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Genomics and Proteomics of the Reproductive System

(T539) EXPRESSION PROFILING OF GENES IN BeWo CELLS FOLLOWING FORSKOLIN INDUCED DIFFERENTIATION.

Rao, A J 1, Neelima , PS1, Yashwanth , R1, Anbalgan , M1, Mahesh , K1, Dahia, Chitra Lekha1, Rao, Rekha M1, 1 Indian Institute of Science, Bangalore, India

ABSTRACT- Differentiation of the BeWo cell line derived from human choriocarcinoma has been widely used as an in vitro model for studying trophoblast fusion and differentiation. In the present study, we have monitored gene expression changes in differentiating BeWo cells using differential display RT-PCR and microarray analysis. Following 72 hrs of 10 M forskolin treatment, DD RT-PCR analysis showed the differential regulation of several transcripts. The differential expression of these transcripts was also confirmed by RT-PCR and northern blot analysis. Microarray analysis revealed the up-regulation of hCG, hCS, Secretory Leucocyte Protease Inhibitor (SLPI) and down regulation of PCNA and Cyclin D1, thus validating the experimental system. Furthermore, several candidate genes that could influence trophoblast differentiation, cell adhesion and cellular proliferation were identified. Genes involved in cellular proliferation include cyclin M3, replication factor 3, signal-induced proliferation-associated gene 1, osteonectin, clusterin, etc clearly indicating a growth-arrested phenotype for the differentiating BeWo cells. Genes that regulate the invasive behaviour of trophoblasts include MMP2, cathepsin K, cystatin B, SLPI and cysteine-rich angiogenic inducer 61, etc. The co-ordinated regulation of proteases and protease inhibitors suggests that these genes play an important role under in vivo conditions in the regulation of trophoblast invasion at the uterine-placenta interface in vivo. Trophoblastic differentiation associated genes included adipose differentiation-related protein, GADD45A, PPAR binding protein, galectin 3, tubulins, collagen, stathmin, etc. These results suggest the usefulness of this model to understand the molecular mechanism of proliferation, invasion and differentiation of placental cells. (Supported by CSIR, ICMR, DST, DBT, Govt. of India; Mellon and Conrad Foundation, USA; and CSIR Emeritus Scientist Fellowship to A.J. Rao).

KEY WORDS: differentiation, BeWo cells, forskolin, microarray



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