Biochemical and Biophysical Research Communications
Stromal interaction molecule 1 is required for neonatal testicular development in mice
Introduction
Spermatogenesis involves a complex series of cellular changes leading to the formation of haploid spermatozoa, including mitotic, meiotic and post-meiotic phases [1]. Sertoli cells play a central role in the development of a functional testis. Sertoli cells are the first cells to differentiate recognizably in the indifferent fetal gonad, an event which enables seminiferous cord formation, and, during early postnatal development, Sertoli cells continue to proliferate, differentiate, and mature as nurse-like cells [2]. Successful spermatogenesis relies on these early events in Sertoli cells because abnormal Sertoli cell proliferation or differentiation can disrupt fertility [3,4].
Stromal interaction molecule 1 (STIM1), is a highly conserved type-I membrane, ER-resident protein, and has been well known containing a luminal EF-hand Ca2+-binding domain and several cytosolic protein-protein interaction domains, and it acts as a Ca2+ sensor and mediates store-operated Ca2+ entry(SOCE) and oxidative stress [[5], [6], [7], [8]]. Global deletion of Stim1 gene in mice is lethal, indicating that STIM1 is indispensable in the organismal physiology of mammals [9]. In our previous study, we found that STIM1 was predominantly expressed in the cytoplasm of Sertoli cells in fetal mice testis, and knockdown of Stim1 gene in fetal testis caused severe disruption in testicular cord development, which was associated with the aberrant oxidative stress [10]. We also successfully identified STIM1 in the proteome profile for neonatal mice testis in our previous research [11], and the bioinformatics analysis showed that STIM1 may play a vital role in testicular development. In the present study, we aimed to study the distribution of STIM1 in mice neonatal testis, and study the effects of its knockdown in vitro.
Section snippets
Animals
Pregnant ICR mice were maintained in a controlled environment under a 12/12-h light/dark cycle at 20–22 °C and 50–70% humidity with food and water available ad libitum. All experiments on mice were approved by the Animal Ethics Committee of Nanjing Medical University.
Western blot analysis
Western blot analysis was performed as described previously, with minor modifications [12]. Briefly, testis lysates were separated by electrophoresis, then transferred to polyvinylidene difluoride (PVDF) membranes (Bio-Rad,
STIM1 expression in neonatal mice testes
We used immunostaining to analyze the distribution of STIM1 in neonatal mice testes. The seminiferous epithelium of newborn mouse testis contains two distinct cell types: germ and Sertoli cells. As shown in Fig. 1, in the seminiferous epithelium of 0.5-day testis, gonocytes are evident in the center of the cords with a spherical nucleus, while in the 7.5-day testis, the gonocytes have migrated to the basement membrane of the seminiferous tubule and differentiated into Spermatogonial Stem cells
Discussion
Here, we report the distribution and function of STIM1 in mammalian neonatal testis. We found that STIM1 was primarily located in the Sertoli cells of mice neonatal testes. Knockdown of Stim1 gene by morpholino in neonatal testicular culture resulted in disrupted Sertoli cell orientation, reduced germ cell survival, marked oxidative stress, and WNT/β-catenin pathway activation. Furthermore, treatment with the antioxidant scavenger, COQ10, resulted in partial rescue of these phenotypes. Overall,
Conflicts of interest (competing financial interests)
The authors have no conflicts of interest to disclose.
Acknowledgments
This work was supported by the Suzhou Key Medical Center (SZZX201505), Jiangsu Provincial Medical Innovation Team (CXTDB2017013), Suzhou Introduced Project of Clinical Medical Expert Team (SZYJTD201708), Suzhou Key Laboratory of male reproduction research (SZS201718), Key Research Fund for Zhenjiang Social Development (SH2016028), Key Research Fund for Zhenjiang Health Science and Technology (SHW2016001), National Natural Science Foundation of China (31701298), Natural Science Foundation of
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These authors contributed equally to the work.