IFIT Proteins Are Involved in CXCL10 Expression in Human Glomerular Endothelial Cells Treated with a Toll-Like Receptor 3 Agonist

Introduction: Various viruses including a novel coronavirus (SARS-CoV-2) can infect the kidney. When viruses invade the glomeruli from the bloodstream, glomerular endothelial cells (GECs) initiate the innate immune reactions. We investigated the expression of interferon (IFN)-induced protein with tetratricopeptide repeats (IFIT) 1/2/3, antiviral molecules, in human GECs treated with a toll-like receptor (TLR) 3 agonist. Role of IFIT1/2/3 in the expression of C-X-C motif chemokine ligand 10 (CXCL10) was also examined. Methods: Human GECs were cultured and stimulated with polyinosinic-polycytidylic acid (poly IC), a synthetic TLR3 agonist. Real-time qPCR, Western blotting, and ELISA were used to examine the expression of IFIT1/2/3, IFN-β, and CXCL10. RNA interference against IFN-β or IFIT1/2/3 was also performed. Results: Expression of IFIT1/2/3 and CXCL10 was induced by poly IC in GECs. The inductions were inhibited by RNA interfering of IFN-β. Knockdown of IFIT1/2/3 decreased the CXCL10 expression. Knockdown of IFIT3 decreased the expression of IFIT1 and IFIT2 proteins. Conclusion: IFIT1/2/3 and CXCL10 were induced by poly IC via IFN-β in GECs. IFIT1/2/3 may increase the expression of CXCL10 which induces lymphocyte chemotaxis and may inhibit the replication of infected viruses. These molecules may play a role in GEC innate immune reactions in response to viruses.

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Cytosolic Sensors of Viral RNA Are Involved in the Production of Interleukin-6 via Toll-Like Receptor 3 Signaling in Human Glomerular Endothelial Cells

Kidney & Blood Pressure Research ◽

10.1159/000498837 ◽

2019 ◽

Vol 44 (1) ◽

pp. 62-71 ◽

Cited By ~ 4

Author(s):

Qiang Liu ◽

Tadaatsu Imaizumi ◽

Tomomi Aizawa ◽

Koji Hirono ◽

Shogo Kawaguchi ◽

...

Keyword(s):

Endothelial Cells ◽

Signaling Pathway ◽

Interleukin 6 ◽

Viral Rna ◽

Pivotal Role ◽

Toll Like Receptor ◽

Polycytidylic Acid ◽

Glomerular Endothelial Cells ◽

Human Glomerular Endothelial Cells ◽

Tlr3 Signaling

Background/Aims: Dysregulation of interleukin-6 (IL-6) production in residual renal cells may play a pivotal role in the development of glomerulonephritis (GN). Given that Toll-like receptor 3 (TLR3) signaling has been implicated in the pathogenesis of some forms of GN, we examined activated TLR3-mediated IL-6 signaling in cultured normal human glomerular endothelial cells (GECs). Methods: We treated GECs with polyinosinic-polycytidylic acid (poly IC), an authentic double-stranded RNA, and analyzed the expression of IL-6 and the cytosolic viral RNA sensors retinoic acid-inducible gene-I (RIG-I) and melanoma differentiation associated gene 5 (MDA5) using reverse transcription quantitative real-time polymerase chain reaction, western blotting, and enzyme-linked immunosorbent assays. To further elucidate the effects of poly IC on this signaling pathway, we subjected the cells to small interfering RNA (siRNA) against TLR3, interferon (IFN)-β, RIG-I, and MDA5. Results: We found that poly IC induced the expression of RIG-I, MDA5 and IL-6 via TLR3/IFN-β signaling in GECs. siRNA experiments revealed that both MDA5 and RIG-I were involved in the poly IC-induced expression of IL-6, with MDA5 being upstream of RIG-I. Conclusion: Interestingly, cytosolic sensors of viral RNA were found to be involved in IL-6 production via TLR3 signaling in GECs. Regional activation of TLR3/IFN-β/ MDA5/RIG-I/IL-6 axis due to viral and “pseudoviral” infections is involved in innate immunity and inflammatory reactions in GECs. We believe this signaling pathway also plays a pivotal role in the development of some forms of GN.

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Toll-like receptor 3 signaling and human glomerular endothelial cells

10.46439/nephrology.1.002 ◽

2020 ◽

Vol 1 (1) ◽

Keyword(s):

Endothelial Cells ◽

Toll Like Receptor ◽

Glomerular Endothelial Cells ◽

Human Glomerular Endothelial Cells

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Endothelial expression of fractalkine (CX3CL1) is induced by Toll-like receptor 3 signaling in cultured human glomerular endothelial cells

Modern Rheumatology ◽

10.1080/14397595.2019.1682768 ◽

2019 ◽

Vol 30 (6) ◽

pp. 1074-1081

Author(s):

Koji Hirono ◽

Tadaatsu Imaizumi ◽

Tomomi Aizawa ◽

Shojiro Watanabe ◽

Koji Tsugawa ◽

...

Keyword(s):

Endothelial Cells ◽

Toll Like Receptor ◽

Glomerular Endothelial Cells ◽

Human Glomerular Endothelial Cells

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Toll-Like Receptor 3 Signaling Contributes to Regional Neutrophil Recruitment in Cultured Human Glomerular Endothelial Cells

Nephron ◽

10.1159/000489507 ◽

2018 ◽

Vol 139 (4) ◽

pp. 349-358 ◽

Cited By ~ 4

Author(s):

Qiang Liu ◽

Tadaatsu Imaizumi ◽

Shogo Kawaguchi ◽

Tomomi Aizawa ◽

Tomoh Matsumiya ◽

...

Keyword(s):

Endothelial Cells ◽

Neutrophil Recruitment ◽

Toll Like Receptor ◽

Glomerular Endothelial Cells ◽

Human Glomerular Endothelial Cells

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Inhibitory effect of anti-malarial agents on the expression of proinflammatory chemokines via Toll-like receptor 3 signaling in human glomerular endothelial cells

Renal Failure ◽

10.1080/0886022x.2021.1908901 ◽

2021 ◽

Vol 43 (1) ◽

pp. 643-650

Author(s):

Riko Sato ◽

Tadaatsu Imaizumi ◽

Tomomi Aizawa ◽

Shojiro Watanabe ◽

Koji Tsugawa ◽

...

Keyword(s):

Endothelial Cells ◽

Inhibitory Effect ◽

Toll Like Receptor ◽

Glomerular Endothelial Cells ◽

Human Glomerular Endothelial Cells

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Expression of Zinc-Finger Antiviral Protein in hCMEC/D3 Human Cerebral Microvascular Endothelial Cells: Effect of a Toll-Like Receptor 3 Agonist

NeuroImmunoModulation ◽

10.1159/000521012 ◽

2021 ◽

pp. 1-10

Author(s):

Mako Okudera ◽

Minami Odawara ◽

Masashi Arakawa ◽

Shogo Kawaguchi ◽

Kazuhiko Seya ◽

...

Keyword(s):

Endothelial Cells ◽

Protein Expression ◽

Zinc Finger ◽

Microvascular Endothelial Cells ◽

Brain Microvascular Endothelial Cells ◽

Toll Like Receptor ◽

Antiviral Protein ◽

Polycytidylic Acid ◽

Microvascular Endothelial ◽

The Brain

Introduction: Invasion of viruses into the brain causes viral encephalitis, which can be fatal and causes permanent brain damage. The blood-brain barrier (BBB) protects the brain by excluding harmful substances and microbes. Brain microvascular endothelial cells are important components of the BBB; however, the mechanisms of antiviral reactions in these cells have not been fully elucidated. Zinc-finger antiviral protein (ZAP) is a molecule that restricts the infection of various viruses, and there are 2 major isoforms: ZAPL and ZAPS. Toll-like receptor 3 (TLR3), a pattern-recognition receptor against viral double-stranded RNA, is implicated in antiviral innate immune reactions. The aim of this study was to investigate the expression of ZAP in cultured hCMEC/D3 human brain microvascular endothelial cells treated with an authentic TLR3 agonist polyinosinic-polycytidylic acid (poly IC). Methods: hCMEC/D3 cells were cultured and treated with poly IC. Expression of ZAPL and ZAPS mRNA was investigated using quantitative reverse transcription-polymerase chain reaction, and protein expression of these molecules was examined using western blotting. The role of nuclear factor-κB (NF-κB) was examined using the NF-κB inhibitor, SN50. The roles of interferon (IFN)-β, IFN regulatory factor 3 (IRF3), tripartite motif protein 25 (TRIM25), and retinoic acid-inducible gene-I (RIG-I) in poly IC-induced ZAPS expression were examined using RNA interference. Propagation of Japanese encephalitis virus (JEV) was examined using a focus-forming assay. Results: ZAPS mRNA and protein expression was upregulated by poly IC, whereas the change of ZAPL mRNA and protein levels was minimal. Knockdown of IRF3 or TRIM25 decreased the poly IC-induced upregulation of ZAPS, whereas knockdown of IFN-β or RIG-I did not affect ZAPS upregulation. SN50 did not affect ZAPS expression. Knockdown of ZAP enhanced JEV propagation. Conclusion: ZAPL and ZAPS were expressed in hCMEC/D3 cells, and ZAPS expression was upregulated by poly IC. IRF3 and TRIM25 are involved in poly IC-induced upregulation of ZAPS. ZAP may contribute to antiviral reactions in brain microvascular endothelial cells and protect the brain from invading viruses such as JEV.

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