Elsevier

Cellular Immunology

Volume 319, September 2017, Pages 53-60
Cellular Immunology

Research paper
MicroRNA-124 alleviates chronic skin inflammation in atopic eczema via suppressing innate immune responses in keratinocytes

https://doi.org/10.1016/j.cellimm.2017.08.003Get rights and content

Highlights

  • MiR-124 expression was downregulated in atopic eczema tissues and in response to inflammatory factor stimulation.

  • MiR-124 directly targeted NF-κB.

  • MiR-124 could reverse the effect of p65 on downstream inflammatory factors.

Abstract

Chronic skin inflammation in atopic eczema is associated with elevated expression of proinflammatory genes and activation of innate immune responses in keratinocytes. MicroRNAs (miRNAs) are short, single-stranded RNA molecules that silence genes via the degradation of target mRNAs or inhibition of translation. Recent studies have demonstrated that miR-124 is associated with regulation of inflammation factors in several diseases. The aim of this study was to investigate the role of miR-124 in skin inflammation of atopic eczema. We showed that miR-124 expression is decreased in chronic lesional skin of patients with atopic eczema, and could be strongly inhibited by IFN-γ and TNF-α. Through Western blot, real-time PCR and luciferase assays, we revealed that miR-124 inhibited the expression of p65, a member of NF-κB family which can regulate many factors involved in the immune response and inflammatory reactions, through direct targeting. Further, upon IFN-γ or TNF-α stimulation, IL8, CCL5 and CCL8 showed to be significantly upregulated by IFN-γ or TNF-α, downregulated by miR-124; the promotive effect of IFN-γ and TNF-α could be partially reversed by miR-124. The levels of IL8, CCL5 and CCL8 could be significantly downregulated by p65 knockdown, upregulated by miR-124 inhibition; the suppressive effect of p65 knockdown could be partially reversed by miR-124. Moreover, contrary to miR-124, p65, IL8, CCL5 and CCL8 mRNA expression was upregulated in chronic lesional skin of patients with atopic eczema, and all inversely correlated with miR-124. Taken together, our data demonstrate that miR-124 controls NF-κB-dependent inflammatory responses in keratinocytes and chronic skin inflammation in atopic eczema; rescuing miR-124 expression presents a promising strategy for atopic eczema treatment.

Introduction

Eczema, a skin inflammatory reaction mediated by antigen-specific T cells [1], occurs in 15%–20% of infants and young children and persists throughout adulthood in 1%–3% of the cases [2]. The most common type of eczema is atopic eczema, in which a dry, itchy rash develops [3]. Recurrent exacerbations, activation of the immune system, and hyperplasia and keratinocyte apoptosis in the chronic phase represent cardinal features of atopic eczema [2], [4], [5].

Skin inflammation during an exacerbation can persist for several weeks and is nearly exclusively complicated with secondary infections that contribute to the activation of innate immune responses and the nuclear factor kappa B (NF-κB) pathway in keratinocytes [6], [7]. Several studies have reported that NF-κB plays a critical role in the production of chemokines in TNF-α and/or IFN-γ-stimulated keratinocytes [8], [9], [10]. IFN-γ can induce several characteristic chemokines in keratinocytes, including CCL5 (also known as RANTES) and CCL8 (also known as MCP-2) [11], [12]. CCL5 promoter polymorphisms and enhanced levels have been shown to be associated with atopic dermatitis [13], [14], [15]. CCL8 mediates the recruitment of IL-5-enriched TH2 cells in the inflamed skin in a mouse model of chronic atopic dermatitis [16]. In addition, the presence of TNF-α could increase the secretion of GM-CSF, CCL5, and IL-10 [17].

Regulation of inflammatory responses in diseases is mediated by coordinated control of gene expression, which is also modulated by microRNAs (miRNAs). MiRNAs silence genes by triggering the degradation of target mRNAs and inhibition of translation [18], [19]. Several miRNAs, including miR-124, function in the regulation of various inflammatory processes [20], [21], [22]. In glioma cells, the restoration of miR-124 can reduce the activation of NF-κB [23]. Additionally, an activated NF-κB-centered inflammatory loop in the highly aggressive non-small cell lung cancer cells leads to down-regulation of miR-124 [24]. The transfection of PC12 neurons with miR-124 counteracted the inhibition of neurites mediated by both recombinant TNF-α and macrophages, suggesting that miR-124 might be a valuable tool for desensitizing neurons to a repulsive inflammatory environment [25]. However, the function of miR-124 in the normal skin and keratinocytes and in atopic eczema has not been previously explored.

Here, we demonstrate that the level of miR-124 is decreased in lesions tissue from patients with atopic eczema and also decreased in keratinocytes in response to inflammation factors, in turn, controls chronic inflammatory processes that are triggered by TNF-α and/or IFN-γ though direct targeting NF-κB and regulation of CCL5, CCL8 and IL-8 expression in keratinocytes.

Section snippets

Tissues, cell lines and cell transfection

We collected 37 paired atopic eczema skin lesional tissues and normal non-lesional tissues with approval of the Ethical Review Committees on Hunan University of Chinese Medicine. All patients signed a written informed consent form and were diagnosed by dermatologists according to the diagnostic standard for atopic eczema. The criteria for diagnosis and disease phase of these patients were Eczema Area and Severity Index (EASI) [26].

For functional studies, pooled, normal human epidermal

MiR-124 expression was downregulated in atopic eczema tissues and in response to inflammatory factor stimulation

Previous studies have reported the association between miR-124 and inflammatory factors [25], [27]. Here we firstly evaluated miR-124 expression in atopic eczema lesional tissues and non-lesional normal tissues (n = 37). Results from real-time PCR assays showed that miR-124 expression was downregulated in atopic eczema tissues, compared to the non-lesional normal tissues (Fig. 1A). Next, we conducted a series of inflammatory factor treatment, including IL-13, IL-1β, IL-17A, IL-4, IFN-γ and TNF-α,

Discussion

Information regarding the role of miRNAs in allergic diseases is limited [30], [31]. The present study demonstrates an essential immune regulatory function of miR-124 in the inflammatory skin of atopic eczema. miR-124 is downregulated in the lesional skin of patients with atopic eczema and inhibits the expression of p65 through direct binding to the 3′UTR of RELA, thus to suppress numerous proinflammatory factors, including IFN-γ- or TNF-α- inducible genes IL-8, CCL5 and CCL8 in human primary

Acknowledgments

This work was supported by the National Natural Sciences Foundation of China (No: 81573985) and Chinese medicine research project of Hunan Province (No: 2016137 and 2016127).

Author disclosure statement

There is no conflict of interest.

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