Biomedical subjects

M Conese

Publications and source records attributed to M Conese.

27 records · Page 2Linked to original sources

PubMed1995

Urokinase/urokinase receptor system: internalization/degradation of urokinase-serpin complexes: mechanism and regulation.

The urokinase-type plasminogen activator (uPA) is secreted as a single-chain inactive zymogen (pro-uPA). Upon its secretion, pro-uPA binds to its glycosylphosphatidylinositol-anchored specific cell receptor (uPAR). The activation of pro-uPA to the active two-chain uPA is accelerated with uPAR-bound pro-uPA and is achieved by plasmin and proteases of other classes like cathepsins G and L. uPAR-bound uPA is susceptible to inhibition by its specific inhibitors (PAI-1, PAI-2, and PN-1). uPA-PAI-1 and uPA-PN-1 complexes, but not free uPA, are readily internalized and degraded through a mechanism that involves the multiligand receptors alpha 2-macroglobulin receptor/low density lipoprotein receptor-associated protein (alpha 2-MR) and epithelial glycoprotein 330 (gp330). Upon uPA-inhibitor internalization, uPAR is itself endocytosed and recycled back to the cell surface. PMA-induced differentiation of myeloid cells is accompanied by inhibition of uPA-PAI-1 internalization/degradation and the down-regulation of alpha 2-MR. The regulation of uPAR and alpha 2-MR levels might be part of the differentiation program of myeloid cells.

Animals↗

PubMed1995

Cultured human mesangial cells produce both type 1 and type 2 plasminogen activator inhibitors.

Cultured human mesangial cells (HMC) derived from normal kidneys have been shown to synthesize tissue-type plasminogen activator (t-PA) and excess amounts of PA inhibitor type 1 (PAI-1). Conflicting results have been obtained concerning the production of urokinase-type PA (u-PA) and efforts to show PA inhibitor 2 (PAI-2) met with failure. We evaluated the fibrinolytic profile of cultured HMC lines obtained from 12 patients with renal carcinoma and one cadaveric kidney donor. Subconfluent cells (third passage) were incubated overnight in serum-free medium. t-PA, u-PA, PAI-1 and PAI-2 antigens were assayed by ELISA methods and PA and PAI activities by amidolytic methods both in conditioned medium (CM) and cell extracts (CE). Besides PAI-1, PAI-2 antigen was detected in all but one HMC lines. At variance with the former, which was largely released in the culture medium, PAI-2 was mainly cell-associated. t-PA antigen was found in all but two cell lines while u-PA antigen was detected in relatively high concentrations in 8 cell lines. PA activity, identified as u-PA by functional and immunological criteria, was measured in CM of six of the eight u-PA producing cell lines, whereas PAI activity was undetectable or very low in CM of all cell lines, suggesting that PAI-1 was largely inactive. Functional assays of cell extracts demonstrated the presence of PA activity, again identified as u-PA, only in samples (five lines) containing u-PA antigen in excess over PAI-2. PAI activity was found instead in the extracts in which the inhibitor was higher than the activator (six lines) and was identified as PAI-2, as it inhibited u-PA but not single-chain t-PA and was neutralized by a polyclonal anti-PAI-2 antibody. The heterogeneous fibrinolytic pattern of HMC lines was confirmed by mRNA analysis of three representative lines. Results were similar when HMC lines at passage five were used, except that the u-PA content was significantly reduced both in CM and CE. These findings indicate that the fibrinolytic profile of cultured HMC is more complex than previously reported. The production of large amounts of PAI-2 may represent an additional control mechanism of proteinase activity.

Blotting, Northern↗

PubMed1994

Protease nexin-1-urokinase complexes are internalized and degraded through a mechanism that requires both urokinase receptor and alpha 2-macroglobulin receptor.

After binding to its receptor (uPAR), active cell-surface urokinase (uPA) is not internalized while the complex formed by uPA with plasminogen activator inhibitor type 1 (PAI-1) is internalized and degraded. Internalization and degradation require binding to uPAR and subsequently an interaction with the alpha 2-macroglobulin receptor (alpha 2-MR). To analyze the generality of this mechanism, we studied the internalization of uPA by recombinant protease nexin-1 (rPN-1), an inhibitor of thrombin, uPA, and plasmin. 125I-uPA.rPN-1 complexes bound specifically to uPAR; internalization occurred efficiently, and its time course was essentially the same as for uPA.PAI-1. Internalization required binding to uPAR since it could be blocked by the anti-uPAR monoclonal antibodies, by the uPAR antagonist amino-terminal fragment of uPA, and by the removal of uPAR by the treatment of cells with phosphatidylinositol-specific phospholipase C. As for uPA.PAI-1, the internalization of uPA.rPN-1 also required alpha 2-MR, since it could be inhibited by the 39-kDa alpha 2-macroglobulin receptor/low density lipoprotein receptor-associated protein, a ligand for the alpha 2-MR. Finally, we show by ligand blot analysis that the uPA.rPN-1 complex, like uPA.PAI-1 but unlike free uPA, bound specifically to both uPAR and alpha 2-MR.

Amyloid beta-Protein Precursor↗

PubMed1994

[Comparative evaluation of fluconazole 50 mg and 100 mg versus itraconazole 100 mg in the treatment of dermatomycoses].

In the above double-blind multicenter study the efficacy and tolerability of 50 and 100 mg doses of fluconazole were compared with 100 mg itraconazole in 178 patients with T. corporis, T. cruris, and T. pedis infections. All patients were submitted to clinical and mycological examination before starting, at weekly intervals during treatment, and 4 and 8 weeks after its conclusion. Duration of the three therapeutic regimes was 15 days for T. corporis and T. cruris, and 30 days for T. pedis infection. The percentage of symptomatic cure was 85% and 86.5%, respectively for 50 and 100 mg fluconazole, and 83% for itraconazole. Mycologic cure was achieved in 81.4% of patients treated with 50 mg fluconazole, 83.3% in those treated with 100 mg fluconazole, and 67.9% in those treated with 100 mg itraconazole. None of the groups showed changes in laboratory parameters. It is concluded that all three treatment schemes had high antimycotic activity, but fluconazole both 50 and 100 mg daily was superior. Both drugs were well tolerated and compliance was good.

Adolescent↗

PubMed1993

A ligand-free, soluble urokinase receptor is present in the ascitic fluid from patients with ovarian cancer.

We have identified a soluble form of the human urokinase plasminogen activator (uPA) receptor (uPAR) in the ascitic fluids from patients with ovarian cancer. After purification of uPAR from the ascitic fluids by ligand-affinity chromatography (pro-uPA Sepharose), the uPAR was initially identified by cross-linking to a radiolabeled amino-terminal fragment of human uPA. The uPAR purified from the ascitic fluid has no bound ligand (uPA), as similar amounts can be purified by ligand-affinity chromatography as by immuno-affinity chromatography. uPAR from ascitic fluids partitions in the water phase after a temperature-dependent phase separation of a detergent extract. It therefore lacks at least the lipid moiety of the glycophospholipid anchor present in cellular-bound uPARs. It is highly glycosylated and the deglycosylated form has the same electrophoretic mobility as previously characterized cellular uPAR from other sources. The immunoreactivity of the purified uPAR from the ascitic fluid is indistinguishable from that of characterized uPAR, demonstrated by Western blotting with three different anti-uPAR monoclonal antibodies. The uPAR was found in 11 of 11 ascitic fluids from patients with ovarian cancer and in elevated amounts in the plasma from 2 of 3 patients. The concentration of soluble uPAR in the ascitic fluid was estimated to range between 1 and 10 ng/ml. Human soluble uPAR, derived from the tumor cells, was also found in the ascitic fluid and serum from nude mice xenografted intraperitoneally with three different human ovarian carcinomas.

Animals↗

PubMed1991

Inhibitory effect of retinoids on the generation of procoagulant activity by blood mononuclear phagocytes.

Retinoids are known to modulate several functions of mononuclear phagocytes. We have studied the effect of retinyl acetate (RAc) and retinoic acid (RA) on the production of procoagulant activity (PCA) by human peripheral blood mononuclear cells stimulated with endotoxin (1 microgram/ml, 4 or 20 h at 37 degrees C). Both compounds caused a dose-dependent reduction in the expression of cell-associated PCA (from 86 to less than 10% of control in the range of concentration comprised between 0.1 and 100 microM). This effect was also observed when the cells were exposed to retinoids for 10 min and washed before challenge with endotoxin, indicating that it is rapid and irreversible. In contrast, incubation of RAc or RA for 3 h at 37 degrees C with cells that have been already stimulated with endotoxin (20 h at 37 degrees C) remained without influence on cell PCA. The inhibitory action of retinoids was also observed when monocyte-enriched (greater than 85%) preparations or highly purified monocyte-derived macrophages (greater than 99%) were used instead of whole mononuclear cells. BW755C, an inhibitor of cyclo-oxygenase and lipoxygenase, reversed the inhibitory effect of retinoids, whereas acetylsalycilic acid, an inhibitor of cyclo-oxygenase, was inactive, suggesting the involvement of a lipoxygenase product. The inhibition of monocyte/macrophage PCA production and the subsequent reduction of cell potential for fibrin deposition might represent one of the mechanisms whereby retinoids exert their antiinflammatory and immunomodulatory activities.

Blood Coagulation↗

PubMed1991

Retinoids inhibit the respiratory burst and degranulation of stimulated human polymorphonuclear leukocytes.

Retinoids exhibit a wide spectrum of activities, including antiinflammatory properties. We have investigated the effect of retinoic acid (RA) and retinyl acetate (RAc) on the production of reactive oxygen metabolites and the release of lysosomal enzymes by human polymorphonuclear leukocytes (PMN). Incubation of PMN with RAc or RA (1-100 microM) caused a dose-dependent inhibition (upto 90%) in O2- production and chemiluminescence induced by phorbol myristate acetate (PMA), N-formyl-methionyl-leucyl-phenylanaline (fMLP), opsonized zymosan or ionophore A23187. Both retinoids (1-100 microM) also inhibited, in a dose-dependent way, degranulation induced by fMLP (upto 85% at the highest concentration of RA). These inhibitory effects appear irreversible, since they persist after the drugs are removed and the cells washed before stimulation. Inhibitors of cyclo-oxygenase activity such as acetylsalicyclic acid and indomethacin did not influence the effects of RAc. In contrast, BW755, an inhibitor of both cyclooxygenase and lipoxygenase, reversed the inhibitory action of RAc, suggesting that the effect of retinoids occurs possibly through the mediation of lipoxygenase products. The modulation of PMN oxidative metabolism and degranulation might help explain the antiinflammatory properties of retinoids.

Calcimycin↗

PubMed1990

Unbalanced coagulation-fibrinolysis potential during L-asparaginase therapy in children with acute lymphoblastic leukaemia.

Treatment of acute lymphoblastic leukaemia (ALL) with L-asparaginase (L-asp) may be associated with thrombotic complications, but the pathogenetic mechanisms of thrombus formation and persistence remain unclear. We studied the procoagulant activity (PCA) of peripheral blood mononuclear cells and some components of the plasma fibrinolytic system in 10 children with ALL undergoing remission induction therapy which includes L-asp. Mononuclear cells obtained 14 days after starting L-asp treatment generated significantly higher amounts of PCA (identified as tissue factor) than cells isolated before the first dose of L-asp and 7 days after the cessation of L-asp administration (p less than 0.01). Augmented PCA coincided with an increase in the plasma D-dimer. The plasma levels of type 1 plasminogen activator inhibitor were found significantly elevated during L-asp therapy (p less than 0.05), whereas plasminogen levels were markedly decreased (p less than 0.05). These findings suggest that, during the course of L-asp treatment, the coagulation-fibrinolysis balance is shifted towards promotion of fibrin formation and deposition. Although it remains to be conclusively established whether L-asp per se or the concurrent administration of multiple chemotherapeutic agents is responsible for these changes, the latter could contribute to the thrombotic complications associated with remission induction therapy for ALL.

Asparaginase↗

PubMed1990

Procoagulant activity of mononuclear phagocytes from different anatomical sites in patients with gynecological malignancies.

Mononuclear phagocytes exhibit complex interactions with cancer cells and might contribute to fibrin formation associated with malignancy through the production of procoagulant activity (PCA). We have studied the PCA of peritoneal macrophages in 8 patients with advanced (stages III or IV) ovarian cancer and of macrophages from regional lymph nodes in 14 patients with limited (stages I or II) uterine cancer; peritoneal and lymph-node macrophages from patients with benign gynecological tumors were used as reference cell populations. In all patients, PCA of blood monocytes was also studied. Peritoneal and lymph-node macrophages obtained from patients with ovarian and uterine cancer, respectively, expressed far higher levels of basal PCA than the corresponding cell populations from patients with benign tumors (p less than 0.001). PCA of blood mononuclear cells from patients with ovarian, but not with uterine cancer, was significantly higher (p less than 0.001) than that of control cells. High levels of D-dimer, a specific product derived from plasmin-induced degradation of stabilized fibrin, were found in all ascitic fluids and in all plasma samples but one from patients with ovarian cancer. In contrast, all controls and all uterine cancer patients but one had normal plasma D-dimer. Our findings suggest that local activation of host macrophages for PCA production might contribute to fibrin formation within the tumoral mass. In advanced cancer, blood monocytes may also be activated to produce PCA and thus contribute to activation of intravascular coagulation and, possibly, to thrombo-embolic complications frequently associated with disseminated malignancy.

Aged↗