F Delarue

Unité Inserm U1077, Caen, Lower Normandy, France

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Publications (49)

  • [Show abstract] [Hide abstract] ABSTRACT: In membranous nephropathy, the development of glomerular lesions is related to the formation of immune complexes at subepithelial sites. These deposits are associated with modifications in the fibrinolytic activity of glomerular cells leading to the appearance of fibrin degradation products in the deposits and the urine. A previous study has shown that immune complexes interact with glomerular epithelial cells (GEC) through the neonatal Fc receptor (FcRn). We therefore determined whether this binding could be responsible for a modification in the fibrinolytic activity of GEC. Endocytosis of heat-aggregated immunoglobulins (AgIgG) in cultured human GEC was studied by immunofluorescence and confocal microscopy. The release of urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor 1 (PAI) by GEC or whole glomeruli was assessed by ELISA, fibrin zymography and Northern blot. Human GEC in culture bound AgIgG that possess characteristics similar to soluble immune complexes and internalized them by 10 min. This process was mediated by FcRn since chicken aggregated IgG (AgIgY), that do not bind FcRn, did not colocalize with AgIgG in GEC. AgIgG but not AgIgY induced a decrease of FcRn expression at the membrane and within the cells. The binding of AgIgG to GEC elicited a dose- and time-dependent increase in the release of uPA activity, as in the uPA protein and mRNA expression without modification in the release of PAI. This process was not abrogated by agents inhibiting endocytosis and/or transcytosis such as cytochalasin B, suggesting an endocytosis-independent uPA regulation. GEC response to AgIgG overload comprises at least two sequential steps: (1) a FcRn-mediated endocytosis; (2) an endocytosis-independent fibrinolytic imbalance leading to plasmin generation which could favor in vivo AgIgG clearance and matrix remodeling.
    Article · Feb 2004 · Nephron Experimental Nephrology
  • C Burckle · F Delarue · JJ Saris · [...] · G Nguyen
    Conference Paper · Sep 2002
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    Genevieve Nguyen · Françoise Delarue · Céline Burcklé · [...] · Jean-Daniel Sraer
    [Show abstract] [Hide abstract] ABSTRACT: Renin is an aspartyl protease essential for the control of blood pressure and was long suspected to have cellular receptors. We report the expression cloning of the human renin receptor complementary DNA encoding a 350-amino acid protein with a single transmembrane domain and no homology with any known membrane protein. Transfected cells stably expressing the receptor showed renin- and prorenin-specific binding. The binding of renin induced a fourfold increase of the catalytic efficiency of angiotensinogen conversion to angiotensin I and induced an intracellular signal with phosphorylation of serine and tyrosine residues associated to an activation of MAP kinases ERK1 and ERK2. High levels of the receptor mRNA are detected in the heart, brain, placenta, and lower levels in the kidney and liver. By confocal microscopy the receptor is localized in the mesangium of glomeruli and in the subendothelium of coronary and kidney artery, associated to smooth muscle cells and colocalized with renin. The renin receptor is the first described for an aspartyl protease. This discovery emphasizes the role of the cell surface in angiotensin II generation and opens new perspectives on the tissue renin-angiotensin system and on renin effects independent of angiotensin II.
    Full-text Article · Jul 2002 · Journal of Clinical Investigation
  • [Show abstract] [Hide abstract] ABSTRACT: Chronic allograft nephropathy is the main cause of long-term kidney graft loss. The plasminogen activator inhibitor type 1 (PAI-1) is a potential fibrogenic molecule whose secretion is regulated by several metabolic, inflammatory, and genetic factors. We aimed to determine whether PAI-1 secretion in renal transplant patients is correlated with the decline in renal function after transplantation. Renal transplant patients (145 male/71 female) were included in the study 1-27 years after transplantation (median of follow-up: 7.35 years). At inclusion, routine clinical and biological data were collected, the 4G/5G polymorphism of the recipient PAI-1 gene was determined, and the PAI-1 plasma level was measured. The mean rate of decline in renal function was -4.26+/-0.30 ml/min/year. By multiple linear regression analysis, the rate of decline in renal function was significantly correlated with proteinuria (P=0.0176), occurrence of late acute rejection episodes (P=0.0001), and PAI-1 plasma level (P=0.0051). In addition, PAI-1 plasma level was also significantly correlated with body mass index (P=0.038), insulin (P<0.0001), platelet count (P<0.0001), and fibrinogen (P=0.024). The PAI-1 gene polymorphism tested did not influence the rate of decline in renal function after transplantation nor the plasma level of PAI-1 antigen. We conclude that PAI-1, whose secretion is determined in large part by metabolic and inflammatory factors, may be implicated in the rate of decline in renal function after transplantation.
    Article · Apr 2002 · Transplantation
  • [Show abstract] [Hide abstract] ABSTRACT: Our previous study demonstrated that the GR is expressed in the human kidney glomerulus. The function of the GR of glomerular cells might be affected by the concentration of intracellular glucocorticoids, which is modulated by 11beta-hydroxysteroid dehydrogenase type 2 (11betaHSD2). Because the expression of 11betaHSD2 in the glomerular cells remains unclear, we used competitive RT-PCR and immunoblotting to detect the expression of 11betaHSD2 mRNA and protein in isolated human glomeruli, in whole kidney cortex as a positive control, and in a human glomerular visceral epithelial cell line. 11betaHSD2 mRNA was detected in all samples. Specific antihuman 11betaHSD2 antibody recognized a single band at 41 kDa, consistent with the molecular mass of human 11betaHSD2, in the samples of the isolated glomeruli and whole kidney cortex. Furthermore, definite 11betaHSD2 enzymatic activity was also determined with the sample of isolated glomeruli. We also performed immunohistochemistry by light and electron microscopy to determine the cellular and subcellular localization of 11betaHSD2 in the human glomeruli. Immunoreactivity of the enzyme was clearly observed in the glomerular visceral epithelial cells and endothelial cells as well as in the distal convoluted tubules and collecting ducts. The subcellular localization of 11betaHSD2 was shown to be endoplasmic reticulum. These results suggest that 11betaHSD2 might play a crucial role in modulating the intracellular concentration of glucocorticoids in human glomerular cells.
    Article · Mar 2002 · Journal of Clinical Endocrinology & Metabolism
  • F Delarue · A Hertig · C Alberti · [...] · J D Sraer
    [Show abstract] [Hide abstract] ABSTRACT: Plasminogen activator inhibitor type 1 (PAI-1) exerts antifibrinolytic and profibrotic activities. Inside the glomerulus, PAI-1 is mainly synthesized by mesangial cells. We hypothesized that thrombin, via its receptor protease activated receptor type 1 (PAR-1), present on the membrane of glomerular cells, is an important mediator of PAI-1 synthesis. Using the technique of Peten et al., we microdissected the glomeruli of 23 kidney transplanted patients admitted in our department from 1993 to 1997, and we followed-up these patients for up to 5 years, with sometimes iterative renal biopsies. With this technique, we also microdissected the glomeruli of three patients who have had a nephrectomy for cancer (control patients). We investigated mRNA expression of the PAI-1, the thrombin receptor PAR-1, the alpha2 chain of type IV (alpha2 IV) collagen, and of a housekeeping gene (cyclophilin) by reverse transcription-polymerase chain reaction. The results were correlated with the renal function and the histological findings classified into acute rejection (9 biopsies), chronic rejection (22 biopsies), or normal (8 biopsies). A significant up-regulation of PAI-1 and alpha2 IV collagen mRNA was observed in acute rejection (P<0.05) when compared to normal kidneys. A positive correlation exists between alpha2 IV collagen mRNA level and the degree of cellular infiltration. A negative correlation was found between the level of mRNA of PAR-1 and the degree of vascular thrombosis (P=0.005) and glomerulosclerosis (P=0.04). A positive correlation was found between the degradation of renal function and the mRNA level of PAI-1 at the time of the renal biopsy (P<0.05). These results suggest that glomerular PAI-1 mRNA may be predictive of the long-term renal graft function.
    Article · Oct 2001 · Transplantation
  • [Show abstract] [Hide abstract] ABSTRACT: To analyse the mechanisms of PAR-1 internalisation, we constructed several PAR-1 mutants and stably expressed them in CHO cells. Our study shows that the Ser(306)-->Ala mutation (S306A), which eliminates a potential site of phosphorylation by PKC in the third intracellular loop of PAR-1, did not change the rate of phosphorylation but reduced the rate of thrombin-induced internalisation of the PAR-1 mutant (58 versus 78% of membrane PAR-1 in 15 min, p<0.005). Deletion of the last 43 amino acid residues of the PAR-1 cytoplasmic tail completely suppressed the thrombin phosphorylation of the mutated receptor and significantly reduced its internalisation upon activation. This deletion also inhibited the PMA-induced and the agonist-independent internalisation of the receptor. The Tyr(371)--> Ala mutation (Y371A), in a NPXXY motif of the seventh transmembrane domain of the receptor had no effect on the receptor behaviour. Our results indicate that both the C-tail and the third intracellular loop are involved in PAR-1 internalisation induced by thrombin while only the C-tail plays a role in the PMA-induced and in the agonist-independent PAR-1 internalisation.
    Article · Jun 2001 · International Journal of Molecular Medicine
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    Isabelle Kazes · Françoise Delarue · Jacqueline Hagège · [...] · Geneviève Nguyen
    [Show abstract] [Hide abstract] ABSTRACT: Matrix metalloprotease 2 (MMP2) is secreted in a latent inactive form (pro-MMP2) that is activated on the cell surface by a membrane-type 1 MMP (MT1-MMP) in the presence of the tissue inhibitor of MMP (TIMP2). In spite of evidence for the synthesis of MT1-MMP shown by immunoblotting, immunocytochemistry and RT-PCR, and of TIMP2, MMP2 was found exclusively in a latent form in human mesangial cells (HMC) serum-free culture medium. On purified membranes of HMC, MT1-MMP was found in a 63 kD latent form and as a faint band of 55 kD. The 55 kD band was also present in the ultracentrifuged conditioned medium and likely represented MT1-MMP cleaved from its transmembrane domain, since Northern blot analysis showed only one transcription product. The addition of urokinase plasminogen activator (uPA, 100 nM) to HMC membranes induced the activation of pro-MMP2 via the activation of latent membrane-associated MT1-MMP as reflected by the cleavage of the 63 and 55 kD forms. In addition, when the conditioned medium was successively incubated with uPA and alpha 2-macroglobulin and analyzed by immunoblotting, MT1-MMP decreased, indicating that the soluble MT1-MMP was in a latent form and was activated by uPA. Our results provide the first evidence, to our knowledge, of the existence of a soluble latent form of MT1-MMP secreted by primary human cells in culture, confirming that MT1-MMP is an ectoenzyme, and show that uPA can regulate MT1-MMP activity in a soluble phase.
    Full-text Article · Jan 1999 · Kidney International
  • G Nguyen · L Bouzhir · F Delarue · [...] · J D Sraer
    [Show abstract] [Hide abstract] ABSTRACT: Most proteases a receptor or a binding site that serves to concentrate the proteolytic activity on the cell surface and to mediate cellular effects. We looked for such a receptor for renin, an aspartyl protease. The binding of recombinant human renin labelled with 125I was studied on primary and immortalized human mesangial cells. The binding of renin was specific, saturable and was characterized by Kd = 0.4 nM and 8,000 sites/cell and Kd = 1 nM and 2,000 sites/cell for primary and immortalized cells, respectively. The binding did not depend on the active site of the enzyme, was not followed by internalization and degradation of renin and did not modify intracellular Ca2+. Stimulation of primary cells with 100 nM induced a significant increase of 3H thymidine incorporation but was not associated with an increase of the cell number. Furthermore, incubation of mesangial cells 24 h with 100 nM renin provoked an increase of tPA and of PAI1 in the conditioned medium. This increase was not modified neither by captopril nor by angiotensin II receptors antagonists. The tPA antigen elevation was confirmed by fibrin zymography showing an increase of tPA/PAI1 complexes. But, surprisingly, the reverse zymogram showed that PAI antigen increase was associated with decreased PAI activity which was due to PAI clivage in an inactive form. PAI clivage by renin required the presence of the cells and could not be obtained by incubating renin and recombinant human PAI alone. When primary mesangial cells were cultured in the presence of a specific inhibitor of renin active site, RO 42-5982, PAI accumulation in the conditioned medium was reduced by 50-60%, suggesting that endogenous renin plays a role in PAI synthesis and/or secretion. The binding of renin does not induce cAMP and cGMP generation. However, in the presence of renin (100 nM and 1 microM) the extent of cGMP generated by CNP (10 and 100 nM) was reduced by 50%. Preliminary results of the renin receptor purification by affinity chromatography indicate that the receptor Mr is about 57 kDa.
    Article · Feb 1998 · Néphrologie
  • Geneviève Nguyen · Françoise Delarue · Jeannig Berrou · [...] · J D Sraer
    [Show abstract] [Hide abstract] ABSTRACT: Some proteases possess a membrane receptor that focalizes their proteolytic activity on the cell surface and may mediate a proliferative effect, such as urokinase on glomerular epithelial cells. Since some hypertensive states are associated with high concentrations of renin and proliferation of arteriolar smooth muscle cells, we asked whether renin, an aspartyl-protease, would bind to mesangial cells that are smooth-muscle derived cells, which would induce their proliferation. The binding of 125I labeled recombinant human renin (125I-R) was studied on human primary mesangial cells and mesangial cells immortalized by transfection with SV40-T antigen. At 37 degrees C, the binding of 125I-R was time dependent and reached a plateau after two hours. 125I-R was found to bind in a saturable and specific manner with a Kd = 0.4 nM and 1 nM and 8,000 and 2,000 binding sites/cell, for primary and immortalized cells, respectively. When binding experiments were performed in the presence RO 42-5892, a synthetic inhibitor of renin, RO 42-5892 could inhibit the specific binding of labeled renin only at concentrations 1,000 times superior to the IC 50, indicating that the renin-mesangial receptor interaction did not depend on the active site of renin. Analysis by SDS-PAGE and autoradiography of cross-linking experiments of 125I-R bound to a membrane preparation showed a band of approximately 110 to 120 kDa, suggesting a Mr of 70 to 80 kDa for the renin receptor. Incubation of mesangial cells with 100 nM renin for 24 hours provoked a 100% increase of 3H thymidine incorporation that was not accompanied by an increase of the cell number, even after a seven day period of incubation. However, the incubation of mesangial cells with renin for 24 hours induced a significant increase (170% of control, P = 0.04) of plasminogen activator inhibitor-1 (PAI1) antigen in the conditioned medium. In conclusion, we have shown that human mesangial cells in culture express a specific receptor for renin, and that the binding of renin increases 3H thymidine incorporation independently of renin enzymatic activity. The absence of cell proliferation, the increase of 3H thymidine incorporation and the increase of PAI1 antigen suggest that the binding of renin can induce mesangial cell activation, which is reflected by a change in the fibrinolytic capacity of the cells. The role of this receptor remains to be determined in nephropathies and hypertensive states associated with high plasma/tissue renin concentrations, hypertrophy of mesangial or smooth muscle cells and extracellular matrix remodeling.
    Article · Jan 1997 · Kidney International
  • [Show abstract] [Hide abstract] ABSTRACT: The use of human glomerular epithelial cells (HGEC) in research has been severely restricted by several obstacles, which have been circumvented by the generation of T-SV40 immortalized human visceral glomerular epithelial cells (Delarue et al, 1991). In this work, we compared the primary and immortalized HGEC for expression of integrin and some nonintegrin surface receptors. We also studied the adhesion of both types of HGEC to glomerular basement membrane (GBM), type IV collagen (tIV), and its major noncollagenous NC1 domain. The integrins mediating adhesion of HGEC to tIV were also examined. Expression of integrin and some nonintegrin cell surface receptors was analyzed by flow cytometry. Adhesion to GBM, tIV, and its major noncollagenous NC1 domain was studied by direct solid phase cell adhesion assays. Identification of integrins mediating adhesion of HGEC to tIV was achieved by inhibition of cell adhesion using monoclonal antibodies to integrin subunits. The primary and immortalized HGEC share phenotypic characteristics, and alpha3beta1 appeared to be the major integrin present on both HGEC types. The kinetics of binding to GBM, tIV, and its noncollagenous NCI domain were similar in both the primary and immortalized HGEC, although the latter displayed a somewhat weaker binding. Both the primary and immortalized HGEC displayed significantly better adhesion to NC1-alpha3 compared with NC1-alpha1, alpha3beta1 appears to be the major integrin mediating the adhesion of HGEC to tIV. Our studies suggest that alpha3beta1 is the major integrin present on HGEC. This has been confirmed by flow cytometric analysis. In addition, we demonstrated a functional role for this integrin in mediating attachment of HGEC to tIV. Our data also demonstrate a preference in binding of HGEC to alpha3 chains of NC1 compared with alpha1 chains of NC1. These findings were seen in both the primary and immortalized HGEC. The T-SV40 immortalized HGEC can therefore serve as a very useful tool to study glomerular visceral cell biology.
    Article · Apr 1996 · Laboratory Investigation
  • J D Sraer · Françoise Delarue · Jacqueline Hagege · [...] · Eric Rondeau
    [Show abstract] [Hide abstract] ABSTRACT: Kidney International aims to inform the renal researcher and practicing nephrologists on all aspects of renal research. Clinical and basic renal research, commentaries, The Renal Consult, Nephrology sans Frontieres, minireviews, reviews, Nephrology Images, Journal Club. Published weekly online and twice a month in print.
    Article · Feb 1996 · Kidney International
  • A Kanfer · J F Bruch · G Nguyen · [...] · S Uzan
    [Show abstract] [Hide abstract] ABSTRACT: Preeclampsia is characterized by maternal hypercoagulable state and intravascular coagulation, microthromboses in several organs, and impairment of uteroplacental circulation. Excessive fibrin deposition occurs in the placenta, suggesting that disorders of placental coagulation and fibrinolysis physiologic systems may have a role in hemostasis activation. Term placentas were collected from 17 hypertensive/preeclamptic women and from 17 healthy pregnant women, and processed for both histologic and hemostasis studies. Placental fibrinoid deposition was visualized by cresyl-violet staining and quantified by histomorphometric analysis. The content in hemostasis factors was measured on extracts from homogenized placentas treated by a nonionic detergent. The percentage of villi with fibrinoid deposits was higher in the diseased placentas than in controls: 13.2 +/- 11.2 versus 6.75 +/- 2.7% (p < 0.001) for the total amount of deposits; 4.8 +/- 6.7 versus 1.5 +/- 1.0% (p = 0.04) for perivillous fibrinoid deposits, which are considered as histologic markers of intraplacental fibrin. The content in type 2 plasminogen activator inhibitor (PAI-2) antigen was higher in the diseased placentas than in controls: 124 +/- 8 versus 104 +/- 6 ng/mg placental protein (p = 0.046); there was a negative correlation between PAI-2 antigen and thrombomodulin activity (r = -0.57, p = 0.02) in the diseased placentas. No significant differences were found between the two groups for placental procoagulant tissue factor and anticoagulant thrombomodulin activities, and for the content in plasminogen activators and PAI-1 antigens. Placental antifibrinolytic potential is increased in pregnancy-induced hypertension and preeclampsia. This change, and the association of the highest PAI-2 placental concentrations with the lowest concentrations of thrombomodulin, may contribute to the prethrombotic state and to the excessive placental perivillous fibrin deposition observed in these situations.
    Article · Jan 1996 · Laboratory Investigation
  • J Hagège · F Delarue · M N Peraldi · [...] · E Rondeau
    [Show abstract] [Hide abstract] ABSTRACT: Mesangial changes in a variety of pathologic conditions involve mesangial cell proliferation and mesangial matrix remodelling. Heparin has been shown to prevent these processes in vivo. In vitro, heparin interferes with cell growth, proto-oncogene expression, synthesis of specific proteins, and extracellular matrix composition. In some cell types, it seems to interact with intracellular protein kinase C-dependent pathways. The effect of heparin on the mesangial plasminogen activating system (tissue type plasminogen activator, t-PA, and plasminogen activator inhibitor type 1, PAI-1), which is thought to be involved in matrix remodelling, has not been previously reported. Cultured human mesangial cells were stimulated by 10% fetal calf serum (FCS) or 16 nM phorbol myristate acetate (PMA) in the presence or absence of anticoagulant or nonanticoagulant heparins. Cell proliferation, synthesis of t-PA and PAI-1, cell morphology, and PAI-1 matrix deposition were studied using cell counting, [3H]thymidine incorporation, specific t-PA and PAI-1 enzyme-linked immunosorbent assay, Northern blot analysis, light microscopy, immunofluorescence and immunogold silver staining with combined bright-field and epipolarization microscopy. Heparin partially inhibited FCS-stimulated cell growth but not PMA-induced thymidine incorporation. FCS and PMA stimulated t-PA (p < 0.05 and p < 0.01, respectively) and PAI-1 synthesis (p < 0.05 and p < 0.01 respectively). Heparin selectively and partially inhibited FCS-stimulated t-PA, but not PAI-1 synthesis. It has no effect on PMA-stimulated t-PA or PAI-1 synthesis but prevented cell shape-changes induced by PMA, suggesting that heparin inhibits some but not all protein kinase C (PKC)-dependent effects and that heparin block in t-PA synthesis is distal to PKC activation. Heparin decreased PAI-1 matrix accumulation. Similar distal to PKC activation. Heparin decreased PAI-1 matrix accumulation. Similar results were observed with anticoagulant and nonanticoagulant heparin fragments. In human mesangial cells, anticoagulant and nonanticoagulant heparin exert an antiproliferative effect and may prevent mesangial matrix changes by decreasing FCS-stimulated t-PA synthesis and PAI-1 deposition in the matrix. Heparin is able to inhibit PKC-dependent cell shape changes but not PKC-dependent t-PA or PAI-1 synthesis. It also inhibits PKC-independent cell proliferation and t-PA synthesis. These results suggest multiple intracellular sites of action for heparin, unrelated or distal to PKC activation.
    Article · Jan 1995 · Laboratory Investigation
  • C Baudeau · F Delarue · C J Hé · [...] · E Rondeau
    [Show abstract] [Hide abstract] ABSTRACT: MHC class II-encoded molecules HLA-DR, -DP and -DQ play a pivotal role in the human immune response. Their constitutive expression is restricted to a number of immunocompetent cells referred to as antigen-presenting cells. However, gamma-interferon (gamma-IFN) has been shown to induce MHC class II molecule expression in several epithelia. Using flow cytometric analysis, we show here that normal and SV40-transformed human podocytes in culture constitutively expressed gamma-IFN receptors. We also show that MHC class I molecules are constitutively expressed in these cells and that HLA-DR, -DP and -DQ expression, which is not found in unstimulated cells, can be induced by gamma-IFN stimulation. This induction was a time-dependent event, a lag phase of 24-48 h being necessary for MHC class II molecules to become detectable at the cell surface by flow cytometric analysis. Induction of MHC class II molecules in human podocytes also showed a concentration dependence, a plateau being reached at a concentration of 500 IU of gamma-IFN/ml of culture medium. This effect was blunted by coincubation of the cells with an antihuman gamma-IFN receptor monoclonal antibody. HLA-DR expression was associated with specific mRNA accumulation, as detected by Northern blot analysis. By indirect immunofluorescence, the intercellular adhesion molecule 1 was also induced by gamma-IFN stimulation. Induction of DR, DP and DQ in human podocytes may be involved in the pathogenesis of immune glomerulonephritis in man.
    Article · Sep 1994 · Experimental nephrology
  • A KANFER · J.F. Bruch · C FONDACCI · [...] · S UZAN
    Article · Jul 1993 · Placenta
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    [Show abstract] [Hide abstract] ABSTRACT: Human mesangial cells secrete tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor 1 (PAI-1), the latter being secreted in large excess in vitro. We demonstrate that PAI-1 is a major component of the extracellular matrix of cultured human mesangial cells, where its deposition is dependent on cell density. By immunogold silver staining, epipolarization microscopy and dispersive X-ray spectrometry, we have shown that matrix-associated PAI-1 is synthesized by spreading human mesangial cells, as indicated by the time-dependent accumulation of PAI-1 and the inhibitory effect of cycloheximide. Furthermore, by in situ hybridization, PAI-1 mRNA was detected in cultured mesangial cells. t-PA is present inside the cells, or at the cell surface, but is never associated with the extracellular matrix. Exogenous t-PA can remove matrix-associated PAI-1 without affecting cell adhesion. A similar effect was obtained by addition of urokinase-type plasminogen activator (u-PA) but not with fibrinolysis unrelated enzymes. In conclusion, PAI-1 is synthesized by human cultured mesangial cells and is deposited in the extracellular matrix by nonconfluent cells, whereas less PAI-1 is seen between confluent cells. This can explain the absence of detectable PAI-1 in normal human kidney biopsies. t-PA released by mesangial cells can bind and detach matrix PAI-1.
    Full-text Article · Aug 1992 · American Journal Of Pathology
  • [Show abstract] [Hide abstract] ABSTRACT: Human mesangial cells in culture synthesize and secrete plasminogen activator inhibitor 1 (PAI-1) and tissue-type plasminogen activator (t-PA). Phorbol myristate acetate (PMA), a known activator of protein kinase C, induces a three to four-fold increase in t-PA and PAI-1 release over a period of 24 h, whereas cell-associated t-PA and PAI-1 levels remain relatively stable. A similar effect is obtained with oleylacetyl glycerol, a more physiologic protein kinase C activator. The effect of PMA is suppressed in the presence of H7, an inhibitor of cellular protein kinases, and by cycloheximide and actinomycin D, indicating a requirement for de novo protein and RNA synthesis, respectively. Northern blot analysis of PMA-treated cells reveals a rapid and transient increase in PAI-1 mRNA reaching a maximum after 4-8 h, whereas increase in t-PA mRNA levels requires 24 h. Activation of protein kinase A by addition of 8-bromocyclic AMP (8-bromo cAMP) has no significant effect on PAI-1 release but inhibits the PMA-mediated increases in PAI-1 antigen and mRNA. Addition of 8-bromo cAMP alone does not affect t-PA release. When added to PMA-stimulated cells, 8-bromo cAMP inhibits t-PA release in a dose-dependent manner, but causes a superinduction of t-PA mRNA. 8-bromo cAMP also induces a decrease in PMA-stimulated intracellular t-PA release. Similar inhibition is observed after stimulation of endogenous adenylate cyclase with prostaglandin E1 or isoproterenol. This indicates that protein kinase A activation may inhibit PMA-stimulated t-PA release via a post-transcriptional effect, e.g. inhibition of protein synthesis or activation of protein degradation. In conclusion, hormones or mediators which activate protein kinase C can stimulate t-PA and PAI-1 synthesis in human mesangial cells. Protein kinase A activation has no effect on the basal release of PAI-1 and t-PA by human mesangial cells, and, in contrast to endothelial cells, it inhibits both PMA-stimulated PAI-1 and t-PA releases. This cell-specific regulation of t-PA and PAI-1 seems to be mediated by differential transcriptional and post transcriptional mechanisms.
    Article · May 1992 · Biochimica et Biophysica Acta
  • J.M. Rebibou · C.J. He · F Delarue · [...] · J D Sraer
    [Show abstract] [Hide abstract] ABSTRACT: To determine if endothelin 1 (Et1) receptors are present in human glomeruli, and which glomerular cells possess these receptors, 125I Et1 binding to isolated glomeruli and cultured glomerular mesangial and epithelial cells was studied. The latter were identified as podocytes. We demonstrated that Et1 binds specifically and reversibly to isolated human glomeruli and to cultured glomerular mesangial and epithelial cells. Scatchard analysis of competitive inhibition of 125I Et1 binding gave the following results (m +/- SEM, n = 3): isolated glomeruli, Kd = 4.2 +/- 2.1 x 10(-10) M, Bmax = 8.1 +/- 1.2 x 10(10) sites/mg protein; mesangial cells, Kd = 5.2 +/- 1.5 x 10(-10) M, Bmax = 1.87 +/- 0.49 x 10(4) sites/cell; epithelial cells, Kd = 7.2 +/- 1.5 x 10(-10) M, Bmax = 2.46 +/- 0.15 x 10(4) sites/cell. These receptors seem to be functional, since in both mesangial and epithelial cells Et1 induces a rapid and transient increase in intracellular [Ca2+]i. All these results indicate that Et1 may regulate glomerular filtration rate through an autocrine-paracrine pathway on mesangial cells and on podocytes.
    Article · Feb 1992 · Nephrology Dialysis Transplantation
  • E Rondeau · J Cerrina · F Delarue · [...] · J D Sraer
    [Show abstract] [Hide abstract] ABSTRACT: In 10 patients who received a heart lung transplant, TNF-alpha generation by cells collected during bronchioloalveolar lavages (n = 30) and by circulating mononuclear cells was measured. Basal and recombinant IL-2-stimulated productions (50 U/ml) were measured. TNF-alpha concentration was determined by an immunoradiometric assay (IRMA). Circulating mononuclear cells produced at least 4 times less TNF-alpha than BAL cells. Rejection episodes or CMV diseases were not associated with significant changes in TNF-alpha generation. Recombinant IL-2 increased this production in both cell populations but the magnitude of this effect was smaller in BAL cells, suggesting an in vivo preactivation.
    Article · Dec 1991 · La Presse Médicale