Direct conversion of fibroblasts into urothelial cells that may be recruited to regenerating mucosa of injured urinary bladder

Yuta Inoue,Osamu Ukimura,Hideto Taga,Shin-Ichiro Kotani,Makoto Seki,Osam Mazda,Mika Akiyoshi,Tsunao Kishida

doi:10.1038/s41598-019-50388-6

Yuta Inoue, Osamu Ukimura + Show 6 more

Open Access

https://doi.org/10.1038/s41598-019-50388-6

Copy DOI

Journal: Scientific Reports	Publication Date: Sep 25, 2019
Citations: 12	License type: open-access

Affiliation: Kyoto Prefectural University of Medicine

Abstract

Urothelial cells play essential roles in protection of urine exudation and bacterial invasion at the urothelial mucosa, so that defect or damage of urothelial cells associated with urinary tract diseases may cause serious problems. If a sufficient number of functional urothelial cells are prepared in culture and transplanted into the damaged urothelial lesions, such technology may provide beneficial effects to patients with diseases of the urinary tract. Here we found that human adult dermal fibroblasts were converted into urothelial cells by transducing genes for four transcription factors, FOXA1, TP63, MYCL and KLF4 (FTLK). The directly converted urothelial cells (dUCs) formed cobblestone-like colonies and expressed urothelium-specific markers. dUCs were successfully expanded and enriched after serial passages using a specific medium that we optimized for the cells. The passaged dUCs showed similar genome-wide gene expression profiles to normal urothelial cells and had a barrier function. The FTLK-transduced fibroblasts were also converted into urothelial cells in vivo and recruited to the regenerating urothelial tissue after they were transplanted into the bladder of mice with interstitial cystitis. Our technology may provide a promising solution for a number of patients with urinary tract disorders.

Highlights

Urothelial cells form urothelium on the surface of the urinary tract that is composed of renal pelvis, ureter, urinary bladder and proximal urethra
We considered that it might be promising if urothelial cells were generated by the direct conversion technology that induces differentiated tissue cells from another somatic cell lineage without passing through a pluripotent state
It was found that FIT (FOXA1, IRF1 and TP63) and FIH (FOXA1, IRF1 and SHH) induced only a low level of expression of uroplakin 1b (UPK1b) that is a typical urothelial developmental marker[21] (Supplementary Fig. 1A), and neither FIT- nor FIH- transduced cells formed colonies with epithelium-like appearance (Supplementary Fig. 1B)

Summary

Introduction

Urothelial cells form urothelium on the surface of the urinary tract that is composed of renal pelvis, ureter, urinary bladder and proximal urethra. Damaged urothelium in patients could be treated by transplantation of autologous intestine or colon tissue segments, but these gastrointestinal tissues resorb urine, causing many problems including cancer formation[1], metabolic acidosis, infection, stone formation, etc[2] In this context, a lot of efforts have been devoted to search for another source of the cells applicable to transplantation instead of gastrointestinal tissue segments. Transplantation of cells derived from human pluripotent stem cells could cause teratoma formation We considered that it might be promising if urothelial cells were generated by the direct conversion (direct reprogramming) technology that induces differentiated tissue cells from another somatic cell lineage without passing through a pluripotent state. An addition of genes that promote reprogramming of somatic cells into iPS cells may promote direct conversion of some cell types In this context, POU5F1, KLF4 and MYC played crucial roles in directly converting fibroblasts into neural stem cells[11] and chondrogenic cells[12]. We established a procedure of the conversion, and analyzed phenotypes and functions of the resultant directly converted urothelial cells (dUCs)

Methods

Results

Conclusion