Intestinal oxygen and microbiota crosstalk: implications for pathogenesis of gastrointestinal diseases and emerging therapeutic strategies

The advent of high-throughput sequencing in the 21 st century has significantly advanced human microbiome studies, particularly through the use of culture-independent methods to investigate host-associated microbial communities [1]. These studies have shown that microbiome imbalance, or dysbiosis, is associated with various diseases. As the largest microbial community in the body, the colonic microbiota produces metabolites that influence health. Thus, understanding the mechanisms that maintain gut homeostasis and the factors leading to its disruption during dysbiosis is crucial for addressing key questions in human microbiome research [2].

Over the past few decades, the focus of gut microbiota research has continued to evolve. Early studies on invertebrates identified abundant core microbial species, suggesting that defining a healthy gut microbiota might require identifying common core species. However, significant individual differences in human fecal microbiota made it impractical to define gut homeostasis and dysbiosis based on specific core species [3]. Moreover, even with large-scale data analysis from projects like the Human Microbiome Project [4], it remains unclear which features define gut homeostasis or its disruption [5]. These challenges indicate that high-throughput sequence analysis does not provide a straightforward path for defining healthy microbial communities.

Recently, it has been proposed that the host environment controls the healthy microbial community, offering a new approach to quantifying homeostasis [6]. According to this perspective, host-controlled environmental parameters are crucial for microbial growth on body surfaces. For example, oxygen levels in the intestinal lumen are a key driver of dysbiosis. Under physiological conditions, the intestinal lumen of mammals is hypoxic, which is associated with the intestinal epithelium, intestinal blood flow, and intestinal microbes [7]. This hypoxic gradient shapes the gut microbiota, promoting stable colonization by obligate anaerobes [8]. This shift in our understanding of dysbiosis provides a novel starting point for therapeutic strategies to restore microbiome health, suggesting that microbiome homeostasis can be assessed by measuring host functions, such as whether oxygen concentrations along the intestinal axis are within the normal range [9].

Herein, we summarize the role of intestinal oxygen (IO) in modulating host-microbe interactions, with emphasis on the mechanisms through which host IO levels shape microbial homeostasis. While other host factors, such as luminal pH and bile acids, also influence the gut ecosystem, IO remains a central regulator of microbial metabolism and spatial organization. We outline how altered IO reshapes microbial communities, drives metabolic defects, and contributes to intestinal disease. Distinct from previous reviews, we integrate recent advances in epithelial metabolism, HIF signaling, and oxygen regulation to provide a unified framework linking IO homeostasis (IOH), microbial ecology, and disease. Finally, we highlight emerging therapeutic strategies targeting different mechanisms to restore IOH, emphasizing their attractive translational potential.

Of particular recent interest is the interplay between tissue oxygenation and the microbiota. Previous studies have established that the composition of the intestinal microbiome varies along the longitudinal axis of the gut and is related to the radial oxygen gradient distribution. Albenberg et al. [30] used phosphorescence quenching methods to directly demonstrate that host oxygenation affects intestinal lumen oxygenation. When the oxygenation of the host increases, intestinal lumen oxygenation also increases, indicating that oxygen diffuses from host tissues into the intestinal lumen to form a spatial oxygen gradient. Furthermore, after the host tissues recover their physiological state, the pO₂ in the lumen decreases, and the abundance of aerobic microbes adhering to the mucosal surface increases. In summary, these phenomena not only demonstrate the close relationship between the host and the intestinal microbiome but also suggest that both are involved in regulating the local oxygen environment in the gut.

The intestinal bacteria use different energy metabolic pathways in aerobic and anaerobic conditions. In aerobic conditions, bacteria obtain energy through redox reactions. During this process, oxygen acts as an exogenous electron acceptor, substrates undergo phosphorylation or oxidative phosphorylation and ATP (adenosine triphosphate) is produced [31]. In anaerobic conditions, bacteria do not have exogenous electron acceptors and cannot complete the TCA cycle [32], mainly carrying out fermentation, releasing less energy than in aerobic conditions. During this process, pyruvate is converted into acetyl-CoA and formate by pyruvate methylhydroxylase, and then metabolized into CO₂ and H₂ by formate dehydrogenase. In summary, microorganisms that can maximize energy production will dominate the microbial community in which they live.

Furthermore, nitrate and oxygen serve as critical electron acceptors, enabling the host to precisely modulate the composition of the dominant bacterial community within the intestine by controlling the availability of these electron acceptors. For instance, in the hypoxic environment of the colonic lumen (~ 0.6% O₂), obligate anaerobic primary fermenters predominantly utilize endogenous electron acceptors for their metabolic processes. These strict anaerobes, primarily represented by taxa within the classes Bacteroidia (phylum Bacteroidetes) and Clostridia (phylum Firmicutes), form the backbone of the high-density microbial community in the colon [33]. Conversely, the ileum exhibits a slightly elevated average oxygen content compared to the colon (~ 1% O₂), which correlates with an increased abundance of facultative anaerobic bacteria within its microbial community [34]. When oxygen becomes limiting, these bacteria switch to alternative electron acceptors such as nitrate to sustain respiratory metabolism [35].

Nitrate in the ileal lumen is mainly generated by NADPH oxidase 1 (NOX1) and inducible nitric oxide synthase (iNOS), which oxidize luminal or intracellular nitrogen compounds [28, 36]. During intestinal homeostasis, the average luminal nitrate concentration in the ileum of mice has been measured at approximately 6 mM [37]. In mice deficient in iNOS and NOX1 synthesis, a notable increase in the abundance of strictly anaerobic bacteria within the ileal microbiota has been observed, leading to a microbial profile that closely resembles that of the colonic microbiome [28]. Additionally, diet strongly modulates nitrate dynamics. High-fat or low-fiber diets can upregulate epithelial iNOS, elevate luminal nitrate, and favor Proteobacteria expansion, whereas fiber-rich diets enhance butyrate production, which suppresses nitrate synthesis via PPAR-γ activation [38].

Beyond oxygen and nitrate, some inflammatory response products such as S-oxide, N-oxide, and formate also compete for bacterial respiration under microaerobic or inflamed conditions. For example, Enterobacteriaceae can use these byproducts to support their Dimethyl S-oxide respiration, trimethylamine N-oxide (TMAO) respiration and formate oxidation, providing a metabolic advantage when oxygen and nitrate are depleted [39]. While Bacteroidetes and Firmicutes bacteria lack the ability to utilize these by-products, which puts their growth at a disadvantage and ultimately leads to gut dysbiosis. Therefore, the balance among these alternative respiratory substrates plays a pivotal role in defining microbial community structure and metabolic output along the gastrointestinal tract. Figure 2 illustrates the interaction between gut microbiota and IOH.

As previously discussed, many human illnesses occur when the host is unable to properly regulate the IO microenvironment, including ulcerative colitis, colorectal cancer, graft-versus-host disease (GVHD) and irritable bowel syndrome (IBS). These diseases are all marked by dysbiosis, linked to an increased abundance of facultatively anaerobic bacteria in the colonic microbiota. Such organisms can use oxygen or nitrate as respiratory electron acceptors, thereby fueling their growth. Here, we aim to outline the major roles of pathological hyperoxia conditions of gut in the onset and progression of diseases. Impact of pathological hyperoxia on gut diseases is listed in Table 2.

Building on prior discussions, the concept of host-microbiota interactions provides a framework for developing therapeutic strategies that modulate IOH to correct dysbiosis. In the following section, we review recent research on whether the adverse effects of dysbiosis can be mitigated by either inhibiting microbial respiratory pathways that drive community shifts or by enhancing host functions that restrict the availability of oxygen and nitrate in the gut lumen.

Among these strategies, pharmacological approaches have been most extensively investigated. Disruptions in colonic microbiota balance are frequently initiated by compromised mitochondrial function within the colonic epithelium, which perturbs local hypoxic conditions and luminal anaerobiosis [65]. Emerging evidence indicates that drugs that revive mitochondrial bioenergetics in the colonic epithelium can be used to restore gut homeostasis [66]. A key player in enhancing mitochondrial activity within the colonic epithelium is the nuclear receptor peroxisome proliferator-activated receptor gamma (PPAR-γ) [67]. Administration of PPAR-γ agonists, such as 5-aminosalicylic acid (5-ASA), has been shown to bolster mitochondrial efficiency specifically in colonic epithelial cells [45]. Both in patients and mice models of UC, treatment with 5-ASA normalizes the composition of the colonic microbiota by controlling the proliferation of facultative anaerobic bacteria [44, 68]. Additionally, in vitro studies have shown that activation of SIRTUIN 1 can enhance mitochondrial bioenergetics while concurrently lowering levels of circulating trimethylamine N-oxide (TMAO) [69‐71]. These findings suggest that restoring epithelial hypoxia may also reduce the production of harmful microbial metabolites. However, these strategies were not initially designed to target IO regulation. Notably, the observed increase in facultative anaerobes and elevated levels of the luminal oxidative metabolite TMAO suggest that the beneficial effects of these interventions cannot be fully attributed to secondary anti-inflammatory actions. Rather, these findings imply that restoration of epithelial hypoxia may play a direct role in re-establishing IOH and in suppressing the generation of detrimental microbial metabolites.

Another pharmacological approach focuses on inhibiting microbial respiratory pathways. Sodium tungstate (Na₂WO₄) can limit the proliferation of facultative anaerobic bacteria by substituting for molybdenum in key respiratory enzymes [72], thereby preventing their overgrowth and improving IOH-related conditions. For instance, in mouse models of colorectal cancer, sodium tungstate treatment has been shown to inhibit the proliferation of colibactin-producing E. coli, leading to a reduction in polyp formation [57]. In UC models, tungstate selectively restricts Gammaproteobacteria expansion, reducing inflammation [73]. Additionally, limiting the availability of host-derived nitrate in the colonic lumen can prevent the production of harmful metabolites during gut dysbiosis. Aminoguanidine [71], a chemical inhibitor of iNOS, lowers serum TMAO levels in mice fed a high-fat diet with choline by blocking nitrate production. Targeting microbial metabolic pathways therefore represents a complementary strategy to restore intestinal ecological balance.

Finally, dietary interventions or oral probiotics have also been reported to hold great promise for IOH-associated GI disorders. Studies have shown that high-fat, low-fiber diets disrupt the host’s control over gut microbes [71, 74, 75], suggesting that dietary changes could help improve dysbiosis. Zhang et al. reported that supplementing broiler diets with yeast polysaccharides (YPS) increased jejunal villus height and enhanced antioxidant and barrier gene expression, thereby protecting against mycotoxin-induced injury [76]. Probiotic therapy also holds great promise for improving dysregulated gut microbiota. Yang et al. [77]. aggregated Synechocystis sp. PCC6803 (Sp) with Bacillus subtilis (BS) by biomimetic mineralization to form cyanobacteria-probiotics symbionts to efficiently regulate the gut microbiota and reshape the intestinal barrier functions in a murine model of acute colitis. Importantly, probiotics within the symbionts created a local anaerobic environment to activate the [NiFe]-hydrogenase enzyme of cyanobacteria, facilitating the production of H₂ to persistently scavenge elevated reactive oxygen species and alleviate inflammatory factors. Although promising, these approaches remain largely preclinical and require further evaluation in human settings.

Different therapeutic strategies for modulating intestinal oxygen homeostasis vary widely in their levels of supporting evidence and translational readiness. Pharmacological interventions include approved agents with established safety profiles, such as 5-ASA. Although these drugs were not originally developed to target intestinal oxygen homeostasis, some clinical evidence supports their use in relevant contexts. Further clinical validation is needed for all strategies, with particular attention to safety, targeting, and long-term durability. In contrast, most dietary and probiotic interventions are supported mainly by preclinical evidence from animal models. To date, no clinical trials have directly evaluated probiotics or engineered microbial consortia for modulating intestinal oxygen homeostasis in humans. These approaches also face challenges such as uncertain colonization dynamics, host specificity, biosafety concerns, and regulatory barriers related to engineered strains. Nevertheless, regulating the microbiota through host-targeted approaches remains a promising direction because gut dysbiosis is closely linked to a broad spectrum of noncommunicable diseases. Table 3 summarizes potential strategies for reducing abnormal intestinal oxygen concentrations.

Building on the preceding sections, growing evidence underscores the therapeutic potential of targeting IOH. However, several methodological and translational limitations continue to constrain progress. Establishing a physiological range of intestinal oxygen in healthy individuals would greatly aid the diagnosis of dysbiosis, yet such thresholds remain undefined. Current measurement techniques vary widely in spatial resolution and invasiveness, providing only partial or static information. Electrode- and probe-based methods detect local pO₂ but fail to capture luminal heterogeneity [13, 14], whereas chemical markers visualize hypoxia ex vivo without temporal precision [78‐80]. Moreover, gut factors such as enzymatic activity, bile acids, and dietary components can interfere with oxygen sensing, further complicating standardization [7]. The characteristics and potential clinical relevance of these approaches are summarized in Table 4.

Translational evidence in humans also remains limited. Most mechanistic insights into the relationship between epithelial bioenergetics, oxygen leakage, and microbial dysbiosis arise from animal models, leaving their human relevance uncertain. Quantitative human data correlating mucosal oxygen gradients with epithelial metabolism and microbial respiratory activity remain scarce. In addition, therapeutic strategies that modulate intestinal oxygen—such as PPAR-γ agonists, sodium tungstate, iNOS inhibitors, or engineered probiotics—require further validation regarding safety, tissue targeting, and durability of effect. Despite these challenges, IO regulation remains a promising direction for microbiome-based therapeutics. Metagenomic markers that track microbial respiratory gene abundance in feces may offer valuable non-invasive insights into IOH status and guide the development of oxygen-informed treatment strategies [52, 81].

Intestinal oxygen serves as a central determinant of gut microbial composition and metabolic function. Disruption of IOH alters microbial respiration and redox balance, driving dysbiosis and promoting gastrointestinal pathologies. Although growing evidence underscores the therapeutic potential of targeting IOH, several barriers still hinder its clinical translation, including the lack of standardized, non-invasive methods to quantify IO and the limited causal understanding of host-microbe oxygen interactions in humans. Despite these challenges, preclinical studies consistently demonstrate that restoring IOH can re-establish microbial balance and improve mucosal resilience. Pharmacological, dietary, and probiotic strategies that modulate oxygen utilization or promote local anaerobiosis have shown encouraging results, underscoring the feasibility of oxygen-informed interventions. Future efforts should focus on establishing robust IO metrics, integrating redox readouts with multi-omics biomarkers, and validating targeted oxygen-based therapies in clinical settings to translate experimental insights into practical treatment strategies.