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細胞學：非整倍體的植入前基因篩查是不科學的
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關於線粒體替代療法，幹細胞和CRISPR用於人類繁殖的一系列評論
引用自ivf.net
* PGT-A is a superstition of Genetics, and it is not according to the biological trait as Genetics described. Thus PGT-A is not a true genetic test but a DNA test without correct genetic results.
Human normal babies are derived from mosaic embryos with normal and aneuploid cells. Most
aneuploid cells in human embryos do not contain hereditary function after helping implantation
and differentiation. The above phenomena can only be explained by Cytohetics.
Ke-Hui Cui M.D., Ph.D.
Savannah, Georgia, 31405, U.S.A.
October 19, 2020
Email: khcui72@hereditics.net
Edited by Dr. YongYan Cui
Abstract
The presence of aneuploid cells in human embryogenesis is an important and beneficial phenomenon of embryonic physiology in implantation and differentiation. Preimplantation genetic screening (PGS) is screening for embryonic aneuploid cells that does not take into account the parent’s history, i.e. biological trait. When “mosaicism” embryos were transferred, they developed into 221 normal karyotype babies. The embryo’s DNA results did not correspond to the baby’s genetic results. In contrast, preimplantation genetic diagnosis (PGD), which accounts for the parent’s genetic history, maintained their embryo’s genetic results to live birth (>99.0%). The positive predictive value between PGS and PGD is significantly different (p<0.0001). Thus, PGS is not a true Genetic test but a DNA test when neglecting the principal component (biological trait) of Genetics. The failure of PGS in hereditary diagnosis is explained by Hereditics (the study of heredity). The patient’s genetic history is closely related to heredity control system (checkpoint and licensing system) in the cytoplasm. And patients’ hereditary diseases result from the combination of defects in both DNA and heredity control system. In PGS, parents without family history of hereditary disease rarely have defects in the hereditary control system. This is why most mosaic embryos will follow their parent’s normal biological trait and produce normal babies.
Keywords: preimplantation genetic screening; preimplantation genetic test; aneuploidy; mosaicism; hereditics; heredity.
History of PGS
In an early case of preimplantation genetic determination (PGD), a male embryo was misdiagnosed to be a female embryo by polymerase chain reaction (PCR) techniques. Thereafter, Dr. Delhanty and Dr, Handyside began to use fluorescent in-situ hybridization (FISH) techniques for sex determination to prevent X-linked hereditary diseases1. With the use of FISH techniques, a lot of human embryos were reported to be aneuploid. In 1997 Dr. Verlinsky claimed that “60% of human embryos are aneuploid”. Is it possible that such a high percentage of embryos are aneuploid when more than 99% of people in the world are normal karyotyping? Severe debate about “aneuploidy” and FISH techniques was initiated in the American Society for Reproductive Medicine (ASRM) meeting in 1997. At the same time, preimplantation genetic screening (PGS) (i.e. preimplantation genetic testing for aneuploidy, PGT-A) was prematurely introduced as a clinical tool for expectant families. The aim of PGS would be: First, “diagnosing and preventing inherited disease” by PGS “may appear useful”2,3. Second, if all embryos obtained by in vitro fertilization (IVF) were screened for aneuploidy before transfer, implantation and pregnancy rates would improve, and miscarriage rates would decrease4. FISH techniques, in which one side of the chromosomes (DNA) adhere to a glass slide and a long-DNA-sequence probe is used, are full of errors. Consequently, array comparative genomic hybridization (aCGH) and next generation sequencing (NGS) replaced FISH techniques. The “aneuploidy” problem is still present in human embryos. After blastocyst biopsy, people started to report, “60% of human embryos have ‘mosaicism”. However, “aneuploidy” and “mosaicism” were myths in the field of human reproduction.
In the 2015 ASRM meeting, Reprogenetics (company) recommended the use of blastocyst biopsy for PGS and elective single embryo transfer (eSET). eSET is highly accepted by society as a means to produce singletons, which is better for the mother’s and child’s health. However, performing PGS involves extremely traumatic techniques including add-on assisted hatching, laser radiation, embryo biopsy, and freezing for every embryo - a majority of which are originally healthy embryos. Given these risks, the combination of PGS and eSET may not ultimately benefit the embryo. Despite this, in 2016, there was a large-scale trend towards performing PGS in the USA. The clinical results in the USA in 2016 did not reflect improved outcomes but instead indicated worsened clinical outcomes with PGS. According to National Summary data from the Centers for Disease Control and Prevention (CDC) in the USA, in the <35 years old group who had embryo transfers, live birth rates were 47.5% in 2014, 46.5% in 2015, and 45.7% in 2016. Miscarriage rates (pregnancy rate minus live birth rate) did not decrease (2014, 7.1%; 2015, 7.3%; 2016, 7.2%)5-7. The second aim of PGS (“implantation and pregnancy rates would improve and miscarriage rates would decrease”) waned when faced with 2015 and 2016 U.S. national data 8.
At the same time, transfer of “mosaicism” embryos produced 221 babies with normal karyotyping9-22. This fact challenged the practice of PGS. How could “mosaicism” embryos confirmed by genetic techniques produce babies with normal rather than mosaic karyotyping? Geneticists failed to answer the question. The PGS-diagnosed aneuploid cells with abnormal DNA structures were not inherited in babies. In the 2019 ASRM meeting, Dr. Rechitsky reported that a total of 800 babies were born in the world after mosaic embryo transfer. Only one baby (i.e. 1/800 = 0.1%) was reported to be chromosomally abnormal23. Thus, there is no difference between the aneuploidy rate in live birth between these PGS-diagnosed aneuploidy embryos and naturally derived embryos (<0.1% mosaicism in fetus)24. With these data, the first aim of PGS (“diagnosing and preventing inherited disease”) was questionable25. The embryo’s genetic results (“mosaicism”) in PGS do not correspond to the baby’s genetic results.
Amidst the turmoil of PGS, the U.S. Congress issued “Personhood” concerns about PGS in March, 2019. It presented four legitimate concerns: First, concern about the concepts of aneuploid embryo and mosaicism embryo. Second, concern about accuracy and applicability of PGS diagnosis. Third, concern about discarded embryos, which is related to ethical questions and humanity. Fourth, concern about safety of PGS. This paper will focus on the first two concerns.
Before discussing the first two concerns, it is necessary to introduce the basic theory of heredity, which is referred to as Hereditics26: Hereditics is the study of heredity. It includes Genetics (study of genes and DNA) and Cytohetics (study of cytoplasmic heredity). The study of expression of genes is known as Epigenetics. The study of expression of cytoplasm, especially in differentiation of cytoplasm, is called “Epicytohetics”.
PGS is not scientific, and it is pointless in heredity
Heredity control system in every human cell
Some cells (especially some dying cells) are aneuploidy. They exist in our bodies, in fetuses, and in embryos and they die continuously. The temporary existence of aneuploid cells is a physiological mosaic phenomenon, rather than a pathological condition – mosaicism27. Cytohetics (study of cytoplasmic heredity) recognizes: Every cell contains a heredity control system, called a checkpoint and licensing system, mainly in its cytoplasm28-34. The checkpoint and licensing system is named for its biological function to control DNA replication, centrosome duplication and spindle assembly. The checkpoint and licensing system is also called the heredity control system when considering its heredity function. It prevents aneuploid cells from passing through the mitotic exit to split and to be inherited in progeny cells32. Thus, the mitotic exit (in the checkpoint and licensing system) functions as a “switch” of heredity in the cytoplasm30. When it is “turned on”, one cell can divide into two cells, and heredity from cell to cell will happen. In animal, cell to cell heredity of an embryo will at last lead to life to life heredity. When the mitotic exit is “turned off” by heredity control system, the aneuploid cell in late embryonic stage will not divide and will then undergo apoptosis, thus ending the said mosaic phenomena35. In contrast, the normal cells continuously pass through the mitotic exit, divide, and grow until an embryo becomes a life - fetus and then a human being without mosaicism. Only when the checkpoint and licensing system of an embryo is defective will the said pathological aneuploid cells in the embryo continuously pass mitotic exit to divide and develop into a baby with aneuploidy or mosaicism27. No self-correction of the defective checkpoint and licensing system was reported, and thus hypothesis of “self-correction” of aneuploidy is questionable27,36,37. The aneuploid cells are degenerated by apoptosis rather than still surviving by "self-correction". An embryo should not be defined as “mosaicism” or “aneuploidy” only based on the presence of aneuploid cell(s) on laboratory results. These cells only represent mosaic phenomena, most of which (799 normal babies/800 babies = 99.9%) is temporary physiological aneuploid cells due to the existing normal heredity control system.
Blastocyst biopsy is not scientific in PGS due to new loose checkpoint and licensing system in fast blastocyst differentiation
The embryo at the 8-cell stage is totipotent. When an embryo grows to the blastocyst stage, the cells have differentiated. Differentiation creates a significant difference between an inner cell mass (ICM) and trophectoderm38. Differentiation also leads to a significant difference in the checkpoint and licensing systems between the ICM and trophectoderm. The ICM, which will later develop into the germ tract, will maintain stricter licensing of transcription and translation39. However, the trophectoderm cells, which will develop into the chorion, will produce new and different loose licensing components40-42. Changes of checkpoint regulator PLK4 were found to be associated with mitotic aneuploidy in 15388 human trophectderm samples. The change of PLK4 are related to the new change of mitotic fidelity43,44. The new loose checkpoint and licensing system allows the trophectoderm cells to differentiate and produce syncytial and aneuploid cells45. In eukaryote (such as in parasite and fungi species), syncytia usually occur with aneuploidy to provide beneficial effects in evolution for adaptation to stress46-49. It is common phenomena that pervasive syncytial and aneuploid cells coexist with normal cells in normal human liver with both beneficial and detrimental consequence50,51. Thus, having many different kinds of aneuploid cells with syncytial cells in the differentiation of trophectoderm is normal phenomenon in evolution and in human physiology. It is not a pathological problem but is normal results from the new loose checkpoint and licensing system in trophectoderm differentiation52. Experimental results showed that 12 embryos biopsied on day 3 were confirmed as normal 2N (i.e. diploid). When they later grew into blastocysts and were analysed as individual cells, they all (except one arrest) contained different kinds of aneuploid cells. This demonstrates that the number of aneuploid cells in blastocysts is significantly higher than that in day 3 embryos due to normal differentiation. In that experiment, 10 of 21 blastocysts contained 4N chromosomes (47.6%) that differentiated to syncytial cells accompanying with all kinds of aneuploid cells53,54. This is a beneficial and benign phenomenon of differentiation in human embryos. Some aneuploid cells coexisting with syncytial cells have a more invasive function for villi to implant into the mother’s endometrium55,56 because “aneuploidy-independent modulation of the microtubule cytoskeleton enhances directional migration” and invasive strength57,58. The aneuploid cells in blastocysts and villi develop into a sharp angular shape, which can be seen under the microscope, and facilitate rapid invasion and implantation into the mother’s endometrium58. Without these aneuploidy cells, implantation will fail and miscarriage will result. The fast invasion allows adequate nutrition to support the fast-growing embryo and fetus, and prevent abnormal functioning of the ICM and embryonic soma. In another study, 42 human blastocysts had their individual cells tested. Almost all embryos (95%) contained some aneuploid cells59. It was assumed that up to 100% embryos contain more or less aneuploid cells60-62. The embryos containing some aneuploid cells are a healthy and physiological phenomenon in the blastocyst stage. These aneuploid cells and syncytial cells in the villi will not result in fetal mosaicism35. In 72,472 pregnancies, chromosomal mosaicism was noted in 2.1% of chorionic villi samples, and <0.1% [(75 – 14) / 72472] in the fetus24. “Overall, the proportion of aneuploid cells is progressively depleted from the blastocyst stage onwards”35. Performing blastocyst biopsy will therefore lead to the finding of more aneuploid cells, and lead to the misdiagnosis of pathological “mosaicism” and unjustifiable discard of normal embryos.
The basic principle of hereditary diseases in Genetics (i.e. patient’s hereditary history) is neglected in PGS
Heredity is the basic principle of hereditary diseases. All geneticists know: in chromosomal abnormalities and genetic diseases, the patient’s history is a valuable source of predicting the probability (such as 25% or 50%) of the offspring’s inheritance of the disease. Patient’s history reveals patient’s biological trait. However, PGS is used to detect aneuploid cells in the general population and does not take into account the patient’s history (or biological trait). In over 800 normal babies born from “mosaicism” embryos, there was only one aneuploid baby23. The PGS positive predictive value (PPV) for aneuploidy was 0.1% (1/800). PGS designers assumed that aneuploid cells would be inherited to offspring because DNA is a hereditary material2,3. However, the majority of aneuploid cells in human embryos are produced by the loose checkpoint and licensing system and are the physiological aneuploid cells, which do not have heredity characteristic40,43,52. This is why PGS repeatedly performed on the same embryos (biopsied on day 3, 5, and 12, or up to stem cells, and up to babies) very rarely have the same results36,37.
Biological inheritance refers to the passing of genetic traits from parents to their offspring. Preimplantation genetic diagnosis (PGD), which is performed based on the patient’s hereditary history, (i.e. following genetic traits) is highly successful. The defective checkpoint and licensing system in these PGD parents has an extremely high likelihood of being inherited to the embryos. Because the inherited defective system is unable to recognize DNA abnormalities, those abnormal embryonic cells will continue dividing and produce babies with the said hereditary diseases27,30. Thus, considering the principle of heredity (based on the patient’s history), the PPV of PGD in diagnosing hereditary diseases in embryos with parents with known hereditary disease is extremely high. In the ESHRE PGD Consortium data from 2011 to 2012, 17,721 PGD cycles resulted in 3,755 cycles with positive heartbeats. Only 34 misdiagnosed cases (0.9%) were reported63. This data supports that PGD has a 99% PPV. It contrasts with a 0.1% PPV when PGS does not take into account the parent’s family history. There are statistically significant differences (chi-squared test p<0.0001) between PGD and PGS in PPV. Based on statistics, if a clinical test cannot obtain a 95% correct diagnosis rate, the clinical diagnosis is insignificant and does not provide any meaningful clinical diagnoses. Thus, PGS is not qualified as a diagnostic test in heredity. Inferred by above theory of Hereditics, there are two heredity components in Genetics:
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Genetics alone cannot fully explain the failure of PGS
Genetics accounts for the genes that produce proteins. However, Genetics does not explain how those proteins in the checkpoint and licensing system function as mitotic exit in heredity control system27,30. However, Cytohetics (study of cytoplasmic heredity) explains that mosaic phenomenon will not turn into mosaicism because the checkpoint and licensing system in most embryos (99.9%) is normal rather than defective26. Mosaicism and hereditary diseases result from the combination of defects in both DNA and heredity control system26,27.
Why do most aneuploid cells (99%) in the blastocysts of PGS produce normal karyotyping babies? Many papers described that this phenomenon cannot be explained with our existing knowledge of Genetics. Concerns of false positive results were considered10,11,38,64,65. False positive and false negative results exist at a very low rate in PGS66. It is possible that some artifacts from messy blastocyst biopsy techniques and from the uneven distribution of aneuploid cells can lead to false positives even with advanced molecular technique NGS for PGS14,16,67,68. However, similar genetic test results of PGD with parent’s history has confirmed: it is not possible that any genetic test will produce 99% false positive results, but only about 1% false positive63. A systematical problem in PGS is: neglect of parents’ hereditary history. Accurate DNA results do not translate to an accurate diagnosis of babies that will become (or inherit) aneuploidy24. It was reported that aneuploid embryos diagnosed by NGS also produced 31 normal karyotyping babies19. The advanced NGS genetic diagnosis failed 100% in its heredity prediction in this report was not due to “false positive diagnoses” but due to high efficient functioning of the heredity control system in the aneuploid cells24,35. These findings suggest that genetics and genetic tests cannot fully explain the intricacies of heredity or biological trait in PGS. Genetics is not absolutely equal to heredity under the condition of neglecting family history.
Whether most aneuploid cells contain hereditary characteristics depends on whether their checkpoint and licensing system is normal27,30, and this can be determined based on the patient’s hereditary (family) history (i.e. biological trait). Without considering the patient’s history, PGS becomes a DNA test rather than a true Genetic test of hereditary diagnosis. Thus PGS cannot diagnose aneuploid embryos69. However, when taking the patient’s history into account, PGD becomes a test of both genetic and hereditary diagnosis63. The tool of accurate genetic test is excellent for the object of PGD but is incompetent for the object of PGS. Only recognition of Hereditics (i.e. both Genetics and Cytohetics) and therefore recognition of the importance of patient’s history allows PGT to be a genetic test for hereditary diagnosis. According to four components of Hereditics and two components in Genetics, PGS is not a true Genetic test due to PGS neglecting biological trait of the patient's family from Genetics.
Most aneuploid cells in early embryos have some normal and beneficial cellular functions in human embryos as a result of long evolutionary history
A. Maintaining embryo function. Most aneuploid cells in the embryos may regulate the embryo’s internal environment before undergoing apoptosis. These regulatory mechanisms include maintaining stable temperature, absorbing and transferring nutrition, buffering outside mechanical stress, and maintaining spatial configuration and polarity58.
B. Faster implantation. As mentioned above.
C. Aneuploid cells are related to apoptosis during organ and tissue differentiation. The development of a healthy child requires not only very high rates of proliferation and differentiation, but also apoptosis, which is a crucial mechanism for morphogenesis and the development of the inner organs. Changes in the regulation and rate of apoptosis in more or lesser level have deleterious effects on the trophoblast and consequently the developing embryo and fetus70,71. Without aneuploid cells and without apoptosis, there will be no differentiation, and no healthy human babies. In mammals, cellular proliferation, differentiation, and death accompany early placental development. Programmed cell death is a critical driving force behind organ sculpturing, including mammals, amphibians, and insects as a result of long evolutionary history72. In PGS, 110 human embryos that were biopsied on day 3 and had whole embryo analysis with individual cells on day 5, only one embryo (1%) consistently showed 100% 2N cells. However, that embryo was arrested with 12 cells on day 4 53,62. This confirmed that all growing normal human embryos are mosaic. Without aneuploid cells and apoptosis, the embryo could not grow further. Those rare blastocysts consistently containing 100% 2N cells are not healthy and cannot survive long, due to the lack of function at the time of differentiation.
Possibility of aneuploidy arises from meiotic or mitotic nondisjunction
Aneuploidy arises from meiotic or mitotic nondisjunction73. In meiotic events, it occurred in 5.2% of fresh oocytes74, and 4.5% of human sperm75. In embryos with meiotic nondisjunction, all of the cells in the said embryos will have uniform aneuploidy, which were about 3.9% in all of the analysed embryos (6 identical aneuploid embryos from 154 analysed human embryos)53,54,59. Most of them will arrest in early embryonic stages or will not be able to implant after transfer23. About 96% (100% -3.9%) of human embryos produce aneuploid cells by mitotic nondisjunction59-61. Most of the aneuploid cells in these embryos (99%) contain normal heredity control system and will arrest after blastocyst stage24,35.
Limited representability of aneuploidy rate in PGS diagnosis
In PGS, one cell is biopsied from an 8-cell embryo or 5 cells are biopsied from a blastocyst to represent the aneuploid rate of the whole embryo (which contains about one hundred cells). The faithful representability is about 12.5% (1/8) in a blastomere biopsy or 5% (5/100) in a blastocyst biopsy76. If FISH has a correct diagnosis rate of 95% in the biopsied cells77, the faithful representability of the biopsied cells in the whole blastocyst is 4.75% (0.05X0.95). If NGS is 100% sensitive67, the faithful representability is 5.0% (0.05X1.00). Thus, the improvement of genetic techniques68 does not greatly improve the representability of aneuploidy rate in PGS diagnosis.
When embryos have undergone repeated biopsy and testing, PGS showed discordant results38. The reason may be: different aneuploid cells in the same embryo are distributed unevenly in different locations11,65, or the heredity control system changes to looser or stricter condition as mentioned above. This is a problem of representability of the biopsied cells. Five-cell results cannot represent the hereditary condition of the whole embryo, or the embryo’s genetic results do not represent the baby’s genetic results. Thus the diagnosis of “euploid”, “mosaic” and “aneuploid” based on a percentage of aneuploid cells found by PGS12,68,78 is subjective.
Falsification of the “advantage” of PGS
The “advantage” of PGS has be focused for a long time in the second aim of PGS: “implantation and pregnancy rates would improve, and miscarriage rates would decrease”. Many prominent papers have shown the “advantage” of PGS, which provided an opportunity to be studied once more:
In advanced maternal age (AMA), oocyte nondisjunction increases significantly74. Thus, AMA was the first target of PGS. A report showed that the PGS group had a 37% pregnancy rate and 22.5% implantation rate while the control group yielded 27% pregnancy rate and 10.2% implantation rate79. However, the conclusion that PGS “has immediate impact on the ongoing implantation rate, especially in patients aged ≥38 years” was premature. It was contradicted by its later description: “The pregnancy rate per started cycles does not differ between the study and the control groups”. And PGS “leads to a statistically significant reduction in the number of cycles transferred (78% versus 93% in the controls; P<.001)” 79. Another study reported: “More than 3000 clinical cycles indicates the positive impact” of PGS for AMA patients80. Two years later, the same author, Dr. Verlinsky reported contrary conclusion: in older women, PGS “has a high cancellation rate and low cycle outcome”81. More papers confirmed that PGS significantly reduced the rates of ongoing pregnancies and live births in women of AMA82-84.
Another misperception of PGS stems from the confusion between PGS and PGD. In a study of “PGD cycles”, using the results of 7.8% (183/2359) PGD cycles to represent the results of 92.2% (2176/2359) PGS cycles was inappropriate85. The take-home baby rate after PGD was 23% (42/183) per cycle rather than 81.4% as reported in the abstract85. Using the advantage of PGD to describe PGS, or mixing up the PGD and PGS results86 is confusing in science.
Studies comparing the PGS data with general frozen embryo transfer (FET) data (as a control group)87-89 are also not convincing. The general FET embryos are the leftover embryos after fresh transfers, and therefore of lower quality and not suitable to be used as a control for comparison.
The most common method is to compare the PGS group with the control group by “per transfer” rather than by “per cycle”. No embryo transfer (ET) is quite usual in PGS. “No ET” rates can vary significantly: 17.2%90, 20.7%91, 25% (2003 CDC data)92, 31.0%93, 37.4%94, 52.8%95. High transfer cancellation rates correlated with reduced live-birth rates96. The patients per cycle is the total number of patients with intention to treat (ITT), while patients per transfer is a selected group within the whole group. Only using a selected group to show the advantage in the whole group receiving treatment is misleading. On the whole, when PGS was compared “versus morphology as selection criteria” for embryo transfer, PGS “did not improve overall pregnancy outcomes in all women, as analyzed per embryo transfer or per ITT”8,97,98.
Conclusion
PGS (PGT-A) is a large-scale practice that does not follow the principle of heredity and Genetics (i.e. the patient’s hereditary history). It screens aneuploid cell(s) at the embryonic differentiation stage in an attempt to diagnose hereditary diseases. It operates under the assumptions that: 1. the DNA of aneuploid cells always contain hereditary characteristics that lead to abnormal babies; 2. aneuploid cells only harm rather than help implantation; and 3. only embryos containing 100% euploid cells (normal DNA) are the best embryos. These assumptions are incorrect according to the above review. PGS is a type of misunderstood “Genetics” that lacks scientific basis in embryo differentiation and evolution. It is based on the wrong theory that DNA is the unique hereditary material in human beings, and neglects hereditary function of cytoplasm which is called the checkpoint and licensing system. Thus PGS is a kind of DNA tests rather than a true Genetic test. However, taking into account the patient’s history, PGD is not only a test of genetic diagnosis but also a test of hereditary diagnosis. In PGS, while human “mosaicism” embryos are transferred, they follow their parent’s biological trait and develop to normal karyotyping babies in a rate of 99.9%.
References:
1. Griffin, D. K., Wilton, L. J., Handyside, A. H., et al. Duel fluorescent in situ hybridization for simultaneous detection of X and Y chromosome-specific probes for the sexing of human preimplantation embryonic nuclei. Hum Genet. 1992, 89: 18-22.
2. Munné, S., Dailey, T., Sultan, J., et al. Diagnosing and preventing inherited disease: The use of first polar bodies for preimplantation diagnosis of aneuploidy. Hum Reprod. 1995, 10: 1014-1020.
3. Verlinsky, Y., Cieslak, J., Freidine, M., et al. Diagnosing and preventing inherited disease: Pregnancies following pre-conception diagnosis of common aneuploidies by fluorescent in-situ hybridization. Hum Reprod. 1995, 10: 1923-1927.
4. Munné, S., Lee, A., Rosenwaks, Z., et al. Diagnosis of major chromosome aneuploidies in human preimplantation embryos. Hum Reprod. 1993, 8: 2185-2191.
5. CDC. 2014, https://ftp.cdc.gov › pub › publications › art › ART-2014-Clinic-Report-Full. Page 21.
6. CDC 2015, ftp.cdc.gov › pub › art › 2015 › ART_9999_2015_Fertility_Clinic_Report. Page 21.
7. CDC. 2016, ftp.cdc.gov › pub › Publications › art › ART-2016-Clinic-Report-Full. Pp. 22.
8. Paulson, R. J. Outcome of in vitro fertilization cycles with preimplantation genetic testing for aneuploidies: let’s be honest with one another. Fertil Steril. 2019, 112: 1013-1014.
9. Scott, R. T. Jr., Ferry, K., Su, J., et al. Comprehensive chromosome screening is highly predictive of the reproductive potential of human embryos: a prospective, blinded, nonselection study. Fertil Steril. 2012, 97: 870-875.
10. Greco, E., Minasi, M. G, Florentino, F. Healthy babies after intrauterine transfer of mosaic aneuploid blastocysts. N Engl J Med. 2015, 373: 2089–2090.
11. Gleicher, N., Vidali, A., Braverman, J., et al. Accuracy of preimplantation genetic screening (PGS) is compromised by degree of mosaicism of human embryos. Reprod Biol Endocrinol. 2016, 14, 54. doi: 10.1186/s12958-016-0193-6.
12. Maxwell, S. M., Colls, P., Hodes-Wertz, B., et al. Why do euploid embryos miscarry? A case-control study comparing the rate of aneuploidy within presumed euploid embryos that resulted in miscarriage or live birth using next-generation sequencing. Fertil Steril. 2016, 106: 1414–1419.e5.
13. Inoue, N, Lopez, R., Delgado, A., et al. Mosaic embryo transfer after oocyte in vitro maturation in combination with non-invasive prenatal testing (NIPT)-first report of a euploid live birth. J Assist Reprod Genet. 2017, 34: 1199-1205.
14. Fragouli, E., Alfarawati, S., Spath, K., et al. Analysis of implantation and ongoing pregnancy rates following the transfer of mosaic diploid-aneuploid blastocysts. Hum Genet. 2017, 136: 805–819.
15. Lledo, B., Morales, R., Ortiz, J. A., et al. Implantation potential of mosaic embryos. Syst Biol Reprod Med. 2017, 63: 206-208.
16. Victor, A. R., Tyndall, J. C.,, Brake, A. J., et al. One hundred mosaic embryos transferred prospectively in a single clinic: exploring when and why they result in healthy pregnancies. Fertil Steril. 2019, 111: 280-293.
17. Zhang, L., Wei, D., Zhu, Y., et al. Rates of live birth after mosaic embryo transfer compared with euploid embryo transfer. J Assist Reprod Genet. 2019, 36: 165-172.
18. Spinella, F., Biricik, A., Minasi, M. G., et al. Impact of chromosomal mosaicism in IVF outcomes: experience from two hundred mosaic embryos transferred prospectively. pgdis.org › docs › PGDIS2019_GenevaMaterials_0419. OC-07.
19. Shen, X., Chen, D., Fu, Y., et al. Retrospective PGT-A analysis for multiple displacement amplification products of embryos corresponding to 104 live births by next-generation sequencing (NGS). pgdis.org › docs › PGDIS2019_GenevaMaterials_0419. P-34.
20. Filippini, G., Fiandanese, N., Risch, L., et al. Two healthy babies born after implantation of multiple chromosomally rescued oocytes. pgdis.org › docs › PGDIS2019_GenevaMaterials_0419. P-36.
21. Patrizio, P., Shoham, G., Shoham, Z., et al. Worldwide live births following the transfer of chromosomally “abnormal” embryos after PGT/A: results of a worldwide web-based survey. J Assist Reprod Genet. 2019, 36: 1599-1607.
22. Zore, T., Kroener, L. L.,, Wang, C.,, et al. Transfer of embryos with segmental mosaicism is associated with a significant reduction in live-birth rate. Fertil Steril. 2019, 111: 69-76.
23. Munné, S., Spinella, F., Grifo, J., et al. Clinical outcomes after the transfer of blastocysts characterized as mosaic by high resolution Next Generation Sequencing- further insights. Eur J Med Genet. 2019, Aug 21: 103741.
24. Grati, F. R., Gallazzi, G., Branca, L., et al. An evidence-based scoring system for prioritizing mosaic aneuploid embryos following preimplantation genetic screening. Reprod Biomed Online. 2018, 36: 442-449.
25. Gleicher, N., Kushnir, V. A., Barad, D. H. How PGS/PGT-A laboratories succeeded in losing all credibility. Reprod Biomed Online. 2018, 37: 242-245.
26. Cui, K. H. Origin of life, heredity, differentiation and human reproduction: control of overpopulation and prohibition of subhuman reproduction. Letter to FDA, CDC and ASRM on March 15th, 2019.
27. Faesen, A. C., Thanasoula, M., Maffini, S., et al. Basis of catalytic assembly of the mitotic checkpoint complex. Nature. 2017, 542: 498-502.
28. Kramer, A., Neben, K., Ho, A.D. Centrosome replication, genomic instability and cancer. Leukemia. 2002, 16: 767-775.
29. Haase, S.B., Winey, M., Reed, S. I. Multi-step control of spindle pole body duplication by cyclin-dependent kinase. Nat Cell Biol. 2001, 3: 38-42.
30. Xie, S. Q., Xie, B., Lee, M. Y., et al. Regulation of cell cycle checkpoints by polo-like kinases. Oncogene. 2005, 24: 277-286.
31. Blow, J. J., Dutta, A. Preventing re-replication of Chromosomal DNA. Nat Rev Mol Cell Biol. 2005, 6: 476-486.
32. Bettencourt-Dias, M., Glover, D. M. Centrosome biogenesis and function: Centrosomics brings new understanding. Nat Rev Mol Cell Biol. 2007, 8: 451-463.
33. Musacchio, A., Salmon, E. D. The spindle- assembly checkpoint in space and time. Nat Rev Mol Cell Biol. 2007, 8: 379-393.
34. Shen, Z., Prasanth, S. G. Emerging players in the initiation of eukaryotic DNA replication. Cell Div. 2012, 7: 22.
35. Bolton, H., Graham, S. J. L, Van der Aa, N., et al. Mouse model of chromosome mosaicism reveals lineage-specific depletion of aneuploid cells and normal development potential. Nat Commun. 2016, 7: 11165.
36. Munné, S., Velilla, E., Colls, P., et al. Self-correction of chromosomally abnormal embryos in culture and implications for stem cell production. Fert Steril. 2005, 84: 1328-1334.
37. Barbash-Hazan, S., Frumkin, T., Malcov, M., et al. Preimplantation aneuploid embryos undergo self-correction in correlation with their developmental potential. Fertil Steril. 2009, 92: 890-896.
38. Popovic, M., Dheedene, A., Christodoulou, C., et al. Chromosomal mosaicism in human blastocysts: the ultimate challenge of preimplantation genetic testing? Hum Reprod. 2018, 33: 1342-1354.
39. Haig, D. Transposable elements: Self-seekers of the germline, team-players of the soma. Bioessays. 2016, 38: 1158-1166.
40. Lin, H. C., Wu, J. T., Tan, B. C., et al. Cul4 and DDB1 regulate Orc2 localization, BrdU incorporation and Dup stability during gene amplification in Drosophila follicle cells. J Cell Sci. 2009, 122: 2393-2401.
41. McCoy, R. C. Mosaicism in preimplantation human embryos: when chromosomal abnormalities are the norm. Trends Genet. 2017, 33: 448-463.
42. Jones, R. N., Erhard, S. A., Malham, M. R., et al. Expression and characterization of the zebrafish orthologue of the human FOLR1 gene during embryogenesis. Gene Expr Patterns. 2017, 25-26: 159-166.
43. McCoy, R. C, Demko, Z., Ryan, A., et al. Common variants spanning PLK4 are associated with mitotic-origin aneuploidy in human embryos. Science. 2015, 348: 235-238.
44. Bahce, M., Cohen, J., Munné, S. Preimpllantation genetic diagnosis of aneuploidy: were we looking at the wrong chromosomes? J Assist Reprod Genet. 1999, 16: 176-181.
45. Biancotti, J. C. and Benvenisty, N. Aneuploid human embryonic stem cells: origins and potential for modeling chromosomal disorders. Regen Med. 2011, 6: 493-503.
46. Parris, G. E. Clinically significant cancer evolves from transient mutated and/or aneuploid neoplasia by cell fusion to form unstable syncytia that give rise to ecologically viable parasite species. Med Hypotheses. 2005, 65: 846-850.
47. Rancati, G., Pavelka, N., Fleharty, B., et al. Aneuploidy underlies rapid adaptive evolution of yeast cells deprived of a conserved cytokinesis motor. Cell 2008, 135: 879-893.
48. Storchova, Z. Ploidy changes and genome stability in yeast. Yeast. 2014, 31: 421-430.
49. Berman, J. Ploidy plasticity: a rapid and reversible strategy for adaptation to stress. FEMS Yeast Res. 2016, 16: 1-5.
50. Duncan, A. W., Taylor, M. H., Hickey, R. D., et al. The ploidy-conveyor of mature hepatocytes as a source of genetic variation. Nature. 2010, 467: 707-710.
51. Duncan, A. W., Hanlon Newell, A. E., Smith, L., et al. Frequent aneuploidy among normal human hepatocytes. Gastroenterology. 2012, 142: 25-28.
52. Hong, Y. L., Stambrook, P. J. Restoration of an absent G1 arrest and protection from apoptosis in embryonic stem cells after ionizing radiation. PNAS. 2004, 101: 14443-14448.
53. Baart, E. B., Van Opstal, D., Los, F. J., et al. Fluorescence in situ hybridization analysis of two blastomeres from day 3 frozen-thawed embryos followed by analysis of the remaining embryo on day 5. Hum Reprod. 2004, 19: 685-693.
54. Baart, E. B., Martini, E., van den Berg, I., et al. Preimplantation genetic screening reveals a high incidence of aneuploidy and mosaicism in embryos from young women undergoing IVF. Hum Reprod. 2006, 21: 223-233.
55. Enders, A. C., Hendrickx, A. G., Schlafke, S. Implantation in the rhesus monkey: initial penetration of endometrium. Am J Anat. 1983, 167: 275-298.
56. Red-Horse, K., Zhou, Y., Genbacev, O., et al. Trophoblast differentiation during embryo implantation and formation of the maternal-fetal interface. J Clin Invest. 2004, 114: 744-754.
57. Ogden, A., Rida, P. C. G., Aneja, R. Heading off with the herd: How cancer cells might maneuver supernumerary centrosomes for directional migration. Cancer Metastasis Rev. 2013, 32: 269-287.
58. Huber, F., Schnau, J., Ronicke, S., et al. Emergent complexity of the cytoskeleton: from single filaments to tissue. Adv Phys. 2013, 62: 1-112.
59. Daphnis, D. D., Delhanty, J. D. A., Jerkovic, S., et al. Detailed FISH analysis of day 5 human embryos reveals the mechanisms leading to mosaic aneuploidy. Hum Reprod. 2005, 20: 129-137.
60. Munné, S., Grifo, J., Cohen, J., et al. Chromosome abnormalities in human arrested preimplantation embryos: a multiple-probe FISH Study. Am J Hum Genet. 1994, 55: 150-159.
61. Magli, M. C., Gianaroli, L., Fortini, D., et al. Impact of blastomere biopsy and cryopreservation techniques on human embryo viability. Hum Reprod. 1999, 14: 770-773.
62. Chatzimeletiou,, K., Morrison, E. E., Panagiotidis, Y., et al. Comparison of effects of zona drilling by non-contact infrared laser or acid tyrode's on the development of human biopsied embryos as revealed by blastomere viability, cytoskeletal analysis and molecular cytogenetics. Reprod Biomed Online. 2005, 11: 697-710.
63. De Rycke, M., Goossens, V., Kokkali, G., et al. ESHRE PGD Consortium data collection XIV-XV: cycles from January 2011 to December 2012 with pregnancy follow-up to October 2013. Hum Reprod. 2017, 32: 1974-1994.
64. Esfandiari, N., Bunnell, M. E., Casper, R. F. Human embryo mosaicism: did we drop the ball on chromosomal testing? J Assist Reprod Genet. 2016, 33: 1439-1444.
65. Orvieto, R. Preimplantation genetic screening- the required RCT that has not yet been carried out. Reprod Biol Endocrinol. 2016, 14: 35.
66. Hanson, C., Hardarson, T., Lundin, K., et al. Re-analysis of 166 embryos not transferred after PGS with advanced reproductive maternal age as indication. Hum Reprod. 2009, 24: 2960-2964.
67. Fiorentino, F., Bono, S., Biricik, A., et al. Application of next-generation sequencing technology for comprehensive aneuploidy screening of blastocysts in clinical preimplantation genetic screening cycles. Hum Reprod. 2014, 29: 2802-2813.
68. Munné, S., Blazek, J., Large, M., et al. Detailed investigation into the cytogenetic constitution and pregnancy outcome of replacing mosaic blastocysts detected with the use of high-resolution next-generation sequencing. Fertil Steril. 2017, 108: 62-71.
69. Cram, D. S., Leigh, D., Handyside, A., et al. PGDIS position statement on the transfer of mosaic embryos 2019. RBMO. 39, Issue S1, e1-e4.
70. Huppertz, B., Herrler, A. Regulation of proliferation and apoptosis during development of the preimplantation embryo and the placenta. Birth Defects Res C Embryo Today. 2005, 75: 249-261.
71. Dumortier, J. G., Maitre, J. L. A developmental checkpoint synchronizes morphogenesis and cellular differentiation in mammalian embryos. Med Sci (Paris) 2018, 34: 205-207.
72. Jurisicova, A., Detmar, J., Caniggia, I. Molecular mechanisms of trophoblast survival: from implantation to birth. Birth Defects Res C Embryo Today. 2005, 75: 262-280.
73. Conlin, L. K., Thiel, B. D., Bonnemann, C. G., et al. Mechanisms of mosaicism, chimerism and uniparental disomy identified by single nucleotide polymorphism array analysis. Hum Mol Genet. 2010, 19: 1263-1275.
74. Dailey, T., Dale, B., Cohen, J. et al. Association between nondisjunction and maternal age in meiosis-II human oocytes. Am J Hum Genet. 1996, 59: 176-184.
75. Templado, C., Vidal, F., Estop, A. Aneuploidy in human spermatozoa. Cytogenet Genome Res. 2011, 133: 91-99.
76. Simpson, J. L, Rechitsky, S., Kuliev, A. Next-generation sequencing for preimplantation genetic diagnosis. Fertil Steril. 2013, 99: 1203-1204.
77. Munné, S., Weier, H. U. Simultaneous enumeration of chromosomes 13, 18, 21, X, and Y in interphase cells for preimplantation genetic diagnosis of aneuploidy. Cytogenet Cell Genet. 1996, 75: 263-270.
78. Tsuiko, O., Zhigalina, D. I., Jatsenko, T., et al. Karyotype of the blastocoel fluid demonstrates low concordance with both trophectoderm and inner cell mass. Fertil Steril. 2018, 109: 1127-1134.
79. Gianaroli, L., Magli, M. C., Ferraretti, A. P., et al. Preimplantation diagnosis for aneuploidies in patients undergoing in vitro fertilization with a poor prognosis: identification of the categories for which it should be proposed. Fertil Steril. 1999, 72: 837-844.
80. Kuliev, A., Verlinsky, Y. The role of preimplantation genetic diagnosis in women of advanced reproductive age. Curr Opin Obstet Gynecol. 2003, 15: 233-238.
81. Taranissi, M., El-Toukhy, T., Gorgy, A., et al. Influence of maternal age on the outcome of PGD for aneuploidy screening in patients with recurrent implantation failure. Reprod Biomed Online. 2005, 10: 628-632.
82. Platteau, P., Staessen, C., Michiels, A., et al. Preimplantation genetic diagnosis for aneuploidy screening in patients with unexplained recurrent miscarriages. Fertil Steril. 2005, 83: 393-397.
83. Mastenbroek, S., Twisk, M., van Echten-Arends, J., et al. In vitro fertilization with preimplantation genetic screening. N Engl J Med. 2007, 357: 9-17.
84. Hardarson, T., Hanson, C., Lundin, K., et al. Preimplantation genetic screening in women of advanced maternal age caused a decrease in clinical pregnancy rate: a randomized controlled trial. Hum Reprod. 2008, 23: 2806-2812.
85. Verlinsky, Y., Tur-Kaspa, I., Cieslak, J., et al. Preimplantation testing for chromosomal disorders improves reproductive outcome of poor-prognosis patients. Reprod Biomed Online. 2005, 11: 219-225.
86. Hou, W., Xu, Y., Li, R., et al. Role of aneuploidy screening in preimplantation genetic testing for monogenic diseases in young women. Fertil Steril. 2019, 111: 928-935.
87. El-Toukhy, T., Kamal, A., Wharf, E., et al. Reduction of the multiple pregnancy rate in a preimplantation genetic diagnosis programme after introduction of single blastocyst transfer and cryopreservation of blastocysts biopsied on day 3. Hum Reprod. 2009, 24: 2642-2648.
88. Van Heertum, K. PGT for aneuploidy improves perinatal outcomes compared with FET alone: an analysis of the 2014 and 2015 SART data. 2019 ASRM O-73.
89. Bhatt, S. Pregnancy outcomes following in vitro fertilization frozen embryo transfer (IVF-FET) with or without preimplantation genetic testing for aneuploidy (PGT-A) in women with recurrent pregnancy loss (PRL): a SART-CORS study. 2019 ASRM O-76.
90. Munné, S., Fischer, J., Warner, A., et al. Preimplantation genetic diagnosis significantly reduces pregnancy loss in infertile couples: a multicenter study. Fertil Steril. 2006, 85: 326-332.
91. Findikli, N., Kahraman, S., Saglam, Y., et al. Embryo aneuploidy screening for repeated implantation failure and unexplained recurrent miscarriage. Reprod Biomed Online. 2006, 13: 38-46.
92. Mersereau, J. E., Plunkett, B. A., Cedars, M. I. Preimplantation genetic screening in older women: a cost-effectiveness analysis. Fertil Steril. 2008, 90: 592-598.
93. Pagidas, K., Ying, Y., Keefe, D. Predictive value of preimplantation genetic diagnosis for aneuploidy screening in repeated IVF-ET cycles among women with recurrent implantation failure. J Assist Reprod Genet. 2008, 25: 103-106.
94. Ercelen, N., Turtar, E., Gultomruk, M., et al. Successful preimplantation genetic aneuploidy screening in Turkish patients. Genet Mol Res. 2011, 10: 4093-4103.
95. Liang, L., Wang, C. T., Sun, X., et al. Identification of chromosomal errors in human preimplantation embryos with oligonucleotide DNA microarray. PLoS One. 2013, 8, e61838.
96. Ginsburg, E. S., Baker, V. L., Racowsky, C., et al. Use of preimplantation genetic diagnosis and preimplantation genetic screening in the United States: a Society for Assisted Reproductive Technology Writing Group paper. Fertil Steril. 2011, 96: 865-868.
97. Munné, S., Kaplan, B., Frattarelli, J. L., et al. Preimplantation genetic testing for aneuploidy versus morphology as selection criteria for single frozen-thawed embryo transfer in good-prognosis patients: a multicenter randomized clinical trial. Fertil Steril. 2019, 112: 1071-1079 e7.
98. Schattman, G. L. Preimplantation genetic testing for aneuploidy: It's déjà vu all over again! Fertil Steril. 2019, 112: 1046-1047.