Establishment and characterization of immortalized bovine male germline stem cell line

Abstract

Male germline stem cells (mGSCs) are unique adult germ cells with self-renewal potential and spermatogenesis function in the testis. However, further studies are needed to establish a long-term cultural system of mGSCs in vitro, especially for large animals such as bovine mGSCs. In this study, we first established a stable immortalized bovine male germline stem cell line by transducing Simian virus 40 (SV40) large T antigen. The proliferation of these cells was improved significantly. These cells could express spermatogonial stem cell (SSC)-specific markers, such as PLZF, PGP9.5, VASA, LIN28A, and CD49F, both in the mRNA and protein levels. Additionally, these cells could be differentiated into three germ layer cells to enter meiosis, form colonies, and proliferate in the seminiferous tubules of busulfan-induced infertile mice. The immortalized bovine mGSCs maintain the criteria of mGSCs.

Analysis of Long Non-Coding RNA and mRNA Expression Profiling in Immature and Mature Bovine (Bos taurus) Testes

“…These two types of cells were cultured in DMEM/F12 (HyClone), supplemented with 10% FBS (Gibco), 100 U/ml penicillin, and 100 μg/ml streptomycin (HyClone), at 37°C in a 5% CO 2 atmosphere. The Bovine mGSCs (male germline stem cells) were derived from an immortalized cell line (Lei et al, 2017). The cells were harvested for RT-qPCR, and the primer details were shown in Table S14 .…”

Reelin regulates male mouse reproductive capacity via the sertoli cells

“…Reeler mice exhibit impaired motor coordination, tremors, and ataxia; these conditions are attributed to the abnormal localization of neurons in the brain during development . Reelin is not only involved in the regulation of neurodevelopmental disorders, but also controls the development, maintenance, and plasticity of neural networks . However, the function of RELN in other types of tissues or cells, especially those of the sperm, is investigated, although one of its receptors ApoER2 plays a crucial role in the sperm development .…”

“…20 Reelin is not only involved in the regulation of neurodevelopmental disorders, 21 but also controls the development, maintenance, and plasticity of neural networks. 22 However, the function of RELN in other types of tissues or cells, especially those of the sperm, is investigated, although one of its receptors ApoER2 plays a crucial role in the sperm development. 5 Meanwhile, the mechanism by which Reelin regulates the productive capacity of male mammals remains unidentified.…”

Culture bovine prospermatogonia with 2i medium

“…During the last decade, efforts have been made to optimise the culture system of bovine spermatogonia from different aspects including media and serum (Izadyar et al., 2003), feeder cells (Nasiri et al., 2012; Oatley et al., 2004), extracellular matrix (Akbarinejad et al., 2015; Kim et al., 2014), hormones (Narenji Sani et al., 2013) and growth factors (Aponte et al., 2006, 2008; Sahare et al., 2016). Long‐term maintenance of bovine undifferentiated spermatogonia (Oatley et al., 2016) and generation of undifferentiated spermatogonial cell lines from bulls were also reported (Lei et al, 2017; Suyatno et al., 2018). Nevertheless, the culture system of bovine undifferentiated spermatogonia still needs to be improved.…”