Analysis of birth defects among children 3 years after conception through assisted reproductive technology in China

BACKGROUND : Previous studies inconsistently suggest that assisted reproduction technology (ART) may increase the risk of birth defects in children. METHOD(S) : Live birth infants, conceived by in vitro fertilization fresh embryo transfer (IVF), intracytoplasmic sperm injection fresh embryo transfer (ICSI), or frozen‐thawed embryo transfer (FET) in Reproductive Center of Tongji Hospital (Wuhan, China) between 1997 and 2008, were followed up at birth and after 3 years. Preterm pregnancy, multiple pregnancy, sex ratio (male/female), congenital malformation were compared. RESULT(S) : A total of 4,236 children were born after ART (IVF 2,543, ICSI 908, FET 785). Compared with IVF, the rate of preterm pregnancy and sex ratio in ICSI were lower (p < 0.05); the rate of multiple pregnancy in ICSI and FET were all lower than IVF (p < 0.05). Congenital defects were comparable in all groups at birth. In total, 2,908 children participated in the second follow‐up from 34 months to 60 months with an average of 40 months, and the cases of birth defects had doubled (3 years: 5.16%, birth: 2.22%). The birth defect rate in boys conceived through ICSI was significantly higher than the IVF group after 3‐year follow‐up (ICSI boys: 8.62%, IVF boys: 5.21% [p < 0.05]), even though there was no significant difference at birth. CONCLUSION(S) : Compared with IVF, FET may not increase risk of birth defects. Children conceived through ICSI, especially males, had higher rates of congenital malformations that were inapparent at birth. So longitudinal monitoring may provide insights into the risks of ART. Birth Defects Research (Part A) 97:744–749, 2013. 2013. © 2013 Wiley Periodicals, Inc.     

Sequential analysis of zona thickness during in vitro culture of human zygotes: Correlation with embryo quality,age, and implantation

Zona pellucida thickness was measured daily in zygotes and cleavage stage embryos. Measurements were performed on a Nikon inverted microscope equipped with Hoffman modulation optics, using an ocular micrometer. Zona thickness of each zygote/embryo was measured four times, the zygote/embryo was then “rolled over,” and four more measurements were repeated for a total of eight. The zygotes/embryos were photographed daily and the measurements repeated on the prints. Subsequently, the mean zona thickness for each stage was calculated. A total of 81 patients (mean age 33.8 ± 4.2) participated in the study. A total of 427 embryos were evaluated. Categorical data differences between groups were evaluated by ANOVA and multiple linear regression. For nominal data, the Kruskal-Wallis test was applied; when P < 0.05 the differences were considered to be significant. We found that the average zona thickness on day 1 of in vitro culture was 17.7 ± 0.14 μm; 16.3 ± 0.14 μm on day 2 and 14.9 ± 0.14 μm on day 3 (P < .0001). When the zona thickness was analyzed in relation to the number of blastomeres on day 3 of culture, there was a highly significant correlation with blastomere number (P < .0001). Similarly, there was a highly significant correlation with embryo grade (P < .005) and fragmentation (P < .001). The data were also analyzed for embryos transferred that resulted in a successful pregnancy, revealing that embryos in a pregnancy cycle had significantly thinner zonae pellucidae (P < .0001), when compared to embryos that were not transferred or from nonconceptual cycles. The average zona thickness also decreased with age, and was most apparent after 35 years. Changes in zona thickness correlated with the number of blastomeres, grade, fragmentation, age and were more evident in embryos transferred from cycles resulting in successful pregnancies. Therefore, zona pellucida measurements should be included in the overall assessment of embryo quality, since this information may be useful in the selection of optimal embryos for transfer. Mol. Reprod. Dev. 47:99–104, 1997. © 1997 Wiley-Liss, Inc.     

生物杀虫剂-昆虫病原线虫的培养技术

昆虫病原线虫是新型的生物杀虫剂,对钻蛀及土栖性害虫防效较好,具有部分替代化学农药的潜力。昆虫病原线虫的产业化培养是昆虫病原线虫商业化应用的基础。本文介绍了目前世界上常用的昆虫病原线虫的培养方法,包括活体培养、半固体培养和液体培养的技术,这些技术的应用有助于昆虫病原线虫的种质保存及培养,为拓展昆虫病原线虫的产业化生产和应用奠定了基础。     

A simple,high-yield method for obtaining multipotential mesenchymal progenitor cells from trabecular bone

In vitro cultures of primary, human trabecular bone-derived cells represent a useful system for investigation of the biology of osteoblasts. Our recent discovery of the multilineage mesenchymal differentiation potential of trabecular bone-derived cells suggests the potential application of these cells as mesenchymal progenitors for tissue repair and regeneration. Such applications are crucially dependent on efficient cellisolation protocols to yield cells that optimally proliferate and differentiate. In this study, we describe a simple, high-yield procedure, requiring minimal culture expansion, for the isolation of mesenchymal progenitor cells from human trabecular bone. Moreover, these cells retain their ability to differentiate along multiple mesenchymal lineages through successive subculturing. Cell populations isolated and cultured as described here allow the efficient acquisition of a clinically significant number of cells, which may be used as the cell source for tissue-engineering applications.     

“试管婴儿”技术30年——回顾和展望

1978年第一例试管婴儿诞生以及随后近30多年来该领域不断取得的突破性进展,促进了以体外受精为基础的辅助生殖技术在临床的广泛应用,并取得了很好的疗效。本文将系统回顾近30多年来辅助生殖技术在临床和实验室两方面建立的包括:控制性超排卵、序贯培养、精子胞浆内注射以及植入前遗传学诊断和筛查等主要技术突破,并对医源性多胎、新技术的安全性等目前存在的主要问题及可能的解决方案进行探讨。     

Assisted reproductive technology and the risk of pediatric cancer: A population based study and a systematic review and meta analysis

BackgroundThere is controversy whether exposure to assisted reproductive technology (ART) is associated with increased risk of pediatric cancer.We aimed at calculating the overall risk of pediatric cancers after ART in a large cohort of exposed women; and to conduct a systematic review and meta- analysis of cohort studies examining overall risk of pediatric cancers after ART.MethodsAll children born in Israel who were members of Maccabi Health Services (MHS) between 1999 and 2016 after ART, were linked to the Israeli Registry of Childhood Cancer (IGS) to identify those with cancer diagnosed before 16 years of age. In parallel we conducted a systematic review and meta-analysis of observational cohort studies with more than 5000 ART- exposed cases that measured pediatric cancer after ART.ResultsIn the cohort study, the risk ratio for pediatric cancer after ART in general was 0.95 (95% CI, 0.76–1.19). The RR was 1.09 (95% CI, 0.79–1.48) for IVF treatments. Meta- analysis of 13 cohort studies with a total of 750,138 women exposed to ART (with 1152 pediatric cancers) and 214,008,000 unexposed controls (with 30,458 pediatric cancers) did not reveal increased risk for pediatric cancers (RR 0.99; 95% CI, 0.85–1.15).ConclusionsBased on very large numbers, ART in general, and IVF in particular, are not associated with overall increased risk of pediatric cancer.     

Noninvasive preimplantation genetic testing in assisted reproductive technology: current state and future perspectives

Invasive genetic screening of pre-implantation embryos via biopsied trophectoderm (TE) cells has been in use for more than 20 years, while its benefits in selecting euploid embryos remain controversial. Recent advances in the ability to process embryonic cell-free DNA (cfDNA) from blastocoel fluid (BF) and spent culture media (SCM) of blastocysts in a manner similar to that of a biopsied TE sample provide a potential alternative holding great promise for obtaining cytogenetic information of the embryos without intrusive biopsy of traditional biopsy-based pre-implantation genetic testing (PGT). Several studies have reported even higher diagnostic accuracy in non-invasive PGT (ni-PGT) than conventional PGT. However, there are still several technical challenges to be overcome before ni-PGT can be accepted as a reliable genomic information source of embryo. In this review, we have summarized the emergence and current state of ni-PGT, and discussed our own perspectives on their limitations and future prospect. There is still a long way to go before truly wide clinical application of ni-PGT.     

SMARThivVLmos: a complexity-free and cost effective dynamic model technology for monitoring HIV viral load in resource-poor settings

We design a "simple" and "low cost" model technology for monitoring HIV viral load in resource-poor settings: SMARThivVLmos. Cost and complexity are the major challenges to the developing world, in monitoring HIV patients viral load. We have previously demonstrated in our SMARThivPack model that cost and complexity of laboratory monitoring of HIV patients, may be reduced not only at a first technology development level, but also at a second technology implementation, and at a third global coordination levels. In our SMARThivPack model, the P24 HIV viral load monitoring system passed both the "cost" and the "complexity" tests. However, compared to other alternative viral monitoring systems such as the Cavidi EXAVIR, the sensitivity of the P24 system is too low. Here we describe a dynamic model technology that overcomes the sensitivity barrier of the P24 system while maintaining simplicity and low cost.     

SMARThivCD4mos: a complexity-free and cost effective model technology for monitoring HIV patients CD4 number in resource-poor settings

We design a "simple" and "low cost" model technology for monitoring HIV patients CD4 number in resource-poor settings: SMARThivCD4mos. Cost and complexity are the major challenges to the developing world in transferring and implementing HIV surveillance technologies. We previously proposed a "three tests" combo kit model for improving HIV patients monitoring standards in resource-poor settings. From the pool of recommended alternative CD4 counting technologies, our "three tests" combo kit model retained the Capcellia technology for its "simplicity". However, compared to other CD4 counting technologies, such as Dynabeads, the Capcellia technology is "expensive". Here, we describe a cost reduction strategy of the Capcellia technology.     

Holographic imaging of unlabelled sperm cells for semen analysis: a review

Male reproductive health in both humans and animals is an important research field in biological study. In order to characterize the morphology, the motility and the concentration of the sperm cells, which are the most important parameters to feature them, digital holography demonstrated to be an attractive technique. Indeed, it is a label‐free, non‐invasive and high‐resolution method that enables the characterization of live specimen. The review is intended both for summarizing the state‐of‐art on the semen analysis and recent achievement obtained by means of digital holography and for exploring new possible applications of digital holography in this field.

Quantitative phase maps of living swimming spermatozoa.     

Review: Preimplantation genetic diagnosis (PGD) as a reproductive option in patients with neurodegenerative disorders

Preimplantation genetic diagnosis (PGD) was introduced in the late 1980s and represents an option for couples at risk of transmitting an inherited, debilitating or neurological disorder to their children. From a cleavage or blastocyst stage embryo, cell(s) are collected and then genetically analyzed for disease; enabling an unaffected embryo to be transferred into the uterus cavity. Nowadays, PGD has been carried out for several hundreds of heritable conditions including myotonic dystrophy, and for susceptibility genes involved in cancers of the nervous system. Currently, advanced molecular technologies with better resolution, such as array comparative genomic hybridisation, quantitative polymerase chain reaction, and next generation sequencing, are on the verge of becoming the gold standard in embryo preimplantation screening. Given this, it may be time for neurological societies to consider the published evidence to develop new guidelines for the integration of PGD into modern preventative neurology. Therefore, the main aim of this review is to illustrate the option of PGD to enable conception of an unaffected baby, and to assist clinicians and neurologists in the counseling of the patient at risk of transmitting an inherited disease, to explore the genetic journey throughout in vitro fertilization IVF with PGD.     

Seasonal culture of dormant reproductive buds ofSalix tetrasperma: Analysis of the flowering process

The flowering process in a female tree ofSalix tetrasperma was analysed by culturing its reproductive buds at different developmental stages during the dormant period on a chemically defined medium and examining the nature of sprouts produced by them. Buds at the upper eight nodes of the actively growing shoots developing in an acropetal sequence were cultured in separate lots. While all the buds collected from the 1st and 2nd nodes of the branches from the top downwards were vegetative and produced shoots, a considerable number of those collected from the 3rd and 4th nodes were reproductively determined and produced catkins. All the buds obtained from the 5th node and below were reproductive. Reproductive buds were cultured at regular time intervals during the dormant period. Freshly formed buds cultured in March during the spring growth flush produced catkins and were therefore reproductively determined. However, such a determination was not tantamount to flowering, as the floral meristems present in the axils of catkin bracts remained quiescent. Floral meristems of the buds cultured during April to August developed into small vegetative shoots. This was followed by the crucial period during September to December when the hitherto vegetative sprouts of the floral meristems showed a gradual transition into ovaries (female flowers) resulting in fertile catkins. Catkins produced from buds cultured in January and February produced well-developed ovaries.     

PP2Cdelta (Ppm1d, WIP1), an endogenous inhibitor of p38 MAPK, is regulated along with Trp53 and Cdkn2a following p38 MAPK inhibition during mouse preimplantation development

Preimplantation embryos utilize mitogen-activated protein kinase signaling (MAPK) pathways to relay signals from the external environment to prepare appropriate responses and adaptations to a changing milieu. It is therefore important to investigate how MAPK pathways are regulated during preimplantation development. This study was conducted to investigate whether PP2Cdelta (Ppm1d, WIP1) is expressed during mouse preimplantation development and to determine the influences of p38 MAPK inhibition on expression of Trp53 (p53), Ppm1d, (WIP1), and Cdkn2a (p16) during mouse preimplantation development. Our results indicate that Trp53, Ppm1d, and Cdkn2a mRNAs and TRP53 and PP2Cdelta proteins are expressed throughout mouse preimplantation development. Treatment of 2-cell embryos with SB220025 (potent inhibitor of p38 MAPK alpha/beta/MAPK 14/11) significantly increased Trp53, Ppm1d and Cdkn2a and Mapk14 mRNA levels at 12 and 24 hr. Treatment of 8-cell embryos with SB220025 for 12 hr increased Trp53, Ppm1d, and Cdkn2a mRNA levels, but not Mapk14 mRNA levels. Treatment of 8-cell embryos for 24 hr increased Trp53, and Ppm1d mRNA levels, but decreased Cdkn2a and Mapk14 mRNA levels. Therefore, blockade of p38 MAPK activity is associated with embryo stage specific influences on Trp53, Ppm1d, Cdkn2a, and Mapk14 expression during mouse preimplantation development. These results define downstream targets of p38 MAPK during preimplantation development and indicate that the p38 MAPK pathway regulates Trp53, Ppm1d, and Cdkn2a expression. This study increases our understanding of the mechanisms controlling preimplantation development and of the interactions between preimplantation embryos and their culture environments.     

Effects of oxygen tension and humidity on the preimplantation development of mouse embryos produced by in vitro fertilization: analysis using a non-humidifying incubator with time-lapse cinematography

To examine the effects of oxygen tension and humidity on early embryonic development, the preimplantation development of mouse embryos produced by in vitro fertilization was assessed by time-lapse cinematography to evaluate morphokinetic development with higher precision. Zygotes were produced from spermatozoa and oocytes from ICR mice and cultured in KSOM under low or high oxygen tension in a non-humidified incubator with time-lapse cinematography (CCM-iBIS). The developmental rates of embryos to the 4-cell and blastocyst stages under lower oxygen tension in CCM-iBIS were significantly higher than those under higher oxygen tension in CCM-iBIS. Ninety-six hours after insemination, a large number of embryos cultured under low oxygen tension developed to the hatching blastocyst stage. Embryonic development was more synchronized under lower oxygen tension. Non-humidified cultures did not affect embryonic development. On average, mouse embryos cultured at lower oxygen tension reached 2-cell at 18 h, 3-cell at 39 h, 4-cell at 40 h, initiation of compaction at 58 h, morula at 69 h, and blastocyst at 82 h after insemination. In conclusion, lower oxygen tension better supports preimplantation development of mouse embryos fertilized in vitro, and non-humidified culture conditions do not influence the embryonic development in vitro.     

Agar culture of Piscirickettsia salmonis, a serious pathogen of farmed salmonid and marine fish

Piscirickettsia salmonis, a serious bacterial pathogen of farmed marine fish, previously considered culturable only in eukaryotic cell-culture systems, was grown for the first time on agar and broth containing enhanced levels of cysteine, thus greatly increasing the potential for isolation, in vitro culture and study of this organism. Virulence towards Atlantic salmon following passage on agar media was retained in a controlled laboratory trial. Of the studied temperatures, optimal growth on agar was observed at 22 degrees C.     

Two new C3H mouse ascites tumor cell lines capable of proliferation in vivo and in suspension culture: Morphological,karyological, kinetic,and immunological properties

Summary The establishment of mouse tumor cell lines capable of proliferating in vivo and in suspension culture was undertaken. The MM-46 tumor line, initiated from primary mammary carcinoma arising in a C3H/He mouse, was maintained for over 100 generations in the peritoneal cavities of syngenic mice. At the 50th generation of the tumor suspension, cultures were initiated. The established cell lines, designated TMT-1 and TMT-2, were characterized in vitro and in vivo. The morphological finding indicated that TMT-1 and TMT-2 cells from mice closely resembled the MM-46 tumor cells. The oncogenic potential of the cultured cells was comparable to that of the original ascites tumor. The population doubling time of TMT-1 and TMT-2 cell lines was about 12 hr in mice, whereas the population doubling time of both cell lines lengthened to 20 hr in suspension culture. The increase of doubling time in culture was due to the prolongation of the G₁ period. The cell lines, TMT-1 and TMT-2, whether from culture or mice, possessed colony forming ability in soft agar medium. The colony forming ability of the cells decreased gradually through in vivo passages but it recovered upon recultivation of the cells from mouse to culture. Chromosome analysis and cytotoxicity test by anti-MM antiserum indicated that TMT-1 and TMT-2 cell lines closely resembled and had been derived from MM-46 tumor line. Therefore, it is possible to assay cell survival in vitro after in vivo experiments on these cells.     

Consequences of bovine oocyte maturation, fertilization or early embryo development in vitro versus in vivo: implications for blastocyst yield and blastocyst quality.

The aim of this study is to examine the effect of bovine oocyte maturation, fertilization or culture in vivo or in vitro on the proportion of oocytes reaching the blastocyst stage, and on blastocyst quality as measured by survival following vitrification. In Experiment 1, 4 groups of oocytes were used: (1) immature oocytes from 2-6 mm follicles; (2) immature oocytes from > 6 mm follicles; (3) immature oocytes recovered in vivo just before the LH surge; and (4) in vivo matured oocytes. Significantly more blastocysts developed from oocytes matured in vivo than those recovered just before the LH surge or than oocytes from 2-6 mm follicles. Results from > 6 mm follicles were intermediate. All blastocysts had low survival following vitrification. In Experiment 2, in vivo matured oocytes were either (1) fertilized in vitro or (2) fertilized in vivo by artificial insemination and the resulting presumptive zygotes recovered on day 1. Both groups were then cultured in vitro. In vivo fertilized oocytes had a significantly higher blastocyst yield than those fertilized in vitro. Blastocyst quality was similar between the groups. Both groups had low survival following vitrification. In Experiment 3a, presumptive zygotes produced by in vitro maturation (IVM)/fertilization (IVF) were cultured either in vitro in synthetic oviduct fluid, or in vivo in the ewe oviduct. In Experiment 3b, in vivo matured/in vivo fertilized zygotes were either surgically recovered on day 1 and cultured in vitro in synthetic oviduct fluid, or were nonsurgically recovered on day 7. There was no difference in blastocyst yields between groups of zygotes originating from the same source (in vivo or in vitro fertilization) irrespective of whether culture took place in vivo or in vitro. However, there was a dramatic effect on blastocyst quality with those blastocysts produced following in vivo culture surviving vitrification at significantly higher rates than their in vitro cultured counterparts. Collectively, these results indicate that the intrinsic quality of the oocyte is the main factor affecting blastocyst yields, while the conditions of embryo culture have a crucial role in determining blastocyst quality.     

The cost of realized sexual reproduction: assessing patterns of reproductive allocation and sporophyte abortion in a desert moss

The desert moss Syntrichia caninervis exhibits one of the most skewed sex ratios in the plant kingdom, with female individuals far outnumbering male individuals (exceeding 14♀:1♂). The "cost of sex hypothesis" derives from allocational theory and predicts that the sex which is most expensive should be the rarer sex. This hypothesis, which, as considered here represents the realized cost of sexual reproduction, is contingent upon two assumptions that are explored: (1) that male sex expression is more expensive than female sex expression, and (2) that sexual reproduction is resource limited. Using inflorescence biomass and discounting sperm, male sex expression was found to be in the neighborhood of one order of magnitude more expensive than female sex expression, and this difference is reflected in higher numbers of gametangia per male inflorescence, presence of paraphyses in male inflorescences, and a much longer developmental time for male inflorescences. The realized cost of female reproduction from two communities dominated by S. caninervis was found to be lower than the realized cost of male sexual reproduction. Resource-limited reproduction was assessed by determining the frequency of sporophyte abortion, the age distribution of sporophyte abortions, and patterns of sporophyte abortion that may be density dependent. Among ten sexually reproducing populations, abortive sporophytes occurred at a frequency of 0.64. Abortive sporophytes averaged 8% the mass of mature sporophytes, and cohort sporophytes from the same individual female were found to abort in a density-dependent pattern. We conclude that the two assumptions, upon which the cost of sex hypothesis depends, are supported.