A Preliminary Study About CGMP Alleviates ROS Induced Oxidative Stress Damage In Human Spermatozoa

Objective:(1)To investigate the impact of reactive oxygen species (ROS) on sperm motility, cell death, integrity of nuclear DNA and acrosomal reaction of sperm.(2) To investigate the improvement and mechanism of cyclic guanosine monophosphate (cGMP) in sperm damage of ROS induced oxidative stress.Methods:All semen samples were centrifuged on a Percoll gradient to producing sperm suspension. Sperm motility was assessed by computer assisted sperm analysis (CASA). Eosin-Y dye was applied to evaluate sperm death rate. Concentrations of ROS in sperm were measured by H2DCF-DA in combination with flow cytometry. The expression of antioxidant enzymes was detected by RT-PCR. AOT was used to detect sperm DNA integrity. FITC-PSA stain was applied to evaluate the acrosomal reaction of sperm.Results:(1)100μM cGMP significantly increased the cyclic guanosine monophosphate (cGMP).(2)200μM SNP had no effect on the progressive motility of sperm, but SNP, as the same time incubation with LY83583, significantly reduced progressive motility of sperm.(3) Catalase (CAT) and cGMP significantly reversed the inhibition of SNP and LY83583on the progressive motility of sperm.(4)SNP and LY83583treatment increased death rate, and generated a large number of reactive oxygen species (ROS).(5) cGMP sharply induced the CAT and manganese superoxide dismutase (Mn-SOD) expression to inhibit the ROS damage of sperm cells.(6) Progressive motility of sperm declined with the concentration increase of H2O2. cGMP improved the effects of sperm progressive motility inhibited by H2O2. The DNA fragmentation index (DFI) of H2O2was obviously higher than that of control.(7) The DFI of H2O2+cGMP and H2O2+CAT were significantly lower than that of H2O2, cGMP and CAT markedly reversed the damage on the integrity of the DNA by H2O2.(8) Acrosome reaction rate (AR) of cGMP was obviously higher than that of control and H2O2. Meanwhile, SNP and H2O2both had a higher AR than control.Conclusion:(1) low concentration of SNP and H2O2could increase the sperms activity obviously and high concentration of SNP and H2O2could inhibit sperms activity markedly.(2) cGMP may play an important role in sperm protection from ROS. cGMP can induce sperm ROS, cell death and DNA integrity.(3) The antioxidase expression induced by cGMP may take part in regulated redox in sperm.(4) cGMP may promote the occurrence of sperm acrosomal reaction.(5) cGMP may have unique physiological role in vitro sperm, especially in the ART sperm preparation.