In Vivo Evolution of Tumor-Derived Endothelial Cells

The growth of a malignant tumor beyond a certain, limited size requires that it first develop an independent blood supply. In addition to providing metabolic support, this neovasculature also allows tumor cells to access the systemic circulation, thus facilitating metastatic dissemination. The neovasculature may originate either from normal blood vessels in close physical proximity to the tumor and/or from the recruitment of bone marrow-derived endothelial cell (EC) precursors. Recent studies have shown that human tumor vasculature ECs may also arise directly from tumor cells themselves and that the two populations have highly similar or identical karyotypes. We now show that, during the course of serial in vivo passage, these tumor-derived ECs (TDECs) progressively acquire more pronounced EC-like properties. These include higher-level expression of EC-specific genes and proteins, a greater capacity for EC-like behavior in vitro, and a markedly enhanced propensity to incorporate into the tumor vasculature. In addition, both vessel density and size are significantly increased in neoplasms derived from mixtures of tumor cells and serially passaged TDECs. A comparison of early- and late-passage TDECs using whole-genome single nucleotide polymorphism profiling showed the latter cells to have apparently evolved by a process of clonal expansion of a population with a distinct pattern of interstitial chromosomal gains and losses affecting a relatively small number of genes. The majority of these have established roles in vascular development, tumor suppression or epithelial-mesenchymal transition. These studies provide direct evidence that TDECs have a strong evolutionary capacity as a result of their inherent genomic instability. Consequently such cells might be capable of escaping anti-angiogenic cancer therapies by generating resistant populations

Gene expression profiling identifies inflammation and angiogenesis as distinguishing features of canine hemangiosarcoma

<p>Abstract</p> <p>Background</p> <p>The etiology of hemangiosarcoma remains incompletely understood. Its common occurrence in dogs suggests predisposing factors favor its development in this species. These factors could represent a constellation of heritable characteristics that promote transformation events and/or facilitate the establishment of a microenvironment that is conducive for survival of malignant blood vessel-forming cells. The hypothesis for this study was that characteristic molecular features distinguish hemangiosarcoma from non-malignant endothelial cells, and that such features are informative for the etiology of this disease.</p> <p>Methods</p> <p>We first investigated mutations of VHL and Ras family genes that might drive hemangiosarcoma by sequencing tumor DNA and mRNA (cDNA). Protein expression was examined using immunostaining. Next, we evaluated genome-wide gene expression profiling using the Affymetrix Canine 2.0 platform as a global approach to test the hypothesis. Data were evaluated using routine bioinformatics and validation was done using quantitative real time RT-PCR.</p> <p>Results</p> <p>Each of 10 tumor and four non-tumor samples analyzed had wild type sequences for these genes. At the genome wide level, hemangiosarcoma cells clustered separately from non-malignant endothelial cells based on a robust signature that included genes involved in inflammation, angiogenesis, adhesion, invasion, metabolism, cell cycle, signaling, and patterning. This signature did not simply reflect a cancer-associated angiogenic phenotype, as it also distinguished hemangiosarcoma from non-endothelial, moderately to highly angiogenic bone marrow-derived tumors (lymphoma, leukemia, osteosarcoma).</p> <p>Conclusions</p> <p>The data show that inflammation and angiogenesis are important processes in the pathogenesis of vascular tumors, but a definitive ontogeny of the cells that give rise to these tumors remains to be established. The data do not yet distinguish whether functional or ontogenetic plasticity creates this phenotype, although they suggest that cells which give rise to hemangiosarcoma modulate their microenvironment to promote tumor growth and survival. We propose that the frequent occurrence of canine hemangiosarcoma in defined dog breeds, as well as its similarity to homologous tumors in humans, offers unique models to solve the dilemma of stem cell plasticity and whether angiogenic endothelial cells and hematopoietic cells originate from a single cell or from distinct progenitor cells.</p

Deepâ intronic variants in CNGB3 cause achromatopsia by pseudoexon activation

Our comprehensive cohort of 1100 unrelated achromatopsia (ACHM) patients comprises a considerable number of cases (~5%) harboring only a single pathogenic variant in the major ACHM gene CNGB3. We sequenced the entire CNGB3 locus in 33 of these patients to find a second variant which eventually explained the patientsâ phenotype. Fortyâ seven intronic CNGB3 variants were identified in 28 subjects after a filtering step based on frequency and the exclusion of variants found in cis with pathogenic alleles. In a second step, in silico prediction tools were used to filter out those variants with little odds of being deleterious. This left three variants that were analyzed using heterologous splicing assays. Variant c.1663â 1205G>A, found in 14 subjects, and variant c.1663â 2137C>T, found in two subjects, were indeed shown to exert a splicing defect by causing pseudoexon insertion into the transcript. Subsequent screening of further unsolved CNGB3 subjects identified four additional cases harboring the c.1663â 1205G>A variant which makes it the eighth most frequent CNGB3 variant in our cohort. Compound heterozygosity could be validated in ten cases. Our study demonstrates that whole gene sequencing can be a powerful approach to identify the second pathogenic allele in patients apparently harboring only one diseaseâ causing variant.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/152731/1/humu23920_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/152731/2/humu23920.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/152731/3/humu23920-sup-0001-Revised_supplementary_Material_with_track_changes_accepted.pd

Scapula fractures: interobserver reliability of classification and treatment

OBJECTIVES:There is substantial variation in the classification and the management of scapula fractures. The first purpose of this study was to analyze the interobserver reliability of the OTA/AO and the New International Classification of scapula fractures. The second purpose was to assess the proportion of agreement among orthopaedic surgeons on operative or nonoperative treatment. DESIGN:: Web-based reliability study SETTING:: Independent orthopaedic surgeons from several countries were invited to classify scapular fractures in an online survey. PARTICIPANTS:One-hundred and three orthopaedic surgeons evaluated 35 movies of 3DCT-reconstruction of selected scapular fractures, representing a full spectrum of fracture patterns. MAIN OUTCOME MEASUREMENTS:Fleiss' kappa (κ) was used to assess the reliability of agreement between the surgeons. RESULTS:: The overall agreement on the OTA/AO Classification was moderate for the types (A, B, and C, κ = 0.54) with a 71% proportion of rater agreement (PA) as well as for the nine groups (A1 to C3, κ = 0.47) with a 57% PA. For the New International Classification, the agreement about the intra-articular extension of the fracture (Fossa (F), κ = 0.79) was substantial, the agreement about a fractured body (Body (B), κ = 0.57) or process was moderate (Process (P), κ = 0.53), however PAs were more than 81%. The agreement on the treatment recommendation was moderate (κ = 0.57) with a 73% PA. CONCLUSIONS:The New International Classification was more reliable. Body and process fractures generated more disagreement than intra-articular fractures and need further clear definitions