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Ewing sarcoma is a bone tumor most commonly diagnosed in adolescents and young adults. Survival for patients with recurrent or metastatic Ewing sarcoma is dismal and there is a dire need to better understand the mechanisms of cell metastasis specific to this disease. Our recent work demonstrated that microenvironmental stress leads to increased Ewing sarcoma cell invasion through Src activation. Additionally, we have shown that the matricellular protein tenascin C (TNC) promotes metastasis in Ewing sarcoma. A major role of both TNC and Src is mediation of cell–cell and cell-matrix interactions resulting in changes in cell motility, invasion, and adhesion. However, it remains largely unknown, if and how, TNC and Src are linked in these processes. We hypothesized that TNC is a positive regulator of invadopodia formation in Ewing sarcoma through its ability to activate Src. We demonstrate here that both tumor cell endogenous and exogenous TNC can enhance Src activation and invadopodia formation in Ewing sarcoma. We found that microenvironmental stress upregulates TNC expression and this is dampened with application of the Src inhibitor dasatinib, suggesting that TNC expression and Src activation cooperate to promote the invasive phenotype. This work reports the impact of stress-induced TNC expression on enhancing cell invadopodia formation, provides evidence for a feed forward loop between TNC and Src to promote cell metastatic behavior, and highlights a pathway by which microenvironment-driven TNC expression could be therapeutically targeted in Ewing sarcoma.

Visual Abstract, Highlights • PKA-phosphorylation of Hsp20 is elevated in human failing hearts. • Increases in phosphorylated Hsp20 in vivo are associated with fibrotic remodeling and reduced left ventricular function. • The phosphorylated Hsp20 in cardiomyocyte promotes upregulation of IL-6 and its subsequent paracrine actions on the cardiac fibroblast. • Blockade of IL-6 effects ex vivo and in vivo reduces the pro-fibrotic effects of phosphorylated Hsp20. • Targeting phosphorylated Hsp20 in the cardiomyocyte may represent a potential therapeutic strategy to mitigate fibrotic remodeling and preserve function in the failing heart., Summary Cardiomyocyte-specific increases in phosphorylated Hsp20 (S16D-Hsp20) to levels similar to those observed in human failing hearts are associated with early fibrotic remodeling and depressed left ventricular function, symptoms which progress to heart failure and early death. The underlying mechanisms appear to involve translocation of phosphorylated Hsp20 to the nucleus and upregulation of interleukin (IL)-6, which subsequently activates cardiac fibroblasts in a paracrine fashion through transcription factor STAT3 signaling. Accordingly, treatment of S16D-Hsp20 mice with a rat anti-mouse IL-6 receptor monoclonal antibody (MR16-1) attenuated interstitial fibrosis and preserved cardiac function. These findings suggest that phosphorylated Hsp20 may be a potential therapeutic target in heart failure.

Epithelial ovarian carcinoma spreads via shedding of cells and multicellular aggregates (MCAs) from the primary tumor into peritoneal cavity, with subsequent intraperitoneal tumor cell:mesothelial cell adhesion as a key early event in metastatic seeding. Evaluation of human tumor extracts and tissues confirms that well-differentiated ovarian tumors express abundant E-cadherin (Ecad), whereas advanced lesions exhibit upregulated N-cadherin (Ncad). Two expression patterns are observed: "mixed cadherin," in which distinct cells within the same tumor express either E- or Ncad, and "hybrid cadherin," wherein single tumor cell(s) simultaneously expresses both cadherins. We demonstrate striking cadherin-dependent differences in cell-cell interactions, MCA formation, and aggregate ultrastructure. Mesenchymal-type Ncad+ cells formed stable, highly cohesive solid spheroids, whereas Ecad+ epithelial-type cells generated loosely adhesive cell clusters covered by uniform microvilli. Generation of "mixed cadherin" MCAs using fluorescently tagged cell populations revealed preferential sorting into cadherin-dependent clusters, whereas mixing of cell lines with common cadherin profiles generated homogeneous aggregates. Recapitulation of the "hybrid cadherin" Ecad+/Ncad+ phenotype, via insertion of the CDH2 gene into Ecad+ cells, resulted in the ability to form heterogeneous clusters with Ncad+ cells, significantly enhanced adhesion to organotypic mesomimetic cultures and peritoneal explants, and increased both migration and matrix invasion. Alternatively, insertion of CDH1 gene into Ncad+ cells greatly reduced cell-to-collagen, cell-to-mesothelium, and cell-to-peritoneum adhesion. Acquisition of the hybrid cadherin phenotype resulted in altered MCA surface morphology with increased surface projections and increased cell proliferation. Overall, these findings support the hypothesis that MCA cadherin composition impacts intraperitoneal cell and MCA dynamics and thereby affects ultimate metastatic success.