Comparison of the Effects of Single Sperm Cryopreservation and Conventional Cryopreservation on Embryo Culture and Pregnancy in Patients Undergoing Micro-TESE / 中山大学学报(医学科学版)

ObjectiveTo compare the effects of single sperm cryopreservation and conventional cryopreservation on embryo culture and pregnancy in patients undergoing microdissection testicular sperm extraction （micro-TESE）. MethodsA retrospective analysis was done on the patients who underwent micro-TESE at the Reproductive Medicine Center in the Sixth Affiliated Hospital of Sun Yat-Sen University between January 2018 and December 2021. The single sperm cryopreservation group included 39 patients undergoing single sperm cryopreservation and 307 MII oocytes. The conventional cryopreservation group included 39 patients undergoing conventional cryopreservation and 410 MII oocytes. Propensity score matching （PSM） was performed to balance the selection bias. The fertility rate， embryo culture and pregnancy of these two groups were compared. ResultsThere was no statistical difference in age， body mass index （BMI）， years of infertility， basal FSH， basal LH， basal E2， AMH， AFC， number of oocytes retrieved and number of transferred embryos between the two groups （P>0.05）. No significant difference was found in fertilization rate （65.8% vs. 60.49%）， available embryo rate （67.82% vs. 58.87%） and high-quality embryo rate （70.80% vs. 75.34%）. The single sperm cryopreservation group had significantly lower clinical pregnancy rate than conventional cryopreservation group （45.45% vs. 70.0%， P=0.049）. ConclusionIf the patients undergoing micro-TESE have very few sperms， single sperm cryopreservation could be an effective choice to increase the utilization of each sperm and ensure the subsequent sperm retrieval after thawing， but the clinical pregnancy rate is decreased. Due to the limited number of cases， we need a large additional number of cases to observe and analyze.

Study on the correlation between male serum calcitonin gene-related peptide and the embryo culture outcome after in vitro fertilization / 中华检验医学杂志

Objective:To investigate the correlation between serum calcitonin gene-related peptide (CGRP) and the culture outcome of in vitro fertilization embryo in male patients with infertility.Methods:In this study, the randomized samples from 25 male patients who received in vitro fertilization-embryo transfer (IVF-ET) in the 1st Affiliated Hospital of Fujian Medical University from June 2019 to October 2019 were analyzed by multiple linear stepwise regression, with some important clinical outcomes, such as the logarithmic conversion index of serum CGRP, fertilization method, masturbation difficulty, age, infertility duration, and prolactin, as independent variables, while total fertilization rate, normal fertilization rate, high quality embryo rate at day 3, blastocyst formation rate as dependent variables.Results:The Pearson correlation analysis showed that the D3 high-quality embryo rate was related to the normal sperm morphology rate in the primary infertility group ( r=0.537, P=0.048), the blastocyst formation rate was correlated with sperm density ( r=0.760, P=0.002), the CGRP logarithm was correlated with the total fertilization rate ( r=0.693, P=0.006). The logarithmic conversion index of serum CGRP was related to the total fertilization rate and normal fertilization rate in the secondary infertility group ( r=0.614, P=0.042 and r=0.611, P=0.046). In the secondary infertility group, there was a linear relationship between normal fertilization rate and total sperm count, serum CGRP log conversion, and sperm normal morphology rate, with standardized regression coefficients of 0.2, -0.729, and 6.8, respectively. Conclusion:Serum CGRP level, together with total sperm count and normal sperm morphology rate may affects normal fertilization rate in male patients with infertility.

Protein profile as a quality indicator of cryopreserved semen from Tambaqui Colossoma macropomum (Cuvier, 1818) / Perfil protéico como indicador qualitativo do semen criopreservado de Tambaqui Colossoma macropomum (Cuvier, 1818)

Abstract The aim of this study was to evaluate the association between proteins in the seminal plasma of tambaqui Colossoma macropomum (Cuvier, 1818) with seminal quality indicators after thawing. The semen was cryopreserved with a dilution based on BTS with 8% DMSO. A 200 µL sample of semen from each animal was diluted in 800 µL BTS, centrifuged at 800 rpm, and the supernatant was cryopreserved to further analyze of the protein profile of seminal plasma through one-dimensional electrophoresis (SDS-PAGE). After 15 days of cryopreservation, a cryopreserved semen straw was thawed to analyze both qualitative and quantitative parameters. When considering all collections, the SDS-PAGE identified 15 protein bands in the seminal plasma of tambaqui. When the interaction (presence or absence) between proteins observed in the seminal plasma and the post thawed spermatic parameters was evaluated, we observed a great influence of the presence of proteins on spermatic quality. A greater (P<0.05) fertilization rate was observed with the presence of proteins 12, 34, 44, 85, and 90 kDa. Proteins in seminal plasma of tambaqui influenced the spermatic quality after thawing, and thus, they can be utilized as an indicator of sperm quality, especially the proteins with a molecular weight ≤ 50 kDa.

Resumo O objetivo desse estudo foi de avaliar a associação entre a presença de proteínas no plasma seminal do tambaqui Colossoma macropomum (Cuvier, 1818) com indicadores de qualidade seminal pós-descongelamento. O semen foi criopreservado com diluidor a base de BTS com 8% DMSO. Uma amostra de 200 µL de semen de cada animal foi diluída em 800 µL de BTS, e centrifugada em 800 rpm, e somente o sobrenadante foi criopreservado para posterior análise do perfil proteico do plasma seminal, através da eletroforese unidimensional (SDS-PAGE). Decorridos 15 dias da criopreservação, uma palheta com semen criopreservado foi descongelado para análise dos parâmetros quali-quantitativos. Considerando todas as coletas, o SDS-PAGE identificou 15 bandas proteicas no plasma seminal do tambaqui. Quando se avaliou a interação (presença ou ausência) das proteínas encontradas no plasma seminal, com os parâmetros espermáticos pós-descongelamento, observou-se grande influência da presença das proteínas na qualidade espermática. Observou-se maior taxa de fertilização (P<0,05) com a presença das proteínas 12, 34, 44, 85 e 90 kDa. As proteínas do plasma seminal de tambaqui influenciaram na qualidade espermática após descongelamento, podendo ser utilizadas como indicadores para a qualidade espermática após descongelamento, principalmente as proteínas com peso molecular ≤50 kDa.

Protein profile as a quality indicator of cryopreserved semen from Tambaqui Colossoma macropomum (Cuvier, 1818)

Abstract The aim of this study was to evaluate the association between proteins in the seminal plasma of tambaqui Colossoma macropomum (Cuvier, 1818) with seminal quality indicators after thawing. The semen was cryopreserved with a dilution based on BTS with 8% DMSO. A 200 µL sample of semen from each animal was diluted in 800 µL BTS, centrifuged at 800 rpm, and the supernatant was cryopreserved to further analyze of the protein profile of seminal plasma through one-dimensional electrophoresis (SDS-PAGE). After 15 days of cryopreservation, a cryopreserved semen straw was thawed to analyze both qualitative and quantitative parameters. When considering all collections, the SDS-PAGE identified 15 protein bands in the seminal plasma of tambaqui. When the interaction (presence or absence) between proteins observed in the seminal plasma and the post thawed spermatic parameters was evaluated, we observed a great influence of the presence of proteins on spermatic quality. A greater (P 0.05) fertilization rate was observed with the presence of proteins 12, 34, 44, 85, and 90 kDa. Proteins in seminal plasma of tambaqui influenced the spermatic quality after thawing, and thus, they can be utilized as an indicator of sperm quality, especially the proteins with a molecular weight 50 kDa.

Resumo O objetivo desse estudo foi de avaliar a associação entre a presença de proteínas no plasma seminal do tambaqui Colossoma macropomum (Cuvier, 1818) com indicadores de qualidade seminal pós-descongelamento. O semen foi criopreservado com diluidor a base de BTS com 8% DMSO. Uma amostra de 200 µL de semen de cada animal foi diluída em 800 µL de BTS, e centrifugada em 800 rpm, e somente o sobrenadante foi criopreservado para posterior análise do perfil proteico do plasma seminal, através da eletroforese unidimensional (SDS-PAGE). Decorridos 15 dias da criopreservação, uma palheta com semen criopreservado foi descongelado para análise dos parâmetros quali-quantitativos. Considerando todas as coletas, o SDS-PAGE identificou 15 bandas proteicas no plasma seminal do tambaqui. Quando se avaliou a interação (presença ou ausência) das proteínas encontradas no plasma seminal, com os parâmetros espermáticos pós-descongelamento, observou-se grande influência da presença das proteínas na qualidade espermática. Observou-se maior taxa de fertilização (P 0,05) com a presença das proteínas 12, 34, 44, 85 e 90 kDa. As proteínas do plasma seminal de tambaqui influenciaram na qualidade espermática após descongelamento, podendo ser utilizadas como indicadores para a qualidade espermática após descongelamento, principalmente as proteínas com peso molecular 50 kDa.

Seminal plasma anti-Müllerian hormone and inhibin B and serum inhibin B in predicting the outcome of routine IVF fertilization / 中华男科学杂志

Objective@#To analyze the correlations of seminal plasma (sp) anti-Müllerian hormone (spAMH) and inhibin B (spINHB) and serum INHB (serINHB) with semen parameters in oligoasthenospermia patients and explore their value in predicting the outcome of routine in vitro fertilization (IVF).@*METHODS@#We obtained the levels of spAMH, spINHB and serINHB as well as semen parameters from 88 infertile males undergoing IVF due to oligoasthenospermia or female uterine tubal factors from August 2016 to February 2017. Using the ROC curve and Pearson's correlation analysis, we examined the effects of the obtained parameters on the fertilization rate and assessed the correlation of the levels of spAMH, spINHB and serINHB with the semen parameters of the patients.@*RESULTS@#Concerning the predictive value for the outcome of IVF, Pearson's correlation analysis showed that the area under the ROC curve (AUC) of spAMH was 0.807 (sensitivity = 84.6%, specificity = 76%, cut-off point = 3.529, P <0.001) and that of spINHB was 0.768 (sensitivity = 84.6%, specificity = 88.7%, cut-off point = 31.117, P = 0.002). The serINHB level was found positively correlated with sperm concentration (r = 0.346, P = 0.001), total sperm count (r = 0.378, P <0.001), sperm motility (r = 0.521, P <0.001), and the percentage of progressively motile sperm (r = 0.343, P = 0.001).@*CONCLUSIONS@#The levels of spAMH and spINHB can be used as laboratory indexes to predict the fertilization rate of routine IVF and are correlated with semen parameters in oligoasthenospermia patients, while that of serINHB has a positive correlation with the semen parameters of the patients.

Oocyte quality of tambaqui (Colossoma macropomum) during the reproductive season / Qualidade ovocitária de tambaqui (Colossoma macropomum) ao longo da estação reprodutiva

The study aimed to analyze the Colossoma macropomum reproductive behavior and quality of the female gametes throughout the reproductive season. The experiment was carried out in Pimenta Bueno - Rondônia State (Northern Brazil) during the reproductive season (2010-2011) using 36 females. Each sampling was performed on a 15 ± 5 days interval. Female gametes were collected by stripping and the following analyses were performed: weight of oocytes released (g); productivity index, fertilization and hatching rate. During the sampling period was verified effect (p < 0.05) of collecting time into the season for oocytes weight, productivity index and fertilization rate. Although the period 3 (December) did not differ significantly from other periods, it showed better parameters for the quality of C. macropomum oocytes.

O estudo foi conduzido com o objetivo de analisar o comportamento reprodutivo da espécie Colossoma macropomum, quanto à qualidade de seus gametas femininos ao longo da estação reprodutiva. O experimento foi executado em Pimenta Bueno-Rondônia durante a estação reprodutiva do C. macropomum. Utilizaram 36 fêmeas durante a estação de 2010-2011. Cada coleta apresentou um intervalo de 15±5 dias. Através da extrusão foram coletados os gametas femininos e realizadas as seguintes análises ao longo da estação: peso de oócitos liberados (g); índice de produtividade; taxa de fertilização e eclosão. Durante a estação 2010-2011 foi verificado efeito (p < 0,05) de período (coleta) dentro da estação para peso de oócitos, índice de produtividade e taxa de fertilização. Apesar do período 3 (coleta – mês de dezembro) não ter diferenciado significativamente de alguns períodos, foi o que apresentou os melhores parâmetros estabelecidos para a qualidade dos oócitos de C. macropomum.

A insemination method to prevent fertilization failure-intracytoplasmic sperm injection in half oocytes of retrieval:A study of 364 cycles / 重庆医科大学学报

Objective:To investigate the indications of half-ICSI and the value of half-ICSI to prevent fertilization failure. Methods:The indications of the cases in the group of fertilization failure(fertilization rate was 0%)or lower fertilization rate(fertilization rate less than 30%) were compared with those in the group(fertilization rate more than 65%). Results:The fertilization rate of IVF(in vitro fertilization) in the half-ICSI was lower than that of conventional IVF. The indication of the borderline semen quality in the groups of fertilization failure and lower fertilization rate were 51.2% and 63.3% ,respectively.Compared with the normal fertilization rate group, the indications in the groups of fertilization failure or lower fertilization rate were sig- nificantly differen(tP

Comparison of IVF Outcomes in Patients with Endometriosis According to Severity / 대한불임학회지

OBJECTIVE: To evaluate the impact of endometriosis on IVF-ET cycles and to compare IVF outcomes between stage I/II and stage III/IV endometriosis. METHODS: We analyzed 697 patients (1,199 cycles) with endometriosis (stage I-II: 638 cycles, stage III-IV: 561 cycles) and 325 pts (459 cycles) with tubal factor as controls between January 1994 and April 2004. Pts with endometriosis were diagnosed by laparoscopy and medical and surgical treatment were done in 353 cycles (55.3%) and 466 cycles (83.1%) of stage I-II/stage III-IV endometriosis. Cycles with age>35 years or FSH>20 mIU/mL or severe male factor infertility were excluded. RESULTS: The number of retrieved oocytes (9.97+/-7.2 vs. 13.4+/-7.9 (p<0.0001)), total number of embryos (6.5+/-4.8 vs. 9.1+/-5.6 (p<0.0001)), and good quality embryos (2.43+/-1.6 vs. 2.74+/-1.7 (p=0.013)) significantly decreased in stage III-IV endometriosis than in control. But pregnancy rate of stage III-IV endometriosis was comparable with control (35.7% vs. 36.8%). Fertilization rate and number of total embryos were lower in stage I-II endometriosis than in control (64.8+/-22.9 vs. 70.8+/-20.8 (p<0.0001), 7.6+/-5.0 vs. 9.1+/-5.6 (p<0.0001)). In patients with medical and surgical treatment of endometriosis, pregnancy rate and live birth rate was significantly lower in stage I-II than in stage III-IV endometriosis (29.2 vs. 36.2 (%), p=0.045, 23.9 vs. 31.5 (%), p=0.043). There was no difference in the mean age, but the duration of infertility was significantly longer (56.5+/-26.3 vs. 46.9+/-25.8 (mon), p<0.0001) and fertilization rate was lower (64.7+/-23.3 vs. 70.5+/-22.7 (%), p=0.001) in stage I-II than stage III-IV endometriosis. CONCLUSION: We suggest that IVF should be considered earlier in patients with minimal to mild endometriosis because of significantly decreased fertilization rates.

Comparative Analysis of Pregnancy Outcomes after In Vitro Fertilization with Intracytoplasmic Sperm Injection (IVF-ICSI) between Obstructive and Non-obstructive Azoospermia / 대한불임학회지

OBJECTIVE: To compare the pregnancy outcomes after in vitro fertilization with intracytoplasmic sperm injection (IVF-ICSI) between obstrucvtive and non-obstrucvtive azoospermia. METHODS: From January 1994 to December 2002, 524 patients with obstructive azoospermia (886 cycles) and 163 patients with non-obstructive azoospermia (277 cycles) were included in this study. Microsurgical epididymal sperm aspiration (MESA) or testicular sperm extraction (TESE) in obstructive azoospermia and TESE in non-obstructive azoospermia were perfomed to retrieve sperm, which was used for ICSI and then fertilized embryos were transferred. The results of ICSI-fertlization rate (FR), clinical pregnancy rate (CPR), clinical abortion rate (CAR) and delivery rate (DR)- were statistically analysed in obstructive versus non-obstructive azoospermia. RESULTS: There were no differences in the number of retrieved oocytes, injected oocytes for ICSI and oocyte maturation rate. FR was significantly higher in obstructive than non-obstructive azoospermia (71.7% vs. 61.1%, p<0.001). There was no difference in CPR per embryo transfer cycle. After pregnancy was established, however, CAR was significantly higher in non-obstructive than obstructive azoospermia (25.6% vs. 12.5%, p=0.004). DR per clinical pregnancy cycle was significantly higher in obstructive than non-obstructive azoospermia (78.0% vs. 64.4%, p=0.012). In the karyotype ananlysis of abortus, abnormal karyotypes were found in 75.0% (6/8) of obstructive and 55.6% (5/9) of non-obstructive azoospermia. CONCLUSION: Our data show significantly higher FR in obstructive than non-obstructive azoospermia. Though there was no differrence in CPR, CAR was significantly higher in non-obstructive than obstructive azoospermia. The abortion may be related to the abnormal karyotype of embryo, but further investigations are necessary to elucidate the cause of clinical abortion in azoospermia.

Comparison of Intracytoplasmic Sperm Injection and Partial Zona Dissection followed by Insemination in Hamster Oocytes / 대한불임학회지

OBJECTIVES: This study was to investigate the fertilization rate after intracytoplasmic sperm injection (ICSI) or partial zona dissection (PZD) of human and hamster sperm into hamster oocyte in in vitro fertilization (IVF). In addition, the possibility of clinical application was evaluated by the comparison of usefulness and difference of these method. MATERIALS AND METHODS: Hamster immature oocytes were obtained from oviducts superovulated by PMSG and hCG, and hamster sperms were obtained from epididymis. The freezed human sperms were thawed before use. Fertilization were confirmed by two pronuclei, one pronucleus, swollen sperm head or/and two polar bodies at 7~8 hour after ICSI or PZD. RESULTS: The fertilization rates after ICSI and PZD of human sperm to hamster oocyte were 3.6%, 64.2%, 73.6%, and 55.6% for negative control, positive control, ICSI, and PZD respectively, suggesting that ICSI only showed improved fertilization rate (p<0.01). The fertilization rates after ICSI and PZD of hamster sperm to hamster oocyte were 11.1%, 51.2%, 39.6%, and 72.7% for negative control, positive control, ICSI, and PZD respectively, suggesting that PZD only showed improved fertilization rate (p<0.01). PZD showed significantly higher fertilization rate than ICSI (p<0.05). CONCLUSIONS: As for the fertilization rate by ICSI and PZD using hamster oocyte in IVF, ICSI technique was considered to be more useful for human sperm and PZD technique for hamster sperm. Therefore, ICSI technique was considered more appropriate for experimental application using human sperm and hamster oocyte.

Intracyplasmic Sperm Injection in Patients with Past History of Failed or Poor Fertilization in Previous IVF - ET Cycles : Comparison with Patients with Severe Male Factor / 대한산부인과학회잡지

OBJECTIVE: The purpose of this study was to determine whether intracytoplasmic sperm injection(ICSI) could overcome the defects of oocytes in IVF-ET patients with previous fertilization failure by conventional fertilization technique. Design: Retrospective study Materials and METHODS: A total of 119 ICSI cycles in 57 IVF-ET patients performed from May, 1995 to December, 1997 was enrolled. Subjects were divided into two groups: FR group included 66 ICSI cycles in 35 patients with normal sperm who underwent ICSI due to past history of failed or poor fertilization in the previous IVF-ET cycles, and OAT group included 53 ICSI cycles in 22 patients with severe oligoasthenoterato- zoospermia(OAT) which was defined as sperm concentration < 20 million/ml, mo#dlity < 30% and normal morphology < 4% by strict morphologic criteria. The outcomes of ICSI were analyzed and compared in both groups. RESULTS: The age of female patients, basal serum FSH level, and the numbers of oocytes retrieved and metaphase II oocytes were all comparable in both groups. The fertilization rate after ICSI was similar in both groups(68.7+/-25.3% vs. 67.7+/-24.5%), as were the cleavage rate of normally fertilized oocytes(93.1+/-21.4% vs. 89.3+/-21.6%), the number of embryos transferred(4,00+/-1.98 vs. 4.64+/-2.10), and cumulative embryo score(CES) indicating the quality of embryos(47.3+/-33.2 vs. 54.1+/-33.2). The implantation rate(4.3+/-10.5% vs. 3.8+/-11.0%) and the clinical pregnancy rate per cycle(15.2% vs. 13.2%) were also comparable in both groups. CONCLUSIONS: Although it has been shown that there is a higher risk of chromosomal abnormalities in oocytes from IVF-ET patients with pevious failed or poor fertilization, higher implantation and clinical pregnancy rates wer#e not observed in patients with OAT following ICSL Therefore, the functional defect of sperm such as loss of capacitation, defect of aaasome reaction, and abnormality of nucleus decondensation should be also considered in patients with previous failed or poor fertilization.

Fertilization rate during intracytoplasmic sperm injection procedure comparing 6 and 12 O’clock polar body position.

A Prospective study was designed to compare fertilization rate between the . and 12 o’clock position of the polar body during the intracytoplasmic sperm injection (ICSI) procedure. A total of 128 oocytes were injected by placing the first polar body at 6 o’clock and 12 o’clock alternately. Normal fertilization rates were 70.31 and 71.87 percent respectively. There was no statistical significance between the two groups.

Microassisted Fertilization of Human Oocytes with Intracytoplasmic Sperm Injection in In VitroFertilization Program / 대한산부인과학회잡지

In spite of much progress in in vitro fertilization and embryo transfer(IVF-ET) program,the pregnancy rate remains at 20~30%, and the endometrial implantation rate per embryotransferred at 10%. Although IVF-ET is widely applied in the treatment of coupleswith male factor infertility, it may fail in many infertile couples with normal semen parameters,and certain couples cannot be accepted for standard IVF-ET due to unfertilization orextremely low fertilization rate of oocytes. Recently, several procedures of microassistedfertilization(MAF) using micromanipulation have been introduced, and pregnancies and birthshave been obtained after intracytoplasmic sperm injection(ICSI).This clinical study was performed to develop and establish ICSI as an effective procedureof MAF in infertile couples who could not be accepted for standard IVF-ET becauseof extremely impaired semen characteristics(Group A) and because of failure in fertilizationof extremely low fertilization rate of oocytes with the conventional fertilization technique inthe previous IVF-ET cycles(Group B). From March, 1995 to December, 1996, a total of 114cycles of IVF-ET with ICSI in 65 infertile patients were included in study group, and theoutcomes of ICSI were analyzed according to fertilization rate, cumulative embryo score(CES), and pregnancy rate.In Group A, 34 patients were evaluated with semen score such as number of totalmotile sperms, and then divided into 4 groups accordingly. In 62 ICSI cycles, the numberof oocytes retrieved after controlled ovarian hyperstimulation(COH) was 12.4+/-6.8, and thenumber of oocytes optimal for ICSI procedure was 8.8+/-5.5. The fertilization rate of 65.7+/-23.6% could be obtained after ICSI. The number of embryos transferred was 4.4+/-2.2 withthe mean CES of 50.5+/-34.3 in ICSI cycles. The overall pregnancy rate was 24.2%(15/62)per cycle and 44.1%(15/34) per patient. There were no significant differences in the pregnancyrates among 4 groups. Although more mature oocytes were retrieved, the fertilizationrate was significantly lower in Group A-1 compared with Group A-IV. However, semenscore did not clearly affect the outcomes of ICSI in couples with severe male factor infertility.In Group B, the number of oocytes retrieved after COH was 10.5+/-6.1 in 49 previouscycles, and 10.8+/-5.7 in 52 ICSI cycles. In ICSI cycles, the number of oocytes optimal forICSI procedure was 8.5+/-5.1 with the fertilization rate of 72.4+/-22.5%. The number ofembryos transferred was 1.4+/-2.4 in previous cycles, and 4.7+/-1.8 with the mean CES of 50.4+/-29.9 in ICSI cycles. In ICSI cycles, the overall pregnancy rate was 30.8%(16/52) percycles and 51.6%(16/31) per patients.In conclusion, MAF of human oocytes with ICSI is a promising fertilization method forIVF-ET patients, especially with few spermatozoa for the conventional methods of in vitroinsemination and with the past history of failure in fertilization or low fertilization rate inthe previous cycles, and ICSI using micromanipulation procedures applied to human oocyteswill provide a range of novel techniques which may dramatically improve the pregnancy ratein IVF-ET program and contribute much to the effective management of infertile couples.