Alessandro Schuffner

Cryopreservation-thawing of fractionated human spermatozoa and plasma membrane translocation of phosphatidylserine

OBJECTIVE(S) [1] To evaluate sperm membrane damage during cryopreservation-thawing by the assessment of phosphatidylserine (PS) translocation and [2] to examine the relationship between reactive oxygen species (ROS) and cryopreservation-related alterations. DESIGN Prospective cohort study. SETTING University-based center. PATIENT(S) Men consulting for infertility and fertile donors (controls). INTERVENTION(S) Semen processing was performed by density gradient separation followed by cryopreservation and thawing. MAIN OUTCOME MEASURE(S) Membrane PS translocation was evaluated with annexin V binding, generation of ROS was detected by chemiluminescence, and motion parameters were assessed by computer analysis. RESULT(S) Annexin V binding was detected in the prefreeze fractions with high and low sperm motility. In the patient group, there were significantly higher postthaw levels of annexin V binding in both fractions when compared with prefreezing values. However, such induction of PS translocation was significantly higher in the fractions with high sperm motility. Significantly higher ROS levels were detected in prefreeze samples of the fractions with low sperm motility. CONCLUSION(S) In the population of men studied, [1] cryopreservation-thawing was associated with induction of membrane PS translocation; [2] postthaw ROS levels were lower than before freezing; and [3] neither annexin V binding results nor the generation of ROS were able to accurately predict sperm cryosurvival rates.

Outcome of intracytoplasmic sperm injection in azoospermic patients : Stressing the liaison between the urologist and reproductive medicine specialist

OBJECTIVES To analyze the outcome of intracytoplasmic sperm injection (ICSI) cycles in infertile couples in whom the main diagnosis of infertility was azoospermia of obstructive and nonobstructive origin. METHODS Eighty-three consecutive ICSI cycles were carried out with retrieved testicular or epididymal spermatozoa, 60 cycles in 32 patients with obstructive azoospermia and 23 cycles in 12 patients with nonobstructive azoospermia. Fifty-four testicular biopsies (testicular sperm extraction) and 18 epididymal aspirations (microepididymal sperm aspiration) were performed.Results. Motile spermatozoa were recovered in 65 cycles (90.3%). In another 3 (4.2%), nonmotile spermatozoa were retrieved. In 4 patients (5.5%), sperm could not be recovered. In 11 cycles, frozen sperm from a previous procedure were used. A significantly lower fertilization rate (64% versus 73%, P = 0.02), clinical pregnancy rate (13% versus 47%, P <0.001), and good embryo quality rates (35% versus 56%, P = 0.009) were observed in patients with nonobstructive azoospermia. In patients with obstructive azoospermia, no significant differences were observed when the outcome was analyzed on the basis of the sperm origin (ie, from testicular sperm extraction or microepididymal sperm aspiration). CONCLUSIONS When combining testicular sperm extraction or microepididymal sperm aspiration with ICSI in patients with obstructive azoospermia, the results in terms of fertilization, implantation, and pregnancy rates were similar to those found in patients with nonazoospermic obstruction who underwent ICSI with ejaculated sperm. Patients with nonobstructive azoospermia had lower fertilization, embryo quality, and pregnancy rates than did those with obstructive azoospermia, probably because of severe defects in spermatogenesis, leading to poor gamete quality. The urologist and reproductive endocrinologist now have an excellent therapeutic option to offer men with previously intractable infertility.

Comparison of various preparation methods for the use of cryopreserved-thawed spermatozoa in insemination therapy.

Different approaches have been implemented to improve the recovery and/or the quality of motile sperm after cryopreservation-thawing. Gradient separation of the motile fraction before freezing offers the possibility of selecting spermatozoa that retain motility for up to 24 h (1). An intrauterine insemination (IUI)-ready cryopreservation method using sucrose and glycerol-based cryoprotectant with Percoll processing produced improved results when compared to conventional cryopreservation (2). Additionally, selection of a highly motile sperm population by swim-up was shown to improve postthaw acrosomal integrity, motility, and other functional parameters (3). The purpose of this prospective study was to assess the impact of different semen processing methods (i.e., samples cryopreserved as whole semen and IUI-ready, and those washed for IUI after thawing) on motility parameters postthaw.