D. Check

Effect of Pentoxifylline Added to Freezing Media on Subsequent Post—Thaw Hypoosmotig Swelling Test and Other Semen Parameters

This study evaluated the effects of pentoxifylline (PTX) on post-thaw semen parameters as well as the hypoosmotic swelling (HOS) test. Fourteen samples were evaluated for volume, count, motility, % grade A sperm, and HOS test. Two aliquots were frozen, one in freezing medium and the other in a 3 mM solution of PTX and freezing medium. Both groups were frozen in liquid nitrogen vapors for 30 min. Thawing was performed at 37 degrees C for 15 min, followed by a wash with 2 parts 0.5% HSA/MHTF to 1 part sample. Pellets were resuspended in 0. MHTF and then evaluated as described above. In addition, motility was evaluated 2 h post-thaw. Following freeze-thaw, the mean motile densities were similar (17.5 x 10 motile/mL vs. 20.4 x 10(6) motile/mL for PTX and control, respectively). Two hours post-thaw, the PTX group had a mean sperm motility of 31.3% vs. 37.7% for the control group (p > .05). There were no significant differences in % grade A sperm in PTX (13.0%) vs. control (12.0%). Similarly, HOS scores did not improve following cryopreservation (43.0% and 50.6% for PTX and control, respectively). Thus, no improvement was found by freezing sperm with PTX.

Effect of shortened exposure time to the critical period for ice crystal formation on subsequent post-thaw semen parameters from cryopreserved sperm

Cryopreservation of human sperm using present methods leads to a reduced fertility potential of the specimen. In many instances this prevents the successful fertilization of the female partner from the frozen-thawed specimens of males whose semen has been cryopreserved prior to surgery, chemo-therapy, or even vasectomy. Furthermore, even though some donor specimens can be successfully used for achieving pregnancies, one needs to place the sperm intrauterine to approach the same pregnancy rates as those of fresh intracervical insemination. The main mechanism considered for sperm damage by cryopreservation is ice crystal formation. The most critical time for forming ice crystals is from 0 to -10 degrees C. In the present study the effect of a modified rapid cryopreservation technique with reduction of exposure time to the 0 to -10 degrees C temperature range was compared to standard freezing procedures on subsequent semen parameters. Though no significant differences were found on post-thaw motile densities or hypoosmotic swelling test scores, a new, equally effective, but more rapid technique for cryopreservation is reported.

Fresh versus frozen seminal plasma for enhancing sperm motility in asthenozoospermic males

Previous data demonstrated improvement in severe asthenozoospermia in men with retrograde ejaculates by adding donor seminal plasma (DSP); interestingly, no such improvement was demonstrated following suspension in Hams F-10 medium. A study was performed to examine whether DSP would also improve asthenozoospermic ejaculates from men with antegrade semen. In addition, the study further explored whether the ameliorative effect of DSP could be maintained after freeze-thawing. The mean increase in motility following fresh DSP in 37 specimens was 102% and was -1.3% for DSP frozen at -20 degrees C and 16.7% for DSP frozen at -196 degrees C. The only statistical difference (using Students t test) was seen in the comparison of fresh DSP to DSP frozen at -20 degrees C (p = .001). Nine of twenty-four men exhibited doubled baseline motility rates following addition of fresh DSP, compared to only 1 of 20 and 1 of 5, respectively, following addition of semen treated with DSP frozen at -20 degrees C and -196 degrees C, respectively. The data suggest that poor motility may improve when DSP is added to the specimen. However, proper quarantine may not be possible because efficacy is lost after freezing.

Improved Results of Thawed Sperm Cryopreserved with Slow Stage Cooling with a Cellevator

There is a need to develop a sperm cryopreservation technique that will allow good pregnancy rates following intrauterine insemination of thawed semen specimens that have been frozen prior to sperm-destructive procedures, such as surgery, chemotherapy, or radiation therapy. A slower cooling rate using a commercial semiprogrammable freezer may provide improved post-thaw motility and hypoosmotic swelling (HOS) test scores. However, the cost of this apparatus precludes it from being used in most andrology centers. This study compares the efficacy of slow stage cooling using an inexpensive cellevator (a device used to freeze lymphocytes) to liquid nitrogen vapor freezing. The semen from 27 males was equally divided and one aliquot was cryopreserved with the cellevator stage cooling and the other with the liquid nitrogen vapor technique. The percent motility and percent grade A sperm post-thaw were significantly higher when cryopreserved with the cellevator than with vapor freezing, as was the mean percentage of sperm showing HOS changes.

EFFECT OF AN INTERMEDIATE HOLD WITH VAPOR FREEZING ON SUBSEQUENT HYPOOSMOTIC SWELLING IN THAWED SPERM

Cryopreservation and thawing of sperm exerts an adverse effect on functional integrity of the sperm membrane as measured by the hypoosmotic swelling (HOS) test. Supercooling using the liquid nitrogen (LN2) vapor technique may damage membranes by favoring ice crystal formation. Slight slowing of the cooling process may allow escape of intracellular fluid. This study was conducted to evaluate modification of the LN2 vapor technique by employing an intermediate hold in a freezer (to slow down the rate of cooling) on HOS scores on specimens thawed 1 month after freezing. Semen samples were obtained from male partners of infertile couples with a requirement of a baseline HOS score < 70% but > or = 60%. The HOS test was performed on the unprepared semen sample prior to freezing and immediately post-thaw 1 month later on the aliquot frozen with LN2 vapors only vs. the equal fraction subjected to an intermediate hold. The mean initial HOS score was 68.5% and was 47% in thawed specimens that had been cryopreserved with and without an intermediate hold. There were no differences in the percentage of specimens exhibiting a > 50% HOS score following vapor freeze (70%) or vapor freeze with a hold (74%). Thus, these data do not demonstrate any advantage of slowing the vapor freezing process by utilizing an intermediate hold.