Habib Tajalli
1,2* 
, Masoud Maleki
3,4, Fatemeh Firouzi Amoudizaj
3,4, Zahra Akbarpour
3,4 , Esmail Safavi
4,5, Reza Shahi
6, Alireza Sotoudeh
1,2, Ahmad Babazadeh Bedoustani
4,71 Research Center of Biophotonics, Tabriz Branch, Islamic Azad University, Tabriz, Iran
2 Department of Physics, Tabriz Branch, Islamic Azad University, Tabriz, Iran
3 Department of Biology, Tabriz Branch, Islamic Azad University, Tabriz, Iran
4 Research Center of Biotechnology, Tabriz Branch, Islamic Azad University, Tabriz, Iran
5 Department of Basic Sciences, Faculty of Veterinary Medicine, Tabriz Branch, Azad Islamic University, Tabriz, Iran
6 Department of Mathematics, Tabriz Branch, Islamic Azad University, Tabriz, Iran
7 Department of Biology, Faculty of Natural Science, University of Tabriz, Tabriz, Iran
Abstract
Introduction: Infertility affects approximately 15% of couples worldwide, with male factors accounting for approximately half of the cases. Low-level laser therapy (LLLT) has been increasingly considered in modern medicine due to its high efficacy, ease of use, and lack of side effects. Evidence suggests that this method can prevent DNA damage in cells and activate key genes related to fertility. This study aimed to investigate the effects of LLLT on sperm production in azoospermic mice using in vitro and in vivo experimental models.
Methods: Adult male NMRI mice (8–9 weeks old, 30–35 g) were divided into the negative control (healthy), positive control (azoospermia via intraperitoneal busulfan, 30 mg/kg), and experimental (azoospermia with 808 nm LLLT at 8 J/cm2) groups (8 mice per group) for in vivo experiments. For the in vitro part, spermatogonial stem cells were cultured from 6 azoospermic mice under control conditions or treated with laser (808 nm LLLT at 4 J/cm2). Morphological examination and real-time polymerase chain reaction were used to assess testicular structure and expression of several genes, such as deleted in azoospermia-like (DAZL), G protein-coupled receptor 125 (GPR125), synaptonemal complex protein 3 (SYCP3), DEAD-box helicase 4 (VASA/DDX4), protamine (PRM), acrosin (ACR), and tripartite motif containing 36 (Haprin/TRIM36).
Results: In vivo, LLLT increased VASA expression, improved germ cell activity, and increased sperm production compared with untreated azoospermic control groups, although these changes were not statistically significant. In vitro, 4 J/cm2 radiation modulated several genes related to spermatogenesis, supporting its role in germ cell differentiation.
Conclusion: LLLT with a wavelength of 808 nm could improve spermatogenesis and sperm production in a mouse model of busulfan-induced azoospermia in vivo and in vitro. These results demonstrated its potential as a supportive treatment for male infertility.