Publication: Involvement of mapk/erk and akt siganling pathways in the proliferation of keratinocytes cocultured with adipose-derived stem cells (AZSS)porous chitosan scaffold (PSC)
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Date
2017
Authors
Halim, Ahmad Sukari
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Abstract
Secretory factors of adipose-derived stem cells have paracrine effects that may have potential in accelerating proliferation of keratinocytes by coculturing both of the cells. This study was conducted to evaluate the indirect effects of ASCs-PCS on the growth of keratinocytes by paracrine activities. Human adipose-derived stem cells and human epidermal keratinocytes were isolated and verified with their specific markers. Distribution and proliferation of ASCs on porous chitosan scaffold were assessed by scanning electron microscope, live/dead assay and Alamar blue assay. Coculture groups were divided into two groups; culture ASCs and culture of ASCs-PCS. Proliferation of keratinocytes in the coculture wells were analysed by Alamar blue assay at 24 hours and 72 hours of coculture while the concentration human epidermal growth factor in culture supernatant were quantified at 72 hours of coculture. Adipose-derived stem cells culture have been established and displayed high expression of CD29, CD73, CD90, CD105 and low expression of CD34. Keratinocytes culture expressed involucrin and cytokeratin 6 as demonstrated by immunocytochemistry. Percentage of proliferation of ASCs within PCS was more than 80% accompanied with spindle-shaped features and good viability. After 72 hours of coculture, the percentage of nHEK in the coculture group was increased due to growth medium. ELISA assay revealed no secretion of EGF from ASCs. This study demonstrated via dual -chamber coculture model, human -derived ASCs and PCS showed no growth stimulatory effect towards the growth of keratinocytes via the tested growth factor, EGF. The proliferation of nHEK in the coculture wells possibly due to other growth factor or other mechanism.