ABSTRACT
Introduction: Nowadays, spermatogonial stem cells [SSCs] cultivation has been used by many researchers as an effective tool for infertility treatments. Oxidative conditions can be effective on cell proliferation and differentiation of these cells. So, the aim of this study was to establish oxidative stress model for antioxidant activity of some drugs investigation during SSCs in vitro culture
Methods: Neonatal NMRI male mice [3-5 day] were used for isolation of SSCs. The cell suspension was prepared by twice enzymatic digestion. The cell suspension contents were spermatogonial and sertoli cells and treated by different doses of H2O2 logarithmic concentrations from 0-100 and muM after 24 hours. To access the optimal dose, extra doses from 10-100 and muM was evaluated. After 2 hours of H2O2 treatment, viability was determined by Trypan blue assay. The data were analyzed using SPSS software and One-way ANOVA test
Results: Our data showed that spermatogonial stem cells colonies appeared after 4 days of isolation. These cells expressed OCT4 and PLZF proteins. Many of spermatogonial stem cells were removed after using higher doses of H2O2. The results showed that 30 and muM concentration of H2O2 could induce oxidative stress in spermatogonial stem cell during in vitro culture
Conclusion: According to this study, 30 and muM concentration of H2O2 can cause cell death lower than 50% of total number of cells and can increase oxidative stress in cultivation of SSCs. This model is a suitable tool for studying some new antioxidant drugs
ABSTRACT
Catsper proteins are responsible for entering Ca[2+] to the cell and play an important role in sperm motility and male fertility. Antioxidants are vital for sperm motility too. Escanbil [Calligonum] extract possess some of the important antioxidant like Catechin and Quercetin. Here we investigated the effects of Escanbil [Calligonum] extract on the sperm parameters and the expressing of Catsper gene in aging male mice. In this animal study, firstly, dose response was performed by using these three doses of Escanbil [Calligonum] [10, 30 and 50 mg/kg]. 5 mice in each group were considered and Intra Peritoneal injection was done for 5 weeks. the sperm parameters analyzed and Terminal deoxynucleotidyl transferase dUTP nick end labeling [TUNEL]staining was done. 30 mg/kg dose was considered as optimum dose. Secondly: fifteen aging male mice [11-13 months] were divided into three groups: control, sham and experiment. The experiments were injected Intra peritonealy with Escanbil [Calligonum] extract [30mg/kg] weekly for up to 5 weeks. The sham group was injected Intra Peritoneal [DMSO]. Sperm parameters were analyzed. Expression of Catsper genes was analyzed by Real time PCR. Our results showed that after Escanbil [Calligonum] treatment [30 mg/kg], the sperm parameters were improved in experimental group [p<0.05]. Our data showed that there was a statistical significance difference between the expressions of Catsper 2, 4 in aging experiment group comparison with aging control group [p<0.05]. We investigated that the Escanbil [Calligonum] extract [30 mg/kg] can improve sperm parameters and change the expression of Catsper genes in aging male mice. This herbal extract can be used as an antioxidant component for clinical usages