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P Ronco

Publications and source records attributed to P Ronco.

At least 55 records · Page 3Linked to original sources
PubMed1995

Segmental distribution of the endocytosis receptor gp330 in renal proximal tubules.

The subcellular distribution and segmental variations in location of gp330, a scavenger receptor for filtered proteins in renal proximal tubules, was analyzed. Kidney tissue from rats (4 different strains), rabbits and humans were analyzed by light- and electron microscope immunocytochemistry, using cryosections or Lowicryl sections from cryosubstituted tissue. Gp330 was located mainly in apical coated pits, small and large endocytic vacuoles and in dense apical tubules in the proximal tubule cells. The labeling density was markedly higher in segments 1 and 2 as compared to segment 3 of the proximal tubule. In addition to the location in the early part of the endocytic pathway, gp330 was also present in lysosomes, especially in segments 1 and 2. The lysosomal labeling was not restricted to the membrane, but was also seen in the matrix. Localization of gp330 in lysosomes was confirmed on sections from purified lysosomal fractions from rat renal cortex. The brush border localization of gp330 in proximal tubules exhibited a characteristic segmental variation. In the initial part of segment 1, there was virtually no brush border labeling. In the remaining part of segment 1 and in segment 2, there was a distinct but sometimes patchy labeling of the brush border. In segment 3, groups of microvilli of approximately 10 as seen in sections were intensively labeled from bottom to tip and there were often more than one of these groups on a single cell, the remaining microvilli were unlabeled. No differences in the cellular and subcellular localization of gp330 were observed between species or rat strains. In conclusion, the present study demonstrates that in addition to its location in the early endocytic and recycling pathway, gp330 is also present in microvilli and the protein and degradation products thereof is present in lysosomes, consistent with its role as a protein scavenger receptor.

Animals↗
PubMed1994

Adenoviral-mediated gene transfer to renal tubular cells in vivo.

The efficient introduction of genetic material into quiescent renal cells is potentially important in the study of renal physiopathology and for gene therapy of kidney related disorders. A replication-deficient adenoviral vector that contained a reporter gene encoding the nuclear beta-galactosidase was either selectively perfused into the renal artery or infused through a retrograde catheter into the pyelic cavity of the left kidney of adult rats. Highly efficient gene transfer was achieved by either route of administration, and nuclear beta-galactosidase activity was detected for two to four weeks following a progressive decline of expression. Genetically-modified cells were identified as proximal tubular cells when the adenoviral vector was selectively perfused via the renal artery, while tubular cells from the papilla and medulla were selectively transduced by retrograde infusion of the viral vector. No obvious cytopathic effect was observed. We conclude that: (i) efficient gene transfer in renal tubular cells can be achieved by adenoviral vectors; (ii) the targeted cell population can be chosen through the route of administration.

Adenoviruses, Human↗
PubMed1994

Autoimmune haemolytic anaemia after ABO-match, ABDR full match kidney transplantation.

We report a case of autoimmune haemolysis after an ABO- and ABDR-identical kidney transplantation which leads to the discussion of the role of cyclosporin A (CsA). A 46-year-old woman with end-stage renal disease and no history of auto-immune disease received an ABO- and ABDR-identical first renal allograft from a cadaver donor. On day 16, while on a heavy sequential immunosuppressive regimen including anti-thymoglobulins, azathioprine (Aza), prednisolone (Pred) and CsA, she developed an autoimmune haemolysis with positive Coombs test, IgM+C type. Elution of antierythrocyte antibodies did not enable us to identify any specificity. Haemolysis lasted 45 days before haemoglobin slowly increased after CsA had been greatly reduced. Direct antiglobulin tests remained positive 5 months after transplantation and became negative the following month. Eight months after the transplantation the patient had a normal haemoglobin level and normal renal function. Although the typing of autoantibodies was not possible, our data suggest that this patient's haemolysis may be related to the clonal development of donor B lymphocytes in the recipient, favoured by an HLA A-B-DR identity and post-transplant CsA therapy, as exceptionally reported in the literature.

ABO Blood-Group System↗
PubMed1994

Antibodies to the 280-kd coated pit protein, target of teratogenic antibodies, produce alterations in the traffic of internalized proteins.

Previous studies have identified two high-molecular weight (280 and 330 kd) glycoproteins expressed by coated pits of the proximal renal tubule and yolk sac and have further established that, in vivo, antibodies to gp280 but not to gp330 induce fetal malformations. In the present study, we report the effect of these antibodies on the endocytic process by yolk sac visceral epithelial cells of rat embryos explanted at day 10 of gestation. Antibodies to gp280 markedly altered development of the yolk sac and embryo, induced malformations, inhibited by 40% the uptake of [14C] sucrose and perturbed the intracellular traffic of internalized proteins. Under control conditions, rat immunoglobulin G present in the culture medium was immunolocalized in lysosomes of epithelial cells, whereas in the presence of antibody, it was detected in small vesicles scattered through the apical cytoplasm. Alterations of the endocytic pathway were confirmed by experiments analyzing the uptake of peroxidase added to the medium for 2 to 60 minutes. The initial compartments of endocytosis visualized by peroxidase were increased in size and abnormal in shape and the transfer of the internalized peroxidase to the lysosomal compartment was delayed. In contrast, antibodies to gp330 had a minimal effect on embryonic development and did not induce fetal malformations. Endocytosis was only modestly altered; uptake of [14C] sucrose was decreased by 25%, and only minor modifications of the intracellular transit of peroxidase could be detected. We suggest that the key role of anti-gp280 antibodies is via trapping of the target antigen in the early endocytic compartment thus preventing its normal function in lysosomal transfer.

Animals↗
PubMed1993

[Tubulo-interstitial renal complications of myeloma].

Tubulo-interstitial lesions are the main causes of renal failure in patients with myeloma, and they sometimes reveal this tumour. They are mainly found in large tumoral mass myelomas and are characterized by tubular lesions and specific casts of unexplained physiopathology. Symptomatic and aetiological treatments must be instituted at an early stage to obtain the recovery, at least partial, of renal function, as observed in more than 50% of the cases. The persistence of renal failure carries a sombre prognosis, which emphasizes the importance of a preventive treatment that is too often omitted. Functional tubular disorders are frequent but seldom result in Fanconi's syndrome. A better understanding of the mechanisms underlying the nephrotoxicity of monoclonal immunoglobulins should permit to reduce the incidence of tubular lesions in myelomas.

Fanconi Syndrome↗
PubMed1993

K+ fluxes mediated by Na(+)-K(+)-Cl- cotransport and Na(+)-K(+)-ATPase pumps in renal tubule cell lines transformed by wild-type and temperature-sensitive strains of Simian virus 40.

The relative contributions of Na(+)-K(+)-ATPase pumps and Na(+)-K(+)-Cl- cotransport to total rubidium (Rb+) influx into primary cultures of renal tubule cells (PC.RC) and cells transformed either with the wild-type or a temperature-sensitive mutant of the simian virus 40 (SV40), were measured under various growth conditions. The Na(+)-K(+)-ATPase-mediated component represented 74% and 44-48% of total Rb+ influx into PC.RC and SV40-transformed cells, respectively. Proliferating transformed cells showed substantial ouabain-resistant bumetanide-sensitive (Or-Bs) Rb+ influx (41-45% of total) which indicated the presence of a Na(+)-K(+)-Cl- cotransport. The Or-Bs component of Rb+ influx was greatly reduced when temperature-sensitive transformed renal cells (RC.SVtsA58) grown in Petri dishes or on permeable filters were shifted from the permissive (33 degrees C) to the restrictive temperature (39.5 degrees C) to arrest cell growth. The ouabain-sensitive Rb+ influx mediated by the Na(+)-K(+)-ATPase, the total and amiloride-sensitive Na+ uptakes were not modified following inhibition of cell proliferation. A similar fall in the Or-Bs influx was obtained when renal tubule cells transformed by the wild-type SV40 (RC.SV) were incubated with the K+ channel blocker, tetraethylammonium (TEA) ion, which we had previously shown to arrest cell growth without affecting cell viability (Teulon et al.: J. Cell. Physiol., 151:113-125, 1992). Reinitiation of cell growth by removal of TEA or return to 33 degrees C of the temperature-sensitive cells restored the Or-Bs component of Rb influx. Taken together, these results indicate that the Na(+)-K(+)-Cl- cotransport activity is critically dependent on cell growth conditions.

Animals↗
PubMed1993

Mineralocorticoid receptors in SV40-transformed tubule cell lines derived from rabbit kidney.

The presence of mineralocorticoid (MR) and glucocorticoid (GR) receptors was investigated in two renal tubular cell lines, derived from primary cultures of isolated rabbit kidney cortical cells infected with the wild-type SV40 virus, which exhibit thick ascending limb (RC.SV2) and collecting tubule (RC.SV3) phenotypes (Vandewalle et al. J. Cell. Physiol. 141, 1989, 203-221). MR and GR were quantified, in cell monolayers and cell cytosolic fractions, with [3H]aldosterone, [3H]dexamethasone and [3H]RU486, an antiglucocorticoid with no affinity for MR. Cytosolic receptors from RC.SV2 and RC.SV3 cells labeled with [3H]aldosterone, [3H]dexamethasone or [3H]RU486 sedimented at approximately 8 S in a 15-40% glycerol gradient. All steroids displaced bound [3H]dexamethasone to the same extent, suggesting that dexamethasone bound to both MR and GR: under the conditions of assay, [3H]aldosterone binds exclusively to MR, and [3H]RU486 to GR. In both RC.SV2 and RC.SV3 cells, [3H]aldosterone bound to one class of high affinity sites (Kd 0.14-0.8 nM; Nmax 8 to 22 fmol/mg protein). In both cell lines, the number of high affinity binding sites for [3H]dexamethasone ranged from 9 to 18 fmol/mg protein with an affinity of 0.5-1.3 nM. Compared to renal cortex, the most striking observation was a marked decrease in [3H]dexamethasone binding in primary cultures and SV40-transformed cells. These results indicate that MR and GR are expressed in two established mammalian kidney tubular cell lines providing new models of cultured renal cells for studies on the physiological effects of corticosteroid hormones.

Aldosterone↗
PubMed1993

Characterization of neutral endopeptidase in vascular smooth muscle cells of rabbit renal cortex.

In addition to biological and clearance receptors for atrial natriuretic factor (ANF), cultured vascular smooth muscle cells (VSMC) from the rabbit renal cortex possess ectoenzymes degrading this hormone. We examined whether neutral endopeptidase (NEP) was implicated in this process. The presence of NEP in VSMC was demonstrated as follows. 1) NEP activity measured from the hydrolysis of a synthetic substrate by intact cells cultured in a medium containing 10% fetal calf serum was 1,609 +/- 65 pmol.min-1 x mg-1 [surface localization of the enzyme was confirmed by low activity (4% of total) in the cytosol; release of NEP activity in the medium was negligible]; 2) a monoclonal antibody directed against rabbit NEP specifically stained VSMC membranes; and 3) mRNA from VSMC hybridized a NEP cDNA probe with a single band as shown by Northern blot analysis. The role of NEP in ANF catabolism was demonstrated by incubating 125I-ANF or unlabeled ANF for increasing periods of time with VSMC in the presence of thiorphan (1-100 microM). Intact hormone estimated by trichloroacetic acid precipitation or radio-immunoassay, respectively, increased markedly compared with control in the presence of this specific inhibitor of NEP. NEP activity was stimulated (x1.6) in quiescent VSMC deprived from serum during 3 days. This effect was dose dependent and was not observed with creatine kinase activity measured as control. NEP expression at the cell surface estimated by sorting of immunostained cells was also increased in the absence of serum. Northern blot analysis showed increases in the mRNA band of NEP with increasing periods of serum deprivation.(ABSTRACT TRUNCATED AT 250 WORDS)

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1993

Epitope specificity of anti-gp330 autoantibodies determines the development of proteinuria in active Heymann nephritis.

In active Heymann nephritis, an experimental autoimmune disease in the rat, gp330 is regarded as the main antigenic target. Immunization with detergent-solubilized renal tubular epithelium (RTE-DOC) has been shown to be less nephritogenic than immunization with crude RTE. In this study immunization with either crude RTE or affinity-purified gp330 did, but immunization with RTE-DOC did not induce proteinuria. Both a possible aberrant subclass distribution of anti-gp330 autoantibodies and the involvement of additional nephritogenic autoantigens such as DPP IV (gp90) or laminin could be excluded. Circulating anti-gp330 autoantibody titers were significantly higher in RTE-DOC-immunized rats than in RTE-immunized animals. In contrast, significantly more antibodies were shown to bind in the glomeruli in the latter group. The time of onset of abnormal proteinuria was shown to be related to the recognition of a particular V8 protease-induced 250 kD fragment of gp330 in Western blots. This study shows that a particular fragment-specific subset of autoantibodies against gp330 is involved in the glomerular damage in Heymann nephritis.

Animals↗
PubMed1993

Interferon-gamma induces dipeptidylpeptidase IV expression in human glomerular epithelial cells.

Because dipeptidylpeptidase IV (DPP IV) is present in vivo on glomerular visceral epithelial cells and possesses immunogenic properties, as shown by the capability of anti-DPP IV antibody to induce the Heymann model of glomerulonephritis, we studied the expression and regulation of DPP IV in cultured human glomerular visceral epithelial cells. DPP IV is an ectoenzyme, as indicated by the rapid detection of the product of the reaction in the incubation medium of intact cells and the staining of paraformaldehyde-fixed cells in the presence of a specific anti-DPP IV antibody. DPP IV activity was inhibited by diisopropylfluorophosphate and phenylmethyl sulphonylfluoride. Its optimum pH was alkaline (7.7-8) and it exhibited a Km value of 0.94 mM. DPP IV expression was induced in cells treated by interferon-gamma (IFN-gamma). The effect was significant after a 3-day treatment with 100 U/ml. It increased with time, reaching a plateau after 11 days, and was dose-dependent with a maximum at a concentration of 1000 U/ml. Staining of the cells with anti-DPP IV antibody was also increased after a 6-day treatment with 100 U/ml IFN-gamma. It was shown by Northern analysis that, after 24 hr of exposure to 500 U/ml of IFN-gamma, DPP IV mRNA transcript was stimulated. Transcriptional activation by IFN-gamma did not require new protein synthesis. Interleukin-1 (IL-1) and cyclic AMP had a small stimulatory effect, whereas dexamethasone and phorbol esters were inefficient. These results suggest that DPP IV of glomerular epithelial cells may be up-regulated by IFN-gamma from activated T lymphocytes in glomerular diseases and during lymphocyte-mediated graft rejection.

Blotting, Northern↗
PubMed1993

["Idiopathic" extracapillary glomerulonephritides without immune deposits are vascularitides: clinical and serologic analysis].

To determine the spectrum of systemic diseases potentially associated with pauci-immune rapidly progressive glomerulonephritis (GN), most of which being considered as idiopathic, we have analyzed extra-renal manifestations, occurrence of extra-glomerular vasculitis and incidence and specificity of ANCAs in 40 patients selected only on histological criteria. Extra-renal symptoms were unexpectedly observed in all patients but one, and were suggestive of vasculitis in 24 of them. Extra-glomerular vasculitis was evidenced in 18 kidney biopsies and four biopsies from other organs. Among the 33 patients with suspected or established vasculitis, 13 had presumed or biopsy-proven Wegener's granulomatosis (WG), three had a macroscopic form of polyarteritis nodosa and 17 had a clinical presentation compatible with the so-called microscopic polyarteritis previously described in the british literature. An additional patient had clinical signs of WG without clinical and histological evidence of vasculitis. ANCAs were detected in 28/33 and 25/34 sera tested by IF and ELISA, respectively: 19 contained anti-myeloperoxidase (MPO) antibodies and 6 had anti-proteinase 3 (Pr3) activity. Anti-MPO and anti-Pr3 antibodies were present in all clinical subgroups but with various incidences: anti-MPO antibodies were surprisingly more often detected (6/12) than anti-Pr3 (4/12) in patients with suspected or histologically proven WG but anti-Pr3 antibodies were nonetheless 3- to 4-fold more frequent in WG than in non-WG systemic vasculitis (1/12) and necrotizing CGN without evidence of extra-renal vasculitis (1/10). These results strongly suggest that pauci-immune necrotizing CGNs belong to the broad spectrum of necrotizing vasculitides affecting glomerular capillaries. This hypothesis is also supported by the good response of patients to immunosuppressive treatments known for their efficacy in vasculitides, whereas these treatments are usually less successful in severe forms of extra-capillary GN with immune deposits.

Glomerulonephritis↗
PubMed1993

[Teratogenesis and autoimmunity: significance of an experimental murine model for human pathology].

Fetal malformations constitute a major problem of public health. Unfortunately the known causes do not account for more than 50% of the cases observed. The potential role of immune mechanisms is suggested by experimental studies in the rat indicating that antibodies reactive with the yolk sac induce fetal malformations. In this study we show that these antibodies are specific for a 280 kDa protein expressed only in the kidney and the yolk sac by cell structures associated with the formation of endocytic vesicles. We further show that a similar protein is expressed in man by the yolk sac, the kidney and the trophoblasts. The possible role in pathology of antibodies against the human protein is discussed.

Animals↗
PubMed1993

Comparative immunochemistry and ontogeny of two closely related coated pit proteins. The 280-kd target of teratogenic antibodies and the 330-kd target of nephritogenic antibodies.

We have previously shown that monoclonal antibodies specific for a 280-kd protein (gp280) concentrated within the coated pits of renal and yolk sac brush border-induced fetal malformations, whereas antibodies specific for gp330, another coated pit protein with a similar distribution, had no deleterious effect on embryonic development. In this study, we show that gp280 and gp330 are closely related proteins, as indicated by: 1) similarities in peptide maps obtained after cyanogen bromide cleavage, 2) immunological cross-reactivity related to a minor contingent of antibodies that do not have teratogenic activity, and 3) asynchronous but related expressions during ontogenesis. During the early stages of development, the expression of the two glycoproteins was limited to (gp330) or predominant in (gp280) the clathrin-coated pits and intermicrovillar areas. In the pre-implantation embryo, gp330 was expressed by trophectodermal cells, which became negative in day-6 embryos trapped in endometrial infoldings. At this stage, gp280 and gp330 were both simultaneously detectable at the apical pole of the first entoblastic cells and remained expressed by the brush border of visceral yolk sac epithelial cells until the end of pregnancy. In addition, gp330 was expressed by amniotic cells and neurectodermal structures. During nephrogenesis, in contrast, the expression of gp280 and gp330 by the intermicrovillar areas of the proximal tubule cell was the result of a complex maturation process. gp280 and gp330 were diffusely distributed in S-shaped bodies in the presumptive areas of the glomerulus, proximal tubule, and distal tubule (gp330). During development of the nephron, the pattern of expression became progressively restricted to the proximal tubule and glomerulus (gp330), and selective localization in the intermicrovillar areas was only achieved in filtrating nephrons.

Animals↗
PubMed1992

Expression of cALLa/NEP on gliomas: a possible marker of malignancy.

First described on pre-B leukemia cells, the common acute lymphoblastic leukemia antigen (cALLa) is also expressed on glioma cells in vitro. Its identity to neutral endopeptidase (NEP) (E.C.3.24.11) was corroborated by our finding that cALLa positive glioma cells had NEP activity. To study cALLa/NEP distribution on glial tumours in vivo, we examined 76 brain tumour biopsies by immunostaining techniques on frozen tissue sections using anti-cALLa (FAH99) and anti-NEP (135 A 3) monoclonal antibodies. We found that 96% of grade 4 gliomas (25/26) expressed NEP. Whereas only 45% (4/9) of grade 3 or anaplastic astrocytomas did. In low grade gliomas, we found 2 positive tumours out of 21 tested (10%). Double immunostaining procedures revealed that NEP was co-expressed with GFAP. However no NEP could be detected on non-glial brain tumours nor on reactive astrocytes. These results suggest that cALLa/NEP expression could be linked to malignant progression of gliomas.

Adolescent↗
PubMed1992

Cell surface aminopeptidase A and N activities in human glomerular epithelial cells.

Cell surface aminopeptidases N (APN) and A (APA) have been characterized on cultured human glomerular epithelial cells and a SV40-transformed cell line derived from them. APN had a wide substrate specificity whereas APA only attacked peptides with an acidic N terminal amino acid. Both enzymes also differed by their sensitivity to divalent cations and to aminopeptidase inhibitors. Phorbolmyristate acetate (PMA) stimulated APN but not APA expression after a lag time of 12 hours. An increase of twice the basal value was observed with 10 ng.ml-1 PMA. This effect was confirmed by immunofluorescence staining using a specific anti-APN monoclonal antibody. Both ecto- and total enzyme activities were stimulated by PMA. The effect of PMA was suppressed by H7, a PKC inhibitor, and cycloheximide, an inhibitor of protein synthesis. Thrombin (1 to 2.5 U.ml-1) and interferon (IFN)-gamma (100 U.ml-1) also stimulated APN activity, the latter after longer exposure of the cells. APA activity was increased by 8-bromo-cAMP and two cAMP-stimulating agents, forskolin and isobutylmethylxanthine (IBMX). A twofold increase above basal value was obtained with 100 microM forskolin after 72 hours of treatment. cAMP-stimulated APA activity was suppressed by cycloheximide. Dexamethasone also stimulated APA activity. The effects of forskolin and dexamethasone were additive. These results demonstrate that APN and APA in glomerular epithelial cells are under different regulations: mitogens and IFN-gamma for APN, cAMP and glucocorticoids for APA. This selective expression may imply possible functional consequences in glomerular diseases.

Aminopeptidases↗