Fertilization, Embryo Quality, and Cryosurvival in In Vitro Fertilization and Intracytoplasmic Sperm Injection Cycles

Purpose: Our purpose was to investigate the influence of semen quality on fertilization, embryo morphology, cleavage, and cryosurvival in conventional in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI)programs. Methods: A retrospective analysis of 513 couples undergoing IVF and 255 couples undergoing ICSI was done. Results: Semen quality influenced fertilization in IVF and abnormal fertilization in IVF and ICSI, but no effects on the development, morphology, implantation capacity, or cryosurvival of embryos were found. Fertilization, embryo quality, and cryosurvival rates were similar after IVF and ICSI. The fertilization rate of mature oocytes in IVF was lower when cytoplasmic immaturity in the oocyte population was frequent. The speed of development of embryos was 2 hr faster after ICSI than after IVF. Two-cell–stage embryos survived best after cryopreservation with propanediol and sucrose on day 2. Conclusions: After fertilization, semen parameters had no effect on the quality or cryosurvival of embryos in either IVF or ICSI.     

The Outcome of Cryopreserved Human Embryos After Intracytoplasmic Sperm Injection and Traditional IVF

Objective: Our objective was to analyze the outcome of cryopreserved embryos obtained after intracytoplasmic sperm injection (ICSI) and in vitro fertilization (IVF) in terms of survival rate, implantation rate (IR), total and clinical pregnancy rate (PR) in a retrospective, comparative study. Methods: Three hundred seventy-five IVF and 463 ICSI surnumerary cleaved embryos, frozen on Day 2 with 1,2-propanediol, were thawed. Results: Thirty-two percent of the thawed IVF embryos survived and 11 pregnancies (8 clinical) were obtained from 68 transfers (16.1%). Fourty-seven percent of the ICSI embryos survived, with 19 pregnancies (18 clinical) from 116 transfers (16.4%). The IR was 8.5% (8/94) in IVF cycles and 10.8% (20/185) in ICSI cycles. Conclusions: A significantly better survival rate of ICSI embryos was observed but with no difference in PR, preclinical, and clinical abortion rate, or IR.     

Relationship of the Human Cumulus-Free Oocyte Maturational Profile with In Vitro Outcome Parameters After Intracytoplasmic Sperm Injection

Purpose: We investigated whether the human oocyte maturational profile at the removal of cumulus/corona cells affects the fertilization rate and subsequent embryo quality after intracytoplasmic sperm injection. Methods: A total of 1011 oocytes from 150 cycles was included in this retrospective analysis. Cumulus-free oocytes that were in prophase or metaphase I of meiosis at the removal of cumulus/corona cells were incubated in vitro until they reached metaphase II (in vitro-matured oocytes) and were then immediately injected with a single spermatozoa. Oocytes that were in metaphase II at the removal of cumulus/corona cells (MII oocytes) received sperm injection after 3–4 hr of preinjection incubation. Results: The fertilization rate of the MII oocytes was significantly higher than that of in vitro-matured oocytes (81 vs 62%; P < 0.001). The cleavage rates were similar in the two groups (MII oocytes, 94%; in vitro-matured oocytes, 91%). However, MII oocytes had significantly higher percentages of good-quality embryos (grade 1–3 embryos, 87 vs 58%, P < 0.001) and embryos with high cumulative embryo scores (score 10–32 embryos, 62 vs 33%, P < 0.001). The mean cumulative embryo score of MII oocytes after fertilization was also higher than that of in vitro-matured oocytes (12.1 ± 3.8 vs 8.8 ± 3.4; P = 0.014). Conclusions: MII oocytes that extruded the first polar body at the removal of cumulus/corona cells had better fertilization rates and embryo morphology than in vitro-matured oocytes that extruded the first polar body following the removal of cumulus/corona cells and in vitro culture.     

两种指征的卵胞浆内单精子注射结局的比较

目的:探讨常规IVF受精失败患者再次周期行ICSI-ET治疗时,卵子因素对胚胎结局的影响。方法:回顾分析因前次IVF-ET中受精障碍或受精率≤30%而行ICSI治疗的38个周期(A组)和因严重精液异常而行ICSI治疗的181个周期(B组)的ICSI结局。结果:A、B组的受精率(FR)、卵裂率(CR)、胚胎利用率(URE)、胚胎着床率(EIR)、临床妊娠率(CPR)和早期流产率(EAR)分别为82.5%vs78.0%(P<0.05),97.5%vs97.6%(P>0.05),76.4%vs73.1%(P>0.05),10.0%vs19.8%(P<0.05),27.0%vs31.8%(P>0.05),40.0%和16.1%(P>0.05)。结论:常规IVF-ET受精失败的患者,再次周期行ICSI-ET治疗,受精率提高,由于卵子异常对胚胎发育的不利影响,部分胚胎着床和着床后的远期发育潜能降低。     

Incidence and Perinatal Outcome of Multiple Pregnancies After Intracytoplasmic Sperm Injection Compared to Standard In Vitro Fertilization

Purpose:Our purpose was to assess the incidence of multiple pregnancies and their obstetric outcome after intracytoplasmic sperm injection.Methods:The study group comprised women who delivered twins or triplets after intracytoplasmic sperm injection and standard in vitro fertilization. The incidence and main perinatal outcome of 140 multiple pregnancies resulting from intracytoplasmic sperm injection or standard in vitro fertilization treatment were analyzed.Results:A total of 60 multiple pregnancies was obtained after intracytoplasmic sperm injection (3.4 ± 1.1 embryos/cycle) and 80 after standard in vitro fertilization (3.3 ± 2.0 embryos/cycle). The incidence of multiple pregnancy, i.e., 22.6 compared to 20.7%, respectively, was calculated. The obstetric outcome of 47 multiple pregnancies after intracytoplasmic sperm injection was 39 twin deliveries at between 27 and 37 weeks of gestation (mean, 36 ± 3.3) and 8 successful triplet deliveries between 26 and 36 weeks of gestation (mean 32.6 ± 2.4). The outcome after regular in vitro fertilization was similar. No major malformations were observed.Conclusions:The results of this study showed that the incidence of multiple pregnancies after intracytoplasmic sperm injection was similar to that after standard, conventional in vitro fertilization. The perinatal outcome did not differ between both groups.     

Reproductive Performance of Patients Undergoing Intracytoplasmic Sperm Injection with 100% Implantation Rate

Purpose: To characterize the differences between twomatched groups of patients treated by ICSI: those pregnantafter all embryos transferred implanted (100% implantationrate) compared with nonpregnant patients. Methods: Twenty-one patients in whom one transferredembryo achieved a singleton pregnancy (group A) and 21pregnant patients to whom two or three embryos were transferredand achieved 11 twin and 10 triplet pregnancies(group B) compared with matched nonpregnant patients(group C and D, respectively). Results: The singleton pregnant patients were significantlyolder than the twin and triplet pregnancy patients. Althougha similar number of human menopausal gonadotropinampules were used in the singleton compared with the twinsand triplets a significantly lower number of oocytes andembryos were achieved at lower levels of estradiol on thehuman chorionic gonadotropin day in the former than inthe latter respectively. No difference was found between thepregnant women and their nonpregnant controls in any ofthe mentioned parameters. Good embryo morphology wasfound in 86% of the embryos in group A compared with62% in group C (P = 0.08) and 92% in group B comparedwith 66% in group D (P < 0.002). Conclusions: The only parameter in which pregnant patientswith 100% implantation rate differ from their nonpregnantcontrols was embryo quality.     

非男性因素不孕体外受精失败行卵胞浆内单精子注射的结局分析

目的:探讨卵胞浆内单精子注射(ICSI)对非男性因素不孕IVF失败患者治疗结局的影响。方法:回顾性分析由于第一周期常规IVF治疗中卵子完全不受精或受精率≤25%,行补救性ICSI的10个周期(补救性ICSI组),以及因前次受精失败而在随后的治疗周期中采取ICSI方法受精的19个周期(后续性ICSI组)的ICSI治疗结局,并以因男方少弱精子症进行第1次ICSI治疗的133个周期为对照组。结果:后续性ICSI组受精率、植入率、妊娠率和分娩率均高于补救性ICSI组,但差异均无统计学意义(P>0.05)。后续性ICSI组优胚率显著高于补救性ICSI组(P<0.05);补救性ICSI组受精率(48.9%)、优质胚胎率(29.2%)、植入率(0%)、妊娠率(0%)、分娩率(0%)均显著低于对照组(分别为72.1%、46.6%、21.2%、45.1%、39.1%);后续性ICSI组受精率、植入率、妊娠率、分娩率分别为55.4%、8.8%、21.1%、15.8%,均低于对照组(P<0.05或P<0.01)。优质胚胎率后续性ICSI组(44.2%)低于对照组,但无统计学差异。结论:对于非男性因素不孕IVF失败患者,ICSI能避免受精失败,但是受精率以及妊娠结局受到卵母细胞隐匿性异常的影响。     

The Outcome of Sperm Retrieval and Intracytoplasmic Sperm Injection in Patients with Obstructive Azoospermia: Impact of Previous Tuberculous Epididymitis

Purpose: Our purpose was to investigate the influence of previous tuberculous epididymitis in patients with obstructive azoospermia on the outcome of sperm retrieval and intracytoplasmic sperm injection (ICSI). Methods: Eighty-eight cycles of ICSI were performed in 44 patients with obstructive azoospermia; 16 cycles (7 patients) with tuberculous obstructive azoospermia and 72 cycles (37 patients) with nontuberculous obstructive azoospermia. Results: The rates of fertilization and embryo cleavage were comparable, and there was no significant difference in the clinical pregnancy rate per fresh transfer between the two groups. The rates of embryo implantation and clinical miscarriage were also comparable. Conclusions: Embryo quality and pregnancy outcome in sperm retrieval and ICSI were comparable in both the tuberculous and the nontuberculous obstructive azoospermia patients. Although there was a preponderance of testicular sperm used in the tuberculous obstructive azoospermia group, our results suggest that previous tuberculous epididymitis in patients with obstructive azoospermia does not affect the outcome of sperm retrieval and ICSI.     

Embryo developmental stage at transfer influences outcome of treatment with intracytoplasmic sperm injection

Purpose: Our purpose was to determine if embryo cell stage at the time of intrauterine transfer correlates with pregnancy rate in patients treated with intracytoplasmic sperm injection (ICSI). Methods: We conducted a retrospective analysis of 455 embryo transfer cycles following ICSI and 304 conventional in vitro fertilization (IVF) and embryo transfer cycles in women aged 40 years or less. Abstracted information included grading of the embryo cell stage and quality at the time of transfer. Results: The overall ICSI pregnancy rate was 30.8%, while that of conventional IVF was 29.3%. However, the ICSI pregnancy rate fell to 9.3% for embryo transfers taking place at the two-cell stage but increased to 35.8% when at least one embryo had more than two cells, and this difference was statistically significant (P≤0.0001). The pregnancy rate following conventional IVF was 22.0% when only two-cell embryos were transferred and 32.0% when at least one of the embryos had more than two cells, but this difference in pregnancy rates was not significant (P>0.05). Conclusions: The stage of embryo development at transfer appears to exert a powerful influence on the successful establishment of pregnancy after ICSI.     

Relationship Between the Sperm Motility Index Assessed by the Sperm Quality Analyzer and the Outcome of Intracytoplasmic Sperm Injection

Purpose: Intracytoplasmic sperm injection (ICSI) has been validated as a useful treatment in severe male-factor patients who could not achieve fertilization and live births by conventional in vitro fertilization treatment. To examine the impact of male factors on ICSI outcome, clinical laboratory data were retrospectively analyzed. Methods: One hundred two cycles of ICSI treatment indicated by severe male-factor infertility were entered into this study. Sperm parameters including sperm motility, sperm concentration, and sperm motility index assessed by the Sperm Quality Analyzer were evaluated. Results: Five hundred seventy-six metaphase II oocytes retrieved were manipulated. The normal fertilization (2 PN) rate per oocyte was 64.9 ± 26.0% (mean ± SD). Of the 99 transfers, 31 clinical pregnancies were obtained, yielding an average pregnancy rate of 31.3% per transfer. The mean sperm motility, sperm concentration, and sperm motility index were 20.3 ± 16.1% (range, 0 to 50%), 18.2 ± 25.1 × 10 ⁶ /ml (range, <1 to 150 × 10 ⁶ /ml), and 31.2 ± 45.0 (range, 0 to 220), respectively. Sperm concentration did not have a significant impact on fertilization rate by ICSI. In four cases, ICSI was performed using totally immotile sperm and the fertilization rate was 43.5%, which was significantly lower than that of some of the other sperm motility groups, and no pregnancy could be achieved. In 14 cases in which the sperm motility index assessed by the Sperm Quality Analyzer was 0, the fertilization rate (50.0%) was significantly lower than in most of the other sperm motility index groups. Conclusions: These findings suggest that in severe malefactor cases with totally immotile sperm or a sperm motility index of 0, the selection of good-quality sperm should be verified before injection.     

男性不育患者Y染色体多态对卵胞浆内单精子注射妊娠结局的影响

目的:探讨Y染色体多态对卵胞浆内单精子注射(ICSI)治疗结局的影响。方法:因男性严重少、弱精或梗阻性无精拟行ICSI治疗的患者术前行外周血染色体检查,以Y染色体多态者作为研究组(35个移植周期),随机选取同期行ICSI治疗,Y染色体正常者作为对照组(48个移植周期),比较组间行ICSI治疗的各项指标。结果:Y染色体多态性组和对照组的获卵数(12.5±6.4个vs11.8±4.2个)、正常受精数(8.1±4.6个vs9.1±3.3个)、卵裂数(7.9±4.6个vs8.9±3.4个)、优质胚胎数(5.3±3.5个vs5.9±2.9个)、临床妊娠率(48.57%vs56.25%)、流产率(11.76%vs25.93%)、新生儿出生孕周(37.07±2.60周vs38.00±1.02周)、体质量(2.95±0.90kgvs3.22±0.54kg)组间均无统计学差异(P>0.05)。结论:Y染色体多态对于ICSI治疗结局无明显影响。     

The Status of Controlled Prospective Clinical Trials for Efficacy of Intracytoplasmic Sperm Injection in In Vitro Fertilization for Non-Male Factor Infertility

Purpose: Intracytoplasmic sperm injection (ICSI) of some sibling oocytes may have a beneficial effect in couples going through in vitro fertilization for causes of infertility not related to the male factor. Our purpose was to critically appraise the randomized controlled studies done in this area and arrive at some recommendations. Methods: The four controlled trials done so far have utilized similar methodology, i.e., they randomly allotted sibling oocytes to ICSI versus standard insemination in patients going through in vitro fertiliztion and embryo transfer. Results: In the first trial reported in 1995 there was no difference in fertilization rate, whereas the later trials reported in 1997, 1999, and 2000 showed improvement with ICSI that reached statistically significant level in the last two studies. Conclusions: Total fertilization failure of an in vitro fertilization cycle can be prevented and fertilization can be improved if half of sibling oocytes are subjected to ICSI.     

In Vitro Fertilization and Intracytoplasmic Sperm Injection for Couples with Unexplained Infertility After Failed Direct Intraperitoneal Insemination

Purpose: The objective was to determine the optimal insemination technique in patients undergoing in vitro fertilization (IVF) after failed direct intraperitoneal insemination (DIPI) and the outcome of intracytoplasmic sperm injection (ICSI) in such cases. Methods: In case–control studies, 53 couples with unexplained infertility who underwent IVF after four failed DIPI cycles were compared with 75 couples with tubal or endometriosis infertility as controls. Thirty couples with unexplained infertility after failing to conceive with DIPI and conventional IVF who underwent ICSI and 58 couples with male-factor infertility as controls also were compared. Fertilization cleavage, embryo quality, implantation, and pregnancy were compared after IVF and after ICSI. Results: There was a significant difference in fertilization rates after IVF between cases of unexplained infertility after failing to conceive with DIPI (40.4%) and patients with tubal or endometriosis infertility (67.9%). There also was a significant difference in total fertilization failure rates between the two groups (30.4% and 3.9%, respectively). There was a slight but significant difference in numbers of fertilized oocytes after ICSI between patients with low fertilization rate undergoing IVF after failing to conceive DIPI (85.8%) and patients with male factor (90.4%). Total fertilization failure was not observed in these cases. Conclusions: Couples with unexplained infertility after failing to conceive with DIPI show a failed fertilization or a low fertilization rate after IVF. However, they demonstrated a good chance of becoming pregnant after subsequent ICSI, even with statistically significant difference in fertilization rate as compared with male-factor cases.     

Assessment of Maximal Fertilization Rates with Intracytoplasmic Sperm Injection

Purpose: Maximal fertilization rates following ICSI wereassessed using two essential steps: immobilization of spermand aspiration of oocyte cytoplasm. Methods: ICSI procedure was performed for couples(N = 42) in whom the male suffered severe infertility or failureof fertilization in previous IVF cycle using different oocytecytoplasmic aspiration and sperm immobilization methods.Outcome in four patient groups was measured by oocytedamage, fertilization rate, and pregnancy rate. Results: Maximal fertilization (90%) were achieved from thegroup which used immobilization of sperm by hard-touching thetail with a pipette and optimal aspiration of oocyte cytoplasm. Conclusions: The results suggest that if the immobilizationof sperm and aspiration of oocyte cytoplasm are handledright during ICSI, this procedure can be expected to yielda 90% fertilization rate.     

Effects of Electric Stimulation on Bovine Oocyte Activation and Embryo Development in Intracytoplasmic Sperm Injection Procedure

Purpose: This study was carried out to investigate the efficacyof electric stimulation before and/or after intracytoplasmicsperm injection (ICSI) on bovine oocyte activation andembryo development. Methods: The oocytes were treated with electric shock before(B), before and after (B&A), and after (A) sperm injection.In each group, sham ICSI (ICSI-s) was performed to excludethe effect of parthenogenesis (B ICSI-s, B&A ICSI-s, and AICSI-s). An electric pulse was applied with a single directcurrent (DC) pulse (0.8 kV/cm, 70 sec). Results: One pronucleus (PN) formation in the B&A ICSI-sgroup was slightly higher than that found in B and B&AICSI group; however, the difference was not significant. TwoPN formation in B&A ICSI group was higher than that foundin sham ICSI groups (P < 0.05). There were no differencesamong treatment groups in the cleavage rate; however, morulaeand blastocyst formation in the B&A embryos wassignificantly higher than that of other groups (P < 0.05)and got pregnant. Conclusions: Electric stimulation before and after injectionwas an effective method in inducing bovine oocyte activationand in sustaining embryo development to the morulae andblastocyst stage.     

Very Low Sperm Count Affects the Result of Intracytoplasmic Sperm Injection

Purpose: The aim was to examine the influence of extremelylow sperm count on intracytoplasmic sperm injection(ICSI) outcome. Methods: Over 1000 consecutive unselected ICSI cycleswere divided into four groups according to spermconcentration of their patients: A, cryptozoospermia, 107 patients; B,sperm concentration of 1×10⁴, 146 patients; C, spermcount of 1×10⁴–1×10⁵, 135 patients; and concentration of 1×10⁵ and < 10×10⁶/ml (control group), 688 patients. Results: A significant decrease in pregnancy rate wasnoticed in the cryptozoospermic group in comparison to thecontrol group (20% vs. 31%). Fertilization rate in group Awas significantly lower in comparison to all other groups,respectively (46% vs. 52%, 54%, 61%). Embryo quality wasinferior in group A in comparison to the control group. Ahigher yet not statistically significant abortion rate wasobserved in the cryptozoospermic group (as well as in groupC) (30%, 27%) compared to the control group (15%). Conclusions: It seems that an extremely low sperm counthas a negative effect on the outcome of ICSI. Neverthelesspatients with cryptozoospermia should not be offered ICSItreatment with the ejaculated sperm before karyotype isestablished.     

梗阻性无精子症患者精子顶体完整性与卵胞质单精子注射治疗结局之间的关系

目的:探讨梗阻性无精子症(OA)患者精子的顶体完整性及其与卵胞质单精子注射(ICSI)治疗临床结局之间的关系。方法:选取梗阻性无精子症患者共37例为试验组,同期进行体外受精治疗且精液常规参数正常的男性33例为对照组,应用荧光标记的豌豆凝集素法(PSA-FITC)检测精子顶体完整性,巴氏染色法分析精子形态,比较试验组与对照组的顶体完整率(AIR)、正常形态率(NFR)、受精率(FR)、卵裂率(CR)及优质胚胎率(OER),并将AIR与FR、NFR与FR进行相关性分析。结果:试验组的AIR、NFR、FR显著低于对照组(P<0.01),CR、OER试验组与对照组相比无统计学差异(P>0.05)。试验组AIR与FR呈显著正相关(r=0.595,P<0.01),NFR与FR显著正相关(r=0.463,P<0.01);对照组AIR与FR显著正相关(r=0.683,P<0.01),NFR与FR呈显著正相关(r=0.205,P<0.01)。结论:梗阻性无精子症患者的精子AIR较低。行皮下附睾抽吸术(PESA)-ICSI的梗阻性无精子症患者精子其AIR高则受精率也会高。     

Frequency and Patterns of Premature Sperm Chromosome Condensation in Oocytes Failing to Fertilize After Intracytoplasmic Sperm Injection

Purpose: The purpose of this study was to evaluate reasonsfor fertilization failure after intracytoplasmic sperminjection as a part of internal quality control and to reviewcorresponding previous data. Methods: One hundred injected but unfertilized oocytes werefixed and examined after Giemsa staining. Results: Three oocytes (3.0%) did not show the presenceof a spermatozoon and two (2.0%) contained pronuclearstructures. An intact spermatozoon was found in 25 cases(25.0%), whereas the sperm nucleus had undergonepremature chromosome condensation (PCC) in 70 cells (70.0%).A modified classification system was established tocharacterize the different PCC patterns. Conclusions: PCC indicates a correct intracytoplasmicinjection and excludes technical problems as a major reasonfor fertilization failure in the present study. A lack of oocyteactivation due to cytoplasmic immaturity is consideredresponsible for the occurrence of PCC. A review of theliterature shows that the role of sperm chromatinabnormalities in the process of nuclear decondensation needs furtherinvestigation.     

卵胞质内单精子显微注射后异常受精的影响因素分析

目的:探讨卵胞质内单精子显微注射(ICSI)后异常受精发生的影响因素。方法:回顾性分析299个ICSI周期,按照是否有异常受精分为异常受精组(n=118)和正常对照组(n=181),异常受精组至少发生1个非2原核(2PN)的受精卵。比较分析异常受精组和对照组临床和实验室资料。结果:异常受精组hCG注射日E2水平(14 097±3 066 pmol/L)高于对照组(1 2461±6 836 pmol/L),差异有统计学意义(P<0.05);异常受精组获卵数及成熟卵数分别为17.8±7.2个、15.3±6.1个,多于对照组的13.6±7.0个、10.2±5.3个,差异均有统计学意义(P<0.01);卵子成熟率(81.7%vs76.4%)、2PN受精率(78.4%vs 86.9%),组间比较差异均有统计学意义(P<0.01);而患者年龄、基础激素水平、促性腺激素(Gn)使用总量、精子来源及质量、着床率、临床妊娠率组间比较,差异均无统计学意义(P>0.05)。结论:卵巢对Gn刺激的高反应性可能与ICSI后异常受精的发生有关。异常受精卵的发生虽对临床结局无明显影响,但减少异常受精发生率,增加卵子利用率,可能会提高累积妊娠率。     

3106个不同来源精子ICSI周期临床妊娠结局分析

目的:分析精子的来源对卵胞质内单精子注射(ICSI)治疗结局的影响。方法:回顾性分析因男性不育行ICSI的3 106个新鲜周期,按精子来源分为:射精组(A组)、附睾穿刺取精(PESA)组(B组)、睾丸穿刺取精(TESA)组(C组)、冻融PESA精子组(D组)及冻融TESA精子组(E组),比较各组ICSI后胚胎发育及妊娠结局情况。结果:C组2PN受精率、卵裂率显著低于A组及B组;B组临床妊娠率、胚胎植入率显著高于A组及C组,A组、B组及C组间分娩率、异位妊娠率、流产率及新生儿畸形率无统计学差异(P>0.05);E组2PN受精率显著低于D组,但B组与D组之间、C组与E组间2PN受精率、优质胚胎率、多胎率、流产率及异位妊娠率均无统计学差异(P>0.05)。结论:PESA/TESA-ICSI、冻融PESA/TESA精子技术是治疗梗阻性无精子症安全有效的方法,建议首先选择附睾取精,并可将剩余PESA/TESA精子冻存。