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Ciprofloxacin and levofloxacin attenuate microglia inflammatory response via TLR4/NF-kB pathway.

ABSTRACT: BACKGROUND:Neuroinflammation is the response of the central nervous system to events that interfere with tissue homeostasis and represents a common denominator in virtually all neurological diseases. Activation of microglia, the principal immune effector cells of the brain, contributes to neuronal injury by release of neurotoxic products. Toll-like receptor 4 (TLR4), expressed on the surface of microglia, plays an important role in mediating lipopolysaccharide (LPS)-induced microglia activation and inflammatory responses. We have previously shown that curcumin and some of its analogues harboring an ?,?-unsaturated 1,3-diketone moiety, able to coordinate the magnesium ion, can interfere with LPS-mediated TLR4-myeloid differentiation protein-2 (MD-2) signaling. Fluoroquinolone (FQ) antibiotics are compounds that contain a keto-carbonyl group that binds divalent ions, including magnesium. In addition to their antimicrobial activity, FQs are endowed with immunomodulatory properties, but the mechanism underlying their anti-inflammatory activity remains to be defined. The aim of the current study was to elucidate the molecular mechanism of these compounds in the TLR4/NF-?B inflammatory signaling pathway. METHODS:The putative binding mode of five FQs [ciprofloxacin (CPFX), levofloxacin (LVFX), moxifloxacin, ofloxacin, and delafloxacin] to TLR4-MD-2 was determined using molecular docking simulations. The effect of CPFX and LVFX on LPS-induced release of IL-1? and TNF-? and NF-?B activation was investigated in primary microglia by ELISA and fluorescence staining. The interaction of CPFX and LVFX with TLR4-MD-2 complex was assessed by immunoprecipitation followed by Western blotting using Ba/F3 cells. RESULTS:CPFX and LVFX bound to the hydrophobic region of the MD-2 pocket and inhibited LPS-induced secretion of pro-inflammatory cytokines and activation of NF-?B in primary microglia. Furthermore, these FQs diminished the binding of LPS to TLR4-MD-2 complex and decreased the resulting TLR4-MD-2 dimerization in Ba/F3 cells. CONCLUSIONS:These results provide new insight into the mechanism of the anti-inflammatory activity of CPFX and LVFX, which involves, at least in part, the activation of TLR4/NF-?B signaling pathway. Our findings might facilitate the development of new molecules directed at the TLR4-MD-2 complex, a potential key target for controlling neuroinflammation.

SUBMITTER: Zusso M

PROVIDER: S-EPMC6637517 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature

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Ciprofloxacin and levofloxacin attenuate microglia inflammatory response via TLR4/NF-kB pathway.

Zusso Morena M Lunardi Valentina V Franceschini Davide D Pagetta Andrea A Lo Rita R Stifani Stefano S Frigo Anna Chiara AC Giusti Pietro P Moro Stefano S

Journal of neuroinflammation 20190718 1

<h4>Background</h4>Neuroinflammation is the response of the central nervous system to events that interfere with tissue homeostasis and represents a common denominator in virtually all neurological diseases. Activation of microglia, the principal immune effector cells of the brain, contributes to neuronal injury by release of neurotoxic products. Toll-like receptor 4 (TLR4), expressed on the surface of microglia, plays an important role in mediating lipopolysaccharide (LPS)-induced microglia act ...[more]

PMID: 31319868

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Project description:Background: Acute lung injury (ALI) is characterized by dysfunction of the alveolar epithelial membrane caused by acute inflammation and tissue injury. Qingwenzhike (QWZK) prescription has been demonstrated to be effective against respiratory viral infections in clinical practices, including coronavirus disease 2019 (COVID-19) infection. So far, the chemical compositions, protective effects on ALI, and possible anti-inflammatory mechanisms remain unknown. Methods: In this study, the compositions of QWZK were determined via the linear ion trap/electrostatic field orbital trap tandem high-resolution mass spectrometry (UHPLC-LTQ-Orbitrap MS). To test the protective effects of QWZK on ALI, an ALI model induced by lipopolysaccharide (LPS) in rats was used. The effects of QWZK on the LPS-induced ALI were evaluated by pathological changes and the number and classification of white blood cell (WBC) in bronchoalveolar lavage fluid (BALF). To investigate the possible underlying mechanisms, the contents of interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), monocyte chemoattractant protein (MCP-1), interleukin-1β (IL-1β), interleukin-18 (IL-18), and immunoregulatory-related factors interferon-γ (IFN-γ) were detected by ELISA. Furthermore, the expression of Toll-like receptor 4 (TLR4), p-IKKα/β, IKKα, IKKβ, p-IκBα, IκBα, p-NF-κB, nuclear factor-κB (NF-κB), NOD-like receptor family pyrin domain containing 3 (NLRP3), cleaved caspase-1, pro-caspase-1, apoptosis-associated speck-like protein containing CARD (ASC), and β-actin were tested by Western blot. Results: A total of 99 compounds were identified in QWZK, including 33 flavonoids, 23 phenolic acids, 3 alkaloids, 3 coumarins, 20 triterpenoids, 5 anthraquinones, and 12 others. ALI rats induced by LPS exhibited significant increase in neutrophile, significant decrease in lymphocyte, and evidently thicker alveolar wall than control animals. QWZK reversed the changes in WBC count and alveolar wall to normal level on the model of ALI induced by LPS. ELISA results revealed that QWZK significantly reduced the overexpression of proinflammatory factors IL-6, TNF-α, MCP-1, IL-1β, IL-18, and IFN-γ induced by LPS. Western blot results demonstrated that QWZK significantly downregulated the overexpression of TLR4, p-IKKα/β, p-IκBα, p-NF-κB, NLRP3, cleaved caspase-1, and ASC induced by LPS, which suggested that QWZK inhibited TLR4/NF-κB signaling pathway and NLRP3 inflammasomes. Conclusions: The chemical compositions of QWZK were first identified. It was demonstrated that QWZK showed protective effects on ALI induced by LPS. The possible underlying mechanisms of QWZK on ALI induced by LPS was via inhibiting TLR4/NF-kB signaling pathway and NLRP3 inflammasome activation. This work suggested that QWZK is a potential therapeutic candidate for the treatments of ALI and pulmonary inflammation.

Project description:Patients with antiphospholipid syndrome have been identified to have higher incidence rates of atherosclerosis (AS) due to the elevated levels of anti‑β2‑glycoprotein I (β2GPI) antibody (Ab). Our previous studies revealed that the anti‑β2GPI Ab formed a stable oxidized low‑density lipoprotein (oxLDL)/β2GPI/anti‑β2GPI Ab complex, which accelerated AS development by promoting the accumulation of lipids in macrophages and vascular smooth muscle cell. However, the effects of the complex on endothelial cells, which drive the initiation and development of AS, remain unknown. Thus, the present study aimed to determine the proinflammatory roles of the oxLDL/β2GPI/anti‑β2GPI Ab complex in human umbilical vein endothelial cells (HUVECs) in an attempt to determine the underlying mechanism. Reverse transcription‑quantitative PCR, enzymy‑linked immunosorbent assay, western blotting and immunofluorescence staining were performed to detect the expressions of inflammation related factors and adhesion molecules. Monocyte‑binding assay was used to investigate the effects of oxLDL/β2GPI/anti‑β2GPI Ab complex on monocyte adhesion to endothelial cells. The results demonstrated that the oxLDL/β2GPI/anti‑β2GPI Ab complex upregulated the expression of Toll‑like receptor (TLR)4 and the levels of NF‑κB phosphorylation in HUVECs, and subsequently enhanced the expression levels of inflammatory cytokines, including TNF‑α, IL‑1β and IL‑6, as well as those of adhesion molecules, such as intercellular adhesion molecule 1 and vascular adhesion molecule 1. In addition, the complex facilitated the recruitment of monocytes by promoting the secretion of monocyte chemotactic protein 1 in HUVECs. Notably, the described effects of the oxLDL/β2GPI/anti‑β2GPI Ab complex in HUVECs were abolished by either TLR4 or NF‑κB blockade. In conclusion, these findings suggested that the oxLDL/β2GPI/anti‑β2GPI Ab complex may induce a hyper‑inflammatory state in endothelial cells by promoting the secretion of proinflammatory cytokines and monocyte recruitment, which was discovered to be largely dependent on the TLR4/NK‑κB signaling pathway.

Dataset Information

Ciprofloxacin and levofloxacin attenuate microglia inflammatory response via TLR4/NF-kB pathway.

Publications

Ciprofloxacin and levofloxacin attenuate microglia inflammatory response via TLR4/NF-kB pathway.

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