Distribution Of Mfn-2in Human Sperm And Its Cryoprotective Potential

BackgroundMitochondria are the main place for synthesis of adenosine triphosphate (ATP) ineukaryotic cells to provide energy. Also, mitochondria can store calcium ion, andmaintain the homeostasis of the calcium ion concentration in cells by the synergy withendoplasmic reticulum, and extracellular matrix, etc. Moreover, mitochondria areinvolved in signal transduction of calcium ions during cell apoptosis, and in control ofapoptosis by regulating membrane potential. Mitochondria are also involved in theregulation of cell growth and cell cycle.Mitochondria are highly dynamic organelles, with continuous fusion and division tomeet the needs of a variety of physiological state. By regulation of mitochondrial fusion,mitochondrial fusion protein2(Mfn-2), maintains the mitochondrial morphology,regulates mitochondrial function and plays an important role in cellular energymetabolism, apoptosis, signal transduction, embryonic development, cardiovasculardisease, cryoprotection, etc.Given that energy metabolism, apoptosis, signal transduction and oxidative stressare crucial factors affecting sperm quality, Mfn-2is likely, through these aspects, toregulate the function of the sperm or to affect sperm quality. Nevertheless, Mfn-2inhuman sperm has not been reported yet. In the present study, we determined the distribution and location of Mfn-2, and explored its cryoprotective potential on humansperm.ObjectivesTo investigate the distribution of Mfn-2in human sperm, and its potential role insperm cryoprotection.MethodsIndirect immunofluorescence and confocal laser scanning microscope were used toobserve the distribution of Mfn-2in human sperm. Human semen of normozoospermicdonors (n=10) were permitted to liquefy at room temperature. Then samples were frozenat4℃（37℃for control group） for1hour,2hours,4hours,8hours and12hoursrespectively. Sperm motility was measured and Western Blot was used to determine theMfn-2expression. Mfn-2levels in frost-resistant group, of which>40%sperm canrecover from the liquid nitrogen frozen program, and frost-sensitive group, of which<40%sperm can recover, were also determined by Western Blot.ResultsThrough the confocal laser scanning microscope, the immunofluorescence can onlybe found in the middle of human sperm tail. This part, which was located between theCervical segment and principle segment with the length of5μm to7μm, wascompletely stained positively. The Mfn-2expression of the group which has been frozenat4℃for4hours was significantly higher than other groups (P<0.05), and so was thesperm motility at this time point. The quantity of Mfn-2expression in frost-resistantgroup was much higher than that in frost-sensitive group (P<0.05).ConclusionMfn-2is distributed in the whole middle of human sperm tail, providing insight forstudying the biological function of Mfn-2in human sperm. The relatively high Mfn-2expression in sperm is likely to play an important role in the recovery of sperm motility after4-hour-freezing at4℃. Mfn-2expression of the sperm from the group which>40%sperm can recover was obviously higher than that of the sperm from the group which<40%sperm can recover, indicating Mfn-2may have a pivotal role in protecting humansperm during freezing.