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Déborah Reynaud

Role of NLRP7 protein in normal and tumor pregnancies

Published on 21 December 2018
Thesis presented December 21, 2018

Abstract:
Complete hydatidiform moles (CHM) are benign precancerous lesions of the placenta that evolve in 5% of cases into a highly proliferative cancer called choriocarcinoma (CC). Numerous studies have reported correlations between the development of recurrent CHM and mutations in the NLRP7 gene. NLRP7 protein belongs the NLRP7-inflammasome, whose activation contributes to the production of mature IL-1ß and IL-18. Most of the work published on NLRP7 was focused on the study of NLRP7 mutations in CHM. Though, no study has characterized its role in normal and tumor placental development. The aim of my thesis project was to characterize the role of this protein in the normal and tumor placenta. Three approaches were used, i) A clinical approach, in collaboration with Casablanca University Hospital; the French reference center of Trophoblastic Gestational Disease and with McGill University. These collaborations allowed for tissue access from MHC and CC patients; ii) An in vitro / Ex vivo approach for the characterization of the role of NLRP7 in key processes of normal and tumor placental development using 2D and 3D culture systems. Two trophoblastic cell types were used, a non-tumor cell line, the HTR-8 Sv/Neo and the JEG3 cells, derived from human CC; iii) An in vivo approach through orthotopic injection of JEG3 cells, invalidated or not for the expression of the NLRP7 gene (ShRNA strategy), in the placenta of gravid mice. The tumor impact of JEG3 following their injection into the uterine horn and the vein of the tail of non-pregnant mice was also examined.The first part of my work showed that NLRP7 protein is abundantly expressed in the normal placenta during the first trimester of pregnancy, that its expression is upregulated by hypoxia, a key parameter in placental development, and that this protein controls key processes of placental development such as proliferation and differentiation. Importantly, I have also demonstrated that NLRP7 plays an important compensatory role in the pathology of intrauterine growth retardation. The second part of my work concerning the role of NLRP7 protein in the placental tumor development demonstrated that i) NLRP7 protein levels are increased in the placenta of MHC and CC patients, and that components of the inflammasome machinery are also deregulated, ii) the JEG3 cells overexpress NLRP7 compared to HTR-8 Sv/Neo, iii) NLRP7 knock-down in JEG3 induced a significant decrease in their proliferation and an increase in their migration and invasion both in the 2D and 3D culture systems. The in vivo study demonstrated that the knock-down of NLRP7 decreased the development and metastasis of human CC in the three tested routes. Immunohistological, RNAseq and antibody-array analyses allowed the characterization of the pathways regulated by the NLRP7 protein in JEG3 cells.Altogether my PhD project characterized the critical role of the NLRP7 protein in normal and tumor placental development and proposes the NLRP7 machinery as a potential target for CC treatment.

Keywords:
Placenta, Hydatiform moles, Choriocarcinoma, Tumor invasion, Metastasis, Nlrp7

On-line thesis.