Phloretin Attenuates Allergic Airway Inflammation and Oxidative Stress in Asthmatic Mice.

Wen-Chung Huang,Li-Wen Fang,Chian-Jiun Liou

doi:10.3389/fimmu.2017.00134

Abstract

Phloretin (PT), isolated from the apple tree, was previously demonstrated to have antioxidative and anti-inflammatory effects in macrophages and anti-adiposity effects in adipocytes. Inflammatory immune cells generate high levels of reactive oxygen species (ROS) for stimulated severe airway hyperresponsiveness (AHR) and airway inflammation. In this study, we investigated whether PT could reduce oxidative stress, airway inflammation, and eosinophil infiltration in asthmatic mice, and ameliorate oxidative and inflammatory responses in tracheal epithelial cells. BALB/c mice were sensitized with ovalbumin (OVA) to induce asthma symptoms. Mice were randomly assigned to the five experimental groups: normal controls; OVA-induced asthmatic mice; and OVA-induced mice injected intraperitoneally with one of the three PT doses (5, 10, or 20 mg/kg). In addition, we treated inflammatory human tracheal epithelial cells (BEAS-2B cells) with PT to assess oxidative responses and the levels of proinflammatory cytokines and chemokines. We found that PT significantly reduced goblet cell hyperplasia and eosinophil infiltration, which decreased AHR, inflammation, and oxidative responses in the lungs of OVA-sensitized mice. PT also decreased malondialdehyde levels in the lung and reduced Th2 cytokine production in bronchoalveolar lavage fluids. Furthermore, PT reduced ROS, proinflammatory cytokines, and eotaxin production in BEAS-2B cells. PT also suppressed monocyte cell adherence to inflammatory BEAS-2B cells. These findings suggested that PT alleviated pathological changes, inflammation, and oxidative stress by inhibiting Th2 cytokine production in asthmatic mice. PT showed therapeutic potential for ameliorating asthma symptoms in the future.

Highlights

Allergic asthma is a complex, chronic inflammatory airway disease
Asthmatic mice treated with PT had significantly reduced numbers of eosinophils and total cells compared to the OVA group (eosinophils: PT5: 6.2 × 105 ± 6.9 × 104, p = 0.57; FIGURE 1 | The effect of phloretin (PT) on airway hyperresponsiveness (AHR) and cell counts in bronchoalveolar lavage fluid (BALF) of asthmatic mice. (A) Changes in AHR (Penh values) with inhalation of increasing methacholine doses (10–40 mg/ml) in normal (N) and OVA-stimulated (OVA) mice, without or with PT (PT5-20) treatment (n = 12 mice/group, measured in three independent experiments). (B) Numbers of inflammatory cells and total cells in Bronchoalveolar Lavage Fluid (BALF) with increasing PT doses; (C) percentage of eosinophils in BALFs from OVA-sensitive mice, treated or untreated with PT
The proportion of eosinophils in BALF derived from the P10 or the P20 group was significantly reduced compared to the proportion in the OVA group (Figure 1C)

Summary

INTRODUCTION

Allergic asthma is a complex, chronic inflammatory airway disease. Patients with sudden, acute asthma attacks exhibit shortness of breath, dry coughing, chest tightness, paroxysmal wheezing due to airway obstruction, troubled breathing, and even death [1]. Several immune cells, including Th2 cells, mast cells, eosinophils, and airway epithelial cells, secrete cytokines, chemokines, and inflammatory mediators that damage lung tissue and lead to the restricted breathing [4]. We measured MDA activity in lung tissues with the lipid peroxidation assay kit, according to the manufacturer’s instructions (Sigma). To detect glutathione levels in lung tissues, we employed a glutathione assay kit, according to the manufacturer’s instructions (Sigma). The cell culture supernatants and BALF were used to measure with specific ELISA kits to detect the concentrations of CCL5, CCL11, CCL24, CCL26, intercellular adhesion molecule 1 (ICAM-1), IL-4, IL-5, IL-6, IL-8, IL-13, MCP-1, and tumor necrosis factor-α (TNF-α), according to the manufacturer’s instructions (R&D Systems, Minneapolis, MN, USA), as previously described [16].

RESULTS

DISCUSSION

ETHICS STATEMENT