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Biomedical subjects

F A Carone

Publications and source records attributed to F A Carone.

At least 55 records · Page 3Linked to original sources
PubMed1988

Decreased de novo synthesis of proteoglycans in drug-induced renal cystic disease.

Cellular and extracellular (tubular basement membrane, TBM) alterations in the proteoglycans (PGs) of the rat renal tubules in diphenylthiazole-induced cystic disease were investigated. The PGs of normal and cystic kidneys were labeled with [35S]sulfate in an organ-perfusion system. Extracted cellular and TBM PGs were characterized by Sepharose CL-6B chromatography before or after treatment with heparitinase (degrades heparan sulfate) or chondroitinase ABC (degrades chondroitin sulfate). Total radioactivities in cellular, TBM, and medium fractions of cystic kidneys were reduced by factors of 9, 7, and 3, respectively. The PGs obtained from cystic and normal kidneys had similar profiles, namely, two peaks of radioactivity with Kav values of 0.26 (Mr = 130,000-150,000) and 0.40 (Mr = 50,000-55,000). The peaks had variable proportions of radioactivity for cellular and TBM fractions. Besides heparan sulfate, an additional 15-20% of chondroitin sulfate was synthesized in all three fractions obtained from cystic kidneys. The PGs synthesized by cystic kidneys had lower charge-density characteristics as compared to controls by DEAE-Sephacel chromatography. The medium fractions contained mostly glycosaminoglycan chains (Kav = 0.47, Mr = 24,000-26,000) of heparan sulfate. Autoradiograms of tissue samples revealed approximately 50% and approximately 60% decreases of grain densities over the cellular and TBM compartments, respectively. This decrease in de novo PG synthesis may have some relationship in the pathogenesis of polycystic kidney disease.

Animals↗
PubMed1988

Basement membrane antigens in renal polycystic disease.

Status of basement membrane antigens in renal polycystic disease was investigated. Antibodies directed against various components of basement membrane, including anti-heparan sulfate proteoglycan, Type IV collagen, laminin, and fibronectin, were employed. Their reactivities with basement membranes of normal and cystic segments of the renal tubules were ascertained by indirect immunofluorescence. The tissues were obtained either from kidneys of patients with adult (autosomal dominant) polycystic disease or from rats with renal cystic change induced by administration of 2-amino-4,5-diphenylthiazole HCl. The human and rat tissues that had undergone cystic change exhibited essentially similar results. A loss of reactivity to anti-heparan sulfate proteoglycan antibodies was observed. The reactivity toward anti-Type IV collagen and laminin either remained unchanged or was focally increased. The reactivity toward fibronectin, normally absent, increased dramatically in the peritubular regions and interstitium. The results indicate that there is an imbalance in various antigenic components associated with renal tubular cystic disease in rat and man, which may have a fundamental role in the pathogenesis of this disorder.

Animals↗
PubMed1988

Explant culture of human polycystic kidney.

Autosomal dominant polycystic kidney disease is characterized by the formation of large fluid-filled epithelial cysts. To obtain renal cyst wall epithelium for in vitro study, we employed an explant culture technique using medium-hydrated collagen gel as the culture substrate. Pieces of excised cyst wall were submerged within collagen gel. Cells of the cyst lining migrated to form a polarized epithelium at the surface of the surrounding collagen gel. Regions of the outgrowth were isolated by microdissection and used as a source of cells for subculture. Cyst-derived epithelium retained the ultrastructural features of the renal cyst of origin. The cells were cuboidal and bore short apical microvilli. Cells often were joined by apical tight junctions. Intercellular channels were narrow and bordered by short microvillus projections at the basolateral membrane. Epithelial cells rested on a densely staining basal lamina. The explants commonly developed small solitary cysts within their wall that were filled with fluid. These mural cysts were lined by a simple epithelium morphologically similar to the cells that lined the explant. Explantation of human renal cyst wall to culture within collagen gel provides a reproducible method to isolate autosomal dominant polycystic kidney disease epithelium for subculture. This method offers an alternative to the use of proteolytic enzymes to establish morphologically stable cultures of cyst lining cells for use in experimental renal cystic disease.

Cell Division↗
PubMed1987

Dosimetry study of Nd:YAG laser damage to canine renal cortex.

A dosimetry study of factors affecting the extent of tissue damage inflicted upon the canine renal cortex by the Neodymium:Yttrium Aluminum Garnet (Nd:YAG) laser was undertaken. Laser parameters and renal tissue conditions were varied independently in duplicate in the following manner: (1) power - 5, 10, 20, 50, 75, 100 watts with a spot size of 1.2 mm; (2) exposure duration - 1, 2, and 4 seconds; (3) kidney perfusion and temperature--renal artery unclamped (perfused) without cooling; renal artery clamped (non-perfused) without cooling; and renal artery clamped with cooling. Five days following application of the laser, the animals were sacrificed and serial sections of the renal cortex were examined for maximum depth and width of tissue damage and ablation. Multiple linear regression analysis of the data indicated a direct linear relationship between the joules (watts X seconds) of energy delivered to the renal cortex and the depth and width of tissue damage and ablation (p less than 0.001 for joule regression coefficient for each variable). Seconds and/or watts alone were not major predictors of the outcome after accounting for joules. Clamping the main renal artery significantly reduced the depth and width of laser damage when compared to the perfused kidney (p less than 0.001 for each variable). The depth of damage was similar in the cooled and the non-cooled non-perfused kidney. These data suggest that increased laser energy and kidney perfusion significantly increase renal cortical laser induced damage. Adjustment of these parameters may permit controlled tumor ablation or tissue incision with minimal damage to adjacent normal tissue.

Animals↗
PubMed1987

Viscoelastic properties of tubule basement membranes in experimental renal cystic disease.

Hereditary and acquired renal cysts develop from tubule segments that enlarge progressively. We measured the deformability of basement membranes surrounding individual normal tubules and cysts to determine if cysts might develop by simple extension of abnormally-deformable basement membrane in response to normal or increased transtubule hydrostatic pressures. Deformability (cm/dyne) was measured in individual tubules and cysts in vitro by a micropipet aspiration technique that related negative pressures within the pipet to the distance the tubule or cyst wall was aspirated into the pipet. Viscoelastic creep was determined from the time-dependent effect of pipet aspiration on membrane deformation. Proximal and collecting tubules, glomerular capsules and cysts were microdissected from controls and animals with acquired (Diphenylthiazole [rats]. Nordihydroguaiaretic acid [rats]), hereditary (C57 BL/6J [cpk/cpk] mice) and spontaneous (CFWw mice) renal cystic diseases. The major resistance to deformation was localized to the basement membrane since collagenase destroyed the elasticity of tubule and cyst walls. Tubule basement membranes adjacent to cysts appeared abnormal by electron microscopy in the animals fed DPT, but measurements of deformability and viscoelastic creep showed no differences between normal and cystic tubules in any animal model. Deformability values of cysts (7.7 +/- 1.1, 10.9 +/- 1.1, 11.2 +/- 0.6, 9.4 +/- 0.8 X 10(-3) cm/dyne in DPT, NDGA, C57 BL/6J and CFWw, respectively) are consistent with the interpretation that high transtubule pressures ranging from 39 to 134 cm H2O would be required if cysts form by simple stretching of the basement membrane secondary to a transepithelial hydrostatic pressure-gradient. Since in vivo measurements of hydrostatic pressures across cyst walls are not high enough we conclude that cysts do not enlarge due to increased deformability of tubule basement membranes.

Animals↗
PubMed1987

Effects of D-amino acid substituents on degradation of LHRH analogues by proximal tubule.

Less than Glu-His-Trp-Ser-Tyr-Gly-Leu-Arg-Pro-Gly-NH2, the luteinizing hormone-releasing hormone, LHRH, is degraded in renal proximal tubules (PT) in vivo (rat) and in vitro (rabbit) to less than Glu-His (2), less than Glu-His-Trp (3), and less than Glu-His-Trp-Ser (4). LHRH may be cleaved by endopeptidases simultaneously at multiple bonds, or initially at Ser4-Tyr5 followed by carboxypeptidase hydrolysis of 4 to 3 and then 2. To distinguish between these mechanisms, [3H]LHRH analogues were incubated with rabbit renal brush-border membranes (BBM), microinfused into PT in vivo or in vitro, and products were analyzed by HPLC. [D-Ser4]LHRH was not cleaved at D Ser4-Tyr5 but yielded less than Glu-His-Trp-D-Ser-Tyr-Gly as the major metabolite plus 2 and 3. [D-Trp6]LHRH was cleaved by BBM and PT to 2 and 3, but not to 4. [D-Ser4, D-Trp6]LHRH was not cleaved by BBM, but was degraded to 2 by PT in vivo. Thus, D-amino acid substituents altered the expected cleavage pattern of these analogues. [3H]LHRH was cleaved by BBM or by endopeptidase-24.11 from porcine PT to metabolites 2, 4, small amounts of 3, and less than Glu-His-Trp-Ser-Tyr-Gly, but cleavage was strongly inhibited by the specific inhibitor phosphoramidon. Thus, normally LHRH may be cleaved in PT by endopeptidase-24.11 to 2 and 4, and by angiotensin I-converting enzyme to 3, its known cleavage site.

Amino Acid Sequence↗
PubMed1987

Differences between in vitro and in vivo degradation of LHRH by rat brain and other organs.

Homogenates of brain, pituitary, liver, lung, ovary, and testes were incubated with [pyro Glu1-3,4-3H]luteinizing hormone-releasing hormone ([3H]LHRH), and the profiles of metabolites generated as a function of time were determined. After 5 min of incubation, 5 was the predominant metabolite in most homogenates. Although the profiles of metabolites varied at different time intervals, metabolites 2, 3, 4, and 5, and in some instances 7 and 9, appeared to form simultaneously and were detectable at 10 min. Neither metabolite 6 nor other larger metabolites formed initially as dominant degradation products. The findings suggest cleavage of LHRH by the simultaneous action of several endopeptidases. After a single vascular transit of [3H]LHRH, metabolites were determined in the venous blood of liver, lung, and brain of rats in vivo. There were no metabolites of [3H]LHRH in venous blood of liver and lung; however, metabolites 2-4 were present in venous blood of the brain. Incubation of rat anterior pituitary cells with [3H]LHRH yielded metabolites 1-4 but not metabolites 5 or 9 as in homogenates. Incubation of [3H]LHRH with porcine follicular granulosa cells resulted in the generation of metabolites 2-7 and 9, similar to the profile in homogenates. Thus, since homogenates contain enzymes of disrupted cells, they do not always reflect mechanisms for in vivo hydrolysis of circulating LHRH. Brain degraded 12.1% of LHRH during a single vascular transit and may account for substantial degradation of the circulating hormone.

Animals↗
PubMed1987

Renal folate absorption and the kidney folate binding protein. II. Microinfusion studies.

Surface proximal convoluted tubules (PCT) in rats were microinfused in situ with [3H]folic acid to study the role of folate binding protein (FBP) in the kidney brush-border membrane for renal conservation and transport of folate [3H]folic acid absorption was linearly related to tubular length of PCT and occurred largely in this segment of the tubule. Unlabeled folate derivatives inhibited [3H]folic acid absorption, the extent of which was dependent on the type of unlabeled folate used and its concentration. At equivalent concentrations, inhibition was most effective with unlabeled folic acid, slightly lower than with 5-methyltetrahydrofolate and least effective with methotrexate. Comparisons between [3H]folic acid absorption before and after infusion of a saturating dose of unlabeled folic acid or repetitive injections of [3H]folic acid into the same tubular site revealed continuous and rapid regeneration of unsaturated folic acid uptake sites with an apparent half-life of 28.75 +/- 8.75 s. Determination of [3H] retained in the tubule at various periods after microinfusion of [3H]folic acid revealed slow cellular disappearance with an apparent half-life of 47.3 +/- 5.4 min. It is proposed that the brush-border FBP functions as a receptor of infused folic acid and that following the binding of the ligand the folic acid/FBP complex undergoes a rapid change that results in the internalization of folic acid and regeneration of unsaturated binding sites at the membrane surface. Internalized folic acid is slowly released into renal capillaries.

Animals↗
PubMed1987

Degradation and transport of AVP by proximal tubule.

High-performance liquid chromatography (HPLC) analysis revealed that [3,4,5-3H-Phe3,Arg8]vasopressin ([3H]AVP) was not degraded by isolated renal brush-border membranes or by a cortical lysosomal fraction in vitro; however, in the presence of 1 mM reduced glutathione, [3H]AVP was degraded by both preparations. Renal cortical homogenates in vitro and luminal peptidases of proximal tubule in vivo degraded [3H]AVP and in both instances yielded phenylalanine, hexapeptide AVP 1-6, heptapeptide AVP 1-7, octapeptide AVP 1-8, and two uncharacterized products X and Y. These data suggest that filtered AVP is reduced in the proximal tubule by a reduced glutathione-dependent transhydrogenase and subsequently cleaved to [3H]Phe by tubular aminopeptidases. Following microinfusion of [3H]AVP into proximal tubules, 15.7% of the label was absorbed. Five and fifteen minutes after infusion of [3H]AVP, sequestration of total label in proximal tubules was 4.5 and 2.1%, respectively, and quantitative electron microscope autoradiography revealed accumulation of grains over apical endocytic vacuoles and lysosomes consistent with endocytic uptake and rapid lysosomal degradation of AVP and/or a large metabolite. Thus, enzymatic cleavage of AVP by luminal and lysosomal peptidases in proximal tubules could involve disulfide bond, C-terminal, and N-terminal loci.

Animals↗
PubMed1987

Diphenylthiazole-induced changes in renal ultrastructure and enzymology: toxicologic mechanisms in polycystic kidney disease?

The mechanism by which various chemicals induce renal cystic disease is unknown. To examine the early events in cystogenesis the ultrastructure and biochemistry of liver and kidney were analyzed after the administration of a chemical that induces renal cyst formation. Special emphasis was placed on examining potential mechanisms that would account for the observed loss of extracellular proteoglycans. Renal cystic disease was chemically induced in rats by feeding 2-amino-4,5-diphenylthiazole (DPT) for up to 4 weeks. After 4 days of feeding, DPT had induced a 4-fold increase in total urine output relative to diet-restricted control groups. Both groups maintained, but did not gain, weight during the feeding schedule. Cyst formation was localized to the medullary collecting tubules. Relative to diet-restricted controls, rats fed DPT exhibited diminished renal and hepatic catalase activity, but elevated activity for UDP-glucuronosyltransferase. Medulla showed an increase in the specific activities of the enzymes galactosyltransferase and sulfatase B. These enzymological findings correlated with ultrastructural observations of a loss of peroxisomes, proliferation of endoplasmic reticulum and enlargement of the golgi apparatus. Serum and urinary levels of inorganic sulfate were significantly increased in DPT-fed rats relative to controls. Tissue levels of UDP-glucuronic acid and adenosine 3'-phosphate 5'-phosphosulfate were not depressed by DPT feeding. Thus, DPT-induced cyst formation and loss of staining for glycosaminoglycans does not involve gross depletions of UDP-glucuronic acid and adenosine 3'-phosphate 5'-phosphosulfate, mutual cosubstrates for Phase II drug conjugation reactions and glycosaminoglycan synthesis.

Animals↗
PubMed1985

Tubular basement membrane changes in 2-amino-4,5-diphenylthiazole-induced polycystic disease.

Polycystic renal disease was induced in rats by feeding 2-amino-4,5-diphenylthiazole. Tubular (TBM) and glomerular basement membranes (GBM) were purified and analyzed for possible structural changes that may be a factor in the development of the tubular dilations and cysts. Changes in the relative quantities of TBM polypeptides were detected by sodium dodecylsulfate polyacrylamide gel electrophoresis. An overall increase in the concentration of high molecular weight components and a decrease in concentration of those of low molecular weight components were observed. Changes which were particularly notable included a twofold increase in a component of Mr = 380,000 and a decrease in one of Mr = 55,000 as analyzed without reduction of disulfide bonds. With reduction of disulfide bonds, the Mr = 380,000 component dissociates, whereas the Mr = 55,000 polypeptide does not, and polypeptides of Mr = 245,000 and 145,000 are observed to does not, and polypeptides of Mr = 245,000 and 145,000 are observed to increase about twofold in concentration (approximate molecular weights were determined using globular protein standards). These changes take place most rapidly from 4 to 8 weeks of drug administration and remain relatively constant between 8 and 16 weeks. If feeding of the drug is discontinued, the distribution of TBM polypeptides returns to normal. These results indicate that tubular basement membrane from animals with 2-amino-4,5-diphenylthiazole-induced polycystic renal disease is abnormal, and this should be considered as a possible contributing factor in the formation of cysts.

Amino Acids↗
PubMed1985

Urinary loss of glucose, phosphate, and protein by diffusion into proximal straight tubules injured by D-serine and maleic acid.

In several models of acute renal failure leakage of glomerular filtrate out of the tubule is an important pathogenetic mechanism; however, bidirectional diffusion of solute to account for certain pathophysiologic features of acute renal failure has received meager attention. Using micropuncture and clearance methods, we assessed sequentially leakage of solutes and inulin across proximal straight tubules (PST) injured by two nephrotoxins. In d-serine-treated rats with extensive necrosis of PST, the basis for glucosuria and tubular leakage of inulin was studied. Glucose absorption by the proximal convoluted tubule and glucose delivery to the PST were normal, but glucose delivery to the distal tubule was increased nearly 8-fold, indicating diffusion of glucose from interstitial to tubular luminal fluid across the necrotic PST. Total kidney inulin clearance was greatly reduced, but single nephron glomerular filtration rate, based on proximal convoluted tubule samples, was normal, indicating tubular loss of inulin. Urinary recovery of [14C]inulin infused into tubular lumina revealed that proximal convoluted tubule and distal tubule were impermeable to inulin and that inulin diffused out of the necrotic PST. The progressive return over 6 days of tubular impermeability for inulin correlated with relining of PST with new cells. In maleic acid-treated rats the site and extent of tubular necrosis and the nature of urinary loss of solutes were studied. Microdissection revealed that maleic acid caused limited necrosis of PST which averaged 7.4% of total proximal tubular length. Increased urinary excretion of protein, phosphate, and glucose and increased tubular permeability to microinfused [14C]inulin occurred with the onset of PST necrosis, and return of these abnormalities to normal correlated with the degree of cellular repair of the PST. Together these data indicate that with severe cellular injury there is bidirectional, concentration-dependent diffusion of solutes between the interstitial and tubular fluids. Thus, experimental or human toxic nephropathies should not be classified as acquired Fanconi's syndrome unless specific tubular transport defects are present.

Acute Kidney Injury↗
PubMed1984

Efficacy of polyglycolic acid (PGA) tubing stents in ureteroureterostomies.

Polyglycolic acid (PGA) tube stented, silastic stented and non-stented anastomoses were compared in 18 ureteroureterostomies performed in 13 dogs. Urinalysis, urine culture, BUN and serum creatinine and intravenous pyelograms were done periodically from 1 day to 6 months postoperatively and ureteral dynamics were performed at 6 months. Normal intravenous pyelograms were observed following 10 of 11 PGA stented, 3 of 4 silastic stented and 2 of 3 unstented anastomoses. PGA stents prevented extravasation and dissolved within 7 days without stone formation. Urodynamic and histopathologic studies revealed no significant differences among the 3 groups. The data suggest that PGA stents are as effective as silastic stents and do not require additional procedures for removal.

Animals↗
PubMed1984

Kallikrein-kinin and renin-angiotensin systems in rat renal lymph.

Rat renal lymph contains 254 +/- 17 ng/ml (means +/- SEM, N = 20) of immunoreactive glandular kallikrein. Like the immunoreactive glandular kallikrein in plasma, it is biologically inactive. Gel filtration of renal lymph reveals profiles for immunoreactive glandular kallikrein, protein, and inhibition of trypsin and kallikrein which resemble those seen for plasma except that high molecular weight plasma components are reduced or missing in renal lymph. In contrast, gel filtration of thoracic lymph reveals immunoreactive glandular kallikrein and protein profiles which are indistinguishable from those seen with plasma. Renin levels are 170-fold higher in renal lymph than in thoracic lymph while angiotensin-converting enzyme levels are only 16% those of thoracic lymph. In keeping with the high renin and low converting enzyme activities, renal lymph contains high levels of angiotensin I. Immunoreactive glandular kallikrein levels in renal lymph, thoracic lymph and plasma do not show the striking differences observed for renin.

Angiotensin I↗
PubMed1984

Reversible changes of tubular cell and basement membrane in drug-induced renal cystic disease.

To delineate changes in the development and regression of renal cystic disease, sequential studies were done in rats fed 2-amino-4,5-diphenyl thiazole (DPT). Kidneys were perfusion fixed with either aldehyde fixative alone or with the addition of ruthenium red (RR). Tissues were processed for light and electron microscopy. Light microscopy sections were stained with alcian blue and eosin and hematoxylin. Initially, cellular proliferation and, later, cystic transformation of collecting tubules were observed. The structural changes in tubular cells preceded alterations in the basement membrane and consisted of an increase in smooth and rough endoplasmic reticulum and free polyribosomes, prominence of Golgi complexes, and an increased number of lysosomes. These findings are suggestive of changes in the biosynthetic, secretory, and degradative mechanisms of the cell. With time, the tubular basement membranes became progressively thicker and laminated with concomitant loss of alcian-blue- and RR-staining. When DPT-treated animals with renal cystic disease were placed on a normal diet, tubular cell and basement membrane morphology and alcian-blue- and RR-staining returned to normal and cystic changes regressed. These findings are compatible with altered synthesis and degradation of tubular basement membranes in this model of cystic disease.

Animals↗