New aspects of posttransplant nephrotic syndrome: clinicopathologic correlations with outcomes.

Although posttransplant nephrotic syndrome is frequent, its structural basis and prognosis have not been clearly defined. The biopsy findings of 54 patients with this disorder posttransplant, among 375 total renal transplant recipients engrafted during a 10-year period, were correlated with clinical follow-up data. The mean patient age was 41.7 +/- 12.3 years, female/male ratio 22/32, and cadaveric/living-related donor ratio 37/17. The nephrotic syndrome developed 3 to 91 months posttransplant. At the onset the mean values of serum creatinine was 2.9 +/- 1.8 mg/dL and proteinuria 4.5 +/- 0.8 g/d. The index biopsy findings showed chronic allograft nephropathy (CAN) in 33; de novo glomerulonephritis (GN) in 6, recurrent GN in 9, and undetermined GN in 6 who had an unknown primary renal disease. Among 21 follow-up biopsies during a mean of 44.3 +/- 28 months the CAN progressed but the GN remained the same. The treatment included augmented steroids alone (n = 1) or in combination with cyclophosphamide (n = 2) and with plasmapheresis (n = 1); angiotensin-converting enzyme inhibitors (ACEI) or angiotensin receptor blockers (ARB) along (n = 5); calcium channel blockers (CCB) alone (n = 24); or the two types of drugs together (n = 22). Complete or partial remission was achieved in 8 and 5, respectively, but nephrotic syndrome recurred in 3 of these patients at 45.1 +/- 18 months later. Sustained remission was more likely in cases of GN (minimal change disease and IgA nephropathy) and ACEI-ARB treatment (P <.01). Graft failure, which occurred in 35 patients, correlated strongly with serum creatinine at onset, being significantly greater in patients with CAN (P <.005). Both remission of the nephrotic syndrome and graft survival were greater among patients with GN as compared to those with CAN.

Nonlupus nephritides in patients with systemic lupus erythematosus: a comprehensive clinicopathologic study and review of the literature.

Renal biopsy specimens from patients with systemic lupus erythematosus (SLE) rarely show changes that are pathogenetically and morphologically unrelated to SLE. The morphology and behavior of these nonlupus nephritides are not well known. Two hundred fifty-two renal biopsies performed on 224 patients with SLE collected from 3,036 native kidney biopsies performed between 1975 and 1998 were reviewed, and those that showed nonlupus nephritides (index biopsies) were selected for studies. Thirteen biopsy specimens with nonlupus nephritides were identified in 13 patients, who belonged to 3 clinically distinct groups. Group I included 6 patients in whom SLE was diagnosed at the time of index biopsies. The index biopsies in these patients showed focal segmental glomerusclerosis (FSGS; 3 cases), Immunoglobulin (Ig) M nephropathy (1 case), and thin basement membrane disease (1 case). The diagnostic features for FSGS included segmental sclerosis involving at least 1 glomerulus, absence of lupus nephritis or other conditions that may cause nonspecific segmental sclerosis of glomeruli such as ischemia or nephrosclerosis, and nephrotic-range proteinuria. There was uniform, global, diffuse and marked thinning of the glomerular basement membrane in the case of thin basement membrane disease. Group II included 3 patients in whom SLE was diagnosed 2 to 9 years before the time of index biopsies and SLE was active at the time of biopsy. The index biopsies in these patients showed FSGS (2 cases) and hypertensive nephrosclerosis (1 case). Group III included 4 patients in whom SLE was diagnosed 5 to 36 years before the time of index biopsies and SLE was inactive at the time of biopsy. The index biopsies in these patients showed 1 case each of amyloidosis, FSGS, hypertensive nephrosclerosis, and allergic acute tubulointerstitial nephritis. Previous renal biopsies, performed in 5 patients, showed IgM nephropathy (1 case), diffuse proliferative lupus GN (1 case), focal proliferative lupus GN (1 case), and mesangial proliferative lupus GN (2 cases). Follow-up biopsies, performed in 3 patients, confirmed the diagnosis of FSGS (2 cases) and hypertensive nephrosclerosis (1 case) noted in the index biopsies. Nonlupus nephritides may occasionally be encountered in SLE patients, regardless of clinical or serologic disease activity. These renal lesions display a broad morphologic spectrum in which FSGS seems most frequent. Renal biopsy plays a crucial role in identifying these lesions, which may have prognostic and therapeutic implications distinct from those of lupus nephritis.

Mechanism of chronic obstructive uropathy: increased expression of apoptosis-promoting molecules.

BACKGROUND: We have demonstrated that renal tubular and interstitial cells undergo pronounced apoptosis during the course of chronic obstructive uropathy (COU). Apoptosis is a complex cellular process consisting of multiple steps, each of which is mediated by families of related molecules. These families may include receptor/ligand molecules such as Fas, Fas ligand, tumor necrosis factor receptor-1 (TNFR-1), and TNF-related apoptosis inducing ligand (TRAIL); signal transduction adapter molecules such as Fas-associated death domain (FADD), TNFR-1 associated death domain (TRADD), receptor-interacting protein (RIP), Fas-associated factor (FAF), and Fas-associated phosphatase (FAP); or effector molecules such as caspases. However, the mechanism of tubular cell apoptosis, as well as the pathogenetic relevance of these apoptosis-related molecules in COU, remains poorly understood. METHODS: Kidneys were harvested from sham-operated control mice and mice with COU created by left ureter ligation sacrificed in groups of three at days 4, 15, 30, and 45. To detect apoptotic tubular and interstitial cells, in situ end labeling of fragmented DNA was performed. To detect the expression of apoptosis-related molecules, ribonuclease protection assay was used with specific antisense RNA probes for Fas, Fas ligand, TNFR-1, TRAIL, FADD, TRADD, RIP, FAF, FAP, and caspase-8. Immunostaining for Fas, Fas ligand, TRAIL, TRADD, RIP, and caspase-8 was also performed. To assess the role of these molecules in COU-associated renal cell apoptosis, the frequencies of apoptotic tubular and interstitial cells were separately quantitated for each experimental time point, and their patterns of variation were correlated with those of apoptosis-related molecules. RESULTS: The obstructed kidneys displayed increased apoptosis of both tubular and interstitial cells. Tubular cell apoptosis appeared at day 4 after ureter ligation, peaked (fivefold of control) at day 15, and decreased gradually until the end of the experiment. In contrast, interstitial cell apoptosis sustained a progressive increase throughout the experiment. Apoptosis was minimal at all experimental time points for control and contralateral kidneys. Compared with control and contralateral kidneys, the ligated kidneys displayed a dynamic expression of mRNAs for many apoptosis-related molecules, which included an up to threefold increase for Fas, Fas ligand, TNF-R1, TRAIL, TRADD, RIP, and caspase-8, and an up to twofold increase for FADD and FAP, but there was little change for FAF. These mRNAs increased between days 4 and 15, decreased until day 30, but then increased again until day 45. The rise and fall of mRNAs between days 4 and 30 paralleled a similar fluctuation in tubular cell apoptosis in that period. The subsequent increase of mRNAs was correlated with a continuous rise of interstitial cell apoptosis. We demonstrated a positive immunostaining for Fas and Fas ligand in the tubular cells at early time points as well as in interstitial inflammatory cells at later time points. Although increased expression of TRAIL, TRADD, RIP, and caspase-8 was noted in tubular cells, there was no staining for these molecules in interstitial cells. CONCLUSION: The current study documents a dynamic expression of several molecules that are known to mediate the most crucial steps of apoptosis. It implicates these molecules in COU-associated renal cell apoptosis and in the pathogenesis of this condition. It also lays the foundation for interventional studies, including genetic engineering, to evaluate the molecular control of apoptosis associated with COU.

Peritubular capillary loss is associated with chronic tubulointerstitial injury in human kidney: altered expression of vascular endothelial growth factor.

Chronic tubulointerstitial injury (CTI) including tubular atrophy and interstitial fibrosis represents one major determinant for the progression of chronic renal disease regardless of cause. Although peritubular capillaries (PTCs) are essential to maintain the normal structure and function of renal tubules, little is known about the role of PTCs in the development of CTI. The integrity of PTCs seems to be regulated by growth factors. Vascular endothelial cell growth factor (VEGF) has recently been recognized as a potent regulator of angiogenesis, vascular survival, and vascular permeability. Knowledge of the role of VEGF in renal disease is still rudimentary, and its role in CTI has not been explored. We analyzed the morphologic changes of PTCs and correlated them with other morphologic parameters of CTI in 32 human kidneys with various types of chronic tubulointerstitial disease. The VEGF expression was immunohistochemically evaluated. Compared with normal kidney, PTC loss (41% to 55% of control) and reduced size of PTCs (55% to 88% of control) were noted in kidneys with CTI. The PTC density was positively correlated with the proximal tubular density (r = 0.66, P <.0001), proximal tubular size (r = 0.54, P <.001), and negatively correlated with interstitial volume (r = -0.84, P <.0001). Compared with normal kidney, where podocytes were the only cell type that constantly expressed VEGF, an interesting pattern of increased VEGF expression by renal tubules, especially morphologically intact or hypertrophic ones, was shared by all cases with CTI. Loss of VEGF in sclerotic glomeruli was noted. PTC injury is pathogenetically linked to tubular atrophy, tubular loss, and interstitial fibrosis in human kidneys with CTI and might be a key factor for the progression of chronic tubulointerstitial disease. The characteristic and uniform pattern of altered VEGF expression in kidneys with CTI may result from ischemia induced by PTC loss and represent a protective mechanism against further PTC injuries. HUM PATHOL 31:1491-1497.

Focal segmental glomerulosclerosis in a 32-year-old kidney allograft after 7 years without immunosuppression.

In kidney allografts, focal segmental glomerulosclerosis (FSGS) has been described as recurrent, de novo, or a histological variant of chronic transplant glomerulopathy. We describe a unique case of de novo FSGS in a renal transplant not accompanied by any feature of rejection in a patient who had not been immunosuppressed for several years. A 58-year-old woman received a histoidentical living-related kidney transplant for end-stage renal disease due to chronic pyelonephritis. Twenty-four years after the transplant she voluntarily discontinued all immunosuppressive medication. Seven years later she presented with nephrotic syndrome, mild renal failure, and positive serology for hepatitis C virus (HCV) antibody. The kidney transplant biopsy disclosed de novo FSGS. Features of acute or chronic rejection, including chronic transplant glomerulopathy, were not seen. The pathogenesis of this lesion is probably related to sustained and prolonged glomerular hyperfiltration; alternatively, HCV infection may have triggered or accelerated the appearance of FSGS.

FK506-associated thrombotic microangiopathy: report of two cases and review of the literature.

BACKGROUND: FK506 is a recently developed immunosuppressant that has been useful in improving the survival of transplanted organs. Among the numerous adverse side effects of FK506, thrombotic microangiopathy (TMA) stands out as an infrequent but severe complication. METHODS: We report two cases of FK506-associated TMA and review the 19 previous reported cases. RESULTS: From these 21 cases, the reported incidence of FK506-associated TMA is between 1% and 4.7%. It is more frequent in females, and the mean age at presentation is 47 years. Eighty-one percent of the cases occurred in patients with kidney allografts, and the remaining patients had liver, heart, or bone marrow transplants. Clinically, TMA was diagnosed at an average interval of 9.3 months from the time of transplantation. Patients may be asymptomatic or may present with the full-blown picture of hemolytic uremic syndrome. All patients had an elevated serum creatinine level but did not always show signs of hemolysis. Trough levels of FK506 were not predictive for the development of TMA, but generally a reduction of drug dose correlated with kidney function improvement and disappearance of the hemolytic picture. The renal allograft biopsy provided a conclusive diagnosis in all 17 cases in which this procedure was performed. Treatment, which mainly consisted of reduction or discontinuation of FK506, anticoagulation, and/or plasmapheresis with fresh-frozen plasma exchange, resolved TMA in most patients (57%). However, in one of these patients (5%), the graft was subsequently lost due to causes unrelated to TMA, such as acute or chronic rejection. Despite treatment, one patient (5%) lost the graft due to acute rejection and persistent TMA, and three other patients (14%) who had bone marrow, heart, and liver transplants, died of multiple organ failure, probably unrelated to TMA. In the remaining four patients (19%), response to treatment was not reported. CONCLUSIONS: TMA must be considered in organ transplant patients treated with FK506 whenever kidney function deteriorates, even in the absence of microangiopathic hemolytic anemia. Although TMA usually responds to treatment, it may, in rare cases, lead to loss of kidney function or even the patient's death.

Cell apoptosis and proliferation in obstructive uropathy.

Distinct patterns of cell proliferation and apoptosis have been recognized for tubular, interstitial, and glomerular cells in chronic obstructive uropathy (OU). In many experimental models of OU, tubular cell apoptosis develops quickly after ureter ligation, peaks between 7 and 24 days postobtruction (about 30-fold of control), and tapers thereafter. Apoptosis initially involves the dilated collecting ducts, but subsequently spreads to other tubules. Tubular cell apoptosis probably accounts for renal tissue loss in OU because a direct correlation between its degree and the decline in dry kidney weight is well-documented. Pronounced tubular cell proliferation occurs shortly after ureter ligation, peaks at about day 6 (60-fold above control), and quickly subsides to baseline. Because the peak of tubular cell proliferation immediately precedes the onset of tubular cell apoptosis, a pathogenetic link may exist between these two processes. Interstitial cell apoptosis occurs with an increasing frequency throughout the course of OU (up to 35-fold above control). Interstitial cell proliferation appears in a bimodal pattern with the early peak coinciding with that of tubular cell proliferation and consisting mostly of fibroblasts, whereas the later peak consists mostly of inflammatory cells. Glomerular cell apoptosis and proliferation are not different from control, which explains, in part, the structural integrity of the glomeruli throughout the disease course. Although the general pathways of cell apoptosis and proliferation are well known, the molecular control of these processes in OU is poorly understood. In addition, whether apoptosis or proliferation of tubular and interstitial cells is differentially regulated remains to be studied. However, several molecules known to be activated or overexpressed in kidney with OU may modulate cell apoptosis and proliferation. The relevant functions of these molecules include induction of apoptosis (angiotensin II, reactive oxygen species, jun-N-terminal kinase, p53), inhibition of the cell cycle (transforming growth factor-beta, p21), inhibition of apoptosis (clusterin, epidermal growth factor, insulin-like growth factor, bcl-2, osteopontin), or promotion of interstitial fibroblast proliferation (platelet-derived growth factor).

Chronic obstructive uropathy in severe combined immunodeficient (SCID) mice: lymphocyte infiltration is not required for progressive tubulointerstitial injury.

Progressive renal injury in humans and experimental animal models is characterized by tubular atrophy, infiltration of mononuclear inflammatory cells, and interstitial fibrosis. Permanent unilateral ureter ligation represents a reproducible model for investigating mechanisms of progressive kidney injury, and in the rat is characterized by tubular epithelial cell proliferation followed by apoptosis and progressive infiltration of monocytes and lymphocytes. Nevertheless, whether monocytes or lymphocytes play a dominant role in causing tubulointerstitial damage remains to be elucidated. In the current study, a model of chronic obstructive uropathy in the mouse is established and the role of lymphocyte infiltration in the evolution of the tubule and interstitial alterations is investigated. Permanent ligation of the left ureter in wild-type (C3H/HeJ) mice resulted in progressive atrophy of tubules and interstitial fibrosis compared with the contralateral kidney over a 30-d period. Immunoperoxidase studies on frozen sections taken from kidneys at 0, 3, 10, 20, and 30 d after ureter ligation showed that the tubulointerstitial injury was accompanied by a marked and progressive increase in interstitial macrophages and T lymphocytes, with no appreciable increase in B lymphocytes. No increase in inflammatory cells was detected in contralateral kidneys over the same time frame. The significance of T lymphocyte infiltration was examined by comparing the degree of tubular atrophy and interstitial fibrosis and the nature and quantity of the inflammatory infiltrate in wild-type mice and C3HSMn.C-Scid/J (SCID) mice subjected to permanent left ureter ligation. SCID mice have genetic defects in immunoglobulin and T cell receptor gene rearrangements and are devoid of circulating mature B and T lymphocytes. Wild-type and SCID mice developed tubular atrophy and interstitial volume expansion in the ligated kidney to the same degree and at the same rate. SCID mice developed a prominent and marked monocyte/macrophage infiltrate in the ligated kidney, which was essentially equal to that in wild-type mice. In contrast, consistent with the known absence of mature lymphocytes in SCID mice, there was essentially no T lymphocyte infiltration into the ligated kidney of SCID mice. These results demonstrate the effective establishment of the model of maintained unilateral ureter ligation in mice, which is readily applicable to genetic mutant strains thus allowing for specific investigation of the role of individual components of the inflammatory response in progressive tubulointerstitial injury. These studies further demonstrate that lymphocyte infiltration is not required for progressive tubular atrophy and increased interstitial fibrosis after maintained unilateral ureter ligation.

p38 kinase activity is essential for osmotic induction of mRNAs for HSP70 and transporter for organic solute betaine in Madin-Darby canine kidney cells.

In renal cells, hypertonicity induces genes for heat shock proteins (HSP70, alpha B-crystallin), as well as enzymes and transporters directly involved in the metabolism and transport of protective organic osmolytes. While heat shock proteins are induced by many stresses including osmotic stress, the induction of the osmolytes genes appears to be specific to osmotic stress. These two adaptive mechanisms allow kidney cells to survive and function in the hypertonic environment that exists on routine basis in kidney medulla. In mammalian cells, hypertonicity induces three mitogen-activated protein kinase pathways: ERK (extracellular regulated kinase), JNK (Jun N-terminal kinase), and p38. ERK activation by osmotic stress is a consistent finding in many cells, but it is not essential for transcriptional regulation of mRNA for transporter of organic osmolyte betaine. While the growth of yeast cells on NaCl-supplemented medium is dependent on HOG1 pathway, it is still unclear which pathway mediates the adaptation to osmotic stress in mammalian cells. Here, we show that inhibition of p38 kinase activity, using the specific inhibitor SB203580 (4-(fluorophenyl)-2-(4-methylsulfonyl-phenyl)-5-(4-pyridyl) imidazole), abolishes the hypertonicity-mediated induction of mRNAs for HSP70 and betaine transporter in Madin-Darby canine kidney cells. The inhibition is dose-dependent and correlates with the in situ activity of native p38 kinase, determined as MAPKAPK-2 activity in cell extracts. As reported previously, the activities of ERK-1 and -2 were not affected by SB203580, but surprisingly, inhibition of native p38 kinase activity correlates with up-regulation of native JNK-1 activity in osmotically stressed cells. p38 mRNA is induced by hypertonic stress and is attenuated with p38 kinase inhibition. We also find that thermal induction of HSP70 mRNA is not affected by p38 kinase inhibition. Such findings suggest that p38 kinase activity is essential for the induction of genes involved in the adaptation of mammalian cells to osmotic stress and that the increased activity of JNK-1 during p38 kinase inhibition is consistent with regulation of JNK-1 by p38 kinase in osmotically stressed cells. In addition, the transduction pathways mediating HSP70 mRNA induction by different stresses appear to be divergent; osmotic induction of HSP70 is p38 kinase-dependent, while thermal induction is not.

Plasma membrane calcium ATPase isoform expression in cultured rat mesangial cells.

The maintenance of intracellular ionized calcium (iCa2+) in the submicromolar range is important for mesangial cell (MC) function, and, as in most mammalian cells, plasma membrane Ca(2+)-ATPases (PMCA) play an important role in the homeostatic process. Molecular studies have demonstrated four PMCA isoforms, each with multiple splice variants. The present study examines the expression of PMCA isoforms and calmodulin-binding region splice variants in cultured MC from Sprague-Dawley rats and from spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats before and after the onset of hypertension in SHR. Using reverse transcription-polymerase chain reaction (RT-PCR) and Southern blot analyses, we have demonstrated PMCA1, -3, and -4, but not PMCA2, to be present in MC from these rat strains. Splice variant analysis revealed PMCA1a and -1b, PMCA3a, -3b, and -3c, and PMCA4a and -4b to be expressed in MC from all three strains. The relative quantities of PMCA1 and PMCA4 mRNA were not different in age-matched SHR vs. WKY rats, correlating with similar iCa2+ measurements. The expression of all three isoforms declined with age in SHR and WKY.

Renal abnormality of calcium handling in spontaneously hypertensive rats.

Hypercalciuria has been observed in human and experimental hypertension. The distal convoluted tubule (DCT) is the site of fine regulation for Ca2+ excretion. We assessed the cellular factors responsible for Ca2+ reabsorption in the DCT of spontaneously hypertensive rats (SHR). Vitamin D3 dependent Ca-binding protein 28k (CaBP28k), a factor involved in Ca2+ influx, and plasma membrane Ca ATPases (PMCA), a factor involved in Ca2+ efflux, were studied in hypertensive 16-week-old SHR and normotensive Wistar-Kyoto rats (WKY). mRNA levels for CaBP28k, PMCA 2 and PMCA 4 were not different in the two strains. However, CaBP28k protein was more abundant, and PMCA protein was less abundant in SHR than in WKY. PMCA antibody recognized only DCT in the two strains. In conclusion, decreased PMCA in DCT may be responsible for the hypercalciuria in SHR.

Ca-ATPase and bone cell mineralization.

The plasma membrane Ca-ATPases play an important role in the regulation of intracellular Ca2+ ion concentration by pumping Ca2+ out of the cell into the extracellular fluid at the expense of ATP. These pumps potentially play an important role in the delivery of Ca2+ during mineralization of hard tissues such as bone. The properties of the plasma membrane Ca2+ pump are compared with those of the sarco(endo)plasmic reticulum and the factors regulating pump function are presented. The different gene products for the plasma membrane Ca-ATPases are described as well as their known functional significance. Particular attention is paid on the plasma membrane Ca2+ pumps present in mineralizing tissues and evidence supporting a role for these pumps in the transcellular delivery of Ca2+ during the process of mineralization is also presented.

Plasma membrane and sarcoplasmic reticulum Ca-ATPase and smooth muscle.

Calcium ions (Ca2+) are involved in the regulation of many cellular activities. The Ca-ATPase(s) of the plasma membrane and of the endoplasmic reticulum play an important role in controlling the intracellular Ca2+ concentration. Therefore, it is not unexpected that these enzymes are modulated by different factors. The activity of the plasma membrane Ca-ATPase is modified by the amount of negatively charged phospholipids surrounding the enzyme. The regulation of the endoplasmic reticulum Ca-ATPase depends on the phosphorylation of phospholamban by cAMP- and cGMP-dependent protein kinase. These two different Ca2+ transport ATPases are present in both visceral and vascular smooth muscle, but tissue- and species-dependent differences in their relative amount have been observed. In this article we will review the characteristics of Ca-ATPases of the smooth muscle.

Cell calcium and arterial blood pressure.

In hypertensive humans and animals, intracellular calcium (Ca2+) has been found to be elevated in several cell types, including platelets, red blood cells, and smooth muscle cells. The messenger role of calcium requires its maintenance within cells at very low (submicromolar) ionic concentrations. This transcellular gradient is maintained by the reversible complexing of calcium to specific proteins. Intracellular free calcium can increase through an enhanced influx, release from intracellular stores, or a reduced efflux of calcium from the cell. Calcium channels are found to be increased in activity in some studies of hypertensive animals, but not in others. Sequestration of calcium within the cell has also been implicated in the increased cytosolic calcium in hypertension. Plasma membrane adenosine triphosphatases (ATPases) play a significant role in calcium extrusion; and the activity of these pumps has been found to be decreased in several human and animal studies. A single cause of the cellular calcium alterations in hypertension is not completely clear.

Renal neoplasm in acquired cystic kidney disease.

The development of renal cell neoplasms ranging from adenoma to metastatic carcinoma is the most serious complication of acquired cystic kidney disease (ACKD). A comprehensive review of the pertinent literature shows that there is up to 50-fold increased risk of renal cell carcinoma in ACKD compared to the general population. The ACKD-associated renal cell carcinoma is seen predominantly in males, occurs approximately 20 years earlier than in the general population, and is frequently bilateral (9%) and multicentric (50%). Acquired cystic kidney disease-associated renal cell carcinoma is frequently asymptomatic (86%), but may be associated with bleeding, abrupt changes in hematocrit, fever, and flank pain or rarely with hypoglycemia, hypercalcemia, or metastases at presentation. Computed tomography seems to provide a better diagnostic yield than sonography or magnetic resonance imaging; nevertheless, large (up to 8 cm) tumors not visualized by any imaging techniques have been reported. It is generally agreed that there is a need for regular screening of symptomatic ACKD patients for early detection of renal cell carcinoma; however, whether screening is needed for asymptomatic patients remains controversial. Nephrectomy is indicated for tumors larger than 3 cm. Management for tumors smaller than 3 cm with persistent symptoms, such as back pain or hematuria, remains controversial, but nephrectomy may be recommended since many of these tumors turn out to be unequivocal renal cell carcinoma. Asymptomatic tumors smaller than 3 cm should be serially screened, and tumor enlargement may be an indication for nephrectomy. Acquired cystic kidney disease-associated renal cell carcinoma accounts for approximately 2% of deaths in renal transplant patients. A median length of survival of approximately 14 months and a 5-year survival rate of 35% are comparable to the same data for renal cell carcinoma in the general population. Successful renal transplant probably decreases the risk of renal cell carcinoma in ACKD patients, but this preliminary observation needs confirmation. The development of ACKD-associated renal carcinoma is a continuous process with evolving phenotypic expression, including damaged renal tubule, simple cyst, cyst with atypical lining, adenoma, and, finally, carcinoma. The pathogenesis of this continuous process is not entirely known, but growth factor-induced compensatory growth of tubular epithelium initiated by the changes of end-stage kidney disease, and probably perpetuated by activation of proto-oncogenes, seems to be the most significant factor.

Differential expression of plasma membrane calcium pump mRNA isoforms in rat osteoblast-like cells.

This study was conducted to identify the plasma membrane Ca2+ transport ATPase (PMCA) mRNA isoforms expressed in rat osteoblast-like cells as PMCAs are likely to participate in calcium deposition in bone. We designed oligonucleotide primers for each PMCA isoform based on rat cDNA sequences in order to develop reverse transcription polymerase chain reaction (RT-PCR) assays. Rat kidney total RNA was used to validate the assays as we have shown that each isoform is present in kidney. When used in RT-PCR assays each primer pair gave rise to a single major product of the appropriate size. Southern blot analysis of the PCR products with oligonucleotide probes specific for each isoform revealed that each probe hybridized only to the expected product. Reamplification of purified PCR products with probe and antisense primers gave rise to products of appropriate size, further confirming the identity of the products. Using these primers we have identified the presence of transcripts for PMCA1, 2 and 4 in RNA from UMR-106 osteoblasts and PMCA1 in RNA from ROS 17/2.8 osteoblasts. We conclude that the two major rat cell lines used as models to study osteoblast function differentially express PMCA mRNA isoforms.

Unusual presentation of glomerulocystic kidney disease in an adult patient.

A 38-year-old Latin-American man developed uremic syndrome without any evidence of previous kidney diseases or any other health problems. Ultrasound and CT scan confirmed normal size of the kidneys without evidence of urinary tract obstruction or renal parenchymal cysts. Kidney biopsy showed cystic dilatation of Bowman's space (glomerulocystic kidney disease). The patient was started on hemodialysis. Severe renal dysfunction and uremic symptoms are a rare initial manifestation of glomerulocystic kidney disease. This pathology should be considered in the differential diagnosis of patients with normal size kidney and chronic renal failure.

ATP inhibits the hydrosmotic effect of AVP in rabbit CCT: evidence for a nucleotide P2u receptor.

In rabbit renal cortical collecting tubule (CCT), perfused in vitro at 38 degrees C, ATP in concentrations of 10(-7) M and greater inhibits arginine vasopressin (AVP)-stimulated osmotic water permeability (Pf). The P1-purinergic receptor antagonist 8-phenyltheophylline did not attenuate the inhibitory action of ATP, and the poorly hydrolyzable ATP analogue, 5'-adenylylimidodiphosphate (AMP-PNP), mimicked the effect of ATP, arguing against an effect of ATP on a P1 receptor or the "P site." Purinergic receptor agonists inhibited AVP-stimulated Pf with the following rank order efficacy: ATP = ADP = UTP = AMP-PNP = alpha, beta-methylene-ATP > 2-methylthio-ATP >> AMP > adenosine, consistent with the pharmacology of a "nucleotide" receptor subtype. Pertussis toxin pretreatment attenuated the action of 10(-5) and 10(-6) MATP; however, 10(-4) MATP failed to inhibit the hydrosmotic action of forskolin or 8-bromoadenosine 3',5'-cyclic monophosphate. Pretreatment with the phosphodiesterase inhibitor RO20-1724 or indomethacin did not inhibit the action of ATP. Staurosporin and 3,4,5-trimethoxybenzoic acid 8-(diethylamino)octyl ester significantly attenuated the inhibition of Pf by lower concentrations of ATP. These data suggest that ATP activates nucleotide receptors on the CCT, mobilizing intracellular Ca2+, which inhibits the hydrosmotic action of AVP.

Reconstitution and partial purification of calcium transport activity from rat kidney cortex.

An ATP-dependent Ca2+ uptake system from rat renal cortical basolateral membranes was solubilized with Triton X-100 and reconstituted into liposomes with lecithin. In the presence of Mg2+, Ca2+ uptake in the reconstituted vesicles was time and ATP dependent and was inhibited by vanadate. Ca2+ uptake in basolateral membrane vesicles depleted of endogenous calmodulin was enhanced by exogenous calmodulin and depressed by R-24571. This sensitivity to calmodulin and R-24571 was lost upon reconstitution in the presence and absence of leupeptin. Vesicles containing Ca2+ uptake activity were separated by gradient centrifugation after Ca2+ was taken up and accumulated as calcium phosphate in the vesicles. This resulted in Ca2+ uptake activity that was enriched 25 times. However, Ca(2+)-dependent adenosinetriphosphatase (ATPase) activity was not enriched significantly. This Ca(2+)-ATPase had two kinetic forms for Ca2+: one was a high-affinity low-capacity form; the other had a low affinity and high capacity. The Ca(2+)-ATPase activity also had two kinetic forms for ATP. All kinetic forms were inhibited by Mg2+. Vanadate, calmodulin, and R-24571 had no effects on Ca(2+)-ATPase activity. A protein doublet of Ca(2+)-dependent hydroxylamine-sensitive phosphorylated intermediates was demonstrated at 125 and 136 kDa in the purified vesicles. This doublet was not altered by addition of leupeptin throughout the purification.

Evidence for a fourth rat isoform of the plasma membrane calcium pump in the kidney.

This study was conducted to identify plasma membrane Ca(2+)-transporting ATPases present in rat kidney. Characterization of the cDNAs of the plasma membrane Ca(2+)-ATPases revealed a family of proteins with regions of highly conserved amino acid sequence. To examine the extent of the diversity of rat renal plasma membrane Ca(2+)-ATPases, we used the polymerase chain reaction to detect additional gene products in rat kidney mRNA that shared these conserved regions. Sequences corresponding to three previously known rat plasma membrane Ca(2+)-ATPases were obtained. In addition, we found sequence corresponding to a new putative plasma membrane Ca(2+)-ATPase. Our results demonstrate that the rat kidney contains at least four different plasma membrane Ca(2+)-ATPases and the complexity of this multigene family is greater than previously thought.