An Oxygen Equilibrium at the Host-Microbial Interface Determined by Phosphorescent Nanoprobe Technology

Abstract

Although the colonic lumen is known to be largely devoid of oxygen, the mechanism by which this anaerobic environment is maintained remains unclear due to the inability of available technology to quantify oxygen in the intestinal tract. Indeed, most microbes in the gut are obligate anaerobes. Accurate quantification of oxygen in the gut is essential to elucidate the host’s relationship with its densely populated intestinal microbial community and may be relevant to various disease states in humans, including inflammatory bowel disease (IBD). For instance, multiple studies have demonstrated significant alterations in the gut microbiota in IBD, notably phylum level increases in Proteobacteria and Actinobacteria, which contain predominantly oxygen tolerant organisms, and a decrease in oxygen-sensitive Firmicutes. Phosphorescent nanoprobe (Oxyphor G41), whose phosphorescence decay time is exquisitely sensitive to environmental oxygen, was administered to mice either via the drinking water to measure intestinal luminal oxygen or intravascularly via the tail vein to quantify tissue oxygen. A fiber-optic time domain phosphorometer2 was used to excite G4 and detect phosphorescence at λmax = 635 nm and λmax = 810 nm, respectively. A laparotomy was performed under isoflurane anesthesia and the decay pattern of G4 phosphorescence was measured in various organs. 100% oxygen was delivered via the anesthesia apparatus for several minutes to determine the effect of host oxygenation on luminal oxygen content. Intravenous injection of G4 led to signal detection in all tissues, but ingested G4 was only detected at intestinal segments containing fecal material. The partial pressure of oxygen (pO2) was remarkably lower in the cecal feces than in the adjacent tissue. Inspiration of pure O2 led to a rapid, dramatic increase in cecal tissue pO2 and a delayed, more gradual increase in luminal pO2 (Figure 1). Both effects were reversible once animals were returned to room air. This technical advance has revealed a novel complexity in hostmicrobial interactions - namely that oxygenation of host colonic tissue affects the level of oxygen in the feces. Thus, the anaerobic nature of the intestinal lumen is not static but rather a dynamic equilibrium. We hypothesize that oxygen released by colonic tissue is consumed by the gut microbiota, thus creating an anaerobic luminal environment. This concept is supported by our data in human subjects indicating enrichment of aerobic and facultative anaerobic bacteria adherent to the rectal mucosa compared to the feces where obligate anaerobic bacteria predominated (Figure 2). We believe that the colonic oxygen equilibrium may have great relevance to IBD where the dysbiotic microbiota composition may, in part, result from an alteration in the redox potential of the colonic environment due to hyperemia, bleeding, and/or the oxidative nature of the inflammatory response. Studies are underway to further characterize the relationship between oxygen levels in the gut and the “dysbiotic” microbiome in IBD.