Non-Invasive Mitochondrial Modulation

Recent studies from the Sanderson and Hüttemann laboratories led to the development of a novel, non-invasive therapeutic strategy to target ischemic brain injury. The foundation of our technology is based on our research discovery that discrete near infrared (NIR) light wavelengths directly modulate mitochondrial function in the brain by changing the activity of the mitochondrial enzyme Cytochrome c Oxidase (COX).

(A) Isolated regulatory-competent bovine COX separated into its subunits on a high-resolution urea/SDS-PAGE Coomassie-stained gel. Subunits are indicated in roman numerals. (B) Representative scan of wavelength-dependent COX activity identifying the 750 nm and 950 nm wavelength as inhibitory regions. (C) Effect of NIR emitted by LED diodes confirms that 750 nm and 950 nm NIR inhibit COX in vitro while 810 nm NIR activates the enzyme. Data were obtained over a 3-min interval of irradiation and normalized to non-irradiated samples (n=4; *p<0.05). (D) NIR irradiation modulates oxygen consumption rate (OCR) in a wavelength specific manner. 750 nm and 950 nm NIR reduce OCR below the basal respiration rate whereas 810 nm NIR increases mitochondrial OCR (n=4; *p<0.05).

The neuroprotective effect of NIR was first tested in our adult rat model of global brain ischemia (bilateral carotid-occlusion + systemic hypotension). Following transient global brain ischemia in rats, hippocampal CA1 neurons are particularly sensitive, and a near-complete loss is observed at 3-7 days after reperfusion . Our studies defined the neuroprotective effect of NIR in terms of its ability to prevent degeneration of CA1 neurons (A & B) and to preserve hippocampal function. Non-invasive irradiation of the brain was performed by direct illumination through the scalp and skull applied at the onset of reperfusion and maintained for 2h. All wavelengths or wavelength combinations identified in vitro to reduce COX activity were evaluated for neuroprotection using a randomized and blinded study design. After 14 days, brain sections were stained for neuron counting, together with immuno-fluorescence to detect neurons, microglia, and astrocytes (A & B). Untreated animals subjected to I/R showed an 86% loss of neurons in the CA1 hippocampal region. In contrast, in rats treated with 950 nm or 750 nm NIR, neuronal death was profoundly attenuated: 83% of hippocampal neurons remained viable with 950 nm NIR vs 14% in untreated controls (B) . For the inhibitory NIR-treated groups, % viable neurons ranged from 83% to 65% depending on the wavelength(s) that were applied. We identified: (i) two wavelength/combinations that show maximum and statistically equivalent neuroprotection: 950 nm, and 750/950 nm (83%±10% and 77%±15% viable neurons, respectively); and (ii) one wavelength (810 nm) that provided no protection. Importantly, the protection in NIR treated rats coincided with improved spatial learning (Sanderson et al, Sci Rep, 2018).