The Effects Of Chitosan Film Culture On The Expression Of IL-4 And IL-10 In MSCs

Objective: To investigate the effects of chitosan film culture and conventional plate culture on the expression of anti-inflammatory related genes such as IL-4 and IL-10 in mesenchymal stem cells(MSCs).Methods : In May 2017,umbilical cord specimens of three full-term normal parturients were collected from the Department of Obstetrics and Gynecology,Renmin Hospital of Wuhan University.The primary MSCs were isolated from umbilical cord tissue.The membrane substrates were prepared with chitosan powder as the MSCs culture substrate.And then the primary MSCs were inoculated on the chitosan film and the traditional polyethylene culture plate respectively.After 72 hours,two groups of cells were collected.The expression of LIF,IL6 ST,IL-10,TSG6,COX-2,STC-1,IL-1RN,IL-4,IL-13 m RNA and their proteins were detected by Real-time PCR,Western blot and immunofluorescence.Results: Compared with the traditional two-dimensional adherent culture method,the MSCs cultured with chitosan film grow in suspension,and the cell volume is obviously smaller,which can aggregate to form 3D multicellular sphere structure.The expression levels of anti-inflammatory related genes LIF,IL6 ST,IL-10,TSG6,COX-2,STC-1,IL-1RN,IL-4 and IL-13 m RNA and their proteins in chitosan film-cultured MSCs were significantly higher than those in the conventional culture group,and the difference was statistically significant(P<0.01).Conclusion: Compared with MSCs in conventional culture,MSCs cultured by chitosan film could form multicellular spheres,and the m RNA and protein expression of anti-inflammatory genes in 3D spheres were significantly increased.