SIGNIFICANT IMPROVEMENT IN SPERM MOTILITY IN SEMEN SAMPLES FROM ASTHENOZOOSPERMIC MEN BY USING CONTROLLED INCUBATION PROTOCOLS WITH SINGLE CONTINUOUS CULTURE MEDIUM (CSCM®)

Abstract

To test different incubation periods and to assess an appropriate commercially available medium to improve the performance of semen samples from asthenozoospermic men. Prospective study Semen samples (progressive motility <32%) from 54 asthenozoospermic patients (men aged 21-45 years) were used in the study after signed the informed consent form. Fresh semen samples separated by a discontinuous density gradient or by simple washing, were incubated in different culture media and then analyzed. Four different culture media were studied: a) Single Continuous Culture Medium (CSCM®) containing 10% Human Serum Albumin (HSA®); b) Sperm Rinse medium (SR®) supplemented with fructose and carnitine; c) In Vitro Fertilization Medium (G-IVF®) with fructose and 15% Synthetic Serum Substitute (SSS®) and; d) a control group in which Human Tubal Fluid (HTF®) supplemented with 15% SSS® was used. The last one represents the most commonly used medium by labs worldwide. Samples were then incubated at 37°C in a 5% CO2 atmosphere for 2-hour (defined as the optimal incubation time in previous study). Sperm parameters were then analyzed both in fresh samples and after incubation in different media: manual sperm analysis; motility via Sperm Computer Analysis (SCA®); sperm chromatin integrity via Sperm Chromatin Structural Assay (SCSA®); Reactive Oxygen Species level (ROS) by chemiluminescent detection and mitochondrial activity by staining with 3.3′-diaminobenzidine (DAB). Data were analyzed using the IBM SPSS software. Analysis of variance (ANOVA) and independent student t-tests were used to evaluating statistical significance (P<0.05). CSCM® medium demonstrated a significant increase in post-incubation motility (progressive motility = 24.55±13.97%; total motility = 41.17±19.06%) compared to pre-incubation parameters (progressive motility = 8.33±6.82; total motility = 26.61±18.10%; P<0.025). For all other culture media, no significant differences were observed in seminal parameters. Asthenozoospermic men may have significant improvement in their semen samples in the andrology lab after a 2-hours incubation in CSCM®+HSA® medium. This technique has proven to be very attractive because of its efficiency, low cost and ease of reproduction.