#164 : In-Vitro Vitamin D Treatment on Human Sperm Improves Capacitation, Acrosome Reaction, and Protein Expression of Sperm-Oocyte Activation Factors (SOAF)

Background and Aims: Although many studies have been done to investigate the relationship between serum vitamin D and male infertility, the effect of in-vitro vitamin D treatment on sperm remains unclear, particularly the effect on sperm capacitation, hyperpolarisation, sperm-oocyte fusion and oocyte activation. This study aims to examine whether in-vitro vitamin D treatment on sperm can improve the condition of sperm parameters and further increase fertilisation and pregnancy rate.

Method: Fifty men (N=50, fertile n=26; infertile men n=24, were recruited during their visit to fertility clinics for semen analysis. Each washed semen sample was aliquoted and divided into control and in-vitro vitamin D treatment groups.

For in-vitro vitamin D treatment, semen samples were treated with 1nM in-vitro for 1 hour at 37°C. Intracellular calcium released, capacitation status and mitochondria membrane potential were measured after incubation in both the control and in-vitro vitamin D treatment groups. Furthermore, immunofluorescence of sperm-origin SOAF, Phospholipase C Zeta (PLC$\zeta$), was done to investigate fluorescence intensity.

Results: Compared to the control, higher intracellular calcium release was noted following in-vitro vitamin D treatment, represented by higher Fluo-4AM staining (MD=25043, P=0.02). In the treatment group, sperm capacitation rate and CTCF were higher when Chlortetracyclin (CTC) (MD=4.7%, P=0.01) and immunofluorescence of phosphotyrosine (MD=0.03, P=0.01) were examined. More hyperpolarised mitochondria membranes in the treatment group showed by JC-1 staining (P=0.01) compared to the control group. The red-to-green, fluorescent ratio is 1.62 in the control group and 2.24 in the treatment group. Moreover, immunofluorescence of PLC$\zeta$ shows that the treatment improves this protein’s expression (MD=8.013×10${}^{-3}$, P=0.02). Thus, in-vitro vitamin D enhances the functionality of sperm by increasing intracellular calcium ions, sperm capacitation, hyperpolarisation and protein expression.

Conclusion: Adding vitamin D in in-vitro fertilisation (IVF) in media culture is beneficial as capacitation, sperm membrane polarisation and SOAF protein expression were increased and therefore contributed to successful IVF.