Glycogen Synthase Kinase-3 Isoform Variants and Their Inhibitory Phosphorylation in Human Testes and Spermatozoa.

PURPOSE: To clarify (phospho-) glycogen synthase kinase-3 (GSK3) isoform variants in the germline and soma of human testes and spermatozoa. MATERIALS AND METHODS: GSK3 isoform variants in normospermatogenic and Sertoli cell-only (SCO) testicular biopsies and spermatozoa were examined. RESULTS: In normospermatogenic testes, GSK3α and GSK3ß variants 1 and 2 different in low complexity region (LCR) were expressed and their levels were decreased in SCO testes. GSK3ß variant 3 was only expressed in SCO testes. GSK3ß as well as GSK3α, the dominant isoforms in testes were decreased in SCO testes. In normospermatogenic testes, GSK3ß were found in spermatogonia and markedly decreased in meiotic germ cells in which GSK3α was dominant. p-GSK3α/ß were marginal in spermatogonia and early spermatocytes. In SCO testes, GSK3α/ß immunoreactivity in seminiferous epithelia was weaker than those of normospermatogenic testes whereas p-GSK3α/ß(Ser) immunoreactivity was visibly increased in Sertoli cells. GSK3α was dominant in ejaculated spermatozoa in which GSK3α and p-GSK3α(Ser) were found in the head, midpiece, and tail. In acrosome-reacted spermatozoa, GSK3α was found in the equatorial region of head, midpiece, and tail, and p-GSK3α(Ser) was only found in midpiece. During sperm capacitation, p-GSK3α(Ser) was significantly increased together with phosphotyrosine proteins and motility. CONCLUSIONS: In human male germ cells, GSK3 isoforms different in LCRs switch from GSK3ß to GSK3α during meiotic entry, suggesting the isoform-specific roles of GSK3α and GSK3ß in meiosis and stemness or proliferation of spermatogonia, respectively. In dormant Sertoli cells of SCO testes kinase activity of GSK3 might be downregulated via inhibitory phosphorylation. In spermatozoa, inhibitory phosphorylation of GSK3α might be coupled with activation of motility during capacitation.

Abnormal Human Sperm Parameters Contribute to Sperm DNA Fragmentation in Men with Varicocele.

PURPOSE: This study was performed to evaluate and compare threshold sperm parameters and sperm DNA fragmentation index (DFI), and further analyzed whether sperm DFI could be predicted from sperm parameters in men with varicocele. MATERIALS AND METHODS: A total of 157 semen samples underwent both semen analysis and sperm DNA fragmentation (SDF) testing in men with varicocele. Sperm parameters were assessed using the World Health Organization guidelines. SDF testing was performed using the Halosperm kit. Sperm parameters and sperm DFI results were compared. RESULTS: The overall sperm parameter results and sperm DFI showed normal values; however, the morphology value was at the lower limit of normal. High sperm DFI was associated with significantly lower motility and viability (p<0.001, respectively). Sperm motility and morphology were significantly higher in the higher sperm count group compared to the lower sperm count group (p<0.05), while sperm DFI was higher in the lower sperm count group (p<0.05). Sperm count and viability and sperm DFI were significantly associated with the quality of sperm motility (p<0.001). Sperm motility and sperm DFI were significantly different (p<0.001) between normal and abnormal sperm viability groups. Between normal and abnormal sperm morphology groups, sperm count, motility, and sperm DFI showed significant differences (p<0.001). CONCLUSIONS: In this study, a correlation between SDF and sperm parameters was confirmed in men with varicocele. SDF may be contributing factors to sperm motility, viability, and morphology. Abnormal sperm count, motility, and viability showed high sperm DFI. Therefore, lower sperm parameters were indicative of increasing SDF in men with varicocele.

Comparison of clinical outcomes between in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) in IVF-ICSI split insemination cycles.

OBJECTIVE: This study aimed to compare the clinical outcomes between in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) in sibling oocytes. Additionally, we evaluated whether the implementation of split insemination contributed to an increase in the number of ICSI procedures. METHODS: A total of 571 cycles in 555 couples undergoing split insemination cycles were included in this study. Among them, 512 cycles (89.7%) were a couple's first IVF cycle. The patients were under 40 years of age and at least 10 oocytes were retrieved in all cycles. Sibling oocytes were randomly allocated to IVF or ICSI. RESULTS: Total fertilization failure was significantly more common in IVF cycles than in ICSI cycles (4.0% vs. 1.4%, p<0.05), but the low fertilization rate among retrieved oocytes (as defined by fertilization rates greater than 0% but <30%) was significantly higher in ICSI cycles than in IVF cycles (17.2% vs. 11.4%, p<0.05). The fertilization rate of ICSI among injected oocytes was significantly higher than for IVF (72.3%±24.3% vs. 59.2%±25.9%, p<0.001), but the fertilization rate among retrieved oocytes was significantly higher in IVF than in ICSI (59.2%±25.9% vs. 52.1%±22.5%, p<0.001). Embryo quality before embryo transfer was not different between IVF and ICSI. Although the sperm parameters were not different between the first cycle and the second cycle, split insemination or ICSI was performed in 18 of the 95 cycles in which a second IVF cycle was performed. CONCLUSION: The clinical outcomes did not differ between IVF and ICSI in split insemination cycles. Split insemination can decrease the risk of total fertilization failure. However, unnecessary ICSI is carried out in most split insemination cycles and the use of split insemination might make ICSI more common.

Serum and seminal plasma insulin-like growth factor-1 in male infertility.

OBJECTIVE: Growth hormone and its mediator, insulin-like growth factor-1 (IGF-1), have been suggested to exert gonadotropic actions in both humans and animals. The present study was conducted to assess the relationship between serum IGF-1 concentration, seminal plasma concentration, and sperm parameter abnormalities. METHODS: A total of 79 men were enrolled in this study from December 2011 to July 2012 and were prospectively analyzed. Patient parameters analyzed included age, body mass index, smoking status, urological history, and fertility history. Patients were divided into four groups based on their semen parameters: normal (A, n=31), abnormal sperm motility (B, n=12), abnormal sperm morphology (C, n=20), and two or more abnormal parameters (D, n=16). Patient seminal plasma and serum IGF-1 concentrations were determined. RESULTS: Patient baseline characteristics were not significantly different between any of the groups. The serum IGF-1 levels in groups B, C, and D were significantly lower than the levels in group A; however, the seminal plasma IGF-1 levels were not significantly different between any of the groups. CONCLUSION: Men with abnormal sperm parameters had significantly lower levels of serum IGF-1 compared with men with normal sperm parameters. Seminal plasma IGF-1 levels, however, did not differ significantly between the groups investigated here. Further investigations will be required to determine the exact mechanisms by which growth hormone and IGF-1 affect sperm quality.

Effects of maternal age on embryo quality and pregnancy outcomes using testicular sperm with intracytoplasmic sperm injection.

OBJECTIVE: The aim of this study was to evaluate the influence of maternal age on fertilization, embryo quality, and clinical pregnancy in patients undergoing intracytoplasmic sperm injection (ICSI) using testicular sperm from partners with azoospermia. METHODS: A total of 416 ICSI cycles using testicular spermatozoa from partners with obstructive azoospermia (OA, n=301) and non-obstructive azoospermia (NOA, n=115) were analyzed. Female patients were divided into the following age groups: 27 to 31 years, 32 to 36 years, and 37 to 41 years. The rates of fertilization, high-quality embryos, clinical pregnancy, and delivery were compared across maternal age groups between the OA and NOA groups. RESULTS: The rates of fertilization and high-quality embryos were not significantly different among the maternal age groups. Similarly, the clinical pregnancy and delivery rates were not significantly different. The fertilization rate was significantly higher in the OA group than in the NOA group (p<0.05). Age-group analysis revealed that the fertilization and high-quality embryo rates were significantly different between the OA and NOA groups in patients aged 27 to 31 years old, but not for the other age groups. Although the clinical pregnancy and delivery rates differed between the OA and NOA groups across all age groups, significant differences were not observed. CONCLUSION: In couples using testicular sperm from male partners with azoospermia, pregnancy and delivery outcomes were not affected by maternal age. However, women older than 37 years using testicular sperm from partners with azoospermia should be advised of the increased incidence of pregnancy failure.

Artificial oocyte activation in intracytoplasmic sperm injection cycles using testicular sperm in human in vitro fertilization.

OBJECTIVE: Artificial oocyte activation (AOA) is an effective method to avoid total fertilization failure in human in vitro fertilization-embryo transfer (IVF-ET) cycles. AOA performed using a calcium ionophore can induce calcium oscillation in oocytes and initiate the fertilization process. We evaluated the usefulness of AOA with a calcium ionophore in cases of total fertilization failure in previous cycles and in cases of severe male factor infertility patients with non-motile spermatozoa after pentoxifylline (PF) treatment. METHODS: The present study describes 29 intracytoplasmic sperm injection (ICSI)-AOA cycles involving male factor infertility at Cheil General Hospital from January 2006 to June 2013. Patients were divided into two groups (control, n=480; AOA, n=29) depending on whether or not AOA using a calcium ionophore (A23187) was performed after testicular sperm extraction-ICSI (TESE-ICSI). The AOA group was further split into subgroups according to sperm motility after PF treatment: i.e., motile sperm-injected (n=12) and non-motile sperm-injected (n=17) groups (total n=29 cycles). RESULTS: The good embryo rate (52.3% vs. 66.9%), pregnancy rate (20.7% vs. 52.1%), and delivery rate (10.3% vs. 40.8%) were lower in the PF/AOA group than in the control group. When evaluating the effects of restoration of sperm motility after PF treatment on clinical outcomes there was no difference in fertilization rate (66.6% vs. 64.7% in non-motile and motile sperm, respectively), pregnancy rate (17.6% vs. 33.3%), or delivery rate (5.9% vs. 16.7%) between the two groups. CONCLUSION: We suggest that oocyte activation is a useful method to ensure fertilization in TESE-ICSI cycles regardless of restoration of sperm motility after PF treatment. AOA may be useful in selected patients who have a low fertilization rate or total fertilization failure.

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

This study was performed to assess and compare the outcomes of testicular sperm extraction (TESE)-intracytoplasmic sperm injection (ICSI) using spermatozoa from fresh and frozen testicular tissue from men with subgroups of non-obstructive azoospermia (NOA). A total of 110 cycles of TESE-ICSI were performed. Patients were classified into one of the following NOA subgroups: hypospermatogenesis (HS), maturation arrest (MA), or Sertoli cell-only syndrome (SCO). Laboratory (fertilization, cleavage stage of embryo, and good quality embryo) and clinical (pregnancy, clinical pregnancy, implantation, and delivery) outcomes were assessed. No statistically significant differences were observed in any of the other measured parameters between the three subgroups of NOA. No significant differences in laboratory outcomes were observed between spermatozoa from fresh and frozen testicular spermatozoa; however, statistically significant differences were observed in the pregnancy and implantation rates between groups (p < 0.05). The outcomes of using spermatozoa retrieved from fresh testicular tissue in each of the three subgroups were also compared; although clinical outcomes showed low results, no significant differences were observed between the three subgroups. Similarly, no significant differences were observed in spermatozoa retrieved from frozen testicular tissue. Once spermatozoa have been successfully obtained, acceptable laboratory outcomes can be achieved for NOA, whether or not the spermatozoa are cryopreserved. However, satisfactory clinical outcomes may be more difficult to achieve as the results showed in each group of fresh and frozen testicular spermatozoa. Therefore, achieving acceptable clinical pregnancy results and efficient cryopreservation of testicular spermatozoa should be considered in patients with NOA.

Observation of sperm-head vacuoles and sperm morphology under light microscope.

OBJECTIVE: The presence of sperm-head vacuoles has been suspected to be deleterious to the outcomes of assisted reproductive technology (ART). It is difficult to accurately distinguish morphologically abnormal sperm with vacuoles under a light microscope. This study was performed to analyze the result of the observation of sperm-head vacuoles using Papanicolaou staining under a light microscope and whether the male partner's age affects these vacuoles. METHODS: Sperm morphology with vacuoles was evaluated using Papanicolaou staining and observed under a light microscope (400×) in 980 men. The normal morphology was divided into three categories (group A, <4% of normal morphology; group B, 4%-14% of normal morphology; and group C, >14% of normal morphology). The criteria for the sperm-head vacuoles were those given in the World Health Organization manual. For the analysis of the age factor, the participants were divided into the following groups: 26-30 years, 31-35 years, 36-40 years, 41-45 years, and 46-50 years. RESULTS: The percentage of sperm-head vacuoles increased with normal sperm morphology (group A vs. groups B, C) (p<0.05). In the case of the age factor, a statistically significant difference was not observed across any of the age groups. CONCLUSION: A majority of the sperm-head vacuoles showed a statistically significant difference among normal morphology groups. Therefore, we should consider the probability of the percentage of sperm-head vacuoles not increasing with age but with abnormal sperm morphology. A further study is required to clarify the effect of the sperm-head vacuoles on ART outcomes.

Do cigarette smoking and obesity affect semen abnormality in idiopathic infertile males?

PURPOSE: This study was conducted to find the relative risk of semen abnormality with respect to smoking history and obesity. MATERIALS AND METHODS: Subfertile or infertile men were enrolled in this study from July 2010 to June 2011. All participants provided their cigarette use information, self-reported weight, height, semen analysis, physical examination, and sexually transmitted disease status. None of the enrolled patients had any specific pathological reason for infertility. Semen abnormality was defined as a condition in which one or more parameters did not satisfy the World Health Organization's criteria. RESULTS: A total of 1,073 male patients were considered for this study. After the application of the inclusion criteria, 193 patients were finally analyzed. These patients were divided into two groups according to semen abnormality: the normal semen group (n=72) and the abnormal semen group (n=121). Baseline characteristics, except age and smoking history, were not significantly different between the two groups. Smoking history and age were risk factors for the semen abnormality of idiopathic infertile male patients. CONCLUSIONS: Smoking and old age were risk factors for semen abnormality. However, obesity did not affect the semen abnormality. Smoking affected semen quality and is therefore expected to play a negative role in conception.

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

PURPOSE: To evaluate the viability of frozen embryos generated by intracytoplasmic sperm injection (ICSI) with frozen testicular spermatozoa. METHODS: A total of 68 fresh embryo transfer (ET) cycles and 85 subsequent frozen-thawed ET (FET) cycles were grouped according to the source of spermatozoa: fresh testicular spermatozoa (TESE) or frozen-thawed testicular spermatozoa (t-TESE). RESULTS: There were no significant differences in the age of female patients, number of oocytes, or fertilization rates in fresh ET cycles with TESE (TESE-fresh ET) versus t-TESE (t-TESE-fresh ET). The rate of embryo survival after thawing (95.7 % vs. 94.0 %) was similar in frozen ET cycles (FET) with TESE (TESE-FET) and with t-TESE (t-TESE-FET). While there were significant differences in the proportion of good quality embryos, no statistical differences were found in the pregnancy or clinical abortion rates between the two groups. Moreover, delivery rates were not significantly different. CONCLUSIONS: Although the proportion of good quality embryos was affected by cryopreservation of testicular tissue, embryo survival rate was not. As well, subsequent pregnancy could be achieved successfully via t-TESE-FET cycles. Therefore, FET is not affected by the cryopreservation of testicular tissue, and avoids further oocyte retrieval and TESE procedures.

Comparison of the clinical outcomes of day 4 and 5 embryo transfer cycles.

OBJECTIVE: The majority of embryo transfers (ETs) to date have been performed on day 3 to reduce the potential risk of developmental arrest of in vitro cultured embryos before ET. Development of sequential media has significantly improved culture conditions and allowed blastocyst transfer on day 5. While day 5 ET provides higher clinical pregnancy outcomes with reduced risks of multiple pregnancies, it still has potential risks of developmental arrest of IVF embryos. The aim of this study was to evaluate the clinical outcomes of day 4 ETs and compare the efficacy of day 4 ET with day 5 ET. METHODS: From 2006 to 2009, a total of 747 fresh IVF-ET cycles were retrospectively analyzed (day 4, n=440 or and day 5, n=307). The cycles with any genetic factors were excluded. The rates of matured oocytes, fertilization, good embryos, and clinical pregnancy of the two groups were compared. The chi-square test and t-test were used for statistical analysis. RESULTS: There were no significant differences between the two groups with respect to the mean age of the females and rates of matured oocytes. The pregnancy outcomes of day 4 ET (40.7%) were similar to those of day 5 ET (44.6%). The implantation rate of day 5 ET (24.2%) was significantly higher than that of day 4 ET (18.4%) (p=0.003). CONCLUSION: Day 4 ET can be chosen to avoid ET cancellation in day 5 ET resulting from suboptimal circumstances in the IVF laboratory, but the decremented quality of embryos for transfer and the decreased pregnancy rate must be taken into consideration.

Cryopreservation of mouse spermatogonial stem cells in dimethylsulfoxide and polyethylene glycol.

Assisted reproductive techniques involving isolation, culture, and transplantation of spermatogonial stem cells (SSCs) have the potential to create transgenic livestock and to treat male infertility caused by cancer treatments such as chemotherapy or radiation. Because stem cells may need to be preserved for several years before reintroduction to the patients' testes, efficient SSC cryopreservation techniques need to be developed. SSCs can reinitiate spermatogenesis in recipient testes after freezing; however, optimal cryopreservation protocols have not been identified. The objective of this study was to develop an efficient cryopreservation method for SSCs using permeable cryoprotectant agents (PCAs) or additive cryoprotectant agents (ACAs). To identify an efficient cryopreservation method, populations of mouse testis cells enriched for SSCs were cultured in vitro and frozen using conventional freezing media containing various PCAs or ACAs for 1 wk or 1, 3, 6, 12, or 24 mo. Additionally, various molecular weights and concentrations of polyethylene glycol (PEG) were evaluated. Recovery rate, culture potential, and stem cell activity were significantly greater for cells frozen in 2.5% PEG with a molecular weight of 1000 compared to other treatment groups. These cells also retained the ability to colonize recipient testes, generate normal spermatogenesis, and contribute to viable offspring. The systematic analysis of many cryoprotectant agents indicates that 2.5% PEG (molecular weight 1000) is the most effective agent for efficient SSC cryopreservation.

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

OBJECTIVE: This study was performed to evaluate testicular sperm chromatin condensation using aniline blue-eosin (AB-E) staining and its effects on IVF-ET. METHODS: Chromatin 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. RESULTS: The 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. CONCLUSION: Chromatin 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.

Poor sperm quality affects clinical outcomes of intracytoplasmic sperm injection in fresh and subsequent frozen-thawed cycles: potential paternal effects on pregnancy outcomes.

OBJECTIVE: To evaluate objectively whether poor sperm quality affects sequential events from fertilization to delivery in fresh intracytoplasmic sperm injection (ICSI) and subsequent frozen-thawed embryo transfer (ET) cycles. DESIGN: Retrospective study. SETTING: University-based centers for reproductive medicine. PATIENT(S): For unbiased comparison, 206 cycles were chosen from 1,999 cycles of patients who underwent ICSI-ET and/or subsequent frozen-thawed ET. Cycles met the following criteria: day 3 ET; female age, <40 y; number of retrieved oocytes, >or=5; no split insemination; and no female factors but tubal factor. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): The rates of fertilization, embryo implantation, clinical pregnancy, and delivery and sequential embryonic score (SES) were compared between normal-spermatogenesis patients (NSPs) and defective-spermatogenesis patients (DSPs). RESULT(S): Although sum SES, mean SES, and top SES of transferred embryos on day 3 were similar between NSPs and DSPs, the rates of implantation, clinical pregnancy, and delivery of NSPs were significantly higher than those of DSPs. Furthermore, subsequent ET cycles with frozen-thawed embryos in NSPs and DSPs who failed to achieve pregnancy in their fresh cycles showed that rates of implantation and clinical pregnancy also were significantly lower in DSPs. CONCLUSION(S): Quality of sperm may influence embryo implantation and subsequent pregnancy outcomes without impairment of embryo quality.

Involvement of the Fas-Fas ligand system and active caspase-3 in abnormal apoptosis in human testes with maturation arrest and Sertoli cell-only syndrome.

OBJECTIVE: To provide evidence that the Fas-mediated apoptotic process may participate in developing hypospermatogenesis, such as maturation arrest (MA) and Sertoli cell-only syndrome (SCO). DESIGN: Prospective clinical and descriptive study. SETTING: Hospital-based infertility research laboratory and university laboratory. PATIENT(S): Twenty-two testicular biopsy specimens were obtained for routine clinical purposes from 12 men with nonobstructive azoospermia and from 10 men with obstructive azoospermia. INTERVENTION(S): In situ terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining was used for detection of apoptosis. Reverse transcription-polymerase chain reaction and immunohistochemical analyses were used for detection of testicular expressions of Fas, Fas ligand (FasL), and caspase-3. MAIN OUTCOME MEASURE(S): Apoptotic indices and testicular expressions of apoptosis regulators. RESULT(S): Increased apoptosis of germ cells and Sertoli cells was observed in MA and SCO compared with normal spermatogenesis. In testes with MA, increased immunostaining for FasL was observed in the Sertoli cells and Leydig cells, while intense immunostaining of Fas was observed in primary degenerating spermatocytes. Active caspase-3 immunostaining was detected in the cytoplasm of both Sertoli cells and germ cells. In cases of SCO, expression of Fas, FasL, and active caspase-3 was detected both in Sertoli cells and in hyperplastic interstitial cells. CONCLUSION(S): The current study demonstrates that the expression of FasL is upregulated in the testes of patients with SCO and MA, which suggests that it may be associated with apoptotic elimination or altered maturation of Fas-expressing germ cells through the activation of caspase-3.

Successful testicular sperm extraction in Korean Klinefelter syndrome.

OBJECTIVES: To evaluate the clinical features of Korean Klinefelter syndrome (KS) and the outcome of intracytoplasmic sperm injection using fresh testicular spermatozoa obtained by testicular sperm extraction from these infertile men. Most patients with KS have a nonmosaic 47,XXY karyotype, and about 3% of males with KS are mosaic (46,XY/47,XXY). Great variability is present in the clinical findings. Testicular sperm extraction in conjunction with intracytoplasmic sperm injection has been performed in men with KS in the hope of finding spermatozoa, and some men with KS have had sperm retrieved successfully and achieved pregnancy. METHODS: From January 2000 to January 2002, 42 azoospermic patients were diagnosed with KS after cytogenic evaluation. The karyotypes for all patients were determined by cytologic analysis, including the evaluation of more than 30 peripheral blood lymphocyte metaphases. The patients underwent history and physical examination. The testicular volume was measured using an orchidometer. Semen analysis was performed twice according to the methods recommended by the World Health Organization. Their hormonal profile was also assessed. RESULTS: Our study groups consisted of 36 patients with KS, 11 with mosaic and 25 with nonmosaic KS. Testicular sperm extraction was performed in all 36 patients. In 10 patients (27%), mature sperm were found in the wet preparation and were used for intracytoplasmic sperm injection. No statistically significant correlation was found between sperm extraction and age, serum follicle-stimulating hormone, serum testosterone, or testicular volume. The sperm retrieval rate of the patients with mosaic and nonmosaic KS was 54.5% and 16%, respectively. Intracytoplasmic sperm injection using fresh testicular spermatozoa resulted in a total fertilization rate of 50%. CONCLUSIONS: In our study, patients with mosaic KS had a greater rate of successful sperm retrieval than did those with nonmosaic KS. However, age, serum follicle-stimulating hormone, testosterone level, and testicular volume had no statistically significant relationship with successful sperm retrieval. Testicular sperm obtained from patients with KS induced fertilization and delivery of chromosomally normal children.

Influence of motility on the outcome of in vitro fertilization/intracytoplasmic sperm injection with fresh vs. frozen testicular sperm from men with obstructive azoospermia.

OBJECTIVE: To assess the efficacy of fresh vs. frozen testicular sperm on fertilization and pregnancy using intracytoplasmic sperm injection. DESIGN: Retrospective study. SETTING: Hospital-based infertility research laboratory. PATIENT(S): One hundred sixty patients with obstructive azoospermia undergoing testicular sperm extraction (TESE). INTERVENTION(S): Sections of seminiferous tubule were cryopreserved after TESE. Sperm motility and fertilizing ability were determined after thawing seminiferous tubule sections. MAIN OUTCOME MEASURES: Sperm motility and optimal fertilization and pregnancy rate. RESULT(S): Intracytoplasmic sperm injection was performed using fresh testicular sperm (fresh-sperm group; 84 cases) and thawed seminiferous tubules (thawed-sperm group; 177 cases). The overall fertilization rate was 65.4%, and the pregnancy rate was 34.0%. In the fresh-sperm group, the fertilization rate was 70.9%, and the pregnancy rate was 38.8%. In the thawed-sperm group, the fertilization rate was 62.7%, and the pregnancy rate was 21.7%. Fertilization rates were higher using fresh motile sperm vs. nonmotile sperm (77.0% vs. 29.3%). Pregnancy rates were higher using fresh motile sperm vs. nonmotile sperm (44.3% vs. 20.0%). The fertilization and pregnancy rates of motile vs. nonmotile sperm extracted from the thawed seminiferous tubule were 70.0% vs. 50.9% and 33.9% vs. 27.3%, respectively. Motile spermatozoa could be obtained several hours after thawing in most of the cases. CONCLUSION(S): Optimal fertilization and pregnancy rates were achieved using fresh vs. frozen sperm obtained using TESE when motile sperm were identified. Motile spermatozoa provided superior results to nonmotile sperm and are necessary for optimal fertilization and pregnancy outcomes.

Alternatively spliced variants of the follicle-stimulating hormone receptor gene in the testis of infertile men.

OBJECTIVE: To investigate whether or not alternatively spliced variants of the FSH receptor gene occur in human testis and whether the presence of the splicing variants is associated with spermatogenic defects and serum FSH concentration in infertile men. DESIGN: A prospective case control study. SETTING: An IVF clinic and infertility laboratory at a university hospital. PATIENT(S): Forty-three infertile patients undergoing testicular biopsy. INTERVENTION(S): Total RNA was extracted from the testicular tissues and used for reverse transcriptase-polymerase chain reaction (RT-PCR). MAIN OUTCOME MEASURE(S): Expression pattern was analyzed by nested RT-PCR using primers designed to amplify a fragment of FSH receptor gene. PCR products of splicing variants were cloned and sequenced. RESULT(S): The PCR products showed three kinds of additional bands corresponding to alternatively spliced isoforms of the FSH receptor gene. Exon 9 deleted variant was detected in all patients and inclusion variant of small extra exon was detected in 64% (9/14) of the patients with normal spermatogenesis and 34% (10/29) of the patients with spermatogenic defects. The presence of inclusion variant was not significantly associated with spermatogenic defects but was associated with a low level of serum FSH. On the other hand, exon 6 deleted variant was detected in only one patient having a high level of FSH concentration (30 IU/L) and Sertoli cell only syndrome. CONCLUSION(S): We identified three different types of alternatively spliced variants of the human FSH receptor. However, it is not clear whether or not there is an association between three variants and spermatogenic defects.