Karyotype Aberrations in Action: The Evolution of Cancer Genomes and the Tumor Microenvironment

Cancer is a disease of cellular evolution. For this cellular evolution to take place, a population of cells must contain functional heterogeneity and an assessment of this heterogeneity in the form of natural selection. Cancer cells from advanced malignancies are genomically and functionally very different compared to the healthy cells from which they evolved. Genomic alterations include aneuploidy (numerical and structural changes in chromosome content) and polyploidy (e.g., whole genome doubling), which can have considerable effects on cell physiology and phenotype. Likewise, conditions in the tumor microenvironment are spatially heterogeneous and vastly different than in healthy tissues, resulting in a number of environmental niches that play important roles in driving the evolution of tumor cells. While a number of studies have documented abnormal conditions of the tumor microenvironment and the cellular consequences of aneuploidy and polyploidy, a thorough overview of the interplay between karyotypically abnormal cells and the tissue and tumor microenvironments is not available. Here, we examine the evidence for how this interaction may unfold during tumor evolution. We describe a bidirectional interplay in which aneuploid and polyploid cells alter and shape the microenvironment in which they and their progeny reside; in turn, this microenvironment modulates the rate of genesis for new karyotype aberrations and selects for cells that are most fit under a given condition. We conclude by discussing the importance of this interaction for tumor evolution and the possibility of leveraging our understanding of this interplay for cancer therapy

Marital mediation and a possibility of using a model of transformative mediation

Artykuł przedstawia możliwość zastosowania modelu mediacji transformatywnej z perspektywy przepisów dotyczących mediacji małżeńskiej, zawartych w polskim kodeksie postępowania cywilnego. Ze względu na traumatyczne skutki długotrwałego rozwodu w postępowaniu sądowym oraz niską skuteczność instytucji pojednania małżonków, szczególna uwaga została poświęcona ocenie, czy w obecnym stanie prawnym istnieje możliwość prowadzenia mediacji z elementami terapii małżeńskiej. Celem artykułu jest również zaprezentowanie koncepcji mediacji transformatywnej w świetle przepisów kodeksu postępowania cywilnego.The article describes the institution of marital mediation in Polish civil law. The mediation process is regulated in the Polish Code of Civil Procedure in articles 1831-15 and 436. Mediation is recommended in order to avoid disputes in a family and a lengthy lawsuit in divorce cases. Through mediation the spouses can solve a wide range of problems that are connected with their divorce or functioning of their family. A mediator is obliged to help them to define the issues and improve conflict resolutions. The regulations seem to be not effective because the spouses seldom choose mediation process. Particular focus is given to the results of transformative mediation and its advantages and disadvantages. The author discusses whether there is a possibility of using a model of transformative mediation in Polish civil procedure or not. The aim of the article is to convince the reader that there is a possibility of using transformative mediation in order to achieve a divorce settlement agreement in a peaceful way instead of bearing the mental and financial costs of a divorce suit in court

Micronuclei images and quantification.

<p>FSS exposure results in an increase in CREST-positive micronuclei. Micronuclei were observed in all cell populations and quantified on multiple slides using CREST (green) and DAPI (red) staining for control and adherent cell populations (after three, consecutive, 96 h exposures to FSS). (<b>A</b>) Representative images of cells with either a CREST-positive (CREST+, containing whole chromosomes, left) or a CREST-negative (CREST-, containing chromosome fragments, right) micronucleus (white arrows). (<b>B</b>) Average percentages of micronuclei ± SEM. Importantly, all stages of the disease exposed to FSS displayed a significant increase in CREST-positive (whole chromosome-containing) micronuclei. Asterisks denote statistical significance (Fisher’s Exact, * p< 0.05, ** p< 0.005, *** p< 0.001) for comparison of adherent cells exposed to FSS to corresponding controls.</p

Experimental design.

<p>Overview of the experimental design used in this study. Cells were seeded and either immediately placed on the rotator <b>(A)</b> or allowed to adhere for 4 h before being placed under FSS conditions <b>(B)</b>. Cells were re-seeded at their original density after each 96 hour period of exposure to FSS. Analysis was performed at three different time points corresponding to a total of 96 h, 192 h, or 288h FSS exposure, respectively.</p

Spheroid formation images and size quantification.

<p>FSS induces spheroid formation in tumorigenic ovarian cancer cells. (<b>3.1</b>) Images of differential spheroid formation, adherence and outgrowth of benign (MOSE-E, OCE1), tumorigenic (MOSE-L, SKOV-3), and highly aggressive (MOSE-L_TIC<i>v</i>) ovarian cancer cells at 96 h (A-I) and 288 h (time point 3, K-S) in response to FSS. All representative images were taken at the center of the plates. (<b>3.2</b>) the diameter of the formed spheroids were measured and averaged at each time point to monitor growth over time. Significant growth in spheroid diameter was measured in both FSS-exposed MOSE-L cells. In addition, MOSE-L_{TIC<i>v</i>-imm} cells formed large spheroids that grew over time. Note: the MOSE-L_{TIC<i>v</i>-adh} cells re-attached to the culture dishes with an adherent monolayer outgrowth too large to be measured, but the diameters after 288 h are at least 700μm. Asterisks denote statistical significance (t-test, * p< 0.05, ** p< 0.005, and *** p< 0.001).</p

Actin cytoskeleton and focal adhesion organization.

<p>Actin cytoskeleton organization and focal adhesion number and length change significantly in response to FSS exposure. (<b>A</b>) Changes in actin (green) organization in adherent MOSE-E, COE1, MOSE-L, SKOV-3, and MOSE-L_TIC<i>v</i> cells after three, consecutive 96 h exposures to FSS. (<b>B</b>) FSS effects on vinculin-positive focal adhesions. Nuclei are shown in blue. (<b>C</b>) Quantitation of focal adhesion number and size in controls and after FSS exposure. Asterisks denote statistical significance (t-test, * p< 0.05 and *** p< 0.001).</p

Metaphase images and quantification.

<p>FSS dramatically increases the fraction of MOSE-E cells with near-tetraploid chromosome numbers. (<b>A</b>) Examples of metaphase spreads with near-diploid (left) and near-tetraploid (right) range. (<b>B</b>) Chromosome counts in individual metaphase spreads from the control (black) and FSS-exposed adherent (gray) MOSE-E cells. Upon exposure to FSS, the fraction of near tetraploid cells increased to 76% ± 2.4% compared to 31% ± 5.0% in the control population (<i>χ</i>², p = 0.013).</p

Cell survival.

<p>FSS differentially affects cell viability in ovarian cancer cells of different disease stage. Time-dependent changes in average cell number (x10⁶) ± SEM of MOSE-E (<b>A</b>), OCE1 (<b>B</b>), MOSE-L (<b>C</b>), SKOV-3 (<b>D</b>), and MOSE-L_TIC<i>v</i> (<b>E</b>) cells subjected to FSS of for 96 h periods (time points 1-3) are shown. In each graph, the red line indicates the initial seeding number (2x10⁵). Asterisks denote statistical significance (ANOVA, Tukey’s * p< 0.05, ** p< 0.005, and *** p< 0.001) as compared to the corresponding control group at the same time point.</p