Tetrastigma hemsleyanum ELN-derived miR-396b suppresses pancreatic cancer cell proliferation by targeting IL33

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive malignancies and remains largely refractory to current therapeutic strategies. Plant-derived exosome-like nanovesicles (ELNs) have recently emerged as promising natural nanocarriers capable of delivering bioactive molecules across kingdoms; however, their functional cargos and molecular mechanisms in PDAC remain poorly understood.

ELNs were isolated from Tetrastigma hemsleyanum and characterized by nanoparticle tracking analysis and zeta potential measurement. Cellular uptake was evaluated using PKH67 fluorescence labeling. Small RNA sequencing and RT-qPCR were performed to identify abundant ELN-associated miRNAs. The biological effects of ELNs and miR-396b were assessed by CCK-8 and colony formation assays. Transcriptomic profiling was conducted to identify downstream targets, and IL33 was validated by RT-qPCR, immunoblotting, bioinformatic prediction, and dual-luciferase reporter assays. Rescue experiments were further performed to investigate the functional involvement of the miR-396b/IL33 axis.

T. hemsleyanum-derived ELNs were efficiently internalized by pancreatic cancer cells and inhibited cell proliferation in a dose-dependent manner. Small RNA sequencing identified miR-396b as a highly enriched ELN cargo. Both ELN treatment and miR-396b overexpression significantly reduced IL33 expression. Mechanistically, miR-396b directly bound the IL33 3' untranslated region and suppressed reporter activity, whereas mutation of the predicted binding site abolished this inhibitory effect. Functional rescue experiments demonstrated that inhibition of miR-396b attenuated ELN-mediated growth suppression, while IL33 re-expression partially reversed the anti-proliferative effects induced by miR-396b overexpression, indicating that the miR-396b/IL33 axis contributes substantially to the biological activity of ELNs.

This study identifies miR-396b as a major functional cargo of T. hemsleyanum-derived ELNs and demonstrates that ELN-delivered miR-396b suppresses pancreatic cancer cell proliferation, at least in part, through direct repression of IL33. These findings provide mechanistic insight into a cross-kingdom ELN-miRNA-IL33 regulatory pathway and support the potential of plant-derived ELNs as a bioactive nanoplatform for PDAC intervention. Nevertheless, additional ELN cargos and in vivo validation warrant further investigation.