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Florian Lang - One of the best experts on this subject based on the ideXlab platform.
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Upregulation of Orai1 and STIM1 expression as well as store-operated Ca2+ entry in Ovary Carcinoma cells by placental growth factor.
Biochemical and biophysical research communications, 2019Co-Authors: Khalid N.m. Abdelazeem, Barbora Droppova, Basma Sukkar, Tamer Al-maghout, Lisann Pelzl, Nefeli Zacharopoulou, Nagwa H. A. Hassan, Kamal I. Abdel-fattah, Christos Stournaras, Florian LangAbstract:Abstract Placental growth factor (PlGF) is produced by tumor cells and stimulates tumor growth and metastasis in part by upregulation of hypoxia inducible factor HIF1α. Orchestration of tumor cell proliferation and migration involves oscillations of cytosolic Ca2+ activity ([Ca2+]i). The [Ca2+]i oscillations could be accomplished by triggering of intracellular Ca2+ release followed by store-operated Ca2+-entry (SOCE). Mechanisms accomplishing SOCE include the pore-forming ion channel unit Orai1 and its regulator STIM1. The present study explored whether PlGF influences the expression of Orai1 and STIM1, as well as SOCE and whether this effect impacts on HIF1α expression. To this end, Ovary Carcinoma cells were cultured for 24 h without and with PlGF (10 ng/ml). Orai1, STIM1 and HIF1α transcript levels were quantified utilizing RT-PCR and Orai1, STIM1 and HIF1α protein levels by Western blotting. [Ca2+]i was estimated from Fura-2-fluorescence and SOCE from increase of [Ca2+]i following Ca2+ re-addition after Ca2+-store depletion with extracellular Ca2+ removal and sarcoendoplasmatic Ca2+-ATPase (SERCA) inhibitor thapsigargin (1 μM). As a result, exposure of Ovary Carcinoma cells to PlGF was followed by a significant increase of Orai1 as well as STIM1 transcript and protein levels. PlGF significantly increased store-operated Ca2+-entry following re-addition of extracellular Ca2+, an effect virtually abrogated by Orai1 inhibitor MRS1845 (10 μM). PlGF further increased HIF1α transcript and protein levels, an effect again significantly blunted by MRS1845 (10 μM). In conclusion, PlGF upregulates expression of both, Orai1 and STIM1 thus enhancing store-operated Ca2+-entry with subsequent upregulation of HIF1α.
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Impact of Na+/Ca2+ Exchangers on Therapy Resistance of Ovary Carcinoma Cells.
Cellular physiology and biochemistry : international journal of experimental cellular physiology biochemistry and pharmacology, 2015Co-Authors: Lisann Pelzl, Tamer Al-maghout, Christos Stournaras, Sebastian Schmidt, Zohreh Hosseinzadeh, Kousi Alzoubi, Florian LangAbstract:Background/Aims: According to previous observations, enhanced store-operated Ca2+-entry (SOCE) accomplished by the pore forming ion channel unit Orai1
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Enhanced Orai1 and STIM1 expression as well as store operated Ca2+ entry in therapy resistant Ovary Carcinoma cells
Oncotarget, 2014Co-Authors: Sebastian Schmidt, Christos Stournaras, Guoxing Liu, Guilai Liu, Wenting Yang, Sabina Honisch, Stavros Pantelakos, Arnd Hönig, Florian LangAbstract:// Sebastian Schmidt 1 , Guoxing Liu 1 , Guilai Liu 1 , Wenting Yang 1 , Sabina Honisch 1 , Stavros Pantelakos 1,2 , Christos Stournaras 1,2 , Arnd Honig 3 , Florian Lang 1 1 Department of Physiology, University of Tubingen, D72076 Tubingen, Germany 2 University of Crete Medical School, Department of Biochemistry, Voutes, Heraklion, 71110, Greece 3 Department of Gynecology, University Hospital of Wurzburg, D97080 Wurzburg, Germany Correspondence: Florian Lang, email: // Keywords : Ca2+ release activated Ca2+ channel, SOCE, Akt, SH-6, 2-APB, apoptosis Received : April 22, 2014 Accepted : May 28, 2014 Published : May 28, 2014 Abstract Mechanisms underlying therapy resistance of tumor cells include protein kinase Akt. Putative Akt targets include store-operated Ca 2+ -entry (SOCE) accomplished by pore forming ion channel unit Orai1 and its regulator STIM1. We explored whether therapy resistant (A2780cis) differ from therapy sensitive (A2780) Ovary Carcinoma cells in Akt, Orai1, and STIM1 expression, Ca 2+ -signaling and cell survival following cisplatin (100µM) treatment. Transcript levels were quantified with RT-PCR, protein abundance with Western blotting, cytosolic Ca 2+ -activity ([Ca 2 + ]i) with Fura-2-fluorescence, SOCE from increase of [Ca 2 + ]i following Ca 2+ -readdition after Ca 2+ -store depletion, and apoptosis utilizing flow cytometry. Transcript levels of Orai1 and STIM1, protein expression of Orai1, STIM1, and phosphorylated Akt, as well as SOCE were significantly higher in A2780cis than A2780 cells. SOCE was decreased by Akt inhibitor III (SH-6, 10µM) in A2780cis but not A2780 cells and decreased in both cell lines by Orai1 inhibitor 2-aminoethoxydiphenyl borate (2-ABP, 50µM). Phosphatidylserine exposure and late apoptosis following cisplatin treatment were significantly lower in A2780cis than A2780 cells, a difference virtually abolished by SH-6 or 2-ABP. In conclusion, Orai1/STIM1 expression and function are increased in therapy resistant Ovary Carcinoma cells, a property at least in part due to enhanced Akt activity and contributing to therapy resistance in those cells.
Maria I. Colnaghi - One of the best experts on this subject based on the ideXlab platform.
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folate binding protein distribution in normal tissues and biological fluids from ovarian Carcinoma patients as detected by the monoclonal antibodies mov18 and mov19
European Journal of Cancer, 1994Co-Authors: L Mantovani, Silvana Canevari, Silvia Miotti, Sylvie Ménard, Francesco Raspagliesi, C Bottini, F Bottero, Maria I. ColnaghiAbstract:Folate-binding proteins (FBP), which are molecules relevant in folate metabolism, are overexpressed in ovarian Carcinomas, as detected by the monoclonal antibodies (MAb) MOv18 and MOv19, which recognise two different epitopes of the gp38/FBP. In this paper, features of the FBP such as the distribution on normal tissues and the release in biological fluids of normal and tumour origin have been investigated. Immunohistochemical analyses on frozen sections of normal tissues showed the presence of the gp38/FBP on some epithelia. The reactivity of both the MAb on Fallopian tubes was intense and comparable to that observed on Ovary Carcinoma sections. The kidney, bronchial glands, alveolar epithelium of the lung, oesophagus, stomach, pancreas, breast and thyroid showed different levels of staining. By MOv18/MOv19 double-determinant immunoradiometric assay (DDIRMA), the gp38/FBP was found in soluble form in ascitic fluid, serum and urine of nude mice in which the human Ovary Carcinoma cell line IGROV1 grew as ascitic Carcinomatosis. In human biological fluids, the gp38/FBP was detected in ascites of 60% of ovarian Carcinoma patients, and in 29% of those with other Carcinomas, but not in patients with non-epithelial tumours or with other non-tumoral pathologies. The mean serum arbitrary units (a.u.)/ml values of Ovary Carcinoma patients were significantly different to those of healthy donors or patients with endometriosis (P < 0.005 and P < 0.01, respectively), but not when compared to the sera of lung Carcinoma patients. In addition, the sensitivity of DDIRMA was poor, since only 24% of the Ovary Carcinoma patients were positive with this assay. When a restricted number of cases selected for the presence of tumour cells in the ascites was examined, the percentage of DDIRMA-positive sera and ascites rose to 41 and 94%, respectively. In the urine, a strong reactivity was observed in the samples of both normal and tumour origin.
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Polarization of the α6β4 integrin in ovarian Carcinomas
International journal of cancer, 1993Co-Authors: C Bottini, Silvia Miotti, Patrizia Facheris, Sylvie Ménard, S. Fiorucci, Maria I. ColnaghiAbstract:By immunizing a BALB/c mouse with a human Ovary-Carcinoma cell line (IGROV1), grown intraperitoneally in nude mice, a monoclonal antibody (MAb), designated MAR6, was produced and characterized. Immunofluorescence on the immunizing cell line showed a specific labelling by MAR6 at the cell-to-cell contact points. In addition, MAR6 was found to immunoprecipitate the alpha 6 beta 4 integrin complex. Competition tests with MAbs anti-alpha 6, anti-beta 4, anti-beta 1 sub-units demonstrated that the recognized sub-unit is alpha 6. Indirect immunofluorescence on various cell lines gave MAR6 as positive only on alpha 6-positive lines (IGROV1, OVCAR3, SW626, SKOV3, ME4405, Calu3, N592, MDA468, A431 and HT29). Moreover, on IGROV1 and OVCAR3 Ovary-Carcinoma cells, which normally grow either adhering to the culture flask or forming clumps in suspension in the medium, MAR6 selectively stained the connection points between the cells in clumps, where, in the same position, the presence of the beta 4 sub-unit, laminin and fibronectin was detected. On the contrary, the beta 1 sub-unit was distributed over the whole cell membrane. The same pattern of labelling by these MAbs was observed in 2 cases of ovarian-Carcinoma cells present in ascitic fluids obtained from patients. Immunoperoxidase tests performed on cryosections of various normal tissues showed specific reactivity of MAR6 on basal or basolateral membranes of epithelial cells. On cryosections of ovarian tumors, MAR6 reactivity correlated with the degree of tumor differentiation. Indeed, in benign and well-differentiated tumors, a strong basal or basolateral labelling only of cells surrounding the neoplastic nodules was found. On the contrary, on undifferentiated tumors the inner part of the tumor nodules was also progressively labelled, whereas the staining on the border was weak and discontinuous as a result of the alpha 6 sub-unit dispersion on the tumor cell surface.
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Membrane association and shedding of the GPI-anchored Ca-MOv18 antigen in human Ovary Carcinoma cells.
International journal of cancer, 1992Co-Authors: Silvia Miotti, Silvana Canevari, Saverio Alberti, Patrizia Facheris, Liliana Mantovani, Mara Fornaro, Manuela Stella, Sylvie Ménard, Maria I. ColnaghiAbstract:The antigen recognized by the MOv18 MAb (Ca-MOv18) was recently shown to be a glycosylphosphatidylinositol (GPI)-linked protein. In this report we show that GPI-anchorage is not limited to IGROVI cells nor to other Ovary Carcinoma cell lines, but Ca-MOv18 was also found to be sensitive to phosphatidylinositol-specific phospholipase C (PI-PLC) treatment on fresh ovarian cancer cells. Furthermore, we found a heterogeneous sensitivity of Ca-MOv18 to PI-PLC cleavage, not only among the different cells studied but also in different experiments performed on the same cell line, during extended periods of time in culture. Sensitivity to PI-PLC cleavage was determined by immunofluorescence on live cells and by double-determinant radioimmunoassay of the antigen released in the supernatant. The specificity of the PI-PLC cleavage was demonstrated as follows: (a) TX114 solubilized Ca-MOv18 shifts from the detergent to the aqueous phase after treatment with PI-PLC; (b) on membrane preparations, PI-PLC specifically released a fraction of the antigen, which is distinct from the weakly associated form released by high-salt treatment; (c) Ca-MOv18 from IGROVI expressed the cross-reacting determinant (CRD), which is characteristic of GPI-linked molecules. The absence of CRD expression on the spontaneously released protein and the possibility of artificially inducing antigen shedding during the resynthesis of Ca-MOv18 which follows bacterial PI-PLC treatment are interesting points which need to be further investigated in order to understand the physiology of the Ca-MOv18 tumor antigen.
Christos Stournaras - One of the best experts on this subject based on the ideXlab platform.
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Upregulation of Orai1 and STIM1 expression as well as store-operated Ca2+ entry in Ovary Carcinoma cells by placental growth factor.
Biochemical and biophysical research communications, 2019Co-Authors: Khalid N.m. Abdelazeem, Barbora Droppova, Basma Sukkar, Tamer Al-maghout, Lisann Pelzl, Nefeli Zacharopoulou, Nagwa H. A. Hassan, Kamal I. Abdel-fattah, Christos Stournaras, Florian LangAbstract:Abstract Placental growth factor (PlGF) is produced by tumor cells and stimulates tumor growth and metastasis in part by upregulation of hypoxia inducible factor HIF1α. Orchestration of tumor cell proliferation and migration involves oscillations of cytosolic Ca2+ activity ([Ca2+]i). The [Ca2+]i oscillations could be accomplished by triggering of intracellular Ca2+ release followed by store-operated Ca2+-entry (SOCE). Mechanisms accomplishing SOCE include the pore-forming ion channel unit Orai1 and its regulator STIM1. The present study explored whether PlGF influences the expression of Orai1 and STIM1, as well as SOCE and whether this effect impacts on HIF1α expression. To this end, Ovary Carcinoma cells were cultured for 24 h without and with PlGF (10 ng/ml). Orai1, STIM1 and HIF1α transcript levels were quantified utilizing RT-PCR and Orai1, STIM1 and HIF1α protein levels by Western blotting. [Ca2+]i was estimated from Fura-2-fluorescence and SOCE from increase of [Ca2+]i following Ca2+ re-addition after Ca2+-store depletion with extracellular Ca2+ removal and sarcoendoplasmatic Ca2+-ATPase (SERCA) inhibitor thapsigargin (1 μM). As a result, exposure of Ovary Carcinoma cells to PlGF was followed by a significant increase of Orai1 as well as STIM1 transcript and protein levels. PlGF significantly increased store-operated Ca2+-entry following re-addition of extracellular Ca2+, an effect virtually abrogated by Orai1 inhibitor MRS1845 (10 μM). PlGF further increased HIF1α transcript and protein levels, an effect again significantly blunted by MRS1845 (10 μM). In conclusion, PlGF upregulates expression of both, Orai1 and STIM1 thus enhancing store-operated Ca2+-entry with subsequent upregulation of HIF1α.
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Impact of Na+/Ca2+ Exchangers on Therapy Resistance of Ovary Carcinoma Cells.
Cellular physiology and biochemistry : international journal of experimental cellular physiology biochemistry and pharmacology, 2015Co-Authors: Lisann Pelzl, Tamer Al-maghout, Christos Stournaras, Sebastian Schmidt, Zohreh Hosseinzadeh, Kousi Alzoubi, Florian LangAbstract:Background/Aims: According to previous observations, enhanced store-operated Ca2+-entry (SOCE) accomplished by the pore forming ion channel unit Orai1
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Enhanced Orai1 and STIM1 expression as well as store operated Ca2+ entry in therapy resistant Ovary Carcinoma cells
Oncotarget, 2014Co-Authors: Sebastian Schmidt, Christos Stournaras, Guoxing Liu, Guilai Liu, Wenting Yang, Sabina Honisch, Stavros Pantelakos, Arnd Hönig, Florian LangAbstract:// Sebastian Schmidt 1 , Guoxing Liu 1 , Guilai Liu 1 , Wenting Yang 1 , Sabina Honisch 1 , Stavros Pantelakos 1,2 , Christos Stournaras 1,2 , Arnd Honig 3 , Florian Lang 1 1 Department of Physiology, University of Tubingen, D72076 Tubingen, Germany 2 University of Crete Medical School, Department of Biochemistry, Voutes, Heraklion, 71110, Greece 3 Department of Gynecology, University Hospital of Wurzburg, D97080 Wurzburg, Germany Correspondence: Florian Lang, email: // Keywords : Ca2+ release activated Ca2+ channel, SOCE, Akt, SH-6, 2-APB, apoptosis Received : April 22, 2014 Accepted : May 28, 2014 Published : May 28, 2014 Abstract Mechanisms underlying therapy resistance of tumor cells include protein kinase Akt. Putative Akt targets include store-operated Ca 2+ -entry (SOCE) accomplished by pore forming ion channel unit Orai1 and its regulator STIM1. We explored whether therapy resistant (A2780cis) differ from therapy sensitive (A2780) Ovary Carcinoma cells in Akt, Orai1, and STIM1 expression, Ca 2+ -signaling and cell survival following cisplatin (100µM) treatment. Transcript levels were quantified with RT-PCR, protein abundance with Western blotting, cytosolic Ca 2+ -activity ([Ca 2 + ]i) with Fura-2-fluorescence, SOCE from increase of [Ca 2 + ]i following Ca 2+ -readdition after Ca 2+ -store depletion, and apoptosis utilizing flow cytometry. Transcript levels of Orai1 and STIM1, protein expression of Orai1, STIM1, and phosphorylated Akt, as well as SOCE were significantly higher in A2780cis than A2780 cells. SOCE was decreased by Akt inhibitor III (SH-6, 10µM) in A2780cis but not A2780 cells and decreased in both cell lines by Orai1 inhibitor 2-aminoethoxydiphenyl borate (2-ABP, 50µM). Phosphatidylserine exposure and late apoptosis following cisplatin treatment were significantly lower in A2780cis than A2780 cells, a difference virtually abolished by SH-6 or 2-ABP. In conclusion, Orai1/STIM1 expression and function are increased in therapy resistant Ovary Carcinoma cells, a property at least in part due to enhanced Akt activity and contributing to therapy resistance in those cells.
Francesco Raspagliesi - One of the best experts on this subject based on the ideXlab platform.
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folate binding protein distribution in normal tissues and biological fluids from ovarian Carcinoma patients as detected by the monoclonal antibodies mov18 and mov19
European Journal of Cancer, 1994Co-Authors: L Mantovani, Silvana Canevari, Silvia Miotti, Sylvie Ménard, Francesco Raspagliesi, C Bottini, F Bottero, Maria I. ColnaghiAbstract:Folate-binding proteins (FBP), which are molecules relevant in folate metabolism, are overexpressed in ovarian Carcinomas, as detected by the monoclonal antibodies (MAb) MOv18 and MOv19, which recognise two different epitopes of the gp38/FBP. In this paper, features of the FBP such as the distribution on normal tissues and the release in biological fluids of normal and tumour origin have been investigated. Immunohistochemical analyses on frozen sections of normal tissues showed the presence of the gp38/FBP on some epithelia. The reactivity of both the MAb on Fallopian tubes was intense and comparable to that observed on Ovary Carcinoma sections. The kidney, bronchial glands, alveolar epithelium of the lung, oesophagus, stomach, pancreas, breast and thyroid showed different levels of staining. By MOv18/MOv19 double-determinant immunoradiometric assay (DDIRMA), the gp38/FBP was found in soluble form in ascitic fluid, serum and urine of nude mice in which the human Ovary Carcinoma cell line IGROV1 grew as ascitic Carcinomatosis. In human biological fluids, the gp38/FBP was detected in ascites of 60% of ovarian Carcinoma patients, and in 29% of those with other Carcinomas, but not in patients with non-epithelial tumours or with other non-tumoral pathologies. The mean serum arbitrary units (a.u.)/ml values of Ovary Carcinoma patients were significantly different to those of healthy donors or patients with endometriosis (P < 0.005 and P < 0.01, respectively), but not when compared to the sera of lung Carcinoma patients. In addition, the sensitivity of DDIRMA was poor, since only 24% of the Ovary Carcinoma patients were positive with this assay. When a restricted number of cases selected for the presence of tumour cells in the ascites was examined, the percentage of DDIRMA-positive sera and ascites rose to 41 and 94%, respectively. In the urine, a strong reactivity was observed in the samples of both normal and tumour origin.
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The role of appendectomy in surgical procedures for ovarian cancer
Gynecologic oncology, 1992Co-Authors: Rosanna Fontanelli, D. Paladini, Francesco RaspagliesiAbstract:Abstract To assess the role of appendectomy in the surgical procedures for ovarian cancer, we evaluated retrospectively the clinical charts of 435 patients who underwent surgery after diagnosis of ovarian cancer. The appendix was removed in 160 cases and pathological examination revealed 37 with metastatic implants (23%). All the patients with appendiceal metastases showed advanced disease (stages III–IV) with an incidence of 43%. Ninety-one percent (3134) of the tumors with appendiceal involvement at the staging operation were of the serous cell type and grade II or III. No case with early stage, right Ovary Carcinoma showed appendiceal metastatic foci, denying the existence of a preferential lymphatic pathway. Microscopic involvement was found only in 4 patients with advanced disease (11.7%). No intra- or postoperative complication directly related to the appendectomy was recorded. We conclude, with these results, that appendectomy should be part of the cytoreductive operation for ovarian cancer.
Silvana Canevari - One of the best experts on this subject based on the ideXlab platform.
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Internalization and recycling of ALCAM/CD166 detected by a fully human single-chain recombinant antibody
Journal of Cell Science, 2005Co-Authors: Tiziana Piazza, Emanuela Cha, Francesca Sassi, Andrea Bolognesi, Italia Bongarzone, Silvana Canevari, Letizia Polito, A. Bargellesi, Silvano Ferrini, Marina FabbiAbstract:Activated leukocyte cell adhesion molecule (ALCAM/CD166), a member of the immunoglobulin superfamily with five extracellular immunoglobulin-like domains, promotes heterophilic (ALCAM-CD6) and homophilic (ALCAM-ALCAM) cell-cell interactions. Here we describe a fully human single-chain antibody fragment (scFv) directed to ALCAM/CD166. We selected the I/F8 scFv from a phage display library of human V-gene segments by cell panning and phage internalization into IGROV-I human Ovary Carcinoma cells. The I/F8 specificity was identified as ALCAM/CD166 by matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) peptide mass fingerprinting of the I/F8-immunoprecipitated protein. The I/F8 scFv reacts with the human, monkey and murine ALCAM/CD166 molecule, indicating that the recognized epitope is highly conserved. The I/F8 scFv completely abolished binding of both ALCAM/Fc and CD6/Fc soluble ligands, whereas it did not compete with the anti-ALCAM/CD166 murine monoclonal antibodies J4-81 and 3A6 and therefore recognizes a different epitope. Engagement through I/F8 scFv, 3A6 monoclonal antibody or CD6/Fc ligand induced ALCAM/CD166 internalization, with a kinetics slower than that of transferrin in the same cells. Newly internalized I/F8-ALCAM complexes colocalized with clathrin but not with caveolin and we demonstrated, using surface biotinylation and recycling assays, that endocytosed ALCAM/CD166 recycles back to the cell surface. Such an endocytic pathway allows the efficient delivery of an I/F8 scFv-saporin immunotoxin into tumor cells, as the conjugates are able to selectively kill cell lines expressing ALCAM/CD166. Altogether these data provide evidence of the suitability of the I/F8 scFv for further functional analysis of ALCAM/CD166 and intracellular delivery of effector moieties.
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Interaction of folate receptor with signaling molecules lyn and G(alpha)(i-3) in detergent-resistant complexes from the Ovary Carcinoma cell line IGROV1.
Journal of cell science, 2000Co-Authors: S Miotti, M Bagnoli, A Tomassetti, M I Colnaghi, Silvana CanevariAbstract:Using as a model the Ovary Carcinoma cell line IGROV1, we analyzed the partitioning of the glycosyl-phosphatidylinositol-anchored folate receptor into lipid rafts based on its relative detergent insolubility, with a focus on physically and functionally associated signaling molecules. A variable amount (40-60%) of folate receptor was found in low-density Triton X-100 insoluble complexes together with subunits of heterotrimeric G-proteins and the src-family non-receptor tyrosine kinases p53-56 lyn. In the same fraction the structural component of caveolae, caveolin, was not detected at the protein level, although the corresponding mRNA was detected in trace amounts. Comodulation of folate receptor and signalling molecules was observed in the detergent-insoluble complexes during cell proliferation or induced by phosphatidylinositol-specific phospholipase C treatment or by interaction with anti-folate receptor monoclonal antibodies. Moreover, complexes of folate receptor, lyn and the G(&agr;)(i-3) subunit were immunoprecipitated using either anti-folate receptor or anti-lyn antibodies. In vitro kinase assay of the immunoprecipitates revealed stimulation of phosphorylation of common and specific proteins. In particular, the p53 form of lyn appeared to be enriched and phosphorylated in the anti-folate receptor MOv19 monoclonal antibody immunoprecipitate, whereas a 40 kDa band common to anti-folate receptor and anti-lyn immunoprecipitates was the phosphorylated form of the G(&agr;)(i-3) subunit. These findings point to the functional interaction between folate receptor and associated signaling molecules.
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folate binding protein distribution in normal tissues and biological fluids from ovarian Carcinoma patients as detected by the monoclonal antibodies mov18 and mov19
European Journal of Cancer, 1994Co-Authors: L Mantovani, Silvana Canevari, Silvia Miotti, Sylvie Ménard, Francesco Raspagliesi, C Bottini, F Bottero, Maria I. ColnaghiAbstract:Folate-binding proteins (FBP), which are molecules relevant in folate metabolism, are overexpressed in ovarian Carcinomas, as detected by the monoclonal antibodies (MAb) MOv18 and MOv19, which recognise two different epitopes of the gp38/FBP. In this paper, features of the FBP such as the distribution on normal tissues and the release in biological fluids of normal and tumour origin have been investigated. Immunohistochemical analyses on frozen sections of normal tissues showed the presence of the gp38/FBP on some epithelia. The reactivity of both the MAb on Fallopian tubes was intense and comparable to that observed on Ovary Carcinoma sections. The kidney, bronchial glands, alveolar epithelium of the lung, oesophagus, stomach, pancreas, breast and thyroid showed different levels of staining. By MOv18/MOv19 double-determinant immunoradiometric assay (DDIRMA), the gp38/FBP was found in soluble form in ascitic fluid, serum and urine of nude mice in which the human Ovary Carcinoma cell line IGROV1 grew as ascitic Carcinomatosis. In human biological fluids, the gp38/FBP was detected in ascites of 60% of ovarian Carcinoma patients, and in 29% of those with other Carcinomas, but not in patients with non-epithelial tumours or with other non-tumoral pathologies. The mean serum arbitrary units (a.u.)/ml values of Ovary Carcinoma patients were significantly different to those of healthy donors or patients with endometriosis (P < 0.005 and P < 0.01, respectively), but not when compared to the sera of lung Carcinoma patients. In addition, the sensitivity of DDIRMA was poor, since only 24% of the Ovary Carcinoma patients were positive with this assay. When a restricted number of cases selected for the presence of tumour cells in the ascites was examined, the percentage of DDIRMA-positive sera and ascites rose to 41 and 94%, respectively. In the urine, a strong reactivity was observed in the samples of both normal and tumour origin.
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Membrane association and shedding of the GPI-anchored Ca-MOv18 antigen in human Ovary Carcinoma cells.
International journal of cancer, 1992Co-Authors: Silvia Miotti, Silvana Canevari, Saverio Alberti, Patrizia Facheris, Liliana Mantovani, Mara Fornaro, Manuela Stella, Sylvie Ménard, Maria I. ColnaghiAbstract:The antigen recognized by the MOv18 MAb (Ca-MOv18) was recently shown to be a glycosylphosphatidylinositol (GPI)-linked protein. In this report we show that GPI-anchorage is not limited to IGROVI cells nor to other Ovary Carcinoma cell lines, but Ca-MOv18 was also found to be sensitive to phosphatidylinositol-specific phospholipase C (PI-PLC) treatment on fresh ovarian cancer cells. Furthermore, we found a heterogeneous sensitivity of Ca-MOv18 to PI-PLC cleavage, not only among the different cells studied but also in different experiments performed on the same cell line, during extended periods of time in culture. Sensitivity to PI-PLC cleavage was determined by immunofluorescence on live cells and by double-determinant radioimmunoassay of the antigen released in the supernatant. The specificity of the PI-PLC cleavage was demonstrated as follows: (a) TX114 solubilized Ca-MOv18 shifts from the detergent to the aqueous phase after treatment with PI-PLC; (b) on membrane preparations, PI-PLC specifically released a fraction of the antigen, which is distinct from the weakly associated form released by high-salt treatment; (c) Ca-MOv18 from IGROVI expressed the cross-reacting determinant (CRD), which is characteristic of GPI-linked molecules. The absence of CRD expression on the spontaneously released protein and the possibility of artificially inducing antigen shedding during the resynthesis of Ca-MOv18 which follows bacterial PI-PLC treatment are interesting points which need to be further investigated in order to understand the physiology of the Ca-MOv18 tumor antigen.