L-polyhistidine Inhibit Boar Sperm Apoptosis Induced By Lipopolysaccharide And Its Effects On Sperm Quality

Liquid Storage of boar semen is the basis of artificial insemination technology. Lipopolysaccharide (LPS), which was produced by bacterial lysis in semen liquid-stored condition, not only can impair sperm quality, shorten the life of the sperm, induce sperm apoptosis, but also cause an immune response female genital tract after artificial insemination, and further induce the recruitment of immune cell in female genital tract for swallowing sperm. L-polyhistidine can bind to LPS,whether it inhibit sperm apoptosis induced by LPS still was unknown. In the study, boar semen were added into L-polyhistidine, and then semen quality and sperm apoptosis were examined for exploring whether it effected on semen quality and reduced sperm apoptosis, and was explained its mechanism.Semen from 24 boar were collected and divided into 5 sample pools (n=5), and then diluted to 4×106/mL with BTS dilution. The samples were stored at 17? or 37? for successive experiment. At first, the changes in the concentration of bacterial colonies and LPS concentration in semen were measured at different time-point (0,6,12,36,72,120h) under liquid-stored condition; And then the diluted semen (as the control group), and the semen containing 100?g/mL PMB (PMB-group) or 100?g/mL L-pHis (L-pHis group), were measured the contration of bacteria and LPS, sperm quality(sperm motility index,viability and sperm movement speed) and sperm apoptosis at 0,6,12,36,72,120h poststorage under liquid-stored condition, and were measured quality (sperm motility index, viability and sperm movement speed) and sperm apoptosis 0,1,3,6,9,12,24h post storage at 37? for assessing the effect of PMB and L-pHis on semen quality and apoptosis. Finally, semen were preincubated in BTS medium containning 2?g/mL Anti-CD14 to block the binding of LPS-TLR4, and then assess sperm apoptosis for exploring the possible mechanism of L-pHis inhibiting bore sperm apoptosis.The results showed:(1) The concentration of bacteria rapidly increased from 36 h to 120 h, and itcontinuously significantly increases at 36h (4980±381.58CFU/ml),72h (12920±2833.66CFU/ml),120h (40020±4755.38 CFU/mL) (P<0.05), the average of bacterial concentration is 1.02×104 CFU/mL. The concentration of LPS is no significant change with the time-course (P>0.05), and the concentration of LPS in seminal plasma is 4.70±0.21 EU/mL. (2) The supplement of 100?g/mL L-pHis or 100?g/mL PMB has no effect on the concentration of LPS and can ruduce the concrntation of bacteria under liquid-stored condition, but they could improve semen quality (sperm motility index, sperm speed, sperm viability) under liquid-stored and 37? condition. (3) The additives of 100?g/mL L-pHis or 100?g/mL PMB significantly inhibit sperm apoptosis under liquid-stored or 37? condition (P<0.05). (4) L-pHis could not specifically eliminate LPS in the semen, but the supplement of Anti-CD14 or L-pHis signigicantly inhibit sperm apoptosis (P<0.05), and it's possible reason that L-pHis inhibit sperm apoptosis through blocking the binding of LPS and TLR4.In conclusion:the addition of 100?g/mL L-pHis can improve semen quality effectively, and inhibit sperm apoptosis through blocking apoptosis signal pathway induced by LPS.