Sprayable dynamic Schiff-base hydrogel engineers an oxygen-limited microenvironment for viable Bifidobacterium bifidum delivery and accelerated oral ulcer healing.

Oral ulcers affect a substantial portion of the global population, and current therapies face challenges including drug resistance and treatment dependency.

• Oral ulcers affect over 25% of the global population.
• Current therapeutic approaches are limited by drug resistance and treatment dependency.
• Local delivery of anaerobic strains such as Bifidobacterium bifidum remains challenging because oxygen exposure rapidly reduces bacterial viability.

The sprayable dynamic Schiff-base gelatin/cellulose hydrogel formed via reversible imine crosslinking undergoes in situ gelation and creates an oxygen-limited, moisture-retaining niche.

• The hydrogel is based on reversible imine (Schiff-base) crosslinking between gelatin and cellulose components.
• The network was described as capable of in situ gelation upon spraying onto mucosal surfaces.
• The crosslinking network was characterized as providing deformation tolerance and uniform mucosal adhesion.
• The oxygen-limited microenvironment engineered by the hydrogel was central to preserving anaerobic bacterial viability.

B. bifidum embedded in the hydrogel exhibited sustained release over 2 hours and maintained high activity throughout gelation and degradation.

• Bacterial release was sustained over a 2-hour period.
• High bacterial activity was maintained throughout both the gelation process and subsequent degradation of the hydrogel.
• The oxygen-limited environment created by the hydrogel was credited with preserving bacterial viability during delivery.

In vitro, the hydrogel and B. bifidum synergistically enhanced fibroblast migration.

• Fibroblast migration was enhanced when cells were treated with the B. bifidum-loaded hydrogel compared to controls.
• The effect was described as synergistic between the hydrogel material and the probiotic bacteria.
• This finding was observed in in vitro cell migration assays.

The B. bifidum-loaded hydrogel reduced LPS-induced pro-inflammatory cytokine expression in vitro.

• TNF-α and IL-6 expression were reduced in LPS-stimulated cells treated with the hydrogel and B. bifidum.
• The reduction in inflammatory markers was observed in vitro.
• The synergistic action of the hydrogel and B. bifidum was responsible for the anti-inflammatory effect.

The B. bifidum-loaded hydrogel promoted macrophage polarization toward the M2 phenotype in vitro.

• Macrophage polarization toward the anti-inflammatory M2 phenotype was promoted by the combination of hydrogel and B. bifidum.
• This shift in macrophage phenotype was observed in vitro.
• M2 polarization is associated with pro-healing and anti-inflammatory responses.

In vivo, the B. bifidum-loaded hydrogel markedly accelerated oral ulcer closure and improved tissue regeneration markers.

• The B. bifidum-loaded hydrogel markedly accelerated oral ulcer closure in an in vivo model.
• Improved epithelial regeneration was observed histologically.
• Increased collagen deposition was detected in treated ulcers.
• Elevated expression of α-SMA and collagen I was observed in the treated groups.

In vivo cytokine profiling confirmed a transition toward a pro-healing microenvironment in ulcers treated with the B. bifidum-loaded hydrogel.

• TNF-α and IL-6 levels were decreased in treated ulcers.
• IL-10 levels were increased in treated ulcers, indicating a pro-healing cytokine profile.
• The cytokine profile transition was confirmed through profiling analysis in vivo.

No systemic toxicity was observed with the B. bifidum-loaded hydrogel treatment.

• Systemic toxicity was assessed and no adverse systemic effects were detected.
• The absence of systemic toxicity supports the translational potential of the platform.
• Safety was evaluated as part of the in vivo study.