: Angiotensin II induces oxidative stress in vivo, which contributes to renal injury. This study also demonstrates that angiotensin II induces renal HO activity caused by up-regulation of HO-1 in renal proximal tubules. Finally, angiotensin II directly induces HO-1 in renal proximal tubular epithelial cells in vitro.     

Preconditioning of the distal tubular epithelium of the human kidney precedes nephrocalcinosis

BACKGROUND: Preterm neonates and renal transplant patients frequently develop nephrocalcinosis. Experimental studies revealed that crystal retention in the distal nephron, a process that may lead to nephrocalcinosis, is limited to proliferating/regenerating tubular cells expressing hyaluronan and osteopontin at their luminal surface. Fetal and transplant kidneys contain proliferating and/or regenerating cells since nephrogenesis is not completed until 36 weeks of gestation, while ischemia and nephrotoxic immunosuppressants may lead to injury and repair in renal transplants. This prompted us to investigate the expression of hyaluronan and osteopontin and to correlate this to the appearance of tubular calcifications both in fetal/preterm and transplanted kidneys. METHODS: Sections of fetal/preterm kidneys and protocol biopsies of transplanted kidneys (12 and 24 weeks posttransplantation from the same patients) were stained for osteopontin, hyaluronan, and calcifications (von Kossa). RESULTS: Hyaluronan and osteopontin were expressed at the luminal surface of the epithelial cells lining the distal tubules of all fetal kidneys at birth and in all kidney graft protocol biopsies 12 and 24 weeks posttransplantation. In 7 out of 18 surviving (at least 4 days) preterm neonates crystal retention developed. In renal allografts a striking increase (from 2/10 to 6/10) in tubular crystal retention between 12 and 24 weeks posttransplantation was observed. In addition, crystals were selectively retained in distal renal tubules containing cells with hyaluronan and osteopontin at their luminal surface. CONCLUSION: The results of this study show that luminal expression of hyaluronan and osteopontin preceded renal distal tubular retention of crystals in preterm neonates and renal transplant patients. We propose that the presence of this crystal binding phenotype may play a general role in renal calcification processes.     

Cytoprotective role of heme oxygenase (HO)-1 in human kidney with various renal diseases.

BACKGROUND: We previously reported that glomerular changes in the renal specimen of a human case with heme oxygenase-1 (HO-1) deficiency were mild, but tubulointerstitial injury advanced progressively. This study examined the patterns of HO-1 production in the kidney in various renal diseases. Furthermore, the critical cytoprotective roles of HO-1 were evaluated in the kidney by comparing HO-1 production and expressions of carboxymethyllysine (CML) and pentosidine, both of which are markers of oxidative stress. METHODS: Renal biopsy or autopsy materials were obtained from a total of 74 patients. Degrees of hematuria and proteinuria and the levels of urinary N-acetyl-beta-D-glucosaminidase (NAG), beta2-microglobulin (beta2m), and creatinine were evaluated. Immunohistochemical studies for HO-1, CML, and pentosidine expressions were performed with their specific antiserum. RESULTS: HO-1 staining was observed within tubular epithelial cells in all of the renal diseases, but was not detected within intrinsic glomerular cells. HO-1 staining tended to be more intense within distal tubuli than in proximal tubuli. Within distal tubuli, there was no significant correlation between intensity of HO-1 staining and degree of hematuria or presence of proteinuria. Within proximal tubuli, HO-1 staining tended to be more intense with greater degrees of hematuria, presence of proteinuria, and moderate tubulointerstitial damage. Intense staining of CML and pentosidine was observed within renal tubular epithelial cells only in HO-1-deficient patients. CONCLUSIONS: HO-1 plays important roles in protecting renal tubuli from oxidative injuries, as these cells are constantly exposed to various oxidative stresses. It is suggested that renal tubular epithelia are more susceptible to oxidative stress due to the lack of this critical enzyme in HO-1 deficiency.     

Localization of urinary procoagulant in the human kidney.

By the indirect immunofluorescent and immunoenzymatic techniques with monospecific antiserum against urinary procoagulant (a tissue factor which accelerates blood coagulation), we found the urinary procoagulant in the kidney distributed to the loop of Henle and distal convoluted tubules. In these areas urinary procoagulant was found in association with the luminal and intercellular borders as well as in the cytoplasm of epithelial cells. Both the descending and ascending limbs of Henle were equally stained. The cytoplasmic staining was patchy in distribution among cells of distal tubules and was predominantly localized in the supranuclear areas. Glomeruli, the proximal tubular cells, the vascular wall, and the interstitium were not stained. There was, however, fluorescent staining along the epithelial layers of the Bowman's capsule, which was observed only in the frozen sections. Casts in the distal tubules were also positively stained. These findings suggest that urinary procoagulant is synthesized in the epithelial cells of these particular parts of nephron and is secreted into urine, although its physiologic roles and pathologic significance are not entirely known.     

Heme oxygenase-1 protects rat kidney from ureteral obstruction via an antiapoptotic pathway

This study examined the functional significance of heme oxygenase-1 (HO-1) expression on renal injury induced by ureteral obstruction in the rat kidney. Male Sprague-Dawley rats were divided into three groups, after which unilateral ureteral obstruction (UUO) was performed: untreated (group 1), treated with 30 mg/kg body wt hemin (group 2), and treated with 50 microg/kg body wt zinc (alpha) protoporphyrin eta (ZnPP) and 30 mg/kg hemin (group 3). After 7 and 14 d, histologic changes and the expression of HO-1, Bcl-2, Bad, TGF-beta, and cleaved caspase-3 were examined. Tubular lumens were dilated and epithelial cells were flattened on day 7 after UUO. Interstitial fibrosis and separation of the tubules were markedly increased on day 14. In contrast, the kidneys that were treated with hemin exhibited minimal interstitial fibrosis and flattening of epithelial cells on day 7 and fewer changes on day 14 than in the controls. However, treatment with ZnPP, an inhibitor of HO enzyme activity, eliminated the beneficial effect of hemin on interstitial fibrosis and tubular dilation. Increased HO-1 expression was associated with increased Bcl-2. In the ZnPP-treated rats, Bcl-2 signals were decreased compared with the hemin group. The level of proapoptotic Bad was not changed in any group. The positive cells for cleaved caspase-3 were significantly increased in renal tubular epithelial cells and tubulointerstitial cells in the obstructed rats, and hemin treatment decreased the caspase-3 activation. This study demonstrates that upregulation of HO-1 provides protection against renal injury that follows UUO. This effect is dependent on modulation of the antiapoptotic pathway by HO-1 expression.     

Ureteral obstruction in neonatal mice elicits segment-specific tubular cell responses leading to nephron loss

BACKGROUND: To elucidate the sequence of renal responses leading to nephron loss in obstructive nephropathy, we examined the evolution of segmental nephron cellular changes consequent to chronic unilateral ureteral obstruction (UUO) in the neonatal mouse. METHODS: Neonatal mice were subjected to UUO or sham-operation, and kidneys were harvested 5, 12 or 19 days after surgery. Proximal tubules (PT), distal tubules (DT) and collecting ducts (CD) were identified with lectins. Histomorphometric quantitation was made for cellular necrosis, apoptosis, proliferation, tubular dilatation, tubular basement membrane (TBM) thickening, interstitial collagen, and glomerular maturation. The distribution of hypoxic tissue was determined using pimonidazole as a marker. Additional studies were performed by mechanically stretching monolayer cultures of mouse proximal tubular and collecting duct cells, and measuring apoptosis. RESULTS: Neonatal UUO induced an arrest of glomerular maturation throughout the period of study. Chronic UUO induced hypoxia, tubular necrosis, proliferation, and TBM thickening in the PT, but stimulated apoptosis in the DT and CD. Tubular dilation in the obstructed kidney was most severe in CD and least severe in PT. Tubular cell apoptosis closely paralleled tubular dilation (P < 0.05), and fibrosis surrounding individual tubules also correlated with tubular dilation (P < 0.001). Mechanical stretching of cultured mouse tubular cells induced apoptosis directly proportional to the magnitude of axial strain: apoptosis was consistently greater in CD than in PT cells (P < 0.05). CONCLUSIONS: Following UUO, the co-localization of hypoxia with cellular proliferation, necrosis, and TBM thickening of the PT is consistent with ischemic injury resulting from vasoconstriction. In contrast, a selective dilation of the distal portion of the nephron (DT and CD), which results from the greater tubular compliance there, leads to stretch-induced epithelial cell apoptosis, along with a progressive peritubular fibrosis. Nephron loss in the obstructed developing kidney likely results from complex, segment-specific cellular responses.     

Proximal tubular injury attenuates outer medullary hypoxic damage: studies in perfused rat kidneys

The straight segment (S3) of the proximal tubule is predominantly damaged during renal ischemia-reflow, whereas medullary thick ascending limbs (mTALs) are principally affected in other models of hypoxic acute tubular necrosis (ATN). Since the latter injury pattern largely depends on the extent of reabsorptive activity during hypoxic stress, we hypothesized that proximal tubular damage might attenuate downstream mTAL injury by means of diminished distal solute delivery for reabsorption. In isolated rat kidneys perfused for 90 min with oxygenated Krebs-Henseleit solution, mTAL necrosis developed in 75 +/- 3% of tubules in the mid-inner stripe of the outer medulla. By contrast, S3 segments in the outer stripe were minimally affected, with tubular fragmentation involving some 5 +/- 2% of tubules. In kidneys subjected in vivo to proximal tubular injury and subsequently used for isolated perfusion studies, the injury pattern was inverted: following 20 and 30 min ischemia and reflow for 24 h, S3 fragmentation rose to 18 +/- 16% and 72 +/- 13%, while mTAL damage was reduced to 33 +/- 10 and 24 +/- 8%, respectively. In kidneys subjected in vivo to D-serine S3 necrosis rose to 100%, while mTAL damage fell to 1 +/- 1% (p < 0.001). Substantial S3 tubular collapse (involving approximately 30% of tubules) and inner stripe interstitial hemorrhage were also noted, exclusively in kidneys subjected to ischemia-reflow. Proximal tubular necrosis alone or in combination with collapse inversely correlated with mTAL necrosis (R = -0.51 and -0.72, respectively, p < 0.003). This cogent inverse association might imply that disruption of the proximal nephron attenuates downstream mTAL necrosis by a reduction of distal tubular reabsorptive workload.     

Heme protein-induced chronic renal inflammation: suppressive effect of induced heme oxygenase-1

BACKGROUND: Heme oxygenase (HO) is the rate-limiting enzyme in the degradation of heme; its inducible isozyme, HO-1, protects against acute heme protein-induced nephrotoxicity and other forms of acute tissue injury. This study examines the induction of HO-1 in the kidney chronically inflamed by heme proteins and the functional significance of such an induction of HO-1. METHODS: Studies were undertaken in a patient with chronic tubulointerstitial disease in the setting of paroxysmal nocturnal hemoglobinuria (PNH), in a rat model of chronic tubulointerstitial nephropathy caused by repetitive exposure to heme proteins, and in genetically engineered mice deficient in HO-1 (HO-1 -/-) in which hemoglobin was repetitively administered. RESULTS: The kidney in PNH evinces robust induction of HO-1 in renal tubules in the setting of chronic inflammation. The heme protein-enriched urine from this patient, but not urine from a healthy control subject, induced expression of HO-1 in renal tubular epithelial cells (LLC-PK1 cells). A similar induction of HO-1 and related findings are recapitulated in a rat model of chronic inflammation induced by repetitive exposure to heme proteins. Additionally, in the rat, the administration of heme proteins induces monocyte chemoattractant protein (MCP-1). The functional significance of HO-1 so induced was uncovered in the HO-1 knockout mouse: Repeated administration of hemoglobin to HO-1 +/+ and HO-1 -/- mice led to intense interstitial cellular inflammation in HO-1 -/- mice accompanied by striking up-regulation of MCP-1 and activation of one of its stimulators, nuclear factor-kappaB (NF-kappaB). These findings were not observed in similarly treated HO-1 +/+ mice or in vehicle-treated HO-1 -/- and HO-1 +/+ mice. CONCLUSION: We conclude that up-regulation of HO-1 occurs in the kidney in humans and rats repetitively exposed to heme proteins. Such up-regulation represents an anti-inflammatory response since the genetic deficiency of HO-1 markedly increases activation of NF-kappaB, MCP-1 expression, and tubulointerstitial cellular inflammation.     

Heme oxygenase isoform-specific expression and distribution in the rat kidney

BACKGROUND: The heme oxygenase (HO) genes, HO-1 and HO-2, are the limiting steps in heme degradation and in the regulation of renal heme-dependent enzymes. Previously we reported that selective overexpression of renal HO-1 resulted in a decrease of microsomal heme and the cytochrome P450-dependent arachidonic acid metabolite, 20 HETE, a vasoconstrictor. The present study was undertaken to explore the relative expression and contribution of each of the HO isoforms to HO activity in the rat kidney. METHODS AND RESULTS. Renal HO activity increased above control levels after an injection of the inducers of HO activity, heme or SnCl2. Stannous Mesoporphyrin (SnMP), a nonselective inhibitor of HO, when used alone or in combination with heme or SnCl2, decreased HO activity. Heme alone and combined with SnCl2 decreased the levels of heme content by 13 and 35%, respectively. Western blot analysis showed that both SnCl2 and heme readily induced HO-1 protein, whereas HO-2 was constitutively expressed. Immunohistochemistry showed the distribution of the HO-1 isoform primarily in proximal convoluted tubules. Western blot analysis exhibited relatively higher levels of HO-1 in isolated proximal tubules and relatively higher HO-2 levels in the thick ascending limbs of the loop of Henle and preglomerular arterioles. In vivo administration of HO-1 and HO-2 antisense oligodeoxynucleotides further confirmed that HO-2, but not HO-1, contributed to the basal HO activity; however, following induction of HO with heme, antisense to HO-1, but not to HO-2, inhibited the induced levels of HO activity. CONCLUSION: These results suggest that HO-2 is constitutively expressed in the rat kidney mainly within tubular and arteriolar structures, and its activity may modulate physiological function under basal conditions. On the other hand, the basal levels of expression of HO-1 in the rat kidney are relatively low, and its contribution to HO activity and the regulation of hemoproteins such as cytochrome P450 become apparent only under pathophysiological conditions causing HO induction.     

Heme oxygenase-1 deficiency promotes epithelial-mesenchymal transition and renal fibrosis

Induction of heme oxygenase-1 (HO-1) is associated with potential antifibrogenic effects. The effects of HO-1 expression on epithelial-mesenchymal transition (EMT), which plays a critical role in the development of renal fibrosis, are unknown. In this study, HO-1(-/-) mice demonstrated significantly more fibrosis after 7 d of unilateral ureteral obstruction compared with wild-type mice, despite similar degrees of hydronephrosis. The obstructed kidneys of HO-1(-/-) mice also had greater macrophage infiltration and renal tubular TGF-beta1 expression than wild-type mice. In addition, the degree of EMT was more extensive in obstructed HO-1(-/-) kidneys, as assessed by alpha-smooth muscle actin and expression of S100A4 in proximal tubular epithelial cells. In vitro studies using proximal tubular cells isolated from HO-1(-/-) and wild-type kidneys confirmed these observations. In conclusion, HO-1 deficiency is associated with increased fibrosis, tubular TGF-beta1 expression, inflammation, and enhanced EMT in obstructive kidney disease. Modulation of the HO-1 pathway may provide a new therapeutic approach to progressive renal diseases.     

Expression of mitogen-activated protein kinases in human renal dysplasia

BACKGROUND: We previously reported that the expression of mitogen-activated protein kinases (MAPKs) is developmentally regulated. Dysregulation of MAPKs may lead to kidney malformation. Thus, we investigated the expression of MAPKs in human renal dysplasia, one of the most common kidney malformations. METHODS: Prenatal (gestational ages 20 to 36 weeks, N = 6) and postnatal (2 years old, N = 1) dysplastic kidneys, and normal kidneys (gestational ages 19 to 34 weeks, N = 4) were examined. Immunohistochemical studies were performed using antibodies against extracellular signal-regulated kinase (ERK), p38 MAPK (p38), c-Jun N-terminal kinase (JNK), phospho-MAPKs (P-MAPKs), and proliferating cell nuclear antigen (PCNA). Apoptosis was detected by the TUNEL method. RESULTS: In dysplastic kidneys, proliferation was prominent in dysplastic tubules and also found in cyst epithelia. TUNEL staining was detected in dysplastic tubules and cysts, and occasionally in undifferentiated cells. p38 and anti-phospho-p38 (P-p38) were strongly expressed in dysplastic epithelia, but not detected in normal kidneys at any stage examined. On the other hand, JNK and P-JNK were positive in tubular epithelia of normal kidneys, whereas their expression was barely detectable in dysplastic tubules and cysts. ERK was expressed in all tubular segments, and P-ERK was detected in distal tubules and collecting ducts of normal kidneys. Dysplastic kidney epithelia stained exclusively positive for ERK and P-ERK. CONCLUSIONS: p38 is ectopically expressed, and JNK is down-regulated in dysplastic kidney epithelia. Furthermore, dysplastic epithelia are exclusively positive for ERK and P-ERK. Activated p38 and ERK may mediate hyperproliferation of dysplastic tubules resulting in cyst formation, whereas down-regulated JNK expression may be the cause or the result of an undifferentiated state of dysplastic epithelia.     

Ontogenesis of angiotensin-I converting enzyme in human kidney

The kidney distribution of angiotensin-I converting enzyme (ACE) was studied in 14 fetuses (11 to 30 weeks old) and 7 children (2 days to 13 years old) by immunohistochemistry using specific antibodies to human kidney ACE. Immunohistochemical techniques included indirect immunofluorescence on cryostat sections of frozen tissue, immunoperoxidase and immunofluorescence of fixed tissue embedded in Paraplast, and immunoelectron microscopy. The ACE distribution in the fetal kidneys was independent of the age of the fetus. ACE was detected in two locations: 1) on the basolateral membranes and primary apical microvilli of epithelial cells from early differentiating proximal tubules; the labeling was intense in brush borders of fully developed proximal tubules; and 2) on glomerular endothelial cells; cells were lined by reaction product as soon as capillaries invaded the inferior cleft of the S-shaped body. Tubular ACE distribution was identical in the postnatal kidneys. The staining of the glomerular endothelium was extremely inconstant. The presence of ACE in proximal tubular cells and glomerular endothelial cells at the beginning of nephron differentiation may indicate that it is involved in the development of nephron function and renal hemodynamic.     

Interactions between inducible nitric oxide synthase and heme oxygenase-1 in glomerulonephritis

BACKGROUND: In anti-glomerular basement membrane (GBM) nephritis, inducible nitric oxide synthase (iNOS) and heme oxygenase (HO-1) are co-induced. Moreover, in glomerular mesangial cells iNOS-derived nitric oxide (NO) production stimulates HO-1 while HO-1 activation reduces iNOS expression/activity. These observations prompted us to explore regulatory interactions between iNOS and HO-1 in anti-GBM nephritis. METHODS: Rats with anti-GBM nephritis were pretreated with the iNOS inhibitor l-N6-(1-iminoethyl) lysine (L-NIL) or with the HO-1 inducer hemin. Glomerular HO-1 levels were assessed by Western blot analysis. iNOS activity was assessed by calculating conversion of l-arginine to l-citrulline. RESULTS: iNOS inhibition reduced glomerular HO-1 levels without altering the inflammatory response to anti-GBM antibody induced injury. Induction of HO-1 reduced glomerular iNOS activity. CONCLUSIONS: In anti-GBM nephritis iNOS up-regulates HO-1 presumably via high output NO production. Suprainduction of HO-1 attenuates iNOS activity. This negative feedback interaction points to HO-1 as a target for pharmacologic manipulation to reduce activity of prooxidant heme containing enzymes such as iNOS.     

Expression and distribution of adenosine diphosphate-ribosylation factors in the rat kidney

BACKGROUND: Adenosine diphosphate (ADP)-ribosylation factors (ARFs) are small guanosine triphosphatases involved in membrane traffic regulation. Aiming to explore the possible involvement of ARF1 and ARF6 in the reabsorptive properties of the nephron, we evaluated their distribution along the different renal epithelial segments. METHODS: ARFs were detected by immunofluorescence and immunogold cytochemistry on renal sections, using specific anti-ARF antibodies. RESULTS: ARF1 was detected in proximal and distal tubules, thick ascending limbs of Henle's loops, and cortical and medullary collecting ducts. By immunofluorescence, labeling was mostly localized to the cell cytoplasm, particularly in Golgi areas. By electron microscopy, the Golgi apparatus and the endosomal compartment of proximal and distal tubular cells were labeled. ARF6 immunofluorescence was observed in brush border membranes and the cytoplasm of proximal convoluted tubular cells, whereas it was restricted to the apical border of proximal straight tubules. ARF6 immunogold labeling was detected over microvilli and endocytic compartments of proximal tubular cells. CONCLUSIONS: This study demonstrates the following: (a) the heterogeneous distributions of ARF1 and ARF6 along the nephron, (b) the existence of cytosolic and membrane-bound forms for both ARFs, and (c) their association with microvilli and endocytic compartments, suggesting an active participation in renal reabsorption.     

Ontogenesis of 28 kDa vitamin D-induced calcium-binding protein in human kidney

The kidney distribution of 28 kDa vitamin D-induced calcium binding protein (CaBP) was studied in 15 fetuses (11 to 33 weeks old), six children and adults (12 days to 32 years old) by immunocytochemistry using a specific antibody to rat renal 28 kDa CaBP. Similar results were obtained on frozen and fixed tissues. Kidneys from one adult and three fetuses were studied by immunoelectronmicroscopy for antigen localization at the subcellular level using the indirect immunoperoxidase technique. The 28 kDa CaBP was present in all kidneys from the eleventh week of gestation. At that stage, all deep parts of collecting ducts were homogeneously stained and a few distal tubules located in the deep cortex were intensely labeled. No labeling was observed in the early stage of nephron differentiation (S-body). 28 kDa CaBP distribution changed with kidney maturation. There was a progressive reduction of the deep part of collecting duct labeling and a concomitant increase in the number and intensity of stained distal tubular cells. At the ultrastructural level, 28 kDa CaBP was observed in the cytosol and the nuclear euchromatin. Our study demonstrates the early cellular synthesis of 28 kDa CaBP and its transient expression by deep collecting duct cells during early fetal life, at a time when only a few distal convoluted tubular cells synthetize it.     

A role for extrarenal cells in the regeneration following acute renal failure

BACKGROUND: Recovery of renal function following acute tubular necrosis (ATN) is dependent on the replacement of necrotic tubular cells with functional tubular epithelium. The source of these new tubular cells is thought to be resident renal tubular cells. The discovery of pluripotent bone marrow-derived stem cells has led to a reexamination of the cellular source and processes involved in the recovery from organ injury. METHODS: To test the hypothesis in humans that extrarenal cells participate in the recovery following ATN, we examined the origin of tubular cells in male patients with resolving ATN who had received a kidney transplant from a female donor. Immunohistochmistry of kidney biopsies was performed to identify renal tubular epithelial cells (cytokeratin positive) and leukocytes (CD45 positive). Fluorescent in-situ hybridization was used to detect Y chromosome containing cells with DAPI serving as a nuclear stain. All staining was performed on the same section. RESULTS: The Y chromosome was detected in approximately 40% of tubular cell nuclei in male kidneys (positive control) and in no nuclei of female kidneys (negative control). In male recipients of female kidneys who developed ATN, 1% of tubules contained Y chromosome cells defined by their morphology, positive staining for cytokeratin, and negative staining for CD45. When present, multiple cells in a positive tubule stained for the Y chromosome. No Y chromosome containing tubular cells were seen in similar sex mismatched transplants in male recipients who did not develop ATN, suggesting that recipient derived cells do not routinely repopulate the transplanted kidney. CONCLUSIONS: This proof-of-principle clinical observation demonstrates that extrarenal cells can participate in the regenerative response following ATN. These findings provide rationale for the cellular therapy of acute renal failure.     

Multilocular renal cyst. Immunohistochemical and lectin-binding study.

Multilocular renal cyst is an uncommon lesion of uncertain pathogenesis seen in children and adults. We report the immunohistochemical and lectin-binding profiles of three MRC occurring in adults. All cases had strong and uniform cytoplasmic staining of lining epithelial cells for keratin and binding sites for arachis hypogaea lectin, similar to that seen for the distal convoluted tubules or collecting ducts in normal kidney. However, we found variable expression of other distal nephron markers, including epithelial membrane antigen and Ber-EP4. Furthermore, lining cells in some lesions coexpressed proximal nephron markers such as alpha-1-antitrypsin and lysozyme, as well as binding sites for lotus tetragonolobus lectin. Immunostaining for type IV (basement membrane) collagen demonstrated a continuous subepithelial basement membrane zone and basal laminae surrounding desmin-positive stromal cells. Areas of active collagen synthesis and stromal procollagen deposition were visualized within the interlocular septae using a monoclonal antibody to type I procollagen. Significant proliferative activity was not detected in the lining epithelium or stroma using the anti-proliferating cell nuclear antigen. In conclusion, MRC show aberrant tubular epithelial glycoprotein and glycoconjugate expression, low proliferative activity, and associated activation of interlocular stromal cells.     

Congenital murine polycystic kidney disease

In the current study, the ontogeny of tubular cyst formation was studied in the CPK mouse, a murine strain with autosomal recessive polycystic kidney disease. Utilizing the technique of intact nephron microdissection in addition to standard light and transmission electron microscopy, the earliest morphologic alterations in CPK kidneys were localized in fetal tissue at 17 days of gestation to the distal portion of developing proximal tubules. During disease progression, from birth to 21 days of postnatal age, there was a shift in the site of cystic nephron involvement from proximal tubule to collecting tubules without involvement of other nephron segments. Cysts were enlarged tubular segments which remained in continuity with other portions of the nephron and were not associated with abnormalities in the overall pattern of nephron growth or differentiation. Analysis suggested that alterations in transtubular transport in abnormally shortened proximal tubular segments of juxtamedullary nephrons may have pathogenic importance in the early stages of cyst formation, and that epithelial hyperplasia and cytoskeletal alterations may have a role in progressive proximal tubular cystic enlargement. Cellular hyperplasia of epithelial walls of normally formed tubules was a prominent feature of cyst formation and progressive enlargement in collecting tubules. Such data form the basis for future studies into specific pathophysiological processes which may be operative in specific nephron segments during different stages of cyst formation in the CPK mouse.A preliminary report of this work was presented at the Annual Meeting of the Society for Pediatric Research, Washington DC, USA, May 1986, and has appeared in abstract form (Pediatr Res 20: 446A, 1986)