The Vitro Study Of Induced Pluripotent Stem Cell Which Transfected With Antimicrobial Peptide LL37

Objectives: In this study,we constructed the pGC-FU-LL37-GFP lentivirus and transfected the iPSCs.After tested the transfected iPSCs' ability of promoting angiogenesis,we induced the iPSCs to differentiate into Urothelial cells.Methods:(1)Extracting the target gene of antimicrobial peptide LL37;(2)Constructing and sequencing the identified antimicrobial peptide LL37 into lentivirus vector;(3)iPSCs' transfected experiment and identification;(4)Cell counting,migration and angioplasty experiment were used to confirm the angiogenic and repair effects of the transgenic iPSCs;(5)iPSCs were induced to differentiate into urothelial cells,and the differentiated cells were identified by qPCR;GraphPad Prism 8 software was used to draw statistical results,and IBM SPSS 25.0 statistical analysis software was used for data processing.Results:(1)qPCR confirmed the pCG-FU-LL37-GFP built successfully,and the sequencing results was consistent with the target sequence completely;(2)The target geen was successfully mixed the GFP and expressed stably;(3)The qPCR results indicate LL37 stably expressed in the mRNA level(P<0.01);(4)Antibacterial peptide LL37 was stably expressed in protein level by testing the cells' culture(P<0.05);(5)The transgenic iPSCs' supernatant successfully promoted the quantities of HUVECs after compared with the control group;(6)The migration experiments antibacterial peptide LL37 can effectively shorten the scratches and accelerate wound healing;(7)Angioplasty experiment showed antibacterial peptide LL37 can accelerate the ability of HUVECs' forming vessel tube;(8)The transfected iPSCs were induced to differentiated to epithelioid cells.qPCR and western blot results indicated the Uroplakin III expressed up-regulated.Conclusions:(1)The pGC-FU-LL37-GFP was constructed sucessfully and expressed the target antimicrobial LL37;(2)Transfected iPSCs can promote the HUVECs proliferation,migration and angiogenesis of vascular;(3)The transfected iPSCs were successfully differentiated to urothelial cells.Significance: Our research provided a new direction for the acquisition of urothelium cells.