Bronchial epithelium energetic metabolism and functionality under exacerbation in childhood asthma

Abstract

Introduction

Asthma is the most frequent chronic inflammatory disease in children. The main cause of asthma exacerbations is the viral infection of the bronchial epithelium (BE). Rhinoviruses (RV) are detected in 85 % of asthma exacerbations in children knowing that the prevalence of RVC represents 67.5 % in children and associated with severe exacerbations. In adults, it has been shown that BE energetic metabolism shifts from mitochondrial oxidative phosphorylation towards glycolysis compared to healthy BE [1]. In healthy BE, viral infection of BE promotes a metabolic rewiring in favor for glucose utilization through glycolysis rather than mitochondrial metabolism, impacting the muco-ciliary clearance [2]. We suppose that RV infection enhances BE glycolytic metabolism affecting the BE function in childhood asthma. We will analyse both the energetic metabolism and the functions of BE under RV infections on childhood non-asthma and asthma BE.

Methods

Cell culture.

BE cells were obtained from children bronchi fibroscopic brushing. 100 000 cells were seeded on ALI transwell (0.4 μm pores, Corning Incorporated Transwell, Costar) with ALI medium (StemCell) at basal pole.

RV infection.

We infected BE with RVC MOI 0.1. The mix was removed 1 h after and the experiences were realized 24 h after infection.

Cellular oxygen consumption rate (OCR).

OCR was measured on intact cells at 37 °C in a 2 mL thermostatically monitored chamber (2.5 × 105 cells/mL/run) using an Oroboros O₂k instrument (Oroboros Instruments).

Ciliary beating frequency.

Ciliary beating frequency of BE were measured using videomicroscopy Leica DMi8 (Leica Microsystems) coupled to a high-speed camera sCMOS Flash 4.0 camera (Hamamatsu) available at The Bordeaux Imaging Center. Using MATHLAB program, the images were analyzed after application of the Fourier transform to determine the most represented beat frequency.

Results

Under basal condition, asthmatic children BE were metabolically different from non-asthmatic children BE. Proteomic analysis, OCR and glycolytic enzymes expression indicated that asthmatic BE were using more glucose through glycolysis rather that mitochondrial metabolism. RV infection aggravate the shift within non-asthmatic BE. In association, we observed a defective ciliary beating frequency and efficiency in asthmatic children BE compared to non-asthmatic children BE.

Conclusion

Our study revealed that non-asthmatic and asthmatic children BE appeared metabolically different, in basal conditions but also after RV infection. BE barrier functions seemed less efficient in asthmatic children BE compared to non-asthmatic BE. To go further, we will deepen how RV infection can modulate energetics and how it will affect BE functions.