Anti-oxidant and anti-inflammatory effects of caffeic acid: in vivo evidences in a model of noise-induced hearing loss

Highlights

• Cross-talk between oxidative and inflammatory pathways represents a target for NIHL treatment.

• CA administration decrease cochlear superoxide and lipid peroxidation expression in NIHL.

• Cochlear increased expression of inflammatory markers is counteracted by CA.

• CA administration contributes to the recovery of auditory function in NIHL.

Abstract

Scope

The imbalance of cellular redox status, in conjunction with the activation of inflammatory processes, have been considered common predominant mechanisms of noise-induced hearing loss. The identification of novel natural products as potential therapeuticstargeting oxidative stress and inflammatory pathways is an emerging field. Here, we focused on the polyphenol caffeic acid (CA), the major representative of hydroxycinnamic acids and phenolic acid, in order to investigate its protective capacity in a model of sensorineural hearing loss induced by noise.

Methods and results

Hearing loss was induced by exposing animals (Wistar rats) to a pure tone, 120 dB, 10 kHz for 60 min. By using auditory brainstem responses (ABRs) and immunofluorescence analysis, we found that CA protects auditory function and limits cell death in the cochlear middle/basal turn, damaged by noise exposure. Immunofluorescence analysis provided evidence that CA mediates multiple cell protection mechanisms involving both anti-inflammatory and anti-oxidant effects by decreasing NF-κB and IL-1β expression in the cochlea and opposing the oxidative/nitrosative damage induced by noise insult.

Conclusions

These results demonstrate that the supplementation of polyphenol CA can be considered a valid therapeutic strategy for attenuating noise-induced hearing loss and cochlear damage targeting both inflammatory signalling and cochlear redox balance.

Introduction

Noise-induced hearing loss (NIHL), is a major source of hearing disability in workers and adult population worldwide (Nelson et al., 2005). Furthermore, the impact of noise exposure on the adolescents and young population is a further challenge in the understanding of mechanisms for a targeted therapeutic approach. Since two decades, it is widely accepted that mediator of cochlear noise-induced damage is the excess of free radical oxygen species (ROS) and reactive nitrogen species (RNS) formations that lead to oxidative/nitrosative stress (Henderson et al., 2006; Fetoni et al., 2010, 2015a; Wang and Puel, 2018). However, the entity of cochlear damage cannot be explained with the activation of the oxidative injury, therefore the interest has been recently addressed to the role of ROS generation in leading the production of pro-inflammatory cytokines that can further produce cochlear damage (Keithley et al., 2008; Wakabayashi et al., 2010; Tan et al., 2016). Indeed, noise exposure has been shown to up-regulate cochlear production of cytokines (Fujioka et al., 2006), as interleukin 1-β (IL-1β) and of tumor necrosis factor alpha (TNF-α) (Fuentes-Santamaría et al., 2017), both of which have been observed after ROS generation in the cochlea (Tan et al., 2016). Generation of these pro-inflammatory mediators can occur via activation of the nuclear factor kappa B (NF-κB) signalling cascade, leading to cytokine production (Yamamoto et al., 2009). Similar mechanisms have been reported to occur in drug-induced hearing loss (Rybak et al., 2007; Fetoni et al., 2019) and aging (Fujimoto et al., 2014).

Based on these premises, the identification of novel antioxidant/anti-inflammatory molecules as potential therapeutics is an emerging field of the research in neurodegenerative diseases, including deafness (Linseman et al., 2009; Mancuso et al., 2012). Specifically, increasing evidences strongly support a role for polyphenols, organic compounds abundantly found in plants, in the prevention of redox unbalance as well as of the inflammatory state in cells (Cory et al., 2018). Indeed, literature evidences suggest that consumption of polyphenols can protect against several injuries, as cancers, type 2 diabetes, cardiovascular and neurodegenerative diseases (Cory et al., 2018; Fetoni et al., 2015b; Martín-Peláez et al., 2013). The dominant explanation for these benefits is the “biochemical scavenger theory”, which posits that polyphenol compounds counteract free radicals by forming stabilized chemical complexes, thus preventing further reactions (Sroka and Cisowski, 2003).

More recently, in conjunction with antioxidant properties, anti-inflammatory activities of polyphenols have been reported in acute and chronic inflammation in animal models (Hussain et al., 2016). However, polyphenols are a wide group of molecules and they may differ mainly for their bioavailability due to the composition of colonic microbiota and cell targets (Li et al., 2014; Kawabata et al., 2019). In addition to the evidence of polyphenol direct antioxidant activity, there are indications that they may also act as modulators of cell signalling exhibiting pro-oxidant/anti-oxidant properties depending on dosage and cell context (Fetoni et al., 2015b; Hussain et al., 2016). Herein, among natural compounds we focused on caffeic acid (CA), the major representative of hydroxycinnamic acids and phenolic acid, produced through the secondary metabolism of several vegetables, including olives, coffee beans, fruits, potatoes, carrots and propolis. It is usually found as various simple derivatives such as glycosides, amides, esters and sugar esters (Touaibia et al., 2011) and it is contained in several foods including coffee, one of the most popular beverages consumed by millions of people every day. It has been evaluated for its physiological and pharmacological properties that include antiviral, antioxidant, anti-inflammatory, anticarcinogenic, immunomodulatory, antidiabetic, cardioprotective, antiproliferative and hepatoprotective activity (Gulcin et al., 2006; Touaibia et al., 2011; Kim et al., 2014; Chang et al., 2019; Espíndola et al., 2019). The interest on this polyphenol in a model of damage induced by environmental risk factor, such as noise exposure, is mainly based on the evidence that it can attenuate and reverse the production of pro-inflammatory factors (Hussain et al., 2016) in addition to the modulation of redox unbalance (Li et al., 2014).

Thus, the aim of this study was to determine the protective role of CA in a model of NIHL, given that no effective pharmacologic agents are approved by Food and Drug Administration (FDA) to counteract cochlear dysfunction induced by exposure to environmental noise. In the present study, for the first time we evaluated protective properties of CA in a model of NIHL in vivo, by assessing its antioxidant and anti-inflammatory properties targeting (i) oxidative/nitrosative stress and lipid peroxidation; (ii) endogenous antioxidant responses modulating Nrf2/HO-1 pathway and (iii) inflammation markers as NF-κB and IL-1β.