7.3. O14—A Screening Methodology of the Cell Nuclei Based on Functional Status of the Chromatin in Ziphius cavirostris

Utilizing methods capable of detecting changes in chromatin states as a means to screen cells based on their functional state offers a valuable approach for assessing the effects of pollutants such as PFAS on cetacean species’ cells and evaluating the status of environmental living conditions. In this study, a myogenic cell line obtained from Ziphius cavirostris was examined using a computed set of morphometric parameters to analyze cell nuclear morphology. Additionally, cytogenetic analyses were performed to determine the diploid number, distribution of constitutive heterochromatin (HC), and telomeric regions. Cells were stained with Hoechst 33342 to measure nuclear morphometric parameters through high-throughput screening analysis, combining various parameters such as nuclear length, inverse aspect ratio (Inv/AR), and nuclear intensity. This methodology was employed to assess the cytotoxic effects induced by PFAS in cetacean cells. Cytogenetic techniques, including CBA-banding and FISH-mapping with PNA-telomeric probes, were applied to metaphase chromosomes obtained from myogenic cells. Our model facilitated the grouping and quantification of nuclei into three populations and six groups: the normal population (nuclei in G0 phase, synthesis phase, and mitosis phase), the large population (senescent nuclei), and the small population (nuclei fragmentation). By plotting frequency distributions of Hoechst intensity, we generated cell cycle profiles of nuclei. The combination of cell nuclei counting with cell cycle analysis allowed us to determine the dose-response effect of PFAS. Cytogenetic results confirmed that Ziphius c. has 2n = 42, although several cells exhibited polyploidy, predominantly tetraploidy. CBA-banding revealed large blocks of HC in various chromosomes and chromosome arms. All chromosomes exhibited positive FITC signals in all telomeric regions. This method provides an objective, semi-quantitative tool for screening different nuclear phenotypes depending on the functional status of the chromatin. Cytogenetic analyses revealed characteristic CBA-banding patterns and telomeric regions.