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J J Grantham

Publications and source records attributed to J J Grantham.

At least 73 records · Page 4Linked to original sources
PubMed1991

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.

Animals↗
PubMed1991

Arginine vasopressin stimulates net fluid secretion in a polarized subculture of cyst-forming MDCK cells.

Transepithelial fluid secretion is an important process in the progressive enlargement of certain types of renal cysts. Arginine vasopressin (AVP) increases the rate of cyst formation and expansion in an in vitro model of renal cysts that uses Madin-Darby canine kidney (MDCK) cells grown in a gelled matrix of Type 1 collagen. In this study, it was determined if AVP promoted net fluid secretion by MDCK cells. The rate of volumetric fluid secretion was determined from the net movement of water across epithelial layers of MDCK cells grown on permeable, collagen-coated membranes. AVP in the basolateral medium (but not in apical medium) at concentrations exceeding 10(-9) M caused sustained basolateral to apical transepithelial fluid secretion (approximately 0.6 microL/cm2/h). 1-Desamino-8-D-AVP, a V2 receptor agonist, had a similar effect. The secreted fluid was hyperosmotic compared with the bath (5.7 to 9.7 mosM). Chloride was consistently secreted, but the absolute level in the secreted fluid was variable. Intracellular cAMP content was increased 187% by a 2-h exposure to AVP and 10(-4) M methylisobutylxanthine. Net fluid secretion was augmented by methylisobutylxanthine and theophylline and was inhibited by ouabain, bumetanide, and a sodium-dependent Cl-/HCO3- exchange inhibitor (L-645,695) but was not altered by clonidine, guanabenz, or indomethacin. AVP-induced fluid secretion was not accompanied by a change in transepithelial hydraulic conductivity. It is suggested that AVP stimulates fluid secretion of MDCK epithelial monolayers by activating V2 receptor-mediated adenylate cyclase. The regulation of net fluid secretion by AVP would appear to depend on modulation of solute transport, rather than on water permeability.

Animals↗
PubMed1991

Cellular electrolyte and volume changes induced by acidosis in the rabbit proximal straight tubule.

Cellular acidosis induced either by high Pco2 or by low HCO3- concentrations has been shown to cause cell swelling in isolated, lumen-collapsed, S2 segments of the rabbit proximal tubule (Sullivan et al., Am J Physiol 1990; 258: F831-F839). The swelling is not followed by a volume regulatory response. The ionic basis of the swelling has been investigated by measurement of the cellular K+, Na+, and Cl- content (electron probe) and HCO3- concentration (pH-sensitive fluorescent dye). Cell content of K+, Na+, and Cl- was expressed as a ratio to P content. Exposure to 15% CO2 increased K/P from 0.98 to 1.16, Cl/P from 0.14 to 0.20, and Na/P from 0.09 to 0.11. Cell (HCO3-) increased from 22 to 32 mM. Reduction in bath (HCO3-) from 25 to 5 mM reduced cell (HCO3-) from 24 to 8 mM and increased K/P from 0.75 to 0.90. Na/P fell from 0.13 to 0.09, and Cl/P fell from 0.15 to 0.12. Thus, swelling resulting from acidosis induced by high CO2 was accompanied by an accumulation of K+, Cl-, and HCO3-; that resulting from acidosis induced by a fall in (HCO3-) was combined with an accumulation of K+ and an unidentified anion. To determine if the swelling induced by a fall in pH might be coupled with depolarization of the basolateral membrane, the effect of 1 mM barium was tested. Barium caused cell volume to increase 10.2%. Cell pH rose from 7.38 to 7.56, K/P increased from 0.63 to 0.73, Na/P did not change, and Cl/P rose from 0.17 to 0.20. Cell (HCO3-) increased 10.4 mM. When the pH of the barium-treated tissue was reduced to 7.02 by raising Pco2, additional cell swelling and accumulation of K+ occurred. The effect on cell volume of a reduction of bath (HCO3-) from 25 to 5 mM at constant bath pH was determined. Cell pH was not altered. Cell volume decreased 3% initially and then returned to the control level. When the bath (HCO3-) was restored to 25 mM, cell volume increased 3.9% and then returned to the baseline. Thus, volume regulation was not impaired. It was concluded that a fall in cell pH induces swelling, and this is coupled with an accumulation of K+. This is probably the result of a pH effect on barium-sensitive and barium-insensitive K+ conductance pathways. The nature of the anions that balance the gain in K+ depends on the means used to induce acidosis.

Acidosis, Renal Tubular↗
PubMed1991

Natural history of acquired renal cystic disease in dialysis patients: a prospective longitudinal CT study.

Patients with end-stage kidney disease, particularly those treated with dialysis, develop proliferative processes in their native kidneys that result in the formation of multiple acquired renal cysts, renal adenomas, and carcinomas. Data about these abnormalities have been acquired mainly from retrospective studies. We undertook a longitudinal prospective study in which the native kidneys of 30 dialysis patients were surveyed by serial CT during a 7-year period to study the natural history of acquired renal cystic disease and the frequency of associated complications. Acquired cysts were seen in 87% of patients at the end of the study compared with 57% at the study's onset, and a significant increase was seen in mean renal volume with time. Five patients (17%) developed large hemorrhagic renal cysts and four (13%) developed large perinephric hematomas. Renal cell carcinomas developed in two patients (7%) without renal symptoms. One carcinoma invaded the renal capsule, but did not cause metastases. The other carcinoma was widely metastatic. Our findings are consistent with those of earlier studies documenting an increased prevalence of renal cell carcinoma in dialysis patients as compared with the general population. However, our conclusions are limited because the study sample is small and no control population was studied. We conclude that acquired renal cystic disease in dialysis patients is a progressive disorder often associated with cyst hemorrhage. Dialysis patients may also have an increased prevalence of renal cell carcinoma.

Carcinoma, Renal Cell↗
PubMed1990

Fluorescein transport in isolated proximal tubules in vitro: epifluorometric analysis.

An epifluorometric method was used to quantify the bidirectional fluxes of fluorescein across the basolateral surfaces of nonperfused rabbit tubule segments in vitro. Proximal S2 segments, but not cortical collecting tubules or cortical thick ascending limbs, accumulated fluorescein to levels in cytoplasm over 100-fold greater than in the external medium. The rate of intracellular fluorescein accumulation was dependent on the concentration of the ligand in the external bath. The apparent Km was 10 microM and the Vmax was 623 x 10(-6) mol.min-1.l-1. Probenecid and ouabain inhibited fluorescein accumulation. We conclude that fluorescein is transported into the cytoplasm of proximal tubules by basolateral mechanisms that share features in common with the classical organic anion system. This fluorescent compound offers some unique advantages for the study of the organic anion transport system in intact cells.

Animals↗
PubMed1990

Effect of cellular acidosis on cell volume in S2 segments of renal proximal tubules.

The presence of pH-sensitive transport mechanisms in the basolateral membrane of proximal tubular cells suggests that cell volume and its regulation may be sensitive to changes in cell pH. We have measured the response of cell pH and cell volume to changes in the acid-base composition of solutions bathing isolated, lumen-collapsed, proximal S2 tubular segments taken from the rabbit kidney. Cell pH was determined by measurement of the fluorescence emission of 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein. Cell volume was calculated from measurements of tubular diameter. An increase in CO2 from 5 to 15% reduced cell pH 0.30 units and raised cell bicarbonate concentration ([HCO3]) 10 mM. Cell volume rose to 108.6% of control in 4 min. A decrease in bath [HCO3] from 25 to 5 mM reduced cell pH 0.41 units and cell [HCO3] by 15 mM. Cell volume gradually increased to 105.7% at 8 min. The rate of the regulatory volume decrease after cell swelling on exposure to a 160 mosM solution was determined in the presence of 5 and 15% CO2. The latter reduced the maximum fractional rate of recovery of volume from 0.18 to 0.11 min-1 but did not affect the extent of regulation. We conclude that acidosis causes cell swelling and reduces the rate of volume regulation in response to hypotonic media.

Acidosis↗
PubMed1990

Proximal tubule volume regulation in hypo-osmotic media: intracellular K+, Na+, and Cl-.

This study sought to measure the net loss of intracellular K+, Na+, and Cl- that accompanied isosmotic cell volume regulation in hypotonic media and to determine if electrolyte loss depended on the rate at which the extracellular osmolality was reduced. Isolated nonperfused proximal S2 segments from rabbit kidney cortex were studied in vitro. Gradual lowering of osmolality from 295 to 150 mOsm/kg at a rate of 2 mOsm/kg/min did not cause an increase in tubule cell volume until the medium osmolality decreased below 190 mOsm/kg. By contrast, tubules rapidly bathed in low osmolality media exhibited classical osmometric swelling followed by incomplete volume regulatory decrease. Volume regulation associated with gradual and rapid lowering of osmolality was accompanied by the net loss of intracellular K+, Na+, and Cl- (measured by electron probe); however, the temporal pattern of electrolyte loss depended on the rate of osmotic change. With gradual lowering of osmolality, cell K+ content did not decrease significantly until osmolality was lowered below 200 mOsm/kg, whereas Cl- was lost at the 200 mOsm/kg level and below. With rapid lowering of osmolality, cell K+ content was strikingly decreased at the 200 mOsm/kg level, but Cl- did not change appreciably until osmolality was decreased to 150 mOsm/kg. Cell Na+ content decreased in hypo-osmotic media, but the magnitude was relatively small. During volume regulation that accompanied either gradual or rapid lowering of medium osmolality from 295 to 150 mOsm/kg, intracellular osmolal gap, the difference between medium osmolality and the sum of intracellular concentrations of K+, Na+, and Cl- decreased 87 and 58 mOsm/kg, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Transepithelial water permeability in an in vitro model of renal cysts.

Renal cysts develop from microscopic tubules and may enlarge progressively several thousandfold. Sustained epithelial proliferation, intracavitary fluid accumulation, and extracellular matrix remodeling are central elements in a multistep process that leads to the formation and enlargement of cysts. MDCK cells suspended within medium-hydrated collagen gels grow to form spherical, monolayered, fluid-filled cysts that enlarge steadily. Vasopressin and other agents that increase intracellular levels of cAMP stimulate the rate of MDCK cyst growth and net fluid/solute secretion when added to defined medium in vitro. In this model, net fluid secretion is the only means by which fluid can accumulate within the cyst cavity. We used this cyst-forming line of epithelial cells to evaluate several membrane transport properties that are important in the coupled movements of solute and water in the process of secretory fluid transport. Individual cysts were microdissected from collagen gels, held by a micropipet in a thermostated chamber, and examined at a high magnification by video microscopy. Transepithelial water flow was initiated by rapidly exchanging the bath medium with hyperosmotic solutions. Net water flux, Jv, determined from the initial rate of decrease in cyst diameter, was proportionate to the transmembrane osmotic gradient of NaCl or raffinose; the reflection coefficient for NaCl was indistinguishable from 1.0. Osmotic water permeability (cm3/cm2/osm/min x 10(-6)) was 739 +/- 99 (N = 11) in medium augmented by an NaCl concentration of 100 mosmol/kg. Hydraulic conductivity (Pt), estimated in control cysts, was 6.8 +/- 0.9 microns/s, a value similar to that of medullary and cortical thick ascending limbs of Henle.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

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.

Actins↗
PubMed1989

Chemical modification of cell proliferation and fluid secretion in renal cysts.

We used an in vitro model, MDCK cyst, to determine the extent to which pharmacologic compounds known to inhibit plasma membrane solute transport mechanisms could alter the enlargement of renal epithelial cysts. Solitary MDCK cells cultured within collagen gel undergo clonal growth to form true epithelial cysts in which a single layer of polarized cells (apex toward lumen) encloses a fluid-filled cavity. Repeated observations by light microscopy were used to quantitate the rate of cyst growth in diameter, and demonstrated that cyst enlargement involved an increase in cell number (proliferation) and a net increase in intracystic volume (fluid secretion). Intracyst pressure was greater than the interstitium (6.7 mm H2O +/- 3.1 SD), indicating that fluid entry was secondary to net solute accumulation. Amiloride and seven amiloride analogs that inhibited to different degrees conductive Na+ transport, Na+-dependent H+ transport and Na+-dependent Ca++ transport reversibly decreased the rate of cyst enlargement. The effectiveness of these agents to retard cyst enlargement correlated with their relative potencies to inhibit Na+-dependent Ca++ transport. Morphologic examination indicated that amiloride and amiloride analogs decreased cell proliferation and fluid secretion to the same degree. Ouabain and vanadate (Na+K,ATPase inhibitors), and L-645,695 (Na+-dependent Cl-/HCO3- inhibitor) potently slowed cyst expansion. In contrast to amiloride and amiloride analogs, these agents caused an unusual degree of cellular stratification within the cyst walls, a finding consistent with the notion that fluid secretion was inhibited to a greater extent then cellular proliferation. We conclude that chemical inhibitors of primary and secondary active solute transport can diminish or halt the enlargement of epithelial cysts in vitro by decreasing the rate of cellular proliferation and/or net fluid secretion.

Amiloride↗
PubMed1989

Renal epithelial cyst formation and enlargement in vitro: dependence on cAMP.

Cysts, a common abnormality of kidneys, are collections of urine-like fluid enclosed by a continuous layer of epithelial cells. Renal cysts derive from nephrons and collecting ducts and progressively enlarge as a consequence of epithelial proliferation and transepithelial fluid secretion. The initiation of cyst formation and the factors that control cyst enlargement are unknown. We used an in vitro model of renal cysts to explore the role of the cAMP signal transduction system in the formation and expansion of cysts. MDCK cells, cultured in hydrated-collagen gel, produced polarized monolayered epithelial cysts when intracellular cAMP was increased by prostaglandin E1, arginine vasopressin, cholera toxin, forskolin, or 8-bromoadenosine 3',5'-cyclic monophosphate. All agonists were potentiated by 3-isobutyl-1-methylxanthine, a nucleotide phosphodiesterase inhibitor. The cell proliferation component of cyst enlargement was accelerated by cAMP agonists, as shown by the increased growth of MDCK cells in subconfluent monolayers. The fluid secretion component, reflected by the transepithelial movement of fluid across polarized monolayers of MDCK cells grown on permeable supports, was stimulated by cAMP agonists in the basolateral medium. Chloride levels were higher in the cyst fluid and the secreted fluid than in the bathing medium. We conclude that the development of MDCK cysts is dependent on cAMP. This signal transduction system may be an important modulator of epithelial cell proliferation and transepithelial fluid secretion in the kidney.

1-Methyl-3-isobutylxanthine↗
PubMed1989

Renal epithelial fluid secretion and cyst growth: the role of cyclic AMP.

Transepithelial fluid secretion has been postulated to account for the accumulation of fluid within hereditary and acquired renal cysts, but no such mechanism has been demonstrated in human kidney epithelium. It is shown here that transepithelial fluid secretion was stimulated by prostaglandin E1 (PGE1), forskolin, 8-Br-cyclic AMP, and 1-methyl-3-isobutylxanthine in polarized monolayers of established renal cell lines (MDCK and rat glomerular epithelial cells) and in monolayer cultures derived from the cyst walls of human autosomal dominant polycystic kidney disease and from epithelial cells of normal human renal cortex. Treatment with cyclic AMP agonists caused the same cells, when dispersed within a gel matrix of type I collagen (Vitrogen), to proliferate and form spherical fluid-filled monolayered cysts. Our findings suggest that increased intracellular cyclic AMP levels may have a critical role in the formation and expansion of hereditary and acquired renal cysts.

1-Methyl-3-isobutylxanthine↗
PubMed1989

Volume regulation determinants in isolated proximal tubules in hypertonic medium.

Isolated nonperfused proximal tubules maintained their cell volume at a constant level [isovolumetric regulation (IVR)] when extracellular osmolality was gradually increased above 295 mosmol at 1.5 mosmol/min. Cell volume remained constant between 295 and 361 +/- 7 mosmol in medium containing HCO3- -CO2 as the major buffer. In N-2-hydroxyethyl piperazine-N'-2-ethanesulfonic acid-buffered medium, IVR maintained cell size constant between 295 and 370 +/- 11 mosmol; thus, the volume regulatory process did not require the presence of external HCO3- -CO2. Omission of medium phosphate and sulfate from defined bathing medium had no effect on the volume regulatory process, whereas acetate (5 mM) was required to maintain cell volume constant. Hypertonic IVR was inhibited by acetazolamide (10(-6) M), 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (2.5 x 10(-5) M), and, in medium in which choline replaced all but 25 mM of Na+, amiloride (1 mM). These results suggest the involvement of carbonic anhydrase, Cl-, and HCO3- transport, and Na-H exchange in IVR; and provide indirect support for the view that IVR in hyperosmotic medium depends on the intracellular accumulation of electrolytes resulting from altered rates of membrane ionic exchange mechanisms.

Animals↗
PubMed1989

Effect of amiloride on the compensatory renal growth that follows uninephrectomy in mice.

The increase in contralateral kidney mass that follows the removal of one kidney is caused primarily by the hypertrophy and, to a lesser extent, the hyperplasia of tubule cells. Amiloride inhibits the proliferation of cells in tissue culture environments and diminishes compensatory hyperplasia of liver cells after partial hepatectomy. In the current study, we determined the extent to which amiloride diminished the compensatory increase in contralateral kidney weight and protein content after uninephrectomy. Caworth Farms mice, 3 to 4 months of age, received amiloride by daily intraperitoneal injection for 7 days before the left kidney was removed and for an additional 4 days after nephrectomy. Four days after uninephrectomy, compensatory hypertrophy was quantified by the increase in weight and protein content of the right kidney in comparison with the left (R - L divided by L x 100; [weight] +24.7% +/- 2.9% and [protein] +20.6% +/- 4.4%). Treatment with amiloride reversibly reduced the compensatory increase in kidney weight and protein content (dose required to achieve 50% inhibition, 11.4 and 2.4 mg/kg/day, respectively). Hexamethylene amiloride, an analog with weak affinity for conductive Na+ channels and high affinity for Na+-H+ exchangers, had no effect on compensatory hypertrophy. Spironolactone, a natriuretic and antikaliuretic agent of potency similar to that of amiloride, but with a different cellular mechanism of action, had no effect on compensatory hypertrophy. We conclude that amiloride reversibly blunts the compensatory renal hypertrophy after uninephrectomy in mice by mechanisms that appear to be separate from the drug's effects to block Na+-H+ exchange and transepithelial Na+ and K+ transport.

Amiloride↗