Efficacy of testicular sperm chromatin condensation assay using aniline blue-eosin staining in the IVF-ET cycle

Abstract

ObjectiveThis study was performed to evaluate testicular sperm chromatin condensation using aniline blue-eosin (AB-E) staining and its effects on IVF-ET.MethodsChromatin condensation was analyzed using AB-E staining in 27 cases of testicular sperm extraction. There were 19 cases of obstructive azoospermia (OA) and 8 cases of non-obstructive azoospermia (NOA) in IVF-ET. Mature sperm heads were stained red-pink whereas immature sperm heads were stained dark blue. The percentage of sperm chromatin condensation was calculated from the ratio of the number of red-pink sperm to the total number of sperm analyzed.ResultsThe overall percentages of chromatin condensation in OA and NOA were 31.1±11.2% and 26.3±14.4%, respectively. The fertilization rate was significant higher in OA than NOA (p<0.05); however, the rates of good embryos and clinical pregnancy did not show statistical differences. In OA and NOA, statistical differences were not observed in the rate of chromatin condensation, fertilization, good embryos, and clinical pregnancy between the pregnant group and non-pregnant group.ConclusionChromatin condensation is less stable than OA and showed a low fertilization rate in NOA. While there were no significant differences in chromatin condensation results between NOA and OA, we propose that a pattern of decreased chromatin condensation in NOA is one of the factors of low fertilization results requiring further study.

Effect of testicular spermatozoa on embryo quality and pregnancy in patients with non-obstructive azoospermia

“…It is well known that the early stages of embryo development primarily depend on maternal transcripts, since the major influence of the paternal genome is not felt until the 6-8 cell stage. Sperm with an elevated rate of DNA abnormalities may affect abnormal blastocyst development, implantation failure, and spontaneous pregnancy loss [Carrell et al 2003;Virant-Klun et al 2003;Henkel et al 2004;Tesarik et al 2004a,b;Park et al 2011;Robinson et al 2012;Jodar et al 2013;Sendler et al 2013]. …”

“…Embryos were scored according to the number of blastomeres and the percentage of enucleate fragments. Embryos were graded according to a previously described system [Park et al 2011;Park et al 2014]. Briefly, grade I embryos showed even blastomeres and no fragmentation; grade I-1 embryos showed even blastomeres and 525% fragmentation; grade II embryos showed uneven blastomeres and 525% fragmentation; grade II-1 embryos showed uneven blastomeres and 25%-50% fragmentation; and grade III embryos showed even or uneven blastomeres and !50% fragmentation.…”

Efficacy of cryopreservation of embryos generated by intracytoplasmic sperm injection with spermatozoa from frozen testicular tissue

“…The testicular sperm extraction and preparation procedure was performed as described previously [8,22]. Briefly, a small piece (1 cm 3 ) of extruded testicular tissue was excised and placed in Ham's F-10 medium (Sigma, St. Louis, MO) supplemented with 0.4 % (w/v) human serum albumin (HSA; Sigma).…”

“…Fertilized embryos were transferred to G-I.III medium. Embryos were scored according to the number of blastomeres and percentage of enucleate fragments [22,23]. Embryo grading was classified into five groups, as follows: grade I, even blastomeres, no fragmentation; grade I-1, even blastomeres, fragmentation< 25 %; grade II, uneven blastomeres, fragmentation<25 %; grade II-1, uneven blastomeres, fragmentation 25 % -50 %; grade III, even or uneven blastomeres, fragmentation≥50 %.…”

Pregnancy Outcome of Intracytoplasmic Sperm Injection in Relation to Duration of Cryopreservation of Spermatozoa Obtained through Testicular Sperm Extraction

“…Women included in this study had also received routine infertility work-up. Testicular sperm preparation and freezing & thawing: The testicular sperm extraction and preparation procedure was performed as described by Park et al [14].…”