ZIKA VIRUS ENVELOPE PROTEIN INDUCES MIRNA-MEDIATED ENDOTHELIAL DYSFUNCTION IN HUMAN BRAIN MICROVASCULAR ENDOTHELIAL CELLS

Zika virus (ZIKV) is a neurotropic flavivirus known to cause congenital neurological abnormalities such as microcephaly and encephalitis in adults. The structural envelope (E) protein facilitates the viral entry and is crucial for neuroinvasion. Studies have reported ZIKV crosses the blood–brain barrier (BBB), a highly specialized neurovascular interface which normally restricts the viral entry. However, the molecular mechanisms underlying ZIKV-mediated BBB disruption are poorly understood. In our previous study, we demonstrated ZIKV E protein alters BBB integrity by activating human brain microvascular endothelial cells (hBMECs) and astrocytes, critical components of BBB. We observed overexpression of E protein led to dysregulation of endothelial junction proteins (ZO-1, VE-Cadherin and Occludin), and exhibited inflammatory responses in hBMECs and also resulted in astrogliosis. In our present study, we explored the molecular regulators behind altered hBMECs responses using small RNA sequencing to understand its post-transcriptional regulation. Among differentially expressed (DE) miRNAs, we identified 39 DE miRNAs (35 upregulated, 4 downregulated), including 33 novel miRNAs which have not been previously reported. 5 miRNAs with predicted roles in BBB integrity were validated by RT-PCR. Functional enrichment analyses (GO and KEGG) revealed their association with pathways regulating cell adhesion, junction assembly, vesicle transport, and Wnt/β-catenin signaling. miR-1249-5p was predicted and experimentally validated to target Wnt2b, an important regulator for BBB maintenance. Mimic transfection of miR-1249-5p significantly reduced Wnt2b expression, recapitulating the effects observed in E-transfected hBMECs. Overall, these findings identify novel endothelial miRNA dysregulation in response to ZIKV E protein, providing further insights into ZIKV neuropathogenesis.