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Description
Introduction: Cell culturing of human cells used in oral mucosa and cornea tissue engineering can be optimized by using three-dimensional cell culture platforms1. In general, these systems comprise an extracellular matrix where cells can be cultured in a 3D spatial microenvironment able to simulate the physiological conditions in which cells are found in vivo, including cell-cell and cell-extracellular matrix interactions2. These interactions are essential for mesenchymal stem cells development, whose therapeutic effect is directly related to their paracrine activity. The aim of this study is to evaluate the impact of these systems on cell behaviour of human umbilical cord Wharton’s Jelly Stem Cells (hWJSC).
Methodology: hWJSC were isolated and cultured in 2D conditions (control cells cultured in routine culture medium) and in a 3D system consisting of a part of culture medium and a part of an extracellular matrix (ECM) generated in the laboratory. Different study groups containing increasing concentrations of ECM (25%, 50%, 75% and 100%) were studied. Cells were cultured in each condition were evaluated after 7 days of culture at 37ºC with 5% CO2. Then, total RNA was isolated from each experimental condition and retrotranscribed to cDNA. Real-time qRT-PCR was performed, and the relative expression levels of several genes involved in apoptotic cell death and cell differentiation was determined.
Results: First, we found that hWJSC were able to grow and proliferate in the 3D platform, and their morphology and histological profile was similar to cells cultured in 2D systems. Then, the use of real-time qRT-PCR revealed that the gene expression profile of both types of cells was very similar, with very low expression of caspases and other genes related to cell death by apoptosis at the mRNA level. In addition, the highest concentrations of ECM used in the 3D systems was associated to an increment in certain genes related to cell undifferentiation.
Conclusions: In general, these results suggest that the 3D platform was able to support cell growth and proliferation, and that hWJSC cultured in these systems kept high levels of cell viability and cells maintained their undifferentiated status. These results support the use of ECM-based 3D systems for culturing hWJSC for oral mucosa and cornea tissue engineering purposes.
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