Novel Model System to Optically Quantify Mitochondrial Metabolic Activity

Abstract

Photobiomodulation is a therapeutic technique that has been shown to aid in bone regeneration, traumatic brain injury, and wound care, among others. The main hallmark of photobiomodulation is that it uses a low-energy light in the red or NIR range to impart its benefits. Despite this technique’s wide range of improvements, it is currently unknown how they occur. The leading hypothesis as to why photobiomodulation causes benefits is that the light is absorbed by a protein within the mitochondria, Complex IV, the terminal protein of the electron transport chain. When Complex IV absorbs red or NIR light, it is believed that it increases its rate of activity. This increases the rate at which protons are pumped into the intermembrane space, which in turn allows ATP-synthase to generate ATP at a faster rate. To determine the underlying mechanisms of PBM, I isolated mitochondria and irradiated them with PBM levels of light. I then measured changes in mitochondrial activity by tracking changes in the redox state of cytochrome c with the use of resonance Raman spectroscopy. By individually inhibiting each enzyme within the electron transport chain, I was able to determine how light interacts with each specific protein. Using this technique, I was able to determine the effects that the green light from the resonance Raman laser had on mitochondria. This technique can then be used to determine the effects of PBM on isolated mitochondria at various wavelengths and powers.