Techniques for Detecting Reactive Oxygen Species in Pulmonary Vasculature Redox Signaling

Redox signaling plays important roles in regulating pulmonary vasculature function. Aberrant redox signaling, e.g., overproduction of reactive oxygen species (ROS) that exceeds the capability of cellular antioxidant mechanisms, has been found to alter vasculature function and remodel blood vessel structure, thus contributes to pathological processes of pulmonary vasculature. The regulation of pulmonary vasculature via ROS is a very complicated process with various biological events involved, however, the specific effect of individual ROS and the underlying mechanism still remain unclear. Most of ROS are present as free radical forms with extremely short lifetime, which makes it very difficult to detect the ROS and investigate their bioactivities. Therefore, developing specific and sensitive methods to detect ROS in complex biological system is essential for us to advance our knowledge in pulmonary vasculature regulation. In this chapter, we introduce several commonly used techniques for the detection of ROS in vitro and in vivo, including chemiluminescence-based assay, fluorescence-based assay, cytochrome c reduction method, genetically encoded fluorescent probes, as well as ESR spin trapping technique. We also discuss the advantages, limitations, and recent technical advances of each individual technique as well as their applications in pulmonary vasculature studies. We believe that technical advance in the detection of ROS will provide us with a better understanding on how to maintain normal pulmonary vasculature functions under oxidative stress.