Scanning Electrochemical Microscopy Imaging during Respiratory Burst in Human Cell.

Hiroyuki Kikuchi,Tomokazu Matsue,Ryo Matsuoka,Ankush Prasad,Shigenobu Kasai,Shigeo Aoyagi

doi:10.3389/fphys.2016.00025

Abstract

Phagocytic cells, such as neutrophils and monocytes, consume oxygen and generate reactive oxygen species (ROS) in response to external stimuli. Among the various ROS, the superoxide anion radical is known to be primarily produced by nicotinamide adenine dinucleotide phosphate hydrogen (NADPH) oxidase. In the current study, we attempt to evaluate the respiratory burst by monitoring the rapid consumption of oxygen by using scanning electrochemical microscopy (SECM) imaging. The respiratory burst was measured in a human monocytic cell line (THP-1 cells) derived from an acute monocytic leukemia patient under the effect of the exogenous addition of phorbol 12-myristate 13-acetate, which acts as a differentiation inducer. SECM imaging composed of a microelectrode was used to compare oxygen consumption between normal cellular respiration and during respiratory burst in THP-1 cells. Two-dimensional respiratory activity imaging was performed using XY-scan. In addition, the quantitative evaluation of oxygen consumption in THP-1 cells was performed using a Z-scan. The results obtained show higher consumption of oxygen in cells undergoing respiratory burst. SECM imaging is thus claimed to be a highly sensitive and appropriate technique compared to other existing techniques available for evaluating oxidative stress in human cells, making it potentially useful for widespread applications in biomedical research and clinical trials.

Highlights

Living organisms bear defense mechanisms in which immune cells, such as neutrophils and monocytes, play pivotal roles in responding to and killing foreign bodies that invade the living system (Forman and Torres, 2002; Halliwell and Gutteridge, 2007)
We demonstrated the enhancement of reduction current for H2O2 during the differentiation process of leukocytes and THP-1 cells using microelectrode (Shigenobu et al, 2005; Inoue et al, 2010)
The respiratory burst under phorbol 12-myristate 13-acetate (PMA) addition is known to occur via activation of protein kinase C (PKC), which leads to activation of nicotinamide adenine dinucleotide phosphate hydrogen (NADPH) oxidase (Castagna et al, 1982; Kikkawa et al, 1983)

Summary

Introduction

Living organisms bear defense mechanisms in which immune cells, such as neutrophils and monocytes, play pivotal roles in responding to and killing foreign bodies that invade the living system (Forman and Torres, 2002; Halliwell and Gutteridge, 2007). Phagocytic cells, such as neutrophils and monocytes, produce reactive oxygen species (ROS) during phagocytosis (Forman and Torres, 2002). The main goal of the current study is to introduce scanning electrochemical microscopy (SECM) imaging of oxygen consumption in immune cells for the first time, which is associated with ROS generation during respiratory burst. The quantitative evaluation of respiratory activity of THP-1 cells was performed by Z-scan, and respiratory activity imaging of THP-1 cells was performed by XY-scan

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Methods

Results