Blockade of Autocrine CCL5 Responses Inhibits Zika Virus Persistence and Spread in Human Brain Microvascular Endothelial Cells.

Zika virus (ZIKV) is a neurovirulent flavivirus that uniquely causes fetal microcephaly, is sexually transmitted, and persists in patients for up to 6 months. ZIKV persistently infects human brain microvascular endothelial cells (hBMECs) that form the blood-brain barrier (BBB) and enables viral spread to neuronal compartments. We found that CCL5, a chemokine with prosurvival effects on immune cells, was highly secreted by ZIKV-infected hBMECs. Although roles for CCL5 in endothelial cell (EC) survival remain unknown, the presence of the CCL5 receptors CCR3 and CCR5 on ECs suggested that CCL5 could promote ZIKV persistence in hBMECs. We found that exogenous CCL5 induced extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation in hBMECs and that ERK1/2 cell survival signaling was similarly activated by ZIKV infection. Neutralizing antibodies to CCL5, CCR3, or CCR5 inhibited persistent ZIKV infection of hBMECs. While knockout (KO) of CCL5 failed to prevent ZIKV infection of hBMECs, at 3 days postinfection (dpi), we observed a >90% reduction in ZIKV-infected CCL5-KO hBMECs and a multilog reduction in ZIKV titers. In contrast, the addition of CCL5 to CCL5-KO hBMECs dose-dependently rescued ZIKV persistence in hBMECs. Inhibiting CCL5 responses using CCR3 (UCB35625) and CCR5 (maraviroc) receptor antagonists reduced the number of ZIKV-infected hBMECs and ZIKV titers (50% inhibitory concentrations [IC50s] of 2.5 to 12 µM), without cytotoxicity (50% cytotoxic concentration [CC50] of >80 µM). These findings demonstrate that ZIKV-induced CCL5 directs autocrine CCR3/CCR5 activation of ERK1/2 survival responses that are required for ZIKV to persistently infect hBMECs. Our results establish roles for CCL5 in ZIKV persistence and suggest the potential for CCL5 receptor antagonists to therapeutically inhibit ZIKV spread and neurovirulence. IMPORTANCE Our findings demonstrate that CCL5 is required for ZIKV to persistently infect human brain ECs that normally protect neuronal compartments. We demonstrate that ZIKV-elicited CCL5 secretion directs autocrine hBMEC activation of ERK1/2 survival pathways via CCR3/CCR5, and inhibiting CCL5/CCR3/CCR5 responses prevented ZIKV persistence and spread. Our findings demonstrate that ZIKV-directed CCL5 secretion promotes hBMEC survival and reveals an underlying mechanism of ZIKV pathogenesis and spread. We demonstrate that antagonists of CCR3/CCR5 inhibit ZIKV persistence in hBMECs and provide potential therapeutic approaches for preventing ZIKV persistence, spread, and neurovirulence.