Feedback inhibition of aldose reductase gene expression in rat renal medulla. Galactitol accumulation reduces enzyme mRNA levels and depletes cellular inositol content.

Aldose reductase (AR) is an enzyme responsible for converting glucose into sorbitol and galactose into galactitol. In the renal inner medulla, where sorbitol production plays a role in cellular osmoregulation, AR gene expression has been shown to be osmotically regulated. The present study examined the effects of the accumulation of the AR end product, galactitol, induced by galactose feeding, on AR gene expression and on the balance of other cellular osmolytes, including inositol, in the renal medulla. To differentiate between the effects of excess substrate, product, and intervening osmotic factors, rats were fed either control, galactose, galactose and sorbinil (an AR inhibitor), or control plus sorbinil diets. Renal papillae were assayed for AR mRNA, sodium, urea, galactose, galactitol, sorbitol, inositol, and other organic osmolytes. Galactose feeding resulted in a great accumulation of galactitol and reduction in AR mRNA levels in renal papillae. Associated with these changes was a significant depletion of renal papillary sorbitol, inositol, and glycerolphosphocholine. These effects were largely attenuated by sorbinil. The present findings suggest that renal cellular accumulation of the enzyme's polyol product causes downregulation of AR gene expression. Furthermore, our findings suggest that the inositol depletion associated with sorbitol or galactitol accumulation in various cell types during hyperglycemia may be a function of cellular osmoregulation.

Calcium, cyclic AMP, and MAP kinases: dysregulation in polycystic kidney disease.

Low intracellular calcium, present in untreated polycystic kidney epithelia, results in a proliferative response to cyclic adenosine monophosphate. Treatment with a calcium channel blocker (CCB) caused exacerbation of autosomal dominant polycystic kidney disease in rats. Data regarding use of CCBs in human polycystic kidney disease (PKD) are limited and mixed. Thus, it is premature to extrapolate these findings to human PKD.

Temporal alterations in regional gene expression after nephrotoxic renal injury.

To examine at a molecular level the repair process during recovery from acute renal failure, we determined, after nephrotoxic acute renal injury, levels for several RNAs associated with differentiated functions in the kidney. To delineate changes in different regions, we determined RNA levels separately in cortex, outer medulla, and inner medulla of adult rat kidney. Several RNAs were regionally distributed in control kidneys, including epidermal growth factor (EGF, highest in cortex and outer medulla), aldose reductase (highest in inner medulla), and prostaglandin synthase (highest in inner medulla). Futhermore, RNA for the heat shock protein Hsp70 was present at high levels in the inner medulla of controls. A single intraperitoneal dose of folic acid caused reversible, nonoliguric acute renal failure with impairment of glomerular filtration, tubular sodium reabsorption, and urinary concentration. Fractional excretion of sodium returned to normal by day 3, while serum creatinine, serum urea nitrogen, and urinary osmolality remained abnormal until day 7. During acute renal failure there was a marked, reversible decrease in the regional mRNA levels for EGF, aldose reductase, prostaglandin synthase, and Hsp70. Aldose reductase, prostaglandin synthase, and Hsp70 RNA levels returned to control levels by 7 to 10 days, while EGF remained depressed for a more prolonged period. High levels of aldose reductase, prostaglandin synthase, and Hsp70 RNAs in the inner medulla and their loss/recovery during acute renal failure with a time course similar to alterations in urinary osmolality were consistent with regulation in response to changes in inner medullary osmolality, as has been demonstrated for aldose reductase in normal animals. (ABSTRACT TRUNCATED AT 250 WORDS)

Epidermal growth factor ameliorates autosomal recessive polycystic kidney disease in mice.

C57BL/6J mice homozygous for the cpk gene exhibit an autosomal recessive (AR) form of polycystic kidney disease (PKD), similar to human ARPKD, with massive collecting duct cysts. These cysts are lined by epithelial with an immature phenotype. Since renal expression of epidermal growth factor (EGF) is also significantly decreased in affected mice, we hypothesized that renal EGF is necessary for normal developmental maturation of the collecting duct. To determine if the lack of EGF may be a decisive factor in the initiation and/or growth of collecting duct cysts, we administered exogenous EGF (1 microgram/g body wt subcutaneously) daily for Postnatal Days 3-9 (a critical period for collecting duct maturation) to C57BL/6J-cpk mice. EGF but not sham or albumin treatment retarded the development of PKD, reduced the degree of renal failure associated with the disease, and prolonged the survival of cystic mice. Sulfated glycoprotein-2 gene expression, a marker of immaturity in collecting duct cells, was reduced in cystic kidney by EGF treatment. This finding indicates that EGF treatment was associated with an increase in the maturation of the collecting duct epithelial cells. These findings support the view that decreased EGF may play a significant role in promoting the enlargement of collecting duct cysts in a hereditary model of ARPKD and that PKD involves defective and/or arrested collecting duct cell maturation.

Elevated c-myc protooncogene expression in autosomal recessive polycystic kidney disease.

The polycystic kidney diseases (PKDs) are a group of disorders characterized by the growth of epithelial cysts from the nephrons and collecting ducts of kidney tubules. The diseases can be inherited or can be provoked by environmental factors. To investigate the molecular basis of the abnormal cell growth associated with PKD, c-myc protooncogene expression was studied in a mouse model for autosomal recessive PKD. Homozygous recessive C57BL/6J (cpk/cpk) mice develop massively enlarged cystic kidneys and die from renal failure shortly after 3 weeks of age. Quantitative dot blot and RNA blot hybridization experiments in which whole kidney poly(A)+ RNA was hybridized with a c-myc RNA probe showed a 2- to 6-fold increase in c-myc mRNA at 2 weeks, and a 25- to 30-fold increase in c-myc mRNA at 3 weeks of age in polycystic mice, as compared to normal littermates. c-myc expression was also examined under two conditions in which kidney cell growth was experimentally induced in normal adult mice: compensatory renal hypertrophy and tubule regeneration following folic acid-induced renal cell injury. While compensatory hypertrophy resulted in only a small (less than 3-fold) increase in c-myc, folic acid treatment gave rise after 24 hr to a 12-fold increase in c-myc mRNA. The induction of c-myc by folic acid is consistent with increased cellular proliferation in regenerating tubules. In contrast, polycystic kidneys show only a minimal increase in cellular proliferation over that seen in normal kidneys, while c-myc levels were found to be markedly elevated. Thus, the level of c-myc expression in cystic kidneys appears to be out of proportion to the rate of cell division, suggesting that elevated and potentially abnormal c-myc expression may be involved in the pathogenesis of PKD.

Increased renal expression of monocyte chemoattractant protein-1 and osteopontin in ADPKD in rats.

BACKGROUND: Human autosomal-dominant polycystic kidney disease (ADPKD) is variable in the rate of deterioration of renal function, with end-stage renal disease (ESRD) occurring in only approximately 50% of affected individuals. Evidence suggests that interstitial inflammation may be important in the development of ESRD in ADPKD. Han:SPRD rats manifest ADPKD that resembles the human disease. Homozygous cystic (Cy/Cy) rats develop rapidly progressive PKD and die near age 3 weeks. Heterozygous (Cy/+) females develop slowly progressive PKD without evidence of renal dysfunction until the second year of life, whereas heterozygous (Cy/+) males develop more aggressive PKD with renal failure beginning by 8 to 12 weeks of age. METHODS: To examine the relationship between proinflammatory chemoattractants and the development of interstitial inflammation and ultimately renal failure in ADPKD, we evaluated monocyte chemoattractant protein-1 (MCP-1) and osteopontin mRNAs and proteins in kidneys from Han:SRPD rats. RESULTS: MCP-1 and osteopontin mRNAs, expressed at low levels in kidneys from normal (+/+) animals at all ages, were markedly elevated in kidneys from 3-week-old Cy/Cy animals. In kidneys from heterozygous (Cy/+) adults of either gender, MCP-1 and osteopontin mRNAs were more abundant than normal; MCP-1 mRNA was more abundant in Cy/+ males than in females. Thus, chemoattractant mRNA expression correlated with the development of renal failure in Cy/Cy and Cy/+ rats. Osteopontin mRNA, localized by in situ hybridization, was moderately expressed in the renal medulla of normal animals; however, this mRNA was expressed at very high levels in the cystic epithelia of Cy/+ and Cy/Cy animals. MCP-1 and osteopontin proteins, localized by immunohistochemistry, were weakly detected in +/+ kidneys but were densely expressed in Cy/Cy and in adult Cy/+ kidneys, primarily over cystic epithelium. Increased expression of chemoattractants was associated with the accumulation of ED-1 positive cells (macrophages) in the interstitium of cystic kidneys. CONCLUSIONS: We suggest that proinflammatory chemoattractants have a role in the development of interstitial inflammation and renal failure in ADPKD.

Sequential protooncogene expression in regenerating kidney following acute renal injury.

Following loss of functional renal mass due to acute injury, there is significantly increased proliferation of tubular epithelium to replace injured and necrotic cells. In contrast, following uninephrectomy, the contralateral kidney increases in size primarily by hypertrophy, with little cellular proliferation. We and others have demonstrated only modest increases in renal protooncogene expression following uninephrectomy. In this study, we demonstrate markedly elevated expression of the protooncogenes c-fos, c-myc, c-Ki-ras, and c-Ha-ras following acute renal injury induced by a single large parenteral dose of folic acid. The expression of these genes occurs in a sequential pattern similar to that seen in proliferating cells in culture and in regenerating liver. In addition, we demonstrate elevated levels of histone H4 and beta-actin mRNAs consistent with increased cell proliferation. These data suggest that the molecular mechanisms regulating cell proliferation in the kidney are similar to those in regenerating liver and in cultured cells. In addition, it appears that these events are regulated normally after acute renal injury and following uninephrectomy, since in both instances the levels of protooncogene expression correlate with the degree of cell proliferation. This is in direct contrast to a pathologic renal condition, polycystic kidney disease, in which the level of protooncogene expression is out of proportion to the degree of cell proliferation. Further studies of the molecular correlates of acute renal injury may yield insight into the pathogenesis of this and other clinically important renal disorders.

In vivo osmoregulation of aldose reductase mRNA, protein, and sorbitol in renal medulla.

Sorbitol accumulates in renal medullary cells by synthesis from glucose in a reaction catalyzed by aldose reductase. Medullary sodium and urea are high and vary with urinary concentration. Sorbitol varies similarly, consistent with its role as a compatible intracellular organic osmolyte. We measured renal medullary sodium, urea, sorbitol, aldose reductase (protein and activity), and aldose reductase mRNA in rats treated to change medullary sodium and urea. In untreated Brattleboro rats all measurements were low and increased after 7 days of treatment with arginine vasopressin. In contrast, when normal rats were water deprived for 3 days, urea increased out of proportion to sodium, and sorbitol, aldose reductase, and aldose reductase mRNA were unchanged. After 2 h of diuresis, normal rats had lower medullary sodium and urea and reduced mRNA and sorbitol; however aldose reductase did not change. These data are consistent with previous results from cultured cells in which altered extracellular sodium, but not urea, leads to rapid changes in aldose reductase mRNA and slow changes (days) in aldose reductase. In addition, acute decreases in extracellular sodium increase leakage of sorbitol from cells. We also confirm previous results showing medullary glycerophosphorylcholine correlates best with urea, whereas the sum of all compatible osmolytes correlates best with sodium.

Elevated proto-oncogene expression in polycystic kidneys of the C57BL/6J (cpk) mouse.

Polycystic kidney disease in the C57BL/6J (cpk) mouse is an autosomal recessive disorder which leads to the rapid development of renal cysts and kidney failure during the first 3 to 4 postnatal weeks. Previously, we showed that the cystic kidneys of affected mice have abnormally elevated levels of c-myc mRNA. In the study presented here, it is shown that mRNAs for the proto-oncogenes c-fos and c-Kiras, as well as c-myc, are markedly elevated in cystic kidneys, suggesting that there is a more general abnormality in gene expression associated with the disease. It is also evident that there are two stages to this abnormal proto-oncogene expression. In the first stage, which occurs up through the second postnatal week, there are modest increases in proto-oncogene mRNA which parallel the increased cell proliferation that accompanies cyst growth at this time. In the second stage, which occurs after the second postnatal week, there are markedly elevated levels of proto-oncogene mRNA that are seen at a time when cell proliferation is declining. The development of this latter stage suggests either that there is a fundamental abnormality intrinsic to polycystic kidneys that leads to uncontrolled proto-oncogene expression later in disease progression or that there is a secondary response in the kidney to the progressive renal failure.

Long-term ammonium chloride or sodium bicarbonate treatment in two models of polycystic kidney disease.

Administration of ammonium chloride aggravates, while short-term administration of sodium or potassium bicarbonate lessens the development of polycystic kidney disease in Han:SPRD rats. We have conducted studies to determine whether the protection afforded by the administration of sodium bicarbonate is sustained and prevents development of uremia during chronic administration and whether the effects of the administration of ammonium chloride and sodium bicarbonate are also observed in a different model of polycystic kidney disease, the CD1-pcy/pcy mouse. We found that chronic administration of 200 mM sodium bicarbonate to Han:SPRD rats inhibited cystic enlargement and prevented the subsequent development of interstitial inflammation, chronic fibrosis, and uremia. We also found that, while the administration of ammonium chloride has similar effects in Han:SPRD rats and CD1-pcy/pcy mice, the administration of sodium bicarbonate is only protective in the Han:SPRD rats. This probably reflects differences in these models (predominately involvement of proximal tubules in Han:SPRD rats and of collecting ducts and distal tubules in pcy/pcy mice) and the different location and nature of the renal metabolic responses to the administration of acid or alkaline load.

Gender and the effect of gonadal hormones on the progression of inherited polycystic kidney disease in rats.

Human autosomal dominant polycystic kidney disease (ADPKD) is a common inherited disease and displays a gender dimorphism in renal disease progression. Han:SPRD-Cy rats manifest a form of ADPKD that is similar in many respects to that seen in humans. In Han:SPRD rats, male Cy/+ rats have more prominent renal changes and develop renal failure at an early age, whereas female Cy/+ rats exhibit less severe renal cystic change and have normal renal function until advanced age. To determine whether the male gonadal hormone, testosterone, contributes to this gender dimorphism, males were sham operated or castrated; some castrated rats were repleted with 5alpha-dihydrotestosterone. Female rats were sham operated or ovariectomized before sham operation or testosterone treatment. All treatments started at 4 weeks of age and ended at 10 weeks of age. Renal enlargement, cystic change, and renal function were assessed. In the males, castration reduced renal enlargement and cystic change; testosterone treatment abrogated these effects. Neither of these manipulations affected azotemia in male Cy/+ rats. In the females, testosterone was renotropic for both normal and cystic kidneys. In the Cy/+ females, testosterone treatment caused azotemia and an increase in the severity of the PKD. Ovariectomy blunted the effect of testosterone on cystic kidney enlargement. Testosterone treatment did not completely erase the gender-associated differences in azotemia in the Cy/+ rat. These data confirm the renotropic effects of testosterone and indicate that testosterone influences the progression of renal cystic change in male and female rats with ADPKD.

In vivo and in vitro osmotic regulation of HSP-70 and prostaglandin synthase gene expression in kidney cells.

As a function of the urinary concentrating mechanism, the cells of the renal medulla are exposed to elevated and constantly varying osmolalities and adapt to this environment by selectively expressing certain mRNAs. We evaluated the expression and regulation of two RNAs that may be important in adaptation of rental medullary cells to hyperosmolality. We demonstrate selective, modulated expression in the renal medulla of heat shock protein HSP-70 mRNA and prostaglandin synthase-1 mRNA, with the abundance of these two mRNAs regulated in vivo in concert with changes in medullary sodium and urea. We also determined the abundance of these mRNAs in cultured kidney cells (MDCK) in response to an increase in extracellular osmolality due to selected osmotic agents. HSP-70 and prostaglandin synthase-2 mRNA levels increased when extracellular osmolality was increased to 400-600 mosmol/kg by the addition of NaCl. At 500 mosmol/kg this response was evident at 6 h, was maximal near 24 h, and persisted for a total of 90 days. Prostaglandin synthase-1 mRNA levels in MDCK cells were also increased after chronic exposure to extracellular osmolality. Increased extracellular osmolality caused by agents to which cells are impermeable caused increased levels of HSP-70 and prostaglandin synthase-2 mRNAs, whereas increased extracellular osmolality caused by agents to which cells are permeable did not; thus osmotic regulation involved osmotic water movement. We conclude that the abundance of HSP-70 and prostaglandin synthase-1 mRNAs in the renal medulla is regulated in response to renal medullary osmolality and suggest that this may also be true for other medullary mRNAs yet to be described.

In vivo adenovirus-mediated gene transfer into normal and cystic rat kidneys.

Gene transfer into the mammalian kidney has proved difficult because of the structural complexity of the organ and its low mitotic index. This article describes the use of intra-arterially injected adenovirus to study gene transfer into the rat kidney in vivo. By pre-chilling the kidney, and incubating the virus with the kidney in the cold for extended periods of time, we were able to successfully transfer a beta-galactosidase (beta-gal) reporter gene into the vasculature without ischemic injury to the kidney. Transfer occurred largely in the cortex when cold was used alone, whereas with the use of cold and vasodilators, transfer was accomplished into the outer medulla in both the inner and outer stripes. In the Han:SPRD rat model of autosomal dominant polycystic kidney disease (ADPKD), gene transfer occurred into the vasculature, some epithelial cysts and interstitial cells. This is the first description of substantial in vivo gene transfer into both normal and cystic kidneys. The methodology could find application in the creation of new models of renal disease, for in vivo therapeutic intervention or for genetic modification of an allograft at the time of harvest.

Tumor necrosis factor-alpha mRNA and protein in rat uterine and placental cells.

The pregnant uterus contains TNF-alpha, a potent cytokine with pleotrophic effects. The uterus also contains numerous macrophages, which are well described sources of TNF-alpha. In order to determine if uterine TNF-alpha originated with these macrophages, patterns of macrophage tissue distribution and population densities were first established in rat uterine tissues from early, mid, and late stages of gestation by immunohistology. The potential of these and other uterine and placental cells to synthesize TNF-alpha was then tested by in situ hybridization with the use of biotinylated antisense and sense RNA probes. Although TNF-alpha mRNA was present during all stages of pregnancy, hybridization signals were highest in gestation day 15 tissues. The predominant TNF-alpha mRNA-containing cells were uterine epithelium, decidual cells, and placental trophoblast cells; these cells also contained immunoreactive TNF. Transcription of the TNF gene in the uterus and placenta was also documented by Northern blotting experiments, which showed that the transcript sizes for uterine, placental, and macrophage TNF mRNA were similar. Although stromal cells that were located in macrophage-rich uterine compartments (myometrium, metrial gland) contained TNF-alpha mRNA, the cells did not contain high levels of immunoreactive TNF-alpha. Thus, cells other than macrophages are likely to be the major contributors of TNF-alpha during uncomplicated pregnancy in the rat.

Modification of disease progression in rats with inherited polycystic kidney disease.

The most common inherited form of human polycystic kidney disease (PKD), autosomal dominant PKD (ADPKD), is a leading cause of chronic renal failure, but has a variable clinical presentation, with end-stage renal disease occurring in only 25% to 75%. Several findings are consistent with the idea that factors in addition to the primary mutation can affect the progression of cystic change and chronic renal failure in PKD. Epithelial cell proliferation is a central element in the pathogenesis of renal cysts. We postulated that the superimposition of a growth-promoting stimulus might promote more intense proliferation of cystic epithelial cells in inherited cystic disease. To study this, we subjected Han:SPRD rats, with a form of ADPKD that resembles human ADPKD, from 4 until 10 weeks of age to diets designed to promote tubule cell growth. The diets included supplemental NH4Cl (280 mmol/L in drinking water), limited dietary K+ (0.016% of diet; control diet was 1.1% K+), and increased dietary protein (50%; control diet was 23% protein). Treatments designed to promote cell growth caused more aggressive PKD in males and females, worsened azotemia in males, and resulted in azotemia in females (which normally develop PKD but not azotemia at the ages studied). NH4Cl, K+ restriction, and increased dietary protein each caused greater kidney enlargement in males (kidney weight/body weight ratios increased by 35%, 78%, and 105%, respectively) and worsened azotemia in males (serum urea nitrogen values increased by 63%, 514%, and 224%, respectively); in contrast, decreased dietary protein (4%) caused less severe PKD in males (kidney weight/body weight ratios decreased by 43%) and lessened azotemia in males (serum urea nitrogen values decreased by 49%). Similarly, NH4Cl and K+ restriction caused greater kidney enlargement in females (kidney weight/body weight ratios increased by 206% and 203%, respectively) and caused azotemia in females (serum urea nitrogen values increased by 177% and 430%, respectively). On the basis of these results, we conclude that growth-promoting stimuli can alter the expression of hereditary renal cystic disease. These findings demonstrate that the progression of hereditary renal cystic disease can be altered by factors in addition to the primary genetic defect.

Methylprednisolone retards the progression of inherited polycystic kidney disease in rodents.

Polycystic kidney disease in adult laboratory animals and humans is associated with enlarged kidneys and a progressive decline of renal function, resulting in death from uremia. Interstitial inflammation and fibrosis typically are observed in association with the development of renal insufficiency. To determine whether amelioration of interstitial inflammation and fibrosis may diminish cyst expansion/kidney enlargement and stabilize renal function, we administered methylprednisolone, an anti-inflammatory drug with antifibrogenic effects, to mice and rats with hereditary polycystic kidney disease. The experiment was repeated once for each species. Mice were studied both in America and in Japan. Weanling male and female mice (DBA/FG pcy/pcy [cystic] and +/+ [normal], n = 87 and 20, respectively) and rats (Han:SPRD Cy/+ and +/+, n = 70 and 33, respectively) were administered methylprednisolone (1 to 2 mg/kg/d) in the drinking water for 100 days (mice) or 42 days (rats). Control animals drank distilled water. In normal DBA +/+ mice, methylprednisolone had no effect on serum urea nitrogen (SUN) levels, kidney weight, or kidney/body weight. Untreated male and female mice developed cystic kidneys and azotemia to an equal extent. Methylprednisolone administered in America to mice with renal cystic disease decreased kidney weight, kidney/body weight, SUN levels, volume density of cysts, and severity of interstitial fibrosis. In Japan, methylprednisolone decreased kidney weight and SUN levels of animals with cystic disease, but the effect on kidney/body weight did not reach statistical significance. In contrast to mice, male rats developed more severe renal cystic changes and were more azotemic than female rats. Methylprednisolone administered to male rats with cystic disease decreased SUN levels, kidney weight, kidney/body weight, volume density of cysts, and severity of interstitial fibrosis. Methylprednisolone had no effect on kidney/body weight or SUN levels in female rats with renal cystic disease. In normal Han:SPRD (+/+) rats of both sexes, kidney and body weight were decreased by methylprednisolone, but kidney/body weight and SUN levels were unchanged. On the basis of this study, we conclude that methylprednisolone decreased the extent of renal enlargement, reduced renal interstitial fibrosis, and preserved kidney function in mice and rats with relatively severe forms of inherited polycystic kidney disease.

Hyperoxia amplifies TNF-alpha production in LPS-stimulated human alveolar macrophages.

Human alveolar macrophages (AM) produce a number of inflammatory mediators including tumor necrosis factor (TNF). TNF-alpha has been implicated in several forms of lung injury including that associated with oxygen toxicity. To investigate whether oxygen could induce or augment the release of TNF from AM, we acquired AM from nonsmoking volunteers and determined TNF release after in vitro hyperoxia. Although TNF release was not induced by oxygen exposure alone, if lipopolysaccharide (LPS) stimulation occurred simultaneously, there was significant augmentation by 60 and 95% oxygen over LPS-stimulated AM exposed to 21% oxygen. This increase was paralleled by a significant increase of interleukin (IL)-1 beta. Dimethylthiourea (DMTU), a hydroxyl radical scavenger, inhibited this release. The increase in TNF extracellular concentrations induced by hyperoxia was not associated with significant increases in intracellular concentration or detectable mRNA over LPS-stimulated AM exposed to 21% oxygen. We hypothesize that hyperoxia exposure may alter the LPS-stimulated AM cytoplasmic milieu, thus further enhancing TNF-alpha production by a post-transcriptional mechanism.

Volumetric determination of progression in autosomal dominant polycystic kidney disease by computed tomography.

BACKGROUND: Autosomal dominant polycystic kidney disease (ADPKD) is characterized by progressive renal enlargement and renal failure. We evaluated sequential radiocontrast-enhanced computed tomography (CT) scans to determine the rate of kidney enlargement in patients with ADPKD. METHODS: Ten adult patients with ADPKD (4 men and 6 women) with initial serum creatinine levels 0.05), and 185 +/- 52 mL (P < 0.01), respectively. In 19 individual spherical cysts selected in six patients, the mean initial volume was 15.0 +/- 7.2 mL (range 1.1 to 137 mL), and the average rate of volume increase was 0.52 +/- 0.21 mL/month (P < 0.025, range 0.02 to 4.15 mL/month). In five patients who eventually required dialysis, 11.2 years after the initial CT scan, the initial cyst/kidney volume ratio (combined for both kidneys) exceeded 0.43; four patients with lower cyst/kidney volume ratios had serum creatinine levels <1.5 mg/dL, 8.7 years after the initial CT scan. CONCLUSIONS: On the basis of this preliminary survey of archival material, we conclude that conventional contrast-enhanced CT scans can be used to quantitate volume components of progression in ADPKD. The rates of individual kidney and cyst enlargement are highly variable within and between patients, but overall, the values increase over time. The volume fraction of kidneys comprised of cysts may be a useful indicator of ADPKD progression.

Molecular cloning of cDNA coding for kidney aldose reductase. Regulation of specific mRNA accumulation by NaCl-mediated osmotic stress.

Cells generally respond to long-term hyperosmotic stress by accumulating nonperturbing organic osmolytes. Unlike bacteria, in which molecular mechanisms involved in the increased accumulation of osmolytes have been identified, those in multicellular organisms are virtually unknown. In mammals, during antidiuresis, cells of the renal inner medulla are exposed to high and variable extracellular NaCl. Under these conditions, the cells contain a high level of sorbitol and other osmolytes which help balance the high extracellular osmolality. PAP-HT25 is a continuous line of cells derived from rabbit renal inner medulla. When medium osmolality is increased by raising the NaCl concentration, these cells accumulate sorbitol. The sorbitol is synthesized from glucose in a reaction catalyzed by aldose reductase. When the medium is made hyperosmotic, aldose reductase activity increases because of a larger increase in the amount of enzyme. This increase is produced by the accelerated rate of synthesis of aldose reductase protein. The purpose of the present studies was to examine the mechanism of this increase in aldose reductase protein by measuring the relative abundance of aldose reductase mRNA. A cDNA clone coding for rabbit kidney aldose reductase was isolated. Antisense RNA probes transcribed from this clone hybridized specifically with a 1.5-1.6 kilobase mRNA in Northern blots. Cells grown chronically in hyperosmotic medium had a relative abundance of this specific mRNA which was six times that of cells grown in isoosmotic medium. When cells grown in isoosmotic medium were switched to hyperosmotic medium, the level of aldose reductase mRNA peaked (18-fold) at 18-24 h. The induction of aldose reductase mRNA by osmotic stress was reversible. Our finding of increased abundance of a specific mRNA in direct response to hyperosmotic stress represents the first report of such an effect in animals.

Autosomal recessive polycystic kidney disease in a murine model. A gross and microscopic description.

The C57BL/6J-cpk genetic murine model of autosomal recessive polycystic kidney disease was examined to gain insight into the pathogenesis of renal cystic disease. Fetal through 3-week-old offspring of heterozygote matings were used to study growth parameters and morphology of this genetic form of cystic disease. The kidneys were examined by light and electron microscopy and nephron segments were microdissected. Two phases of cystic disease development were morphologically identified. The first phase in fetal and newborn affected pups was characterized by proximal tubule enlargement and a general increase in the tubular mitotic index. The proximal tubules showed cytologic abnormalities along with an increased necrotic cell index. The later phase, in one through 3-week-old cystic pups, was characterized by progressive enlargement of the kidneys due mainly to cystic change of the collecting ducts and by development of azotemia. Secondary to the azotemia was a stunted body growth. Significant tubular epithelial hyperplasia was not found by mitotic index during the second phase, but an increase in collecting duct cellularity was present. Histone H4 gene expression, which is tightly coupled to DNA synthesis and thus an index of cell proliferation, showed only a minimal increase in cystic kidneys at 1, 2, and 3 weeks of age. Therefore, the degree of cell proliferation necessary to allow the observed tubular enlargement appears to be minimal.