Biology subjects

Holubcova, Z.

Publications and source records attributed to Holubcova, Z..

4 recordsLinked to original sources

bioRxiv2026

Restoring Shugoshin 1 reduces chromosome errors in human eggs

Aneuploidy in human eggs, which rises sharply with age, is a leading cause of infertility, IVF failure, and miscarriage. This age-related aneuploidy is primarily driven by premature sister chromatid separation (PSSC), resulting from loss of the cohesin complex that holds chromatids together. How cohesin is destabilized in the long-lived mammalian oocyte is poorly understood. Here, we show that in mouse oocytes, pericentromeric transcription is essential for maintaining the cohesion protector Shugoshin 1 (SGO1) and PP2A at centromeres, which together safeguard the cohesin subunit REC8. With age, mouse oocytes lose pericentromeric transcription, SGO1, and PP2A, leading to destabilized cohesion and increased PSSC. Supplementing aged mouse oocytes with Sgo1 restores centromeric protection, and reduces PSSC to youthful levels. Aged human oocytes also show reduced SGO1, and SGO1 supplementation reduces the fraction of human eggs with PSSC by approximately half. These findings establish SGO1 supplementation as a potential strategy to preserve chromatid cohesion in aging oocytes.

cell biology↗

bioRxiv2025

Human Serum Albumin as a Hidden Source of Variability in IVF Culture Media

Soluble proteins present in spent culture media (SCM) have been proposed as potential indicators of embryo viability. However, results across studies remain inconsistent, and the identity of reliable protein biomarkers is still unclear. To evaluate the potential of secretome profiling for non-invasive embryo assessment, we applied state-of-the-art mass spectrometry to analyze the protein composition of SCM from individual successful and unsuccessful human embryo cultures, alongside corresponding controls cultured under identical conditions. Surprisingly, both untargeted (nanoLC-TIMS-TOF-MS/MS) and targeted (UHPLC-QqQ-MS/MS) proteomic analyses consistently detected multiple human proteins in unconditioned media, indicating that these signals are not of embryonic origin. Undeclared proteins were also identified in media sampled before their first use, with considerable variability observed across production batches. Supplementation experiments demonstrated that this variability originates from the addition of plasma-derived human serum albumin, which introduces a range of contaminating proteins. In contrast, recombinant albumin did not contribute detectable serum proteins. These findings indicate that the protein background in commercial IVF media is an overlooked variable, not only compromising the reproducibility and interpretation of secretome analyses in research, but also contributing to unstandardized culture conditions in clinical IVF practice. While this study evaluated 13 production lots of monophasic media from two different manufacturers, the results underscore a broader need for transparency and standardization in IVF media composition. The adoption of fully chemically defined IVF media, free from purified serum components, would support more consistent embryo culture conditions and advance the discovery of biomarkers in SCM.

cell biology↗

bioRxiv2023

The ultrastructural nature of human oocytes' cytoplasmatic abnormalities and the role of cytoskeleton dysfunction

BackgroundEgg quality is a limiting factor of female fertility and assisted reproductive technology (ART) success. Oocytes recovered from hyperstimulated ovaries often display morphological anomalies suspected to compromise their fertilization and developmental potential. Knowledge of (ab)normal oocytes intracellular organization is vital to establish reliable criteria for morphological evaluation of oocytes intended for in vitro fertilization (IVF). MethodsTransmission electron microscopy (TEM) was used to investigate the fine morphology of 22 dysmorphic IVF eggs exhibiting different types of cytoplasmic irregularities, namely (1) refractile bodies, (2) centrally-located cytoplasmic granularity (CLCG), (3) smooth endoplasmic reticulum (SER) disc, and (4) vacuoles. The cytoskeleton targeting compounds were employed to address the causative mechanism behind the anomalous cytoplasmic architecture observed in abnormal egg samples. A total of 133 immature oocytes were exposed to chemical inhibitors/control conditions, and their morphology was examined by fluorescent and electron microscopy. ResultsTEM exposed the structural basis of the common oocyte aberrations and revealed that the underlying cause of two of the studied morphotypes was excessive organelle clustering. Inhibition experiments showed that disruption of actin, not microtubules, allows inordinate aggregation of subcellular structures resembling the ultrastructural pattern seen in morphologically abnormal eggs retrieved in IVF cycles. These results imply that actin serves as a regulator of organelle distribution during human oocyte maturation. ConclusionsThe ultrastructural analogy between dysmorphic eggs and oocytes, in which actin network integrity was perturbed, suggests that malfunction of the actin cytoskeleton might be implicated in generating common cytoplasmic aberrations. Knowledge of human oocytes inner workings and the origin of morphological abnormalities is a step forward to more objective egg quality assessment in clinical practice.

cell biology↗

bioRxiv2021

High-resolution 3D reconstruction of human oocytes using FIB-SEM.

The oocyte plays a pivotal role in the reproduction of our species. Nevertheless, its biology remains poorly understood. Electron microscopy is traditionally used to inspect the ultrastructure of female gametes. However, two-dimensional micrographs contain only fragmentary information about the spatial organization of the complex oocyte cytoplasm. Here, we employed the Focused Ion Beam Scanning Electron Microscopy (FIB-SEM) to explore human oocyte intracellular morphology in three dimensions (3D). Volume reconstruction from high-resolution image stacks provided an unprecedented view of ooplasmic architecture. Organelle distribution patterns observed in 9 donor oocytes, representing 3 maturational stages, documented structural changes underlying the process by which the egg acquires developmental competence. 3D image segmentation was performed to extract information about distinct organelle populations. The quantitative analysis of the organelle abundance revealed that mitochondrion occupies ~ 4.26 % of the maturing oocyte cytoplasm. This proof-of-concept study demonstrates the potential of FIB-SEM imaging to study human oocyte morphology.

cell biology↗