Biomedical subjects

M Prat

Publications and source records attributed to M Prat.

At least 37 records · Page 2Linked to original sources

PubMed1998

[Is laparoscopic surgical practice "factual" (evidence based)? Results of a prospective regional survey].

OBJECTIVE: Evidence-based medicine is a growing paradigm in health care. We conducted a prospective study to determine whether laparoscopic surgery is truly evidence-based in everyday practice. METHODS: A prospective regional survey was performed in 11 French hospitals (one university and 10 district hospitals) to ascertain how general laparoscopic surgery was conducted during the last 3 months of 1997. We also searched the electronic databases for original articles on laparoscopic procedures. The methodology of randomized trials was analyzed and procedures were classed by level of evidence. We assumed that an evidence-based procedure was which had been validated by well-designed randomized controlled or prospective trials giving homogeneous results. RESULTS: One half of the procedures performed had been evaluated by randomized controlled trials. Among the 428 laparoscopic procedures, 334 (78%) were found to be evidence-based (CI 74.1-81.9%). Twelve of the 18 indications for laparoscopy (67%) were evidence based (CI: 62.5%-71.5). There was no difference between university teaching hospitals and general district hospitals. CONCLUSION: Contrary to initial criticisms, the practice of laparoscopic surgery appears to be truly evidence-based in the majority of cases.

Evidence-Based Medicine↗

PubMed1998

Expression of Met protein and urokinase-type plasminogen activator receptor (uPA-R) in papillary carcinoma of the thyroid.

Met protein encoded by MET oncogene is the high affinity receptor for hepatocyte growth factor (HGF)/scatter factor (SF). HGF/SF has to be cleaved in its heterodimeric form by the urokinase-type plasminogen activator (uPA) to become active as a ligand for Met receptor. The expression of Met protein and of the high affinity receptor for uPA (uPA-R) was investigated in 39 samples of papillary carcinoma using immunohistochemistry. Reactivity for Met protein was present in 33 of 34 tumours, mostly with a diffuse pattern of staining. Reactivity for uPA-R was present in 78 per cent of papillary tumours and exhibited a pattern of staining similar to that of Met protein. Staining for uPA-R was present in 23 of 25 cases (92 per cent) of papillary carcinoma with prominent sclerosis, and in only 1 of 7 cases (14 per cent) without sclerosis. Peritumoural normal thyroid, follicular adenomas, and follicular carcinomas were negative for Met protein and for uPA-R. Hyperfunctioning tall thyroid cells showed weak membrane reactivity for uPA-R and for Met protein. The findings of immunohistochemistry were confirmed at the mRNA level using in situ hybridization, since the signal for uPA-R and Met RNAs was detected in most tumour cells of five cases of papillary carcinoma.

Adult↗

PubMed1998

Agonistic monoclonal antibodies against the Met receptor dissect the biological responses to HGF.

Hepatocyte growth factor, also known as scatter factor, is a pleiotropic cytokine, which stimulates cell motility, invasion, proliferation, survival and morphogenesis, and induces the expression of specific genes by activating its receptor tyrosine kinase. In this work we have isolated, characterized and used as agonists two monoclonal antibodies (mAbs) directed against the extracellular domain of HGF receptor to investigate the requirements for receptor activation and for the different biological responses. The two mAbs display similar affinities, react with epitopes different from the hepatocyte growth factor binding site, and behave as either full or partial agonists. The full agonist mAb (DO-24) triggers all the biological effects elicited by hepatocyte growth factor, namely motility, proliferation, cell survival, invasion, tubulogenesis and angiogenesis. The partial agonist mAb (DN-30) induces only motility. Only the full agonist mAb is able to induce and sustain the expression of urokinase-type plasminogen activator receptor for prolonged periods of time, while both mAbs up-regulate the constitutive expression of urokinase-type plasminogen activator. Both mAbs activate receptor phosphorylation, which, being strictly dependent on mAb bivalence, requires receptor dimerization. Since simple receptor dimerization is not sufficient to trigger full biological responses, we propose that the region on the ss chain of the receptor recognized by the full agonist mAb is crucial for optimal receptor activation.

Animals↗

PubMed1998

[Epidemiology of hepatitis C virus infection in 1,304 HCV positive patients: variations according to the origin of transmission and year of diagnosis].

UNLABELLED: The evolution of epidemiological data on hepatitis C virus infection is poorly documented and thus the impact of screening is difficult to evaluate. AIM: To study epidemiological variations based on the origin of transmission and the year of diagnosis of hepatitis C virus infection. METHODS: The files of all 1304 patients seen in the hepatology unit of the Rennes University Hospital were analyzed (retrospectively before and prospectively after October 1995) in relation to epidemiological features. RESULTS: Despite widespread screening which is the source of 60% of the diagnoses, the total number of new cases of hepatitis C infection per year has not increased. Compared to patients diagnosed in the first years following the discovery of the virus, patients recently identified were younger (42 +/- 14 years) and frequently drug addicts (40%). Aminotransaminases were normal in 20% of cases. The frequency of cirrhosis has declined (17%). There has been a decrease in the proportion of patients who undergo liver biopsy (50%) and treatment with interferon (one third of patients). CONCLUSIONS: The impact of screening on the number of newly treated patients seems to be lower than previously predicted.

Adult↗

PubMed1997

Activated ras and ret oncogenes induce over-expression of c-met (hepatocyte growth factor receptor) in human thyroid epithelial cells.

Hepatocyte Growth Factor (HGF) receptor, encoded by the protooncogene c-met, is overexpressed in many human tumours, including those of thyroid epithelium. The absence in most cases of any primary structural abnormality of the met gene suggests that overexpression is secondary to mutation of other gene(s). To test this hypothesis we investigated the effect on met expression of two activated oncogenes known to play a major role in thyroid oncogenesis, ras and ret. To minimize the possibility of unknown co-operating events, we introduced these genes directly into normal human thyrocytes in primary culture using amphotropic retroviral vectors and assessed met expression as early as possible in the resulting epithelial colonies. Double immunofluorescence revealed expression of met protein, strictly localized to cells expressing the mutant ras and ret vectors, expression in background normal cells being barely detectable. In contrast, colonies induced to proliferate at a comparable rate by a vector expressing SV40 T showed no increase in met expression. To permit quantitation by Western blotting we also extended these findings to a thyroid cell line (R18) containing a zinc-inducible mutant ras gene. Induction of the oncogene led to a fourfold increase in met protein expression. We conclude that overexpression of met is induced by activation of the ras or ret signalling pathway and not simply by deregulation of the cell cycle per se. The data suggest that the proliferative advantage conferred by these oncogenes may be in part due to the resulting sensitization of tumour epithelium to paracrine HGF secreted by stromal cells.

Antigens, Viral, Tumor↗

PubMed1996

Overexpression of the met/HGF receptor in renal cell carcinomas.

The c-met oncogene encodes the receptor for hepatocyte growth factor/scatter factor (HGF/SF), a multifunctional cytokine able to mediate morphogenesis as well as mitogenesis, motogenesis and invasiveness of epithelial cells. HGF/SF has been implicated in branching tubulogenesis of the developing kidney and in regeneration after renal injury and nephrectomy. We have examined the expression of the met/HGF receptor in normal human kidney and tissues of the genito-urinary tract, and in 50 kidney neoplasms of different histotypes, using monoclonal antibodies (MAbs) against the met/HGF receptor and immunohistochemistry. In normal kidneys, weak staining restricted to the distal tubules was observed. Transitional cell carcinomas were consistently negative, whereas increased expression at various levels was found in 87% of renal cell carcinomas with different cytological features and histological patterns. Western blot analysis of samples showed that the met/HGF receptor found in the malignant cells exhibits features of the normal receptor. The met/HGF receptor is also overexpressed in a renal cell carcinoma cell line, whose motility is triggered by HGF/SF. Our data suggest that expression of the met/HGF receptor may be involved in the onset and progression of renal cell carcinomas.

Antibodies, Monoclonal↗

PubMed1996

Over-expression of hepatocyte growth factor in human Kaposi's sarcoma.

Kaposi's sarcoma is a highly vascularized multifocal tumor which frequently appears as a complication of HIV infection. It has been suggested that a disorder in the cytokine network may contribute to the development of the disease. We examined the expression of several cytokines in human sporadic Kaposi's-sarcoma specimens, as well as in spindle cells cultured from human lesions, and consistently found high levels of expression of hepatocyte growth factor (HGF). In addition, human lesion-derived spindle cells synthesize and secrete biologically active hepatocyte growth factor and express the hepatocyte-growth-factor receptor (c-MET). Moreover, elevated levels of transforming growth factor beta 1 (TGF beta 1) mRNA were found in lesions of human sporadic Kaposi's sarcoma and in lesion-derived spindle cells which also over-express urokinase. Since HGF, TGF beta 1 and urokinase are all involved in capillary-vessel organization, dysregulation of these interacting agents may play a role in the initiation and/or progression of Kaposi's sarcoma, stimulating the growth of spindle cells and recruiting endothelial cells into the lesion.

Base Sequence↗

PubMed1996

Expression of Met protein in thyroid tumours.

Met protein is a transmembrane 190 kD heterodimer with tyrosine kinase activity, encoded by c-MET oncogene. It serves as a high affinity receptor for hepatocyte growth factor (HGF)/scatter factor (SF), a cytokine which stimulates cell proliferation, motility, and invasion. Expression of Met protein was investigated in 116 thyroid tumours using an anti-Met mouse monoclonal antibody (DQ-13) active on paraffin-embedded material. Reactivity for DQ-13 was observed in 77 per cent of papillary carcinomas, in 70 per cent of Hürthle cell tumours, and rarely in other tumours. The staining was either uniformly present throughout the tumour or limited to nests of infiltrating tumour cells. In some Hürthle cell tumours, prominent accumulation of the protein was observed in the Golgi area. Reactivity for Met protein was decreased or absent in poorly differentiated tumours and was not influenced by tumour size, presence of lymph node metastases, or age of the patient. Immunostaining for Ki-67 revealed that cytoplasmic accumulation of Met protein was not associated with enhanced proliferation of tumour cells. Overexpression of Met protein in thyroid papillary carcinoma may result in increased motility of tumour cells, which in turn may account for intraglandular multifocal dissemination and early lymph node metastasis.

Adenocarcinoma↗

PubMed1996

Overexpression and activation of hepatocyte growth factor/scatter factor in human non-small-cell lung carcinomas.

Hepatocyte growth factor/scatter factor (HGF/SF) stimulates the invasive growth of epithelial cells via the c-MET oncogene-encoded receptor. In normal lung, both the receptor and the ligand are detected, and the latter is known to be a mitogenic and a motogenic factor for both cultured bronchial epithelial cells and non-small-cell carcinoma lines. Here, ligand and receptor expression was examined in 42 samples of primary human non-small-cell lung carcinoma of different histotype. Each carcinoma sample was compared with adjacent normal lung tissue. The Met/HGF receptor was found to be 2 to 10-fold increased in 25% of carcinoma samples (P = 0.0113). The ligand, HGF/SF, was found to be 10 to 100-fold overexpressed in carcinoma samples (P < 0.0001). Notably, while HGF/SF was occasionally detectable and found exclusively as a single-chain inactive precursor in normal tissues, it was constantly in the biologically-active heterodimeric form in carcinomas. Immunohistochemical staining showed homogeneous expression of both the receptor and the ligand in carcinoma samples, whereas staining was barely detectable in their normal counterparts. These data show that HGF/SF is overexpressed and consistently activated in non-small-cell lung carcinomas and may contribute to the invasive growth of lung cancer.

Adenocarcinoma↗

PubMed1996

[Organizational models for the hematology areas of the district hospitals of Catalonia].

PURPOSE: The haematological assistance in Catalonia is based upon the district hospitals, in the first step, and the stage III hospitals located usually in higher population nuclei, in the second step. The purpose of this work was to analyse the resources of the "primary haematological assistance" network provided by the district hospitals, to evaluate them and to propose a model for their organisation. MATERIAL AND METHODS: An enquiry was carried out to all members of the Grup de Treball d'Hospitals Comarcals de Catalunya (Catalonia's District Hospitals Task Force). The evaluable data included demographic figures of the population assisted, personnel of each haematological area, organising structure, clinical activity, cytomorphology, blood banks, laboratories and continuous formation activities. RESULTS: The enquiry was answered by 15 of the 21 district hospitals (71.4%) with haematologists in Catalonia. The population assisted in those hospitals is 2,100,000 (ranging between 55,000 an 450,000). All centres are integrated in the National Health network. Eleven of the hospitals analysed have only one haematologist (73.3%). If his dedication is 100% of the time, this would represent a doctor for 105,000 people. The time devoted to work is 690 hours a week for all the population, with a mean of 3,043. Four patients assisted per hour. The total number of hospital beds is 3,353 (50-450), with a mean number of 1 haematologist for every 167.6 beds. The number of patients hospitalized due to blood diseases ranges between 3 and 13 per month. Six of the 15 centres are adjunct to the outpatient clinic. Two centres have a blood bank and 7 have developed an autotransfusion programme. All the centres but one perform oral anticoagulant treatment follow-up, the number of patients assisted ranging from 20 to 210 per week. None of the hospitals has a separate Haematology Service; in most of them haematology is structurally and functionally dependent from Laboratories and in some there is a mixed Laboratory/Internal Medicine functional organisation, depending of the Medical Direction. No haematologist is ever on call specifically for his specialty. Continuous formation activities are carried out in 9 of the 15 centres (60%). COMMENT: Several measures are proposed to improve the haematological assistance, acting on different levels: continuous formation, patient flows and circuits, resident doctors training, anticoagulant treatment network, organisation models, credit cards from the Spanish Association of Haematology.

Adult↗

PubMed1995

In vivo activation of met tyrosine kinase by heterodimeric hepatocyte growth factor molecule promotes angiogenesis.

Hepatocyte growth factor (HGF) is a powerful motogen and mitogen for epithelial cells. The factor is a 90-kD heterodimer composed of an alpha chain containing four kringle motifs and a beta chain showing structural homologies with serine proteases. It is, however, devoid of enzymatic activity. Recently, it has been reported that HGF activates migration and proliferation of endothelial cells and is angiogenic. In this article we discuss (1) the molecular domains of HGF required to activate in vitro and in vivo endothelial cells, studied by use of molecular mutants, and (2) the characteristics of the angiogenic response to HGF in an experimental model system of implanted reconstituted basement membrane (Matrigel). Two groups of mutants were made and used in vitro and in vivo: one with deletions of kringle domains and one with substitution at the cleavage site of the HGF precursor. In vitro, HGF variants containing only the first two (HGF-NK2) or the first three kringles (HGF-NK3) of the alpha chain did not induce proliferation of endothelial cells even if used at concentration 160-fold higher than that optimal for HGF (0.05 nmol/L). High concentrations of these mutants (4 to 8 nmol/L) activated a little endothelial cell motogenic response that was 60% lower than that elicited by HGF. Substitution of Arg 489 with Gln 489 in the HGF precursor generated an uncleavable single-chain factor, unable to induce either endothelial cell migration or proliferation. In vivo, HGF induced a dose-dependent angiogenic response, which was enhanced by heparin.

Animals↗

PubMed1995

Overexpression of c-met protooncogene product and raised Ki67 index in hepatocellular carcinomas with respect to benign liver conditions.

The c-met protooncogene encodes a 190-kd transmembrane tyrosine kinase. This molecule is the receptor for the hepatocyte growth factor (HGF), which is an important mitogen for hepatocytes both in vitro and in vivo. In experimental models, the c-met transcripts appeared strongly expressed by actively proliferating oval cells (OCs). We evaluated the phenotypic modulation of the c-met protooncogene product (c-met pp), in 10 hepatocellular carcinomas (HCCs), 5 focal nodular hyperplasias (FNHs), 4 cases of fulminant hepatitis (FH), and 1 regenerated liver, selected to include the different biological states of hepatocyte (mature normal hepatocytes, transformed ones, and OCs). The supposed mitogenic effect of HGF was analyzed by comparing c-met pp overexpression with the Ki67 index, whereas anti-OV-6 antibody was used for comparison with the presence of OCs. The anti-c-met pp showed a typical plasma membrane-specific staining in all cases. The signal was much stronger in the HCCs than in the benign conditions. The anti-OV-6 monoclonal antibody showed positive immunostaining in many of the cells expressing c-met pp. The percentage of Ki67+ nuclei in high-grade HCCs paralleled c-met protooncogene overexpression. The c-met pp in OV-6+ cells suggests that the paracrine mechanism postulated in experimental models could also apply to human liver.

Biomarkers, Tumor↗

PubMed1994

Immunohistochemistry with antibodies to hepatocyte growth factor and its receptor protein (c-MET) in human brain tissues.

Hepatocyte growth factor (HGF) is a potent mitogen for mature hepatocytes, and also has multifunctional effects on some other cells in various organs. The human c-Met proto-oncogene product has recently been identified as its high-affinity receptor. We examined HGF-like and c-Met protein-like immunoreactivities in the brains of neurologically normal, lacunar stroke and Alzheimer disease (AD) cases. The HGF antibody stained only round cells in the capillaries and astrocytes in the white matter. Positive staining with the antibody to c-Met protein was seen in microglia, predominantly in the white matter. The possibility of interactions between astrocytes and microglia through HGF and its receptor is suggested.

Aged↗

PubMed1994

Generation of a truncated hepatocyte growth factor receptor in the endoplasmic reticulum.

The hepatocyte growth factor (HGF) receptor (p190MET) is a tyrosine kinase composed of two disulfide-linked chains, alpha of 50 kDa and beta of 145 kDa. We have previously described an isoform (p140MET) containing a beta chain of 85 kDa, lacking the cytoplasmic kinase domain. The two receptor variants originate by post-translational processing of a common single-chain precursor of 170 kDa (Pr170). In the endoplasmic reticulum a fraction of Pr170 is cleaved at the cytosolic side generating an intermediate product of 120 kDa (Pr120). This molecule 1) is already detectable after 15 min of pulse labeling, 2) contains high mannose-branched oligosaccharides, and 3) accumulates upon treatments inhibiting the export from the endoplasmic reticulum. A second cleavage, occurring after 30 min of chase in the trans-Golgi network, converts the single-chain precursors Pr170 and Pr120 into the mature heterodimers p190MET and p140MET. This process is inhibited by brefeldin A treatment. Conditions leading to Pr170 accumulation in the endoplasmic reticulum, such as receptor overexpression, induce kinase activation and overproduction of Pr120. Conversely, cells expressing a kinase-defective HGF receptor lack the truncated isoform. The proteolytic cleavage of the cytoplasmic domain may thus represent a safety mechanism aimed at preventing ligand-independent intracellular activation of the HGF receptor kinase.

3T3 Cells↗

PubMed1994

Phosphorylation of serine 985 negatively regulates the hepatocyte growth factor receptor kinase.

The receptor for hepatocyte growth factor/scatter factor (HGF/SF) is an alpha beta tyrosine kinase of 190 kDa which mediates growth and motility in several cell types. We have previously shown that tyrosine autophosphorylation enhances the receptor kinase activity, while serine phosphorylation by protein kinase C or other Ca(2+)-dependent kinase(s) is inhibitory. We now identify Ser985 as the major phosphorylation site for the protein kinases responsible for such inhibition. Both phorbol esters or Ca2+ ionophore treatment induces phosphorylation of the same tryptic phosphopeptide corresponding to the sequence Leu983-Arg987 located in the juxta-membrane domain of the receptor beta chain. Purified protein kinase C phosphorylates in vitro a synthetic peptide (V14S) including Ser985. Trypsin digestion of the phosphorylated V14S generates a single phosphopeptide comigrating in reverse-phase high performance liquid chromatography with the tryptic peptide phosphorylated in vivo. Phorbol ester treatment of cultured cells inhibits the ligand-induced tyrosine autophosphorylation of the receptor. In vitro, Ser985 phosphorylation inhibits the receptor tyrosine kinase activity on exogenous substrates. Substitution of Ser985 by site-directed mutagenesis results in increased tyrosine phosphorylation of the receptor and abolishes down-modulation by protein kinase C. These data show that phosphorylation of Ser985 is a key mechanism for the negative regulation of HGF/SF receptor.

Amino Acid Sequence↗

PubMed1994

Targeting of the SF/HGF receptor to the basolateral domain of polarized epithelial cells.

Scatter Factor, also known as Hepatocyte Growth Factor (SF/HGF), has pleiotropic functions including direct control of cell-cell and cell-substrate adhesion in epithelia. The subcellular localization of the SF/HGF receptor is controversial. In this work, the cell surface distribution of the SF/HGF receptor was studied in vivo in epithelial tissues and in vitro in polarized MDCK monolayers. A panel of monoclonal antibodies against the beta chain of the SF/HGF receptor stained the basolateral but not the apical surface of epithelia lining the lumen of human organs. Radiolabeled or fluorescent-tagged anti-receptor antibodies selectively bound the basolateral cell surface of MDCK cells, which form a polarized monolayer sealed by intercellular junctions, when grown on polycarbonate filters in a two-chamber culture system. The receptor was concentrated around the cell-cell contact zone, showing a distribution pattern overlapping with that of the cell adhesion molecule E-cadherin. The basolateral localization of the SF/HGF receptor was confirmed by immunoprecipitation after domain selective cell surface biotinylation. When cells were fully polarized the SF/HGF receptor became resistant to non-ionic detergents, indicating interaction with insoluble component(s). In pulse-chase labeling and surface biotinylation experiments, the newly synthesized receptor was found exclusively at the basolateral surface. We conclude that the SF/HGF receptor is selectively exposed at the basolateral plasma membrane domain of polarized epithelial cells and is targeted after synthesis to that surface by direct delivery from the trans-Golgi network.

Animals↗

PubMed1994

A monoclonal antibody identifies a novel GPI-anchored glycoprotein involved in epithelial intercellular adhesion.

BD31 mAb, raised against a line of gastric carcinoma cells, reacts with intercellular boundaries of human transformed cells originating from carcinomas or sarcomas growing in epithelial-like clusters as well as in primary cultures of epithelial and endothelial cells. BD31 also reacts with intercellular rims of normal and transformed epithelial tissues and is particularly abundant in glands and fast-growing epithelia but absent in nervous and muscle tissues as well as in blood and in mesenchyme-derived cells. Confocal analysis indicates that BD31 is located in dots at cell-cell contacts but not in basal and apical domains of cultured and in situ epithelial cells. mAb BD31 precipitates a 100 kDa protein from cells labeled with [35S]methionine or [3H]glucosamine as well as from 125I-surface-labeled cells. This glycoprotein resists to trypsin in the presence of Ca2+, releases an 80 kDa fragment in the medium and does not react with antibodies to the conserved cytodomain of known cadherins or, specifically, to the ectodomain of E-cadherin in western blotting; moreover, the lack of cadherin cytodomain and protein removal by phosphoinositide-specific phospholipase indicate its membrane anchoring by a glycosyl-phosphatidylinositol (GPI) moiety. BD31 IgGs do not impair cell-matrix adhesion but induce inhibition of Ca(2+)-dependent aggregation, loss of cell-cell adhesion, scattering of confluent cells and appearance of migratory cell phenotypes in a dose- and time-dependent manner. This novel GPI-anchored glycoprotein may regulate intercellular adhesion by a mechanism involving membrane-associated phospholipases.

Animals↗

PubMed1993

A protein tyrosine phosphatase activity associated with the hepatocyte growth factor/scatter factor receptor.

The receptor for the growth and motility factor, hepatocyte growth factor/scatter factor (HGF/SF), is a transmembrane tyrosine kinase encoded by the MET oncogene. Previous work has shown that receptor phosphorylation on tyrosine is critical for both kinase activation and association with intracellular signal transducers. In this paper, we report that a protein tyrosine phosphatase activity (PTP) coprecipitates with the HGF/SF receptor. The associated PTP activity correlates with the kinase activation of the receptor, increasing up to 5-fold over the basal level after HGF/SF stimulation. The increase is reversible and time- and dose-dependent. A comparable level of activity is associated with constitutively tyrosine-phosphorylated receptors immunoprecipitated from cells where the MET oncogene is amplified and overexpressed. In these cells, a parallel decrease in PTP activity is observed after inhibition of receptor tyrosine phosphorylation following protein kinase C activation. The associated PTP activity is effective in dephosphorylating the HGF/SF receptor. These data show that a protein tyrosine phosphatase is functionally coupled to the HGF/SF receptor.

Cell Line↗