Memorial University of Newfoundland
Graduate Student, Faculty of Medicine
Thesis Title: The Role of Poly (ADP-ribose) Polymerase-1 in Human Trophoblast Differentiation.
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Dr. Daniel MacPhee
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About
The Role of Poly (ADP-Ribose) Polymerase – 1 (PARP-1) in Human Trophoblast Differentiation
1Pater, JA. and 1MacPhee, DJ.
1Memorial University of Newfoundland, St. John’s, NL, CA
Objective: In the fusion pathway of early human trophoblast differentiation, progenitor cytotrophoblast cells within floating chorionic villi proliferate and differentiate by syncytialization to form the overlying multi-nucleated syncytiotrophoblast layer. However, the mechanisms that control these processes are not completely understood. Previous studies conducted in our lab have demonstrated that Poly (ADP-Ribose) Polymerase – 1 (PARP-1), exhibits increased expression as synctialization occurs. Furthermore, PARP-1’s spatial and temporal expression in first trimester human chorionic villi was characterized and was found to be highly expressed in the nucleus of villous cytotrophoblast cells but scarcely detected in the syncytiotrophoblast. Our objective was to examine the role of PARP-1 in trophoblast differentiation.
Methods: The BeWo cell line model will be used in this study because it is a documented model for the study of trophoblast differentiation. BeWo cells will be cultivated in F-12 Ham’s media, however, in fusion assays the cells will be cultured in Ham’s F-12K media containing 25 µM forskolin (fusion media). In our experiments cells will be transfected with either a P3X-FLAG-PARP-1-DNA or Dharmacon PARP-1 ON-TARGETplus siRNA, which overexpresses and silences PARP-1, respectively. Moreover, cells will be treated with a PARP-1 catalytic inhibitor PJ34 to explore the role of PARP-1 catalytic activity in trophoblast differentiation. The transfected and drug treated cells will then be cultured in fusion media. The occurrence of BeWo cell differentiation in transfected and drug treated cells will then be analyzed through western blotting and fluorescence microscopy for the absence or presence of E-cadherin immunolocalization.
