A Zebrafish Model of Roberts Syndrome Reveals That Esco2 Depletion Interferes with Development by Disrupting the Cell Cycle

The human developmental diseases Cornelia de Lange Syndrome (CdLS) and Roberts Syndrome (RBS) are both caused by mutations in proteins responsible for sister chromatid cohesion. Cohesion is mediated by a multi-subunit complex called cohesin, which is loaded onto chromosomes by NIPBL. Once on chromosomes, cohesin binding is stabilized in S phase upon acetylation by ESCO2. CdLS is caused by heterozygous mutations in NIPBL or cohesin subunits SMC1A and SMC3, and RBS is caused by homozygous mutations in ESCO2. The genetic cause of both CdLS and RBS reside within the chromosome cohesion apparatus, and therefore they are collectively known as “cohesinopathies”. However, the two syndromes have distinct phenotypes, with differences not explained by their shared ontology. In this study, we have used the zebrafish model to distinguish between developmental pathways downstream of cohesin itself, or its acetylase ESCO2. Esco2 depleted zebrafish embryos exhibit features that resemble RBS, including mitotic defects, craniofacial abnormalities and limb truncations. A microarray analysis of Esco2-depleted embryos revealed that different subsets of genes are regulated downstream of Esco2 when compared with cohesin subunit Rad21. Genes downstream of Rad21 showed significant enrichment for transcriptional regulators, while Esco2-regulated genes were more likely to be involved the cell cycle or apoptosis. RNA in situ hybridization showed that runx1, which is spatiotemporally regulated by cohesin, is expressed normally in Esco2-depleted embryos. Furthermore, myca, which is downregulated in rad21 mutants, is upregulated in Esco2-depleted embryos. High levels of cell death contributed to the morphology of Esco2-depleted embryos without affecting specific developmental pathways. We propose that cell proliferation defects and apoptosis could be the primary cause of the features of RBS. Our results show that mutations in different elements of the cohesion apparatus have distinct developmental outcomes, and provide insight into why CdLS and RBS are distinct diseases

Modelling and simulation of [18F]fluoromisonidazole dynamics based on histology-derived microvessel maps

Contains fulltext : 97120.pdf (publisher's version ) (Closed access)Hypoxia can be assessed non-invasively by positron emission tomography (PET) using radiotracers such as [(18)F]fluoromisonidazole (Fmiso) accumulating in poorly oxygenated cells. Typical features of dynamic Fmiso PET data are high signal variability in the first hour after tracer administration and slow formation of a consistent contrast. The purpose of this study is to investigate whether these characteristics can be explained by the current conception of the underlying microscopic processes and to identify fundamental effects. This is achieved by modelling and simulating tissue oxygenation and tracer dynamics on the microscopic scale. In simulations, vessel structures on histology-derived maps act as sources and sinks for oxygen as well as tracer molecules. Molecular distributions in the extravascular space are determined by reaction-diffusion equations, which are solved numerically using a two-dimensional finite element method. Simulated Fmiso time activity curves (TACs), though not directly comparable to PET TACs, reproduce major characteristics of clinical curves, indicating that the microscopic model and the parameter values are adequate. Evidence for dependence of the early PET signal on the vascular fraction is found. Further, possible effects leading to late contrast formation and potential implications on the quantification of Fmiso PET data are discussed

Modelling and simulation of the influence of acute and chronic hypoxia on [18F]fluoromisonidazole PET imaging.

Item does not contain fulltextTumour hypoxia can be assessed by positron emission tomography (PET) using radiotracers like [(18)F]fluoromisonidazole (Fmiso). The purpose of this work was to independently investigate the influence of chronic and acute hypoxia on the retention of Fmiso on the microscale. This was approached by modelling and simulating tissue oxygenation and Fmiso dynamics on the microscale based on tumour histology. Diffusion of oxygen and Fmiso molecules in tissue- and oxygen-dependent Fmiso binding were included in the model. Moreover, a model of fluctuating vascular oxygen tension was incorporated to theoretically predict the effects of acute hypoxia. Simulated tissue oxygen tensions (PO(2)) are strongly influenced by the modelled periodical fluctuations (period 40 min, total amplitude 10 mmHg and mean 35 mmHg). Fluctuations led to variations in mean PO(2) of up to 41% and in the hypoxic fraction (PO(2) < 5 mmHg) from 56% up to 65%. Significant Fmiso retention is caused by chronic (87%) as well as acute hypoxia (13%). By simulating Fmiso injection during different phases of the vascular PO(2) fluctuation cycle, it was found that acute hypoxia of an empirically valid magnitude does not influence the reproducibility of PET imaging. Thus, it may be impossible to separate acute and chronic hypoxia from serial PET images

Correlation between tumor oxygenation and (18)F-fluoromisonidazole PET data simulated based on microvessel images

Item does not contain fulltextAbstract Background. Assessing hypoxia with oxygen probes provides a sparse sampling of tumor volumes only, bearing a risk of missing hypoxic regions. Full coverage is achieved with positron emission tomography (PET) using the tracer (18)F-fluoromisonidazole (FMISO). In this study, the correlation between different FMISO PET imaging parameters and the median voxel PO2 (medianPO2) was analyzed. A measure of the median PO2 characterizes the microenvironment in consistency with probe measurements. Material and methods. Tissue oxygenations and FMISO diffusion-retention dynamics were simulated. Transport of FMISO and O2 molecules into and out of tissue was modeled by vessel maps derived from histology of head-and-neck squamous cell cancer xenograft tumor lines. Parameter sets were evaluated for 300 distinct 2 x 2 mm(2) vessel configurations, including medianPO2 and two FMISO PET parameters: FH denotes the sub-regional signal four hours post injection (pi) and FH/P denotes the ratio between FH and the time-averaged signal 0-15 min pi. Correlations between O2 and FMISO parameters were evaluated. A receiver operating characteristics (ROC) analysis was performed, regarding the accuracy of FH and FH/P in identifying voxels with medianPO2 < 2.5 mmHg. Results. In hypoxic sub-regions, the correlation between FH and medianPO2 is low (R(2) = 0.37), while the correlation between FH/P and median PO2 is high (R(2) = 0.99). The ROC analysis showed that hypoxic regions can be identified using FH/P with a higher diagnostic accuracy (YI = sensitivity+ specificity-1 = 1.0), than using FH alone (YI = 0.83). Both FMISO parameters are moderately effective in identifying hypoxia on the microscopic length scale (YI = 0.63 and 0.60). Conclusions. A combination of two FMISO PET scans acquired 0-15 min and four hours pi may yield an accurate measure of the medianPO2 in a voxel (FH/P). This measure is comparable to averaged oxygen probe measurements and has the advantage of covering the entire tumor volume. Therefore, it may improve the prediction of radiotherapy outcome and facilitate individualized dose prescriptions