Expression levels and stoichiometry of Hnf1β,Emx2,Pax8 and Hnf4αinfluence direct reprogramming of induced renal tubular epithelial cells  

作　　者：Xueli Hu Jianjian Sun Meng Wan Bianhong Zhang Linhui Wang Tao P.Zhong 

机构地区：[1]Shanghai Key Laboratory of Regulatory Biology,Institute of Molecular Medicine,School of Life Sciences,East China Normal University,Shanghai 200241,China [2]Guangdong Cardiovascular Institute,Guangdong Provincial People’s Hospital,Guangdong Academy of Medical Sciences,Guangzhou,Guangdong 510100,China [3]Department of Urology,Changhai Hospital,Shanghai 200433,China

出　　处：《Cell Regeneration》2024年第1期180-191,共12页细胞再生(英文)

基　　金：Ministry of Science and Technology of the People’s Republic of China(2018YFA0800103,2018YFA0801004);National Natural Science Foundation of China(NSFC31530044,NSFC31970780,NSFC82202056).

摘　　要：Generation of induced renal epithelial cells(iRECs)from fibroblasts offers great opportunities for renal disease mod-eling and kidney regeneration.However,the low reprogramming efficiency of the current approach to generate iRECs has hindered potential therapeutic application and regenerative approach.This could be in part attributed to het-erogeneous and unbalanced expression of reprogramming factors(RFs)Hnf1β(H1),Emx2(E),Pax8(P),and Hnf4α(H4)in transduced fibroblasts.Here,we establish an advanced retroviral vector system that expresses H1,E,P,and H4 in high levels and distinct ratios from bicistronic transcripts separated by P2A.Mouse embryonic fibroblasts(MEFs)harboring Cdh16-Cre;mT/mG allele are utilized to conduct iREC reprogramming via directly monitoring single cell fate conversion.Three sets of bicistronic RF combinations including H1E/H4P,H1H4/EP,and H1P/H4E have been generated to induce iREC reprogramming.Each of the RF combinations gives rise to distinct H1,E,P,and H4 expres-sion levels and different reprogramming efficiencies.The desired H1E/H4P combination that results in high expres-sion levels of RFs with balanced stoichiometry.substantially enhances the efficiency and quality of iRECs compared with transduction of separate H1,E,P,and H4 lentiviruses.We find that H1E/H4P-induced iRECs exhibit the superior features of renal tubular epithelial cells,as evidenced by expressing renal tubular-specific genes,possessing endocy-totic arrogation activity and assembling into tubules along decellularized kidney scaffolds.This study establishes H1E/H4P cassette as a valuable platform for future iREC studies and regenerative medicine.

关 键 词：iRECs Bicistronic retroviral cassettes Renal transcriptional factors 

分 类 号：R73[医药卫生—肿瘤]