Modulation of differentiation phenotypes of immune/muscle endoderm and neural ectoderm from CHO and P19 cells using soluble factors combined with extracellular matrix.
Authors:Faustino Becerra, Madonna Chico, Lisa Sobelman
Mentor:Dr. James Harber, Professor of Microbiology, Oxnard College
Clinical induced pluripotent stem cell (iPS) technology promises that clinical autotransplantation in a variety of tissues is feasible including previously impracticable organs (including heart, brain and teeth). This project’s goal was to create the three different germ layers (endoderm, mesoderm, and exoderm) from P19 (EC mouse tetracarcinoma) and mesoderm from CHO (chinese hamster ovary fibroblast). In this study, these two cell lines were used with specific factors to further develop an assay for the identification of genes and factors related to cell differentiation and proliferation. Previous tissue regeneration research in this lab showed that CHO cells could display a simple phenotypic morphological change resembling that of the extravasation phase of the immune system response when observed on an extracellular fibrin-fibronectin matrix (FFM), a biomaterial that resembles blood clots. The current study aimed to further knowledge of cell differentiation and movement, in vitro, using a phenotypic based-assay with the following presence/absence conditions in combination: FFM, RA and DMSO. CHO and P19 were observed in single experimental flasks for as long as a month, enough time to see dramatic morphological changes in CHO (parallel alignment of the cells, or cell axis polarity alterations) and in P19, phenotypic departures from its well known cellular differentiation behavior to neuron or cardiac cells. CHO cells also created triangular sail-like tissue connections between several FFM “posts” that were installed in geometric patterns in the tissue culture dishes. The cellular changes that result from treatment with factors (FFM, DMSO, RA) in various configurations implicate signal transduction through the WNT pathway and other ligand driven receptor-dimer pathways in combination. The WNT protein signals transcription of genes affiliated with cell axis polarity and neuron formation. Identifying the genes associated with the observed cell movements would be aided by probing the assays with specific signal transduction inhibitors or the microarray.