The effect of Low-intensity Pulsed Ultrasound Stimulation on Neonate Mouse Spermatogonial Stem Cells

This study presents an efficient, cost-effective method to improve proliferation and colonization of spermatogonial stem cells (SSCs) in vitro. Isolated SSCs from neonate mice were cultured in DMEM culture medium with 10% fetal bovine serum (FBS). In the first phase of the study, the temperature was controlled by low intensity pulsed ultrasound stimulation (LIPUS) of the plate that contained the culture medium. In the next phase, SSCs were stimulated by LIPUS with 200 mW/cm2 with 20% and 40% duty cycle for five days. Proliferation and colonization of SSCs were on the seventh day. LIPUS treatment of mouse SSCs increased the proliferation rate and colonization of SSCs in the experimental groups compared to the control group. Average proliferation rate in the 20% duty cycle group was 1.46±0.06, in the 40% duty cycle group it was 2.00±0.1 and for the control group, it was 1.26±0.06. The average number of colonies in the 20% duty cycle group was 24±7.7, whereas the 40% duty cycle group had 62±1.4 colonies and the control group had an average of 19±5.5 colonies. Average colony diameters were as follows: 186.6±2.07 µm (20% duty cycle group), 185.3±4.4 µm (40% duty cycle group) and 190.0±2.0 µm (control group). Our results showed a significant increase in proliferation rate and number of colonies in the experimental groups compared to the control group (P<0.05), whereas no significant differences were observed between groups in colony diameters. These results suggested that LIPUS treatment can be an efficient, cost-effective method to improve proliferation and colonization of SSCs during in vitro culture.