Photobiomodulation preconditioned human semen protects sperm cells against detrimental effects of cryopreservation.

The biological consequences of semen samples preconditioning with photobiomodulation (PBM) were studied on human sperm cells post cryopreservation. Donated semen samples were collected from 22 married men with normal sperm parameters according to World Health Organization (WHO) criteria. Included samples were divided into control and PBM-preconditioning (one session, 810 nm, diode laser, and 0.6 J/cm2) groups before cryopreservation procedure. Progressive sperm motility (PSM), morphology, viability, sperm mitochondrial membrane potential(MMP), intracellular reactive oxygen species (ROS) and lipid peroxidation of sperm cells were assessed post thawing. PBM preconditioning of cryopreserved semen samples most prominently increased the PSM percentage 30 min post thawing (p = 0.000).Application of PBM before cryopreservation significantly increased the number of viable spermatozoa (p = 0.000), increased significantly the number of spermatozoa with high MMP (p = 0.004) and decreased significantly the number of spermatozoa with low MMP post-thawing(P = 0. 007)compared to control group. Cryopreserved human sperm cells with PBM preconditioning showed significant decrease in the levels of intracellular ROS (47.66 ± 2.14 versus 60.42 ± 3.16, p = 0.002) and lipid peroxidation (3.06 ± 0.13 versus 3.68 ± 0.27, p = 0.05)compared to control group. Our findings, as the first evidence, indicated that PBM-preconditioning of human semen before cryopreservation provides a real and substantial advantage. This might lead to a novel strategy in improving PBM application in the procedures of assisted reproductive technologies.

Comparative Effect of Photobiomodulation on Human Semen Samples Pre- and Post-Cryopreservation.

The primary objective of this study is to evaluate and to compare the effects of photobiomodulation (PBM) on sperm parameters both before and after cryopreservation. In this regard, 24 freshly ejaculated semen samples from normozoospermic men were included in this study. Each semen sample was randomly divided into three groups (1 ml aliquot for each group): the control group (group one) underwent conventional sperm cryopreservation (n = 24), group two underwent pre-freezing PBM exposure (810 nm, diode laser, and 0.6 J/cm2) (n = 24), and group three underwent post freezing and thawing PBM exposure (n = 24). Indicators of sperm quality, including total sperm motility (TSM), progressive sperm motility (PSM), DNA fragmentation, lipid peroxidation levels, apoptosis-like changes, and gene expression levels of protamine (PRM) 1, PRM2, and adducin 1 alpha (ADD1), were investigated in a blinded style. Due to the beneficial effect of pre-freezing PBM therapy, group 2 exhibited the highest TSM and PSM levels compared to groups 1 and 3. At the same time, DNA fragmentation and lipid peroxidation were significantly reduced in the group 2 compared to the group 1 (p = 0.024 p = 0.016, respectively). Evaluation of apoptotic/necrotic changes revealed that parameters including early apoptosis, dead, and necrotic cells decreased in the group 2 compared to the either groups 1 (p = 0. 008, p = 0. 032, p = 0. 02, respectively) or group 3 (p = 0.037, p = 0.108, p = 0.083). There were no significant differences in the expression levels of PRM1, PRM2, and ADD1 among the study groups. Based on our results, PBM therapy prior to cryopreservation, even in the normal semen samples, plays a significant protective role against cryo-damage by preserving the functional parameters of spermatozoa.

Photobiomodulation with 810 nm Wavelengths Improves Human Sperms' Motility and Viability In Vitro.

Background: Enhanced sperm motility is necessary for the successful journey of sperm inside the female genital tract, successful fertilization, and the increased chance of pregnancy. Objective: We investigated the impact of red and near-infrared (NIR) ranges of photobiomodulation (PBM) alone and together on fresh human sperm to validate an optimized PBM protocol that would maximize sperm motility and viability in vitro. Methods: We randomly divided 30 normal human semen samples into 3 different PBM protocols (red, NIR, and red+NIR lasers). Each sample was divided into four subparts, one control group sample and three experimental group samples. Each experimental group received one of the PBM protocols (red, NIR, or red+NIR). Each protocol was adjusted to three energy densities (0.6, 1.2, and 2.4 J/cm2). After exposure to the selected protocol, we determined the percentage of either viable or progressive sperm motility (PSM) and measured the DNA Fragmentation Index (DFI). Results: The NIR and red+NIR lasers at 2.4 J/cm2 energy density significantly increased PSM after 60 min compared with the control groups [least significant difference (LSD) test, p = 0.023 and p = 0.04, respectively]. Samples treated with the red laser at 0.6 J/cm2 had significantly decreased viability compared with the control group (LSD test, p = 0.003). Samples treated with the red+NIR lasers had significantly decreased viability at 0.6 J/cm2 (p = 0.003), 1.2 J/cm2 (p = 0.001), and 2.4 J/cm2 (p = 0.04) energy densities when compared with the control groups. The NIR laser resulted in no significant difference in sperm viability between the control and experimental groups. At 120 min after exposure, treatment with the red+NIR and red lasers at 2.4 J/cm2 density significantly increased DFI compared to the control groups (LSD test, p = 0.000, p = 0.007). Conclusions: In this study, sperm motility, viability, and DFI data confirmed the superiority of the NIR laser at 0.6 J/cm2 energy density compared with the red and red+NIR PBM protocols.

Analysis of meiotic spindle and zona pellucida birefringenceof IVM oocytes in PCOS patients.

BACKGROUND/AIM: Polycystic ovarian syndrome (PCOS) is one of the most common causes of infertility. One of the best therapeutic options for PCOS patients is intracytoplasmic sperm injection (ICSI). In vitro maturation (IVM) can also be a useful technique for these women. The goal of this study was to evaluate both the zona pellucida (ZP) birefringence and meiotic spindle (MS) of in vivo- and in vitro-matured oocytes from PCOS patients using the PolScope system. MATERIALS AND METHODS: Immature oocytes undergoing IVM and MII oocytes were obtained from PCOS patients in an ICSI program. Using PolScope, the presence of MS and ZP birefringence was assessed in both in vivo-matured oocytes (n = 32) and IVM oocytes (n = 24). Oocytes were classified as having highly birefringent (HB) ZP and lowly birefringent (LB) ZP. Furthermore, the rates of fertilization after ICSI were evaluated. RESULTS: The maturation rate was 68.5% after IVM. The percentage of a HB-ZP was significantly higher in the IVM oocytes than in vivo-matured ones (58.3% vs. 31.2%, respectively; P = 0.04). There were similar outcomes for the fertilization rates and MS detection between the two groups (P = 0.80 and P = 0.53, respectively). CONCLUSION: Clinical IVM is a safe technology for the maturation and maintenance of oocyte integrity in PCOS patients. The use of the noninvasive PolScope is recommended for detection of healthy oocytes in ICSI.

Survival Assessment of Mouse Preimplantation Embryos After Exposure to Cell Phone Radiation.

BACKGROUND: Using cellular phone has rapidly increased all over the world. Also, the concern on the possible health hazards of electromagnetic fields (EMF) induced from cell phones to reproduction has been growing in many countries. The aim of this study was to assess the consequences and effects of exposure to the cell phone radiation on the quality and survival rates of preimplantation embryos in mice. METHODS: A total of 40 mice (20 females and 20 males), 6 weeks old and sexually mature BALB/c, were used for control and experimental groups. The ovary burses were removed and the zygotes were dissected in the morning after mating. Next, 2-cell embryos were divided into two groups of control (n=150) and experimental (n=150). EMF (900-1800 MHz) was used for four days in experimental group for 30 min/day in culture at 37°C in a CO 2 incubator. The quality of embryos was recorded daily and the fluorescent staining was used for identification of viable blastocysts. All data were compared by Student's t-test and Mann-Whitney test (p<0.05). RESULTS: The rate of embryo survival to the blastocysts stage was similar in both groups. However, the percentage of dead embryos at the 2-cell stage was significantly higher in EMF-exposed group compared with controls (p=0.03). Also, the loss of cell viability significantly increased in experimental blastocysts (p=0.002). CONCLUSION: The normal embryonic development up to the blastocyst stage indicates that EMF-exposure commonly did not have adverse effect on embryo development in mice. But, it caused loss of blastocysts cell viability.

Developmental competence of immature oocytes aspirated from antral follicles in patients with gynecological diseases.

BACKGROUND: In vitro maturation (IVM) of immature oocytes collected from ovary has been proposed for fertility preservation. In addition, quality of oocytes post IVM is one of the factors determining its developmental competence. By using the non-invasive Polscope system, both meiotic spindle (MS) and zona pellucida (ZP) can be assessed in living oocytes. OBJECTIVE: The aim was to investigate the developmental potential of immature oocytes retrieved from ovarian tissue after IVM, as a method for fertility preservation, in patients with gynecological diseases. MATERIALS AND METHODS: The ovarian cortex from 26 patients with malignant and benign diseases (21-45 years old), were obtained directly from collaborating hospitals, and transported to the IVF center on ice. In total 61 immature oocytes were aspirated, of which 18 (29.5%) were degenerated and discarded. The remaining 43 (70.5%) healthy oocytes were cultured in IVM culture media for 48 hr. The rate of maturity was assessed, and the ZP birefringence and MS were imaged with Polscope technology. RESULTS: Overall 43 immature oocytes underwent IVM technology, of which 30.2% reached viable metaphase II (MII) oocytes. The ovarian tissues of 9 (34.6%) women were lacking oocytes at any stage. During polarized light microscopy examination, MS could be visualized only in one of the MII oocytes, but high ZP birefringence's were observed in the majority of the oocytes post IVM (61.5%). CONCLUSION: Oocytes maturation post IVM from unstimulated ovaries showed a good developmental competence in gynecologic patients. Further studies should be performed to advance the oocyte maturation program, such as co-culture system, for fertility preservation.