J. E. Griepentrog1, A. J. Lewis1, X. Zhang1, J. S. Lee2, M. R. Rosengart1 2University Of Pittsburgh,Department Of Medicine, Division Of Pulmonary, Allergy And Critical Care Medicine,Pittsburgh, PA, USA 1University Of Pittsburgh,Department Of Surgery,Pittsburgh, PA, USA
Introduction:
Pneumonia is a major cause of sepsis and acute lung injury, with three million cases reported annually in the US; it is the sixth most common cause of death. The causal mechanisms of acute alveolar damage include an excessive accumulation of neutrophils in the lung and the subsequent production of inflammatory mediators and neutrophil-mediated oxidant damage. Light has been shown to influence an organism’s response to stress. Circadian proteins, including REV-ERB alpha, are known to attenuate inflammation and regulate mitochondrial biogenesis, and modulating them may enable us to modify the response to sepsis. Our previous animal work demonstrated that blue light enhances bacterial clearance and improves survival during pneumonia. However, the cellular and neurophysiologic mechanisms by which blue light harnesses circadian biology to alter macrophage phagocytic function are unknown.
Methods:
Male C57BL/6 mice were intratracheally administered Klebsiella pneumoniae (6000 CFUs) and immediately exposed to an initial 36 hours followed by 12 hours daily of bright (1400 lux) blue (442nm) or red (617nm), or ambient white fluorescent (400 lux) light. Some mice underwent splenectomy or exposure to the α7 nicotinic AChR antagonist, alpha-bungarotoxin. After 72 hours, mice were euthanized, the lungs lavaged, and the blood/organs harvested. Tissue and blood were analyzed for bacterial colony forming units (CFUs). Lung, spleen, and alveolar macrophages expression of REV-ERB alpha was analyzed by immunofluorescence and immunoblot. Cytokine and BAL protein concentrations were quantified by ELISA and BCA. Statistical analysis was performed by Wilcoxon rank-sum tests at α = 0.05.
Results:
By contrast to either red or ambient white light, blue light reduced lung tissue bacteria (p = 0.06), bacteremia (p = 0.10), and bacterial dissemination into tissues and improved survival (log-rank p = 0.02). Blue light attenuated alveolar neutrophil influx (p = 0.0001), yet preserved the alveolar macrophage population. Either inhibiting cholinergic tone with alpha-bungarotoxin or splenectomy eliminated the protective effects of blue light: Blue vs. Blue with bungarotoxin, lung CFUs (p = 0.004), blood CFUs (p = 0.02), neutrophil influx (p = 0.002). Lung tissue and alveolar macrophage expression of REV-ERB alpha was increased after blue light.
Conclusion:
Blue light functions through a cholinergic pathway and the spleen to augment lung innate immunity and bacterial clearance, while reducing neutrophilic inflammation. The mechanisms appear to involve an augmentation in REV-ERB alpha expression in alveolar macrophages and lung tissue. Further studies are required to determine how altering REV-ERB alpha affects macrophage phagocytic function.