Using the Modified Hypo-Osmotic Swelling Test for the Selection of Immotile Testicular Spermatozoa in ICSI—A Randomized Controlled Study

Abstract

Various methods have been described for the selection of viable among immotile testicular spermatozoa prior to ICSI in cases of total absence of motility. These include the addition of pentoxifyllin, the mechanical touch technique, performing the hypo-osmotic swelling (HOS) test (Jeyendran solution) as well as the modified HOS test (50% culture medium + 50% Millipore grade water). However, these reports have not been confirmed by randomized controlled studies (RCTs). The aim of this work was to conduct a RCT to evaluate the use of the modified HOS test for the selection of viable but immotile spermatozoa in cases of ICSI using testicular sperm (ICSI-TeSE). A randomized controlled trial. All couples treated in our center between January 2002 and August 2004 by ICSI using testicular sperm due to azoospermia (n= 316) were eligible for the study. They included 81 couples whose testicular biopsy (fresh or frozen) revealed total absence of motility and these were included in this study in their first cycle of treatment. The couples were randomized at the time of ICSI using a closed envelope system and allocated to one of two groups. In patients belonging to the first group (no HOS test group), selection of spermatozoa for ICSI was performed on the basis of morphology, while in the second group (HOS test group), the modified HOS was performed and only reactive spermatozoa were used for the injection. The modified HOS test entails placing the immotile spermatozoa in a microdroplet consisting of a 50% culture medium and 50% Millipore grade water. After a maximum of 10 seconds, viable spermatozoa, recognized by their curved or swollen tails, were selected and transferred into another microdroplet of HEPES-buffered medium where they were washed 3 times to re-equilibrate them osmotically, before being transferred to the PVP microdroplet. The tails of the all spermatozoa were crushed before the injection step. Group A consisted of 36 cycles, while group B consisted of 45 cycles. The fertilisation rate was 40.5% in the HOS test group compared to 18.7% in the no HOS test group (P<0.0001). The clinical pregnancy rate was 26.7% in the HOS test group compared to 5.6% in the no HOS test group (P< 0.05). The delivery/ongoing pregnancy rate was 20% in the HOS test group compared to 2.8% in the non-HOS group (P< 0.05). The implantation rate was 22.8% in the HOS test group compared to 5% in the no HOS test group (P< 0.05). The selection of viable among immotile testicular spermatozoa prior to ICSI using the modified HOS test increases the fertilisation rate, the pregnancy rate, the ongoing pregnancy rate and the implantation rate significantly.