Generation and characteristics of human Sertoli cell line immortalized by overexpression of human telomerase

Abstract

Sertoli cells are required for normal spermatogenesis and they can be reprogrammed to other types of functional cells. However, the number of primary Sertoli cells is rare and human Sertoli cell line is unavailable. In this study, we have for the first time reported a stable human Sertoli cell line, namely hS1 cells, by overexpression of human telomerase. The hS1 cells expressed a number of hallmarks for human Sertoli cells, including SOX9, WT1, GDNF, SCF, BMP4, BMP6, GATA4, and VIM, and they were negative for 3β-HSD, SMA, and VASA. Higher levels of AR and FSHR were observed in hS1 cells compared to primary human Sertoli cells. Microarray analysis showed that 70.4% of global gene profiles of hS1 cells were similar to primary human Sertoli cells. Proliferation assay demonstrated that hS1 cells proliferated rapidly and they could be passaged for more than 30 times in 6 months. Neither Y chromosome microdeletion nor tumorgenesis was detected in this cell line and 90% normal karyotypes existed in hS1 cells. Collectively, we have established the first human Sertoli cell line with phenotype of primary human Sertoli cells, an unlimited proliferation potential and high safety, which could offer sufficient human Sertoli cells for basic research as well as reproductive and regenerative medicine.

Generation of a hTERT-Immortalized Human Sertoli Cell Model to Study Transporter Dynamics at the Blood-Testis Barrier

“…The hT-SerCs described in this study were shown to stably overexpress TERT at the transcriptional and translational level compared to the minor expression in primary human SCs. In addition, this cell line retained similar morphological characteristics to the primary human SCs from which they were derived, and cells described in previous studies ( Figure 1 B,C) [ 28 , 29 , 30 , 31 ]. The primary human SCs have been well characterized [ 28 , 29 ], thus presenting a convenient baseline to compare the phenotype of the hT-SerCs.…”

“…Moreover, it has long been thought that SCs cease to proliferate after puberty [ 24 , 25 , 26 , 27 ], thus prompting many studies to isolate and use prepubertal rat SCs. However, recent evidence for the existence of proliferative human SCs post-puberty has challenged this idea [ 28 , 29 , 30 , 31 ]. Although proliferative human SCs can be isolated, the growth of these primary cells is constrained by the Hayflick limit, which defines the maximum number of divisions that a cell can undergo before entering senescence due to critical telomere shortening [ 32 , 33 ].…”

“…This study characterizes a unique, human telomerase reverse transcriptase-immortalized human SC line (hT-SerC) developed for use in studies that evaluate transporter-substrate selectivity at the BTB. Only one other immortalized human SC line (hS1) has been reported; however, the hS1 cell line is not publicly available and its use in transport studies has not been described [ 31 ]. The hT-SerC line described in this study exhibited similar phenotypic characteristics to the primary human SCs from which it was derived and has significantly increased replicative capacity.…”

Induction of Sertoli-like cells from human fibroblasts by NR5A1 and GATA4

“…To determine whether hiSCs reprogrammed with the 5TFs (5F-hiSCs) are similar to human Sertoli cells, we compared the transcriptomes of the AMH:EGFP+ 5F-hiSCs, dH1 cells infected with p2k7 empty virus (dH1-2K7) as negative controls, and primary adult Sertoli cells (aSCs) from human biopsy samples (Wen et al, 2017). We focused our analysis on the differentially expressed genes (DEGs,>2 fold change, p-value<0.05) between 5F-hiSCs and dH1-2K7 and between aSCs and dH1-2K7.…”

“…Primary human skin fibroblast (HSF) was a gift from Professor Ting Chen at National Institute of Biological Sciences, Beijing, and prepared according to the published protocol (Qian et al, 2018). Human adult Sertoli cells were a generous gift from Professor Zheng Li at Shanghai Jiao Tong University and prepared according to the established protocol (Wen et al, 2017). The HPF and HSF cells were maintained at 75 ml culture flask in 15 ml DMEM culture medium (Corning) containing 10% FBS (Gibco), 0.1 mM nonessential amino acids and 1 mM L-glutamine.…”