Endothelial lncRNA-HOXA responses to combined chemical hypoxia and shear stress.

Endothelial cells respond to changes in oxygen availability and mechanical forces by adjusting their transcriptional and secretory profiles, thereby shaping vascular and tissue-level adaptation. In this study, primary human endothelial monolayers were exposed to a chemical hypoxia mimic under static or shear-stress conditions, using a viability-guided experimental design to define non-cytotoxic exposure parameters. Under these conditions, hypoxia-associated responses were observed together with coordinated changes in long non-coding RNAs linked to the HOX genomic region, including HOTAIR and HOTTIP, and in selected members of the HOXA gene cluster. Transcripts related to Notch and BMP signaling showed increased expression, whereas components of the canonical Wnt pathway displayed reduced or stable expression, consistent with a context-dependent transcriptional profile rather than a generalized activation of proliferative programs. Endothelial migration was also altered in scratch-wound assays. Extracellular vesicles released under static and shear-stress conditions exhibited physical characteristics consistent with small vesicle populations, and their RNA cargo reflected transcriptional changes associated with hypoxia and mechanical exposure. Together, these findings provide a descriptive cellular and vesicular framework for how endothelial cells integrate chemical hypoxia and shear stress at the transcriptional and intercellular communication levels. Thus, this work establishes a basis for future functional studies aimed at testing the causal roles of long non-coding RNAs and vesicle-mediated signaling in vascular-stromal interactions within hypoxic and mechanically dynamic tissue microenvironments.