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BACKGROUND AND PURPOSEDehydroepiandrosterone (DHEA) is thought to be an anti-glucocorticoid hormone known to be fully functional in young people but deficient in aged humans. Our previous data suggest that DHEA not only counteracts the effect of cortisol on RACK1 expression, a protein required both for the correct functioning of immune cells and for PKC-dependent pathway activation, but also modulates the inhibitory effect of cortisol on LPS-induced cytokine production. The purpose of this study was to investigate the effect of DHEA on the splicing mechanism of the human glucocorticoid receptor (GR). EXPERIMENTAL APPROACHThe THP1 monocytic cell line was used as a cellular model. Cytokine production was measured by specific ELISA. Western blot and real-time RT-PCR were used, where appropriate, to determine the effect of DHEA on GRs, serine/arginine-rich proteins (SRp), and RACK1 protein and mRNA. Small-interfering RNA was used to down-regulate GRβ. KEY RESULTSDHEA induced a dose-related up-regulation of GRβ and GRβ knockdown completely prevented DHEA-induced RACK1 expression and modulation of cytokine release. Moreover, we showed that DHEA influenced the expression of some components of the SRps found within the spliceosome, the main regulators of the alternative splicing of the GR gene. CONCLUSIONS AND IMPLICATIONSThese data contribute to our understanding of the mechanism of action of DHEA and its effect on the immune system and as an anti-glucocorticoid agent. AbbreviationsDHEA, dehydroepiandrosterone; GNB2L1, guanine nucleotide-binding protein β-2-like 1; GR, glucocorticoid receptor; GRE, glucocorticoid-responsive element; SR protein, serine/arginine-rich protein; RACK1, receptor for activated C kinase 1

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Although recent regulatory approval of splice-switching oligonucleotides (SSOs) for the treatment of neuromuscular disease such as Duchenne muscular dystrophy has been an advance for the splice-switching field, current SSO chemistries have shown limited clinical benefit due to poor pharmacology. To overcome limitations of existing technologies, we engineered chimeric stereopure oligonucleotides with phosphorothioate (PS) and phosphoryl guanidine-containing (PN) backbones. We demonstrate that these chimeric stereopure oligonucleotides have markedly improved pharmacology and efficacy compared with PS-modified oligonucleotides, preventing premature death and improving median survival from 49 days to at least 280 days in a dystrophic mouse model with an aggressive phenotype. These data demonstrate that chemical optimization alone can profoundly impact oligonucleotide pharmacology and highlight the potential for continued innovation around the oligonucleotide backbone. More specifically, we conclude that chimeric stereopure oligonucleotides are a promising splice-switching modality with potential for the treatment of neuromuscular and other genetic diseases impacting difficult to reach tissues such as the skeletal muscle and heart.

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Porcine extracellular matrix (pECM)-derived hydrogels were introduced, in recent years, aiming to benefit the pECM’s microstructure and bioactivity, while controlling the biomaterial’s physical and mechanical properties. The use of pECM from different tissues, however, offers tissue-specific features that can better serve different applications. In this study, pECM hydrogels derived from cardiac, artery, pancreas, and adipose tissues were compared in terms of composition, structure, and mechanical properties. While major similarities were demonstrated between all the pECM hydrogels, their distinctive attributes were also identified, and their substantial effects on cell-ECM interactions were revealed. Furthermore, through comprehensive protein and gene expression analyses, we show, for the first time, that each pECM hydrogel supports the spontaneous differentiation of induced pluripotent stem cells towards the resident cells of its origin tissue. These findings imply that the origin of ECM should be carefully considered when designing a biomedical platform, to achieve a maximal bioactive impact.

COMMUNICATION (1 of 8)Nanoghosts (NGs) is a proprietary novel form of a targeted delivery platform, which is based on nanovesicles that are technologically reconstructed from the whole cytoplasmic membranes of human bone marrow-derived mesenchymal stem cells (MSCs). Initially developed for tumor-targeted drug Nanoghosts (NGs) are nanovesicles reconstructed from the cytoplasmic membranes of mesenchymal stem cells (MSCs). By retaining MSC membranes, the NGs retain the ability of these cells to home in on multiple tumors, laying the foundations, thereby, for the development of a targeted drug delivery platform. The susceptibility of MSCs to functional changes, following their exposure to cytokines or cancer-derived conditioned-media (CM), presents the opportunity to modify the NGs by conditioning their source cells. This opportunity is investigated by comparing the membrane protein composition and the tumor uptake of NGs derived from naïve MSCs (N-NG) against conditioned NGs made from MSCs pre-treated with conditionedmedia (CM-NG) or with a mix of the proinflammatory cytokines TNF-α and IL-1β (Cyto-NG). CM-NGs are found to be more targeted towards immune cells than Cyto-or N-NGs, while Cyto-NGs are the most tumor-targeted ones, with similar immune-targeting capacity as N-NGs but with a higher affinity towards endothelial cells. Proteomic variations were wider in the CM-NGs, with exceptionally higher levels of ICAM-1 compared to N-and Cyto-NGs. From a translational point of view, the data show that the tumor-targeting ability of the NGs, and possibly that of other MSC-derived extracellular vesicles, can be enhanced by simple conditioning of their source cells.