The effect of in-vitro culture on the motility and morphology of fresh and frozen-thawed human testicular spermatozoa obtained from obstructive azoospermic patients and on the motility of testicular spermatozoa obtained from non-obstructive azoospermic patients was evaluated. The outcome of intracytoplasmic sperm injection (ICSI) with fresh and frozen-thawed human testicular spermatozoa was studied. The results showed that significant improvement of sperm morphology and motility was observed in culture of fresh (n = 17) and frozen-thawed (n = 15) testicular sperm samples obtained from patients with obstructive azoospermia. The motility of cultured testicular spermatozoa reached a peak at 72 h without the need for special media. In six of 20 samples obtained from patients with non-obstructive azoospermia, improvement of sperm motility was observed. When only non-motile testicular spermatozoa were cultured, they all remained non-motile (n = 9). In patients with obstructive azoospermia, fertilization rates of 80 and 81% were obtained using ICSI with fresh and frozen-thawed testicular spermatozoa respectively. Clinical pregnancies were observed in four out of nine patients with fresh testicular spermatozoa and two out of five patients after using frozen-thawed spermatozoa. When fresh testicular spermatozoa obtained from patients with non-obstructive azoospermia were used for ICSI, the fertilization rate was 68% and two out of seven patients achieved clinical pregnancies. In conclusion, the morphology and motility of fresh and frozen-thawed testicular spermatozoa in patients with obstructive azoospermia can be significantly improved after in-vitro culture. The outcome of in-vitro culture of testicular spermatozoa in patients with non-obstructive azoospermia is unpredictable. In-vitro culture of non-motile testicular spermatozoa is not successful so far. The outcome of ICSI with fresh and with frozen-thawed testicular spermatozoa was similar.
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