Lactoferrin suppresses LPS-induced expression of HMGB1, microRNA 155, 146, and TLR4/MyD88/NF-кB pathway in RAW264.7 cells

Maryam Nemati,Saeideh Akseh,Maryam Amiri,Hamid Reza Nejabati,Ahmadreza Jodati,Nazila Fathi Maroufi,Yousef Faridvand,Mohammad Nouri

doi:10.1080/08923973.2021.1872616

Abstract

Objective This current study evaluated the underlying mechanisms of LF against the inflammatory microRNAs (miRNAs), HMGB1 expression, and TLR4-MyD88-NF-кB pathway in LPS-activated murine RAW264.7 cells. Methods MTT assay was used to assess cell metabolism and the cell culture levels of the cytokines (TNF-α, IL-6) were evaluated by Enzyme-linked immunosorbent assay (ELISA). The expression of miRNAs was quantified by using qPCR and the expression of HMGB1, TLR4, MyD88, and phosphorylated NF-κB (P-p65) were determined with Western blot and qPCR, respectively. Results The results indicated that LF downregulates IL-6 and TNF-α expression. LF exhibited the degradation of P-p65 and reduced the production of HMGB1, TLR4, and MyD88 in LPS-induced inflammatory response. Importantly, in parallel with the suppression of cytokines and HMGB1-TLR4-MyD88-NF-кB pathway, LF could induce a decrease in inflammatory selected miRNAs, mmu-mir-155, and mmu-mir-146a expression. Conclusions Altogether, these findings provide LF as a prominent anti-inflammatory agent that could modulate HMGB1, mmu-mir-155, mmu-mir-146a, and TLR4/MyD88/NF-кB pathway.

Highlights

This current study evaluated the underlying mechanisms of LF against the inflammatory miRNAs, High mobility group box-1 (HMGB1) expression and Toll-like receptor 4 (TLR4)-MyD88-NF-кB pathway in LPS-activated murine RAW264.7 cells
HMGB1 binds to TLR4 receptor and promotes the NF-κB activity through TLR4/MyD88/NF-кB dependent pathway, thereby; ablation of HMGB1 could be protective against the further production of cytokines and cell damage [6]
The results demonstrated that the levels of HMGB1, TLR4, and Myd88 were blocked in the LPS treatment group (Fig. 4A, 4B, and 4C)

Summary

Introduction

This current study evaluated the underlying mechanisms of LF against the inflammatory miRNAs, HMGB1 expression and TLR4-MyD88-NF-кB pathway in LPS-activated murine RAW264.7 cells. Inflammation can induce complex networks by activating and releasing pro-inflammatory cytokines in response to infections. Lipopolysaccharide (LPS) is a Gram-negative bacteria product, which has been commonly used to induce inflammatory response by activation of macrophage Toll-like receptor 4 (TLR4)[3]. Toll-like receptor (TLR4) is a conserved mediator of inflammation, up-regulate the expression of its downstream effectors, most importantly NF-κB, through MyD88 adaptor [4]. HMGB1 binds to TLR4 receptor and promotes the NF-κB activity through TLR4/MyD88/NF-кB dependent pathway, thereby; ablation of HMGB1 could be protective against the further production of cytokines and cell damage [6]. Targeting HMGB1 may provide an important therapeutic strategy against inflammatory response through TLR4-mediated signaling

Methods

Results

Discussion

Conclusion