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Zika Virus Persistence and Spread in Human Brain Microvascular Endothelial Cells Is Directed by Autocrine CCL5.

Record Type:	Electronic resources : Monograph/item
Title/Author:	Zika Virus Persistence and Spread in Human Brain Microvascular Endothelial Cells Is Directed by Autocrine CCL5./
Author:	Mladinich, Megan Cecilia.
Published:	Ann Arbor : ProQuest Dissertations & Theses, : 2021,
Description:	183 p.
Notes:	Source: Dissertations Abstracts International, Volume: 83-04, Section: B.
Contained By:	Dissertations Abstracts International83-04B.
Subject:	Virology. -
Online resource:	http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=28716490
ISBN:	9798460477630

Zika Virus Persistence and Spread in Human Brain Microvascular Endothelial Cells Is Directed by Autocrine CCL5.
Mladinich, Megan Cecilia.

Zika Virus Persistence and Spread in Human Brain Microvascular Endothelial Cells Is Directed by Autocrine CCL5. - Ann Arbor : ProQuest Dissertations & Theses, 2021 - 183 p.

Source: Dissertations Abstracts International, Volume: 83-04, Section: B.

Thesis (Ph.D.)--State University of New York at Stony Brook, 2021.

This item must not be sold to any third party vendors.

Zika virus (ZIKV) is a mosquito-borne flavivirus (FV) associated with encephalitis, Guillan-Barre syndrome and outbreaks of in utero fetal microcephaly. ZIKV transverses the blood-brain-barrier (BBB) and damages the central nervous system (CNS) by lytically infecting neurons, neural progenitors and astrocytes. In contrast to other FVs, ZIKV is uniquely detected in bodily fluids for up to 6 months, crosses placental barriers and is sexually transmitted. How ZIKV persists in patients after an acute febrile illness and the cells that support persistent ZIKV replication remains an enigma key to clearing ZIKV from patients and preventing ZIKV spread. The endothelial cell (EC) lining of capillaries normally constrains transplacental transmission and forms the BBB, which selectively restricts access of blood constituents to neurons. We found that ZIKV (strain PRVABC59) persistently infects and continuously replicates in primary human brain microvascular ECs (hBMECs) that form the BBB, without cytopathology, for >9 days and following serial hBMEC passages. ZIKV did not permeabilize hBMEC monolayers but was released apically and basolaterally from polarized hBMECs, suggesting basolateral release as a direct mechanism for ZIKV to cross the BBB.Innate and adaptive immune responses normally restrict acute FV replication and spread. ZIKV-infected hBMECs were rapidly resistant to alpha interferon (IFN-α) and transiently induced, but failed to secrete, IFN-β and IFN-λ. Global transcriptome analysis determined that ZIKV constitutively induced IFN regulatory factor 7 (IRF7), IRF9, and IFN-stimulated genes (ISGs) 1 to 9 days post-infection, despite persistently replicating in hBMECs. ZIKV constitutively induced ISG15, HERC5, and USP18, which are linked to hepatitis C virus (HCV) persistence and IFN regulation, chemokine CCL5, which is associated with immunopathogenesis, as well as cell survival factors.We found that CCL5, a chemokine with pro-survival effects on immune cells, was highly and constitutively secreted by ZIKV-infected hBMECs. Although roles for CCL5 in EC survival remain unknown, the presence of CCL5 receptors, CCR3 and CCR5, on ECs suggested that CCL5 could promote ZIKV persistence in hBMECs. We found that exogenous CCL5 induced ERK1/2 phosphorylation in hBMECs, and that ERK1/2 cell survival signaling was similarly activated by ZIKV infection. This suggested that blockade of the CCL5-CCR3/5 pathway may block ZIKV persistence in blood brain barrier ECs. Neutralizing antibodies to CCL5, CCR3 or CCR5 inhibited persistent ZIKV infection of hBMECs. While knock out (KO) of CCL5 failed to prevent ZIKV infection of hBMECs, 3 days post infection (dpi) we observed a >90% reduction in ZIKV-infected CCL5-KO hBMECs and a multi-log reduction in ZIKV titers. In contrast, addition of CCL5 to CCL5-KO hBMECs dose dependently rescued ZIKV persistence in hBMECs. Inhibition of CCL5 responses using CCR3 (UCB35625) and CCR5 (Maraviroc) receptor antagonists reduced the number of ZIKV-infected hBMECs and ZIKV titers (IC50s 2.5-12 µM), without cytotoxicity (CC50 >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 reveal that hBMECs act as a reservoir of persistent ZIKV replication, suggest routes for ZIKV to cross hBMECs into neuronal compartments and define novel mechanisms of ZIKV persistence that can be targeted to restrict ZIKV spread. Further, our results establish roles for CCL5 in ZIKV persistence and suggest the potential for CCL5 receptor antagonists to therapeutically inhibit ZIKV spread and neurovirulence.

ISBN: 9798460477630Subjects--Topical Terms:

642304
Virology.
Subjects--Index Terms:

Chemokines

Zika Virus Persistence and Spread in Human Brain Microvascular Endothelial Cells Is Directed by Autocrine CCL5.
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300 $a 183 p.
500 $a Source: Dissertations Abstracts International, Volume: 83-04, Section: B.
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502 $a Thesis (Ph.D.)--State University of New York at Stony Brook, 2021.
506 $a This item must not be sold to any third party vendors.
520 $a Zika virus (ZIKV) is a mosquito-borne flavivirus (FV) associated with encephalitis, Guillan-Barre syndrome and outbreaks of in utero fetal microcephaly. ZIKV transverses the blood-brain-barrier (BBB) and damages the central nervous system (CNS) by lytically infecting neurons, neural progenitors and astrocytes. In contrast to other FVs, ZIKV is uniquely detected in bodily fluids for up to 6 months, crosses placental barriers and is sexually transmitted. How ZIKV persists in patients after an acute febrile illness and the cells that support persistent ZIKV replication remains an enigma key to clearing ZIKV from patients and preventing ZIKV spread. The endothelial cell (EC) lining of capillaries normally constrains transplacental transmission and forms the BBB, which selectively restricts access of blood constituents to neurons. We found that ZIKV (strain PRVABC59) persistently infects and continuously replicates in primary human brain microvascular ECs (hBMECs) that form the BBB, without cytopathology, for >9 days and following serial hBMEC passages. ZIKV did not permeabilize hBMEC monolayers but was released apically and basolaterally from polarized hBMECs, suggesting basolateral release as a direct mechanism for ZIKV to cross the BBB.Innate and adaptive immune responses normally restrict acute FV replication and spread. ZIKV-infected hBMECs were rapidly resistant to alpha interferon (IFN-α) and transiently induced, but failed to secrete, IFN-β and IFN-λ. Global transcriptome analysis determined that ZIKV constitutively induced IFN regulatory factor 7 (IRF7), IRF9, and IFN-stimulated genes (ISGs) 1 to 9 days post-infection, despite persistently replicating in hBMECs. ZIKV constitutively induced ISG15, HERC5, and USP18, which are linked to hepatitis C virus (HCV) persistence and IFN regulation, chemokine CCL5, which is associated with immunopathogenesis, as well as cell survival factors.We found that CCL5, a chemokine with pro-survival effects on immune cells, was highly and constitutively secreted by ZIKV-infected hBMECs. Although roles for CCL5 in EC survival remain unknown, the presence of CCL5 receptors, CCR3 and CCR5, on ECs suggested that CCL5 could promote ZIKV persistence in hBMECs. We found that exogenous CCL5 induced ERK1/2 phosphorylation in hBMECs, and that ERK1/2 cell survival signaling was similarly activated by ZIKV infection. This suggested that blockade of the CCL5-CCR3/5 pathway may block ZIKV persistence in blood brain barrier ECs. Neutralizing antibodies to CCL5, CCR3 or CCR5 inhibited persistent ZIKV infection of hBMECs. While knock out (KO) of CCL5 failed to prevent ZIKV infection of hBMECs, 3 days post infection (dpi) we observed a >90% reduction in ZIKV-infected CCL5-KO hBMECs and a multi-log reduction in ZIKV titers. In contrast, addition of CCL5 to CCL5-KO hBMECs dose dependently rescued ZIKV persistence in hBMECs. Inhibition of CCL5 responses using CCR3 (UCB35625) and CCR5 (Maraviroc) receptor antagonists reduced the number of ZIKV-infected hBMECs and ZIKV titers (IC50s 2.5-12 µM), without cytotoxicity (CC50 >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 reveal that hBMECs act as a reservoir of persistent ZIKV replication, suggest routes for ZIKV to cross hBMECs into neuronal compartments and define novel mechanisms of ZIKV persistence that can be targeted to restrict ZIKV spread. Further, our results establish roles for CCL5 in ZIKV persistence and suggest the potential for CCL5 receptor antagonists to therapeutically inhibit ZIKV spread and neurovirulence.
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650 4 $a Molecular biology. $3 517296
650 4 $a Biochemistry. $3 518028
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653 $a Chemokines
653 $a Virus
653 $a Replication
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710 2 $a State University of New York at Stony Brook. $b Molecular and Cellular Biology. $3 1682421
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