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Phosphatase and tensin homolog deleted on chromosome 10 (PTEN), a tumor suppressor protein with dual phosphatase activity, is found to be frequently mutated in various cancers. PTEN is post-translationally modulated at various amino acid residues which are crucial for sub-cellular localization as well as its catalytic functions rendering genomic stability. Recent reports suggest that PTEN also acts as a DNA repair protein. But how post-translational modulation of PTEN affects cytological damage and aneuploidy is not studied in detail. Here, we focus on the role of sumoylation of PTEN in context with DNA damage induced cytological damage like micronucleus (MNi), nuclear bud (NB), and nuclear bridge formation. Our data suggest that wild type PTEN but not sumo-dead PTEN significantly reduces cytological damage in PTEN mutant PC3 cells. In case of sumo-dead PTEN, the cytological parameters are increased during 24h recovery time point after DNA damage. Next, we measured the effectiveness of the sumo-dead (PTEN-K254R) mutant on aneuploidy, where we found that sumoylation is essential for maintaining chromosome number. As chromosome number variation in daughter cell is due to multiple spindle pole formation, we qualitatively and quantitatively evaluate the $\gamma$ tubulin pole formation in PTEN-K254R clone transfected cells. We found aberrant pole formation is significantly increased in PTEN-K254R transfected cells compared to wild-type PTEN. Further depletion of sumoylation activity of PTEN increases the expression of phosphorylated form of Aurora kinase A (AURKA) (T288) and PLK1 (T210) proteins with or without nocodazole, a microtubule depolymerizing agent compared to cells expressing wild-type PTEN. Thus, sumoylation of PTEN is essential for maintaining genomic stability.

The identification of viable embryos for transfer is one of the main challenges in reproductive medicine. As chromosomal abnormalities are the major cause of implantation failure, preimplantation genetic testing of aneuploidy plays an important role in embryo selection. To make this approach more efficient, the possibility to retrieve informative DNA through a moderately invasive technique compared to the traditional forms of biopsy is appealing. Blastocoelic fluid is a valuable source of DNA. Its presence, as detected by whole genomic amplification, and the following analysis by comprehensive chromosome screening could add important information on the blastocyst ploidy condition and developmental potential.

Research Question: Does the dose of gonadotropin used for superovulation in IVF affect the proportion of euploid blastocysts obtained after fertilization?

Study Design: Multicentre randomized controlled trial recruiting 57 women who were treated with ovarian stimulation using either 150 or 300 IU Menopur per day. Both groups received GnRH antagonist from day 5 of ovarian stimulation and final oocyte maturation was induced using a leuprolide GnRH (gonadotropin releasing hormone) agonist trigger when three or more follicles reached 17 mm diameter. Oocyte collection was scheduled 36–38 hours post trigger. In vitro fertilization (IVF) or Intracytoplasmic Sperm Injection (ICSI) were performed according to individual unit protocol and embryos were cultured to blastocyst stage. A trophectoderm biopsy was performed on day 5 of embryo culture and used for preimplantation genetic testing for aneuploidy. Euploid embryos were transferred in subsequent frozen embryo transfer cycles with appropriate endometrial preparation.

Results: The number of oocytes obtained from women randomized to 150 IU Menopur was between 3 and 17 (mean = 9), whereas the number of oocytes obtained from women randomized to 300 IU Menopur was between 3 and 24 (mean = 11). There was a positive linear relationship between serum AMH concentration and oocyte yield in both the 150 and 300 IU Menopur groups ($R$ = 0.3359, $R^2$ = 0.1129 and $R$ = 0.3741, $R^2$ = 0.1399). The percentage of euploid to aneuploid embryos in the 150 IU Menopur group was 63% and in the 300 IU Menopur group, the proportion was 75%, which was not significantly different ($p$ = 0.17).

Conclusion: The higher dose ovarian stimulation protocol did not significantly increase the number of oocytes retrieved, nor did the higher dose protocol reduce the proportion of euploid embryos created. This study does not support the hypothesis that use of higher doses of gonadotropin for ovarian stimulation results in a reduction in the proportion of euploid embryos obtained after IVF.

With continued improvements in blastocyst culture, cell sampling approaches, and genetic analysis platforms, the resulting improvements in embryo development and the resolution and accuracy of chromosome analysis have provided valuable insights into the preimplantation embryo. This includes the impact of in vitro culture conditions on chromosomal dynamics. Specifically, through analysis of embryo aneuploidy and mosaicism, a growing number of reports indicate that rates of chromosomal abnormalities can vary between IVF centers. Because differences in mosaicism reflect mitotic errors, this endpoint analysis suggests that IVF laboratory-controlled variables during embryo development may be influencing chromosome separation and segregation. A growing body of literature suggests that culture media may be one variable influencing preimplantation embryo aneuploidy and mosaicism. However, these data are far from definitive in demonstrating cause-and-effect. Whether reported differences may be due to media formulation, use of sequential media or single-step media, or uninterrupted culture approaches is unknown. Importantly, variables directly impacting media performance and embryo development, including pH, temperature, osmolality, and oxygen concentration, must also be considered and make it difficult to isolate the impact of culture media as the sole factor responsible. These IVF laboratory variables will be reviewed and literature suggesting a possible link to mitotic aneuploidy/mosaicism will be discussed.

Background and Aims: MicroRNAs (miRNAs) are abundantly expressed in rapidly growing and undifferentiated cells, for example, embryonic stem cells. This led to the discovery that miRNAs are significantly expressed in human embryos and that intracellular miRNAs expression patterns differ between euploidy and aneuploid embryos. Our study aims to determine the correlation between miR-35b and embryo chromosomal status.

Methods: Thirty-six blastocyst culture media were collected from patients who underwent the In Vito Fertilization (IVF) program followed by Preimplantation Genetic Testing for Aneuploidy (PGT-A) at Dr. Ciptomangunkusumo General Hospital, Jakarta. The trophectoderm cells were biopsied, and half of the spent media culture was evaluated for aneuploidy detection using Next Generation Sequencing (NGS). While the rest of the spent media on day five were analyzed for miR-135b expression using quantitative Real-Time Polymerase Chain Reaction (RT-qPCR). All the data were analyzed using SPSS, and the sensitivity and specificity of the miR-135b were analyzed using the ROC curve.

Results: miR-135b relative expression was detected in blastocyst spent blastocyst media culture and not in culture media without embryo. Based on the average of miR-135b, expression was higher in aneuploidy embryos (7.9(0.01-1-45.57)) than embryos with no aneuploidy detected (0.7(0.003-9.65)) and statistically different with p¡0.05. miR-135 relative expression in spent culture media was 10.72-fold higher in aneuploidy compared with an embryo with a normal chromosome. Based on the validation, the sensitivity value was 71.4%, and the specificity was 68.2%, with a cut-off of the relative expression of miR-135b was 1.185-fold.

Conclusion: miR-135b detected in IVF spent culture media from blastocyst media culture. The significant expression of miR-135b in an aneuploidy embryo makes miR-135b a potential biomarker for predicting the chromosomal quality of blastocyst embryos.