Embryo and fetal gene editing: Technical challenges and progress toward clinical applications.

Gene modification therapies (GMTs) are slowly but steadily making progress toward clinical application. As the majority of rare diseases have an identified genetic cause, and as rare diseases collectively affect 5% of the global population, it is increasingly important to devise gene correction strategies to address the root causes of the most devastating of these diseases and to provide access to these novel therapies to the most affected populations. The main barriers to providing greater access to GMTs continue to be the prohibitive cost of developing these novel drugs at clinically relevant doses, subtherapeutic effects, and toxicity related to the specific agents or high doses required. In vivo strategy and treating younger patients at an earlier course of their disease could lower these barriers. Although currently regarded as niche specialties, prenatal and preconception GMTs offer a robust solution to some of these barriers. Indeed, treating either the fetus or embryo benefits from economy of scale, targeting pre-pathological tissues in the fetus prior to full pathogenesis, or increasing the likelihood of complete tissue targeting by correcting pluripotent embryonic cells. Here, we review advances in embryo and fetal GMTs and discuss requirements for clinical application.

Injectable "Homing-Like" Bioactive Short-Fibers for Endometrial Repair and Efficient Live Births.

The prevalence of infertility caused by endometrial defects is steadily increasing, posing a significant challenge to women's reproductive health. In this study, injectable "homing-like" bioactive decellularized extracellular matrix short-fibers (DEFs) of porcine skin origin are innovatively designed for endometrial and fertility restoration. The DEFs can effectively bind to endometrial cells through noncovalent dipole interactions and release bioactive growth factors in situ. In vitro, the DEFs effectively attracted endometrial cells through the "homing-like" effect, enabling cell adhesion, spreading, and proliferation on their surface. Furthermore, the DEFs effectively facilitated the proliferation and angiogenesis of human primary endometrial stromal cells (HESCs) and human umbilical vein endothelial cells (HUVECs), and inhibited fibrosis of pretreated HESCs. In vivo, the DEFs significantly accelerated endometrial restoration, angiogenesis, and receptivity. Notably, the deposition of endometrial collagen decreased from 41.19 ± 2.16% to 14.15 ± 1.70% with DEFs treatment. Most importantly, in endometrium-injured rats, the use of DEFs increased the live birth rate from 30% to an impressive 90%, and the number and development of live births close to normal rats. The injectable "homing-like" bioactive DEFs system can achieve efficient live births and intrauterine injection of DEFs provides a new promising clinical strategy for endometrial factor infertility.

Effect of Intra Uterine Granulocyte Colony Stimulating Factor vs. Human Chorionic Gonadotropin at Ovum Pick Up Day on Pregnancy Rate in IVF/ICSI Cases With Recurrent Implantation Failure.

OBJECTIVE: Recurrent implantation failure is defined as failure to achieve clinical pregnancy after the transfer of four or more good-quality embryos in a minimum of three fresh or frozen cycles in a woman aged less than 40 years. The objective is to compare between the effect of intrauterine G-CSF, hCG, and saline solution injection (as placebo) at the day of ovum pick-up on clinical pregnancy, chemical pregnancy, implantation, and miscarriage rates in patients with recurrent implantation failure undergoing IVF/ICSI. METHODS: This prospective, double blind, parallel, randomized controlled trial included 150 patients equally divided into 3 groups, each containing 50 individuals. Subjects in Group 1 received intrauterine injections of G-CSF; Group 2: received intrauterine injections of 500 IU of hCG; and Group 3 received intrauterine injections of saline solution as placebo. The primary outcome measure is clinical pregnancy rate. Secondary outcomes are biochemical pregnancy, implantation, and miscarriage rates. RESULTS: Clinical pregnancy, biochemical pregnancy, and implantation rates were highest in the group given G-CSF and lowest in the group administered saline solution; miscarriage rates were not significantly different between the groups. CONCLUSIONS: Intrauterine administration of G-CSF at a dose of 100 µg/1.0 cc at the time of ovum pick-up is associated with better clinical pregnancy, chemical pregnancy, and implantation rates as compared with intrauterine saline solution administration. Further studies are needed to determine the optimum timing of intrauterine administration of G-CSF that achieves the best results, and longer follow-up is needed to determine take-home baby percentages.

Intrauterine injection of HCG before embryo transfer: a parallel, double-blind randomized trial.

RESEARCH QUESTION: Does intrauterine administration of HCG before embryo transfer improve live birth rate during IVF cycles? DESIGN: A parallel, randomized controlled trial conducted between July 2018 and February 2020. Infertile women (n = 181) scheduled for fresh or vitrified-warmed embryo transfer after IVF carried out for any indication were randomized in a 1:1 ratio to receive either HCG (500 IU in 0.1 ml of tissue culture media) or culture media (0.1 ml of tissue culture media) via intrauterine injection 4 min before embryo transfer. In both groups, an intrauterine insemination catheter was used for administering the medication. Primary outcome was live birth, with ongoing pregnancy and clinical pregnancy as secondary outcomes. Analysis was based on intention-to-treat principle. RESULTS: Baseline and cycle characteristics were comparable between the two groups. In the control group, one woman with a confirmed clinical pregnancy was lost to follow-up. Live birth rates were 24% (22/90) in the HCG group versus 19% (17/90) in the control group (RR 1.29, 95% CI 0.74 to 2.27). Clinical pregnancy and ongoing pregnancy rates were 34% versus 26% (RR 1.31, 95% CI 0.84 to 2.04) and 24% versus 19% (RR 1.29, 95% CI 0.74 to 2.27) in the HCG and the control groups, respectively. CONCLUSION: Intrauterine injection of HCG before embryo transfer did not improve live birth rates in women undergoing IVF. As the study was designed to detect a 20% difference between groups, a smaller, clinically important difference could not be ruled out. Treatment outcomes were lower than expected in the control group.

The Effect of Intrauterine Administration of Human Chorionic Gonadotropin (hCG) Before Embryo Transfer During Assisted Reproductive Cycles: a Meta-Analysis of Randomized Controlled Trials.

The fertility success rates of clinical and laboratory-assisted reproductive techniques (ART) remain low, despite major advances. The aim of this study was to conduct a systematic literature review and assess whether the intrauterine administration of human chorionic gonadotropin (hCG) before embryo transfer (ET) improved the clinical outcomes of sub-fertile women undergoing assisted reproduction. The electronic databases PUBMED, EMBASE and Web of Science were systematically searched for randomized controlled trials (RCTs) published from inception to June 2018. The trial data were independently extracted and analyzed using risk ratios (RRs) and 95% confidence intervals (CIs) according to a random- or fixed-effect model (as appropriate), and a meta-analysis was conducted using Review Manager 5.2 software. The meta-analysis included 3241 patients from 12 RCTs, and the combined results demonstrated that intrauterine hCG injection significantly improved the rates of clinical (RR = 1.33; 95% CI: 1.12 - 1.58) and ongoing pregnancy (RR = 1.87; 95% CI: 1.54 - 2.27), compared with controls. However, intrauterine hCG injection had no significant effect on the implantation rate (RR = 1.30; 95% CI: 0.89 - 1.90), abortion rate (RR = 1.06; 95% CI: 0.78 - 1.44), ectopic pregnancy rate (RR = 0.77; 95% CI: 0.17 - 3.42) or live birth rate (RR = 0.99; 95% CI: 0.60 - 1.63). In a subgroup analysis, the intrauterine injection of > 500 IU hCG led to a significant increase in the implantation rate (RR = 1.64; 95% CI: 1.04 - 2.61) relative to controls. Furthermore, the subgroup of women with cleavage-stage ETs who received an intracavity injection of hCG (IC-hCG) exhibited increases in the implantation, clinical pregnancy and ongoing pregnancy rates, compared to women with cleavage-stage ETs and no IC-hCG. The current evidence indicates that intrauterine hCG administration before ET provides an advantage in terms of the clinical pregnancy and ongoing pregnancy rates.

Intrauterine injection of human chorionic gonadotropin before embryo transfer can improve in vitro fertilization-embryo transfer outcomes: a meta-analysis of randomized controlled trials.

OBJECTIVE: To evaluate whether intrauterine injection of hCG before embryo transfer can improve IVF-ET outcomes. DESIGN: Meta-analysis. SETTING: Not applicable. PATIENT(S): Infertile women who underwent IVF-ET and received an intrauterine injection of hCG before ET. INTERVENTION(S): Infertile women treated with or without intrauterine hCG injection before ET. MAIN OUTCOME MEASURE(S): The primary outcomes were live birth rate (LBR), ongoing pregnancy rate (OPR), and clinical pregnancy rate (CPR), and the secondary outcomes were implantation rate (IR) and miscarriage rate (MR). Odds ratios with 95% confidence intervals (CIs) and successful ET rates were pooled to determine the effects of hCG on IVF-ET outcomes. RESULT(S): Fifteen randomized controlled trials (RCTs) with a total of 2,763 participants were included. Infertile women in the experimental group (treated with intrauterine hCG injection before ET) exhibited significantly higher LBR (44.89% vs. 29.76%), OPR (48.09% vs. 33.42%), CPR (47.80% vs. 32.78%), and IR (31.64% vs. 22.52%) than those in the control group (intrauterine injection of placebo or no injection). Furthermore, MR was significantly lower (12.45% vs. 18.56%) in the experimental group than in the control group. CONCLUSION(S): The findings of this meta-analysis indicate that intrauterine injection of hCG can improve LBR, OPR, CPR, and IR after IVF-ET cycles. In addition, different timing and dosages of hCG administration may exert different effects on IVT-ET outcomes. Notably, infertile women treated with 500 IU hCG within 15 minutes before ET can achieve optimal IVF-ET outcomes.

Effect of intrauterine injection of human chorionic gonadotropin before fresh embryo transfer on IVF and ICSI outcomes: a meta-analysis.

PURPOSE: This analysis was performed to evaluate the effects of intrauterine injection of human chorionic gonadotropin (hCG) before fresh embryo transfer (ET) on the outcomes of in vitro fertilization and intracytoplasmic sperm injection. METHODS: Randomized controlled trials (RCTs) were identified by searching electronic databases. The outcomes of live birth, clinical pregnancy, implantation, biochemical pregnancy, ongoing pregnancy, ectopic pregnancy, and miscarriage between groups with and without hCG injections were analyzed. Summary measures were reported as risk ratios (RR) with 95% confidence intervals. RESULTS: Six RCTs on fresh embryo transfer (ET) were included in the meta-analysis. A total of 2759 women undergoing fresh ET were enrolled (hCG group n = 1429; control group n = 1330). Intrauterine injection of hCG significantly increased rates of biochemical pregnancy (RR 1.61) and ongoing pregnancy (RR 1.58) compared to controls. However, there were no significant differences in clinical pregnancy (RR 1.11), implantation (RR 1.17), miscarriage (RR 0.91), ectopic (RR 1.65) or live birth rates (RR 1.13) between the hCG group and control group. CONCLUSION: The current evidence for intrauterine injection of hCG before fresh ET does not support its use in an assisted reproduction cycle.

Efficacy of Intrauterine Injection of Granulocyte Colony Stimulating Factor (G-CSF) on Treatment of Unexplained Recurrent Miscarriage: A Pilot RCT Study.

BACKGROUND: Endometrium undergoes several changes in structure and cellular composition during pregnancy. Granulocyte Colony-stimulating Factor (GCS-F) is an important cytokine with critical role in embryo implantation and pregnancy. The aim of the present study was to evaluate the impact of intrauterine injection of G-CSF in patients who suffer from unexplained recurrent miscarriage (RM). METHODS: In the present randomized clinical trial, a total of 68 patients were randomly allocated into two study groups including intrauterine G-CSF (n=23, 300 µg) injection and control group (n=27, no G-CSF injection). Eighteen out of 68 patients were excluded from the final analysis due to different reasons. All patients were in Ovulation Induction (I/O) cycle. In G-CSF group, intrauterine injection of G-CSF was done twice in the cycle. All enrolled patients were under 40 years old and had at least two unexplained pregnancy losses. Pregnancy was evaluated by titer of ßhCG, presence of gestational sac (implantation) and fetal heart rate (clinical pregnancy) was assessed by vaginal ultrasonography. Student's T test and Mann-Whitney U were used for analysis. The p≤0.05 was determined as statistically significant. RESULTS: No significant differences were observed between the two study groups when the rates of chemical pregnancy (26.1% vs. 29.6%, p=0.781), implantation (26.1% vs. 22.2%, p=0.750), clinical pregnancy (17.4% vs. 11.1%, p=0.689) and abortion (33% vs. 37.5%, p=0.296) were compared. CONCLUSION: In our study, no significant difference was observed between the two study groups when the rates of chemical pregnancy, implantation, clinical pregnancy and abortion were compared.

Effects of intrauterine injection of ethanol on estrous cycle in mice / 中国比较医学杂志

Objective To investigate the effect of uterine cavity injection of absolute ethanol on estrous cycle in mice.Methods Twenty mice with regular estrous cycle were selected by vaginal exfoliated cells staining, and then were injected absolute ethanol into their uterine cavity.The estrous cycle was observed and recorded every day.Ten mice were selected randomly to observe the changes of second estrous cycle every two hours.Results After the injection of absolute ethanol into the uterine cavity of the experimental mice, the estrous cycle was greatly prolonged, and the various degree of disorder was observed in estrus and pre-estrus.Conclusions The disorder was observed in estrous cycle of mice after the injection of absolute ethanol, which maybe provides some references for clinical use of absolute ethanol.

A clinical comparative study of methotrexate intrauterine injection in the treatment for cesarean scar pregnancy / 天津医药

Objective To analyse the clinical efficacy of methotrexate (MTX) combined with intrauterine embryo sac garrotte injection in the treatment of cesarean scar pregnancy (CSP), and discuss its clinical significance. Methods A total of 77 patients with CSP treated in our hospital during June 2013 to December 2016 were selected in this study. Forty patients treated with embryo sac destruction and methotrexate injection were included in the observation group, while 37 cases treated by uterine artery embolization combined with curettage were used as the control group. The time of vaginal bleeding, the time of postoperative blood level of human chorionic gonadotropin (HCG) returned to the normal level, average hospitalization cost and the curative rate were recorded in two groups. All patients were followed up by the outpatient visit. Results In the observation group, the vaginal bleeding time [(22.1±6.7) days vs. (29.5±10.8) days] and treatment cost [(8774.2 ± 714.5) yuan vs. (15258.3 ± 1084.2) yuan] were less than those of the control group (P<0.001). There were no significant differences in the recovery time of HCG [(26.4±9.0) days vs. (25.1±10.4) days] and treatment success rate (87.5%vs. 91.9%) between the two groups (P>0.05). No bleeding or threatened rupture of scar were found in two groups of patients. Conclusion In this study, we take the embryo sac puncture combined with methotrexate injection in the treatment of scar pregnancy. This method has the advantages of low operative difficulty, definite clinical curative effect and low cost

A clinical comparative study of methotrexate intrauterine injection in the treatment for cesarean scar pregnancy / 天津医药

Objective To analyse the clinical efficacy of methotrexate (MTX) combined with intrauterine embryo sac garrotte injection in the treatment of cesarean scar pregnancy (CSP), and discuss its clinical significance. Methods A total of 77 patients with CSP treated in our hospital during June 2013 to December 2016 were selected in this study. Forty patients treated with embryo sac destruction and methotrexate injection were included in the observation group, while 37 cases treated by uterine artery embolization combined with curettage were used as the control group. The time of vaginal bleeding, the time of postoperative blood level of human chorionic gonadotropin (HCG) returned to the normal level, average hospitalization cost and the curative rate were recorded in two groups. All patients were followed up by the outpatient visit. Results In the observation group, the vaginal bleeding time [(22.1±6.7) days vs. (29.5±10.8) days] and treatment cost [(8774.2 ± 714.5) yuan vs. (15258.3 ± 1084.2) yuan] were less than those of the control group (P<0.001). There were no significant differences in the recovery time of HCG [(26.4±9.0) days vs. (25.1±10.4) days] and treatment success rate (87.5%vs. 91.9%) between the two groups (P>0.05). No bleeding or threatened rupture of scar were found in two groups of patients. Conclusion In this study, we take the embryo sac puncture combined with methotrexate injection in the treatment of scar pregnancy. This method has the advantages of low operative difficulty, definite clinical curative effect and low cost

The efficacy of intrauterine injection of human chorionic gonadotropin before embryo transfer in assisted reproductive cycles: Meta-analysis.

OBJECTIVES: A systematic review and meta-analysis to evaluate the effect of human chorionic gonadotropin (hCG) intrauterine injection before embryo transfer on the outcome of in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI). METHODS: Searches of PubMed®, EMBASE®, EBSCO, Web of Science®, SCOPUS® and Cochrane Central Register of Controlled Trials were conducted to retrieve relevant randomized controlled trials (RCTs). Data were extracted and analysed. RESULTS: The meta-analysis included five RCTs (hCG group n = 680; control group n = 707). Intrauterine hCG injection significantly increased rates of biochemical, clinical and ongoing pregnancy compared with controls. There were no between-group differences in implantation or miscarriage rates. CONCLUSION: Women undergoing IVF/ICSI may benefit from intrauterine hCG injection before embryo transfer.

PTEN gene expression in mouse endometria increases during embryo implantation / 第三军医大学学报

Objective To investigate the effect of PTEN gene in the mouse uterus during embryo implantation.Methods Real-time fluorescence quantitative PCR(FQ-PCR) was used to detect PTEN mRNA expressed in the endometria of the nonpregnant mice and the late pregnant mice(day 1,day 3,day 4,day 5 and day 7),with 20 mice sacrificed at each fixed day.Out of another 20 3-day pregnant mice,ten received PTEN antisense oligonucleotide at the horn of uterus and ten received normal saline to count the blastocysts at pregnant day 8.Results The PTEN mRNA/?-actin mRNA in pregnant mice was higher than that of nonpregnant mice,gradually hoisted as days passed by,and reached the highest at pregnant day 5.The number of blastocysts in the mice that received PTEN antisense oligonucleotide was fewer than that received normal saline.Conclusion PTEN persistently expresses in mouse endometria during the early pregnancy and maybe participate in the regulation process of mouse blastodyst implantation.