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Activated protein C upregulates ovarian cancer cell migration and promotes unclottability of the cancer cell microenvironment.

Althawadi H, Alfarsi H, Besbes S, Mirshahi S, Ducros E, Rafii A, Pocard M, Therwath A, Soria J, Mirshahi M - Oncol. Rep. (2015)

Bottom Line: Our results showed that in the OVCAR-3 aPC-induced cells i) an increase in cell migration was noted, which was inhibited when anti-endothelial protein C receptor (EPCR) was added to the culture medium and which may act via MEK-ERK and Rho-GTPase pathways; ii) an increase in threonine, and to a lesser extent tyrosine phosphorylation; iii) cell cycle activation (G1 to S/G2); and iv) a 2-3-fold prolongation of aPTT of normal plasma.In the peritoneal fluid, the sEPCR concentration was 71 ± 23 ng/ml.This binding could also explain the loss of clotting of peritoneal fluids.

View Article: PubMed Central - PubMed

Affiliation: UMR, Paris Diderot, Paris 7 University, Lariboisiere Hospital, INSERM U965, Paris, France.

ABSTRACT
The objective of this study was to evaluate the role of activated protein C (aPC), known to be a physiological anticoagulant, in ovarian cancer cell activation as well as in loss of clotting of cancer ascitic fluid. The effect of aPC on an ovarian cancer cell line (OVCAR-3) was tested in regards to i) cell migration and adhesion with the use of adhesion and wound healing assays as well as a droplet test; ii) protein phosphorylation, evaluated by cyto-ELISA; iii) cell cycle modification assessed by flow cytometric DNA quantification; and iv) anticoagulant activity evaluated by the prolongation of partial thromboplastin time (aPTT) of normal plasma in the presence or absence of aPC-treated ovarian cancer cells. In addition, the soluble endothelial protein C receptor (sEPCR) was quantified by ELISA in ascitic fluid of patients with ovarian cancer. Our results showed that in the OVCAR-3 aPC-induced cells i) an increase in cell migration was noted, which was inhibited when anti-endothelial protein C receptor (EPCR) was added to the culture medium and which may act via MEK-ERK and Rho-GTPase pathways; ii) an increase in threonine, and to a lesser extent tyrosine phosphorylation; iii) cell cycle activation (G1 to S/G2); and iv) a 2-3-fold prolongation of aPTT of normal plasma. In the peritoneal fluid, the sEPCR concentration was 71 ± 23 ng/ml. In conclusion, free aPC binds to membrane EPCR in ovarian cancer cells and induces cell migration via MEK-ERK and Rho-GTPase pathways. This binding could also explain the loss of clotting of peritoneal fluids.

No MeSH data available.


Related in: MedlinePlus

Fibrin formation kinetics on plasma in the presence of PC or aPC bound to OVCAR-3 cells tested by aPTT test. (A) aPTT test of adherent and detached OVCAR-3 cells without PC or aPC as controls. aPTT test of OVCAR-3 cells in (B) detached and (C) adherent conditions, in the presence of PC or aPC. (D) Real-time of aPTT (minutes) in different conditions. OVCAR-3, ovarian cancer cell line; PC, protein C; aPC, active protein C; aPTT, activated partial thromboplastin time.
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f5-or-34-02-0603: Fibrin formation kinetics on plasma in the presence of PC or aPC bound to OVCAR-3 cells tested by aPTT test. (A) aPTT test of adherent and detached OVCAR-3 cells without PC or aPC as controls. aPTT test of OVCAR-3 cells in (B) detached and (C) adherent conditions, in the presence of PC or aPC. (D) Real-time of aPTT (minutes) in different conditions. OVCAR-3, ovarian cancer cell line; PC, protein C; aPC, active protein C; aPTT, activated partial thromboplastin time.

Mentions: The anticoagulant property of OVCAR-3 cells was assessed by measuring the prolongation of aPTT of normal plasma induced by aPC. As shown in Fig. 5A, the effect of OVCAR-3 cells, detached and incubated either with PC or APC, in fibrin polymerization curve of normal plasma is shown. The same experiment was also performed using OVCAR-3 cells in adherent conditions. When both detached (Fig. 5B) and adherent (Fig. 5C) cells, were incubated with aPC, aPTT was prolonged as compared to cells incubated with protein C. As documented in Fig. 5D, PC added to the adherent or detached ovarian cancer cells induced a 2-fold increase in cephalin clotting time, whereas aPC under the same conditions induced a 3.7-fold increase (5.78–11.90 min for PC and 21.42 min for aPC). These results indicate the anti-coagulant action of EPCR-aPC on the OVCAR-3 cells.


Activated protein C upregulates ovarian cancer cell migration and promotes unclottability of the cancer cell microenvironment.

Althawadi H, Alfarsi H, Besbes S, Mirshahi S, Ducros E, Rafii A, Pocard M, Therwath A, Soria J, Mirshahi M - Oncol. Rep. (2015)

Fibrin formation kinetics on plasma in the presence of PC or aPC bound to OVCAR-3 cells tested by aPTT test. (A) aPTT test of adherent and detached OVCAR-3 cells without PC or aPC as controls. aPTT test of OVCAR-3 cells in (B) detached and (C) adherent conditions, in the presence of PC or aPC. (D) Real-time of aPTT (minutes) in different conditions. OVCAR-3, ovarian cancer cell line; PC, protein C; aPC, active protein C; aPTT, activated partial thromboplastin time.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4487670&req=5

f5-or-34-02-0603: Fibrin formation kinetics on plasma in the presence of PC or aPC bound to OVCAR-3 cells tested by aPTT test. (A) aPTT test of adherent and detached OVCAR-3 cells without PC or aPC as controls. aPTT test of OVCAR-3 cells in (B) detached and (C) adherent conditions, in the presence of PC or aPC. (D) Real-time of aPTT (minutes) in different conditions. OVCAR-3, ovarian cancer cell line; PC, protein C; aPC, active protein C; aPTT, activated partial thromboplastin time.
Mentions: The anticoagulant property of OVCAR-3 cells was assessed by measuring the prolongation of aPTT of normal plasma induced by aPC. As shown in Fig. 5A, the effect of OVCAR-3 cells, detached and incubated either with PC or APC, in fibrin polymerization curve of normal plasma is shown. The same experiment was also performed using OVCAR-3 cells in adherent conditions. When both detached (Fig. 5B) and adherent (Fig. 5C) cells, were incubated with aPC, aPTT was prolonged as compared to cells incubated with protein C. As documented in Fig. 5D, PC added to the adherent or detached ovarian cancer cells induced a 2-fold increase in cephalin clotting time, whereas aPC under the same conditions induced a 3.7-fold increase (5.78–11.90 min for PC and 21.42 min for aPC). These results indicate the anti-coagulant action of EPCR-aPC on the OVCAR-3 cells.

Bottom Line: Our results showed that in the OVCAR-3 aPC-induced cells i) an increase in cell migration was noted, which was inhibited when anti-endothelial protein C receptor (EPCR) was added to the culture medium and which may act via MEK-ERK and Rho-GTPase pathways; ii) an increase in threonine, and to a lesser extent tyrosine phosphorylation; iii) cell cycle activation (G1 to S/G2); and iv) a 2-3-fold prolongation of aPTT of normal plasma.In the peritoneal fluid, the sEPCR concentration was 71 ± 23 ng/ml.This binding could also explain the loss of clotting of peritoneal fluids.

View Article: PubMed Central - PubMed

Affiliation: UMR, Paris Diderot, Paris 7 University, Lariboisiere Hospital, INSERM U965, Paris, France.

ABSTRACT
The objective of this study was to evaluate the role of activated protein C (aPC), known to be a physiological anticoagulant, in ovarian cancer cell activation as well as in loss of clotting of cancer ascitic fluid. The effect of aPC on an ovarian cancer cell line (OVCAR-3) was tested in regards to i) cell migration and adhesion with the use of adhesion and wound healing assays as well as a droplet test; ii) protein phosphorylation, evaluated by cyto-ELISA; iii) cell cycle modification assessed by flow cytometric DNA quantification; and iv) anticoagulant activity evaluated by the prolongation of partial thromboplastin time (aPTT) of normal plasma in the presence or absence of aPC-treated ovarian cancer cells. In addition, the soluble endothelial protein C receptor (sEPCR) was quantified by ELISA in ascitic fluid of patients with ovarian cancer. Our results showed that in the OVCAR-3 aPC-induced cells i) an increase in cell migration was noted, which was inhibited when anti-endothelial protein C receptor (EPCR) was added to the culture medium and which may act via MEK-ERK and Rho-GTPase pathways; ii) an increase in threonine, and to a lesser extent tyrosine phosphorylation; iii) cell cycle activation (G1 to S/G2); and iv) a 2-3-fold prolongation of aPTT of normal plasma. In the peritoneal fluid, the sEPCR concentration was 71 ± 23 ng/ml. In conclusion, free aPC binds to membrane EPCR in ovarian cancer cells and induces cell migration via MEK-ERK and Rho-GTPase pathways. This binding could also explain the loss of clotting of peritoneal fluids.

No MeSH data available.


Related in: MedlinePlus