44 Defects in Mitophagy Response in Burns Involved in Poor Regeneration Capacity
Authors: Tamura Takahiko , Shakuo Tomoharu , Kahiwagi Aki , Yasuda Nobuo , Amor Wiem Ben , Morinaga Hiroyuki , Khan Mohammed A , Shinozaki Shohei , Kaneki Masao , Martyn Jeevendra , Yasuhara Shingo 
Published: March 2020

Abstract

Introduction
Wound healing is a biological process directed to the restoration of tissue that has suffered an injury. One of the important functions of wound healing is individual and collective cellular migration. This migration response can largely dictate the outcomes and success of a healing wound. Mitophagy, autophagic degradation of mitochondria, sequesters damaged mitochondria and functions as the essential quality control (QC) system of this organelle. When mitophagy is disturbed, damaged mitochondria are not cleared and will produce superoxide, imposing a risk for cell death. Our previous study documented a compromised mitophagy response in burns, suggestive of poor QC of mitochondria in the severely burned subjects. The relationship between defects in mitophagy response and cellular regeneration capacity was not investigated in detail. We have previously shown that trehalose can serve as an important autophagy modulator and ameliorates the defect in mitophay response. Thus we investigated whether trehalose could improve poor regeneration capacity caused by defects in mitophagy response under burn or oxidative stress in myocytes.

Methods
We examined the effect of trehalose on migration by performing a wound-healing scratch assay. C2C12 myocyte was cultured on 6cm dishes to 95–100% confluence, the cells were treated with or without trehalose for 1 hour before the scratch. The wound was made by scratching a line across the bottom of the dish. By incubating the cells with day 3 burn or sham-burn serum harvested from rats (30%, systemic burn), To simulate the burn injury and/or oxidative stress in diseases, cells were primed with a serum derived from rats with 40% burn injury (or sham-burn control), or with or without hydrogen peroxide treatment. Nine hours later, the gap obtained by scratching a line in C2C12 was observed by time-lapse microscopy.

Results
Oxidative stress decreased the gap closure of C2C12 and delayed wound healing. Under the oxidative stress, the gap closure in C2C12 treated with trehalose is significantly higher than that C2C12 treated without trehalose (54.6 vs 79.1%; p&lt; 0.05). Similarly, burn stress showed a slower the gap closure of C2C12. Among the burn groups with or without trehalose treatment, the gap closure in C2C12 treated with trehalose is also higher than that C2C12 treated without trehalose.

Conclusions
Using a cell culture system simulating the two models of critical illnesses (oxidative stress and burn injury), we found that mitigating the defect of mitophagy response by trehalose treatment can improve poor regeneration capacity under burn or oxidative stress in myocytes.

Applicability of Research to Practice
This is the first study that demonstrated that defects in mitophagy response under disease conditions can serve a novel therapeutic target for enhancing the regeneration capacity in critically ill patients.