Shi Wenhao, Zhao Zhenghao, Xue Xia, Li Qian, Yao Yaxin, Wang Dongyang, Wang Jing, Lu Sijia, Shi Juanzi
The Assisted Reproduction Center, Northwest Women's and Children's Hospital, Xi'an, China.
Department of Clinical Research, Yikon Genomics Company, Ltd., Suzhou, China.
Front Physiol. 2022 Feb 21;13:794210. doi: 10.3389/fphys.2022.794210. eCollection 2022.
Recent studies have demonstrated that both blastocoel fluid (BF) and spent cell culture media (SCM) have potential as materials for non-invasive or less-invasive pre-implantation genetic analysis. BF may allow more opportunity to obtain cell-free DNA from the inner cell mass (ICM), and it has a lower risk of containing contaminant DNA from cumulus cells, sperm and culture media. There are no data regarding the ICM as a gold standard to evaluate the chromosome constitution of BF or SCM for embryo liquid biopsy.
Two hundred eighteen donated human blastocysts were warmed and cultured in blastocyst culture media for 18-24 h. The corresponding SCM was collected, and only clear ICM was biopsied in blastocysts; otherwise, the whole blastocyst (WB) was biopsied. Quantitative PCR was performed to determine the DNA levels in the SCM and BF before and after amplification. ChromInst was used to amplify BF/SCM and blastocyst DNA before sequencing. Chromosomal copy number variation (CNV) was investigated to evaluate the chromosome constitution.
In total, 212 blastocysts were available for SCM and BF collection. The technical success rates (next-generation sequencing data) were 100 and 69.8% (148/212) for SCM and BF, respectively. Among the 148 blastocysts with both SCM and BF data, 101 were euploid and 47 were aneuploid based on ICM ( = 89) or WB ( = 59) analysis as the gold standard. Among all blastocysts, SCM was comparable to BF [specificity: 80.2 versus 61.4% ( = 0.005, χ test); sensitivity: 91.5 versus 87.2% ( = 0.738, χ test); negative predictive value (NPV): 95.3 versus 91.2% ( = 0.487, χ test); positive predictive value (PPV): 68.3% versus 51.3% ( = 0.042, χ test)]. The SCM and BF samples were 83.8% (124/148) and 69.6% (103/148) concordant with the corresponding ICM/WB samples when only two categories, euploid or aneuploid/mosaic, were grouped to calculate the concordance.
Compared with BF, SCM has superior diagnostic performance, and it is non-invasive for embryos.
[http://www.chictr.org.cn], identifier [ChiCTR-BPD-17014087].
最近的研究表明，囊胚腔液（BF）和废弃细胞培养液（SCM）都有潜力作为非侵入性或微创性植入前基因分析的材料。BF可能为从内细胞团（ICM）获取游离DNA提供更多机会，且其含有来自卵丘细胞、精子和培养液的污染DNA的风险较低。目前尚无关于以ICM作为评估BF或SCM用于胚胎液体活检的染色体构成的金标准的数据。
218枚捐赠的人类囊胚经复温后在囊胚培养液中培养18 - 24小时。收集相应的SCM，仅对囊胚中清亮的ICM进行活检；否则，对整个囊胚（WB）进行活检。在扩增前后进行定量PCR以确定SCM和BF中的DNA水平。在测序前使用ChromInst扩增BF/SCM和囊胚DNA。研究染色体拷贝数变异（CNV）以评估染色体构成。
总共212枚囊胚可用于SCM和BF的收集。SCM和BF的技术成功率（下一代测序数据）分别为100%和69.8%（148/212）。在148枚同时有SCM和BF数据的囊胚中，以ICM（n = 89）或WB（n = 59）分析作为金标准，101枚为整倍体，47枚为非整倍体。在所有囊胚中，SCM与BF相当[特异性：80.2%对61.4%（P = 0.005，χ检验）；敏感性：91.5%对87.2%（P = 0.738，χ检验）；阴性预测值（NPV）：95.3%对91.2%（P = 0.487，χ检验）；阳性预测值（PPV）：68.3%对51.3%（P = 0.042，χ检验）]。当仅将整倍体或非整倍体/嵌合体两类分组计算一致性时，SCM和BF样本与相应的ICM/WB样本的一致性分别为83.8%（124/148）和69.6%（103/148）。
与BF相比，SCM具有更好的诊断性能，且对胚胎是非侵入性的。
[http://www.chictr.org.cn]，标识符[ChiCTR - BPD - 17014087]
