Inflammatory bowel disease (IBD), encompassing Crohn's disease and ulcerative colitis, is a chronic autoimmune disorder affecting the entire gastrointestinal tract. Current treatments offer only transient symptom relief, failing to address underlying intestinal barrier disruption and microbiota imbalance. This can lead to severe complications like intestinal fibrosis, characterized by thickened intestinal walls and reduced peristalsis, potentially resulting in life-threatening intestinal obstruction or colorectal cancer. The generation of excess reactive oxygen and nitrogen species (RONS) by activated immune cells contributes to oxidative stress and damages the intestinal barrier. Nanozymes, with their ability to scavenge RONS, are promising therapeutic candidates, but suffer from instability in the acidic gastric environment and short gastrointestinal residence time. Modulating the gut microbiota is another therapeutic strategy, but existing treatments rarely address both IBD and intestinal fibrosis simultaneously. Therefore, a biocompatible platform capable of regulating oxidative stress, balancing gut microbiota, repairing intestinal barriers, and prolonging therapeutic agent retention is urgently needed.

Existing literature highlights the role of oxidative stress and gut microbiota dysbiosis in IBD pathogenesis. Various nanozymes have been explored for IBD treatment, but many are metal-based, unstable in the stomach, and exhibit potential toxicity. The short gastrointestinal residence time of orally administered nanozymes necessitates repeated administration. While modulating the gut microbiota via probiotics or other means has shown promise, most treatments focus on IBD alone, neglecting the fibrotic complication. Hydrogels offer potential as drug delivery systems but often have limitations like poor stability at low pH and complex synthesis. Inulin, a polysaccharide fiber, has shown probiotic benefits and potential in treating various disorders, including IBD.

This study developed a ternary inulin hydrogel (PPy/PFD@Inulin gel) containing polypyrrole (PPy) nanozymes and pirfenidone (PFD). PPy nanozymes were synthesized via aqueous dispersion polymerization. The PPy nanozymes and PFD were incorporated into a heated inulin solution, forming a gel upon cooling. The gel's properties, including stability under varying pH conditions, injectability, and rheological behavior, were characterized using techniques such as TEM, SEM, FTIR, and rheometry. In vitro RONS scavenging capacity was assessed using DPPH, ABTS, O2−, and H2O2 assays. Cytotoxicity and biocompatibility were evaluated using NCM460 cells and a hemolysis test. In vivo biocompatibility and biodistribution were studied in mice. The effects of the gel on intestinal physiology, including gut microbiota composition, were examined using 16S rRNA gene sequencing. Prophylactic and therapeutic efficacy against DSS-induced colitis and intestinal fibrosis were evaluated in mouse models. Intestinal permeability was assessed using FITC-dextran. The expression of tight junction proteins (ZO-1 and occludin) was analyzed using western blotting and immunofluorescence. The antifibrotic mechanism was investigated using human colon fibroblasts (CCD-18Co) and western blotting to analyze fibrosis-related proteins (α-SMA and TGF-β1). RNA sequencing was used to analyze gene expression changes in colon tissues.

The PPy/PFD@Inulin gel exhibited excellent stability across a range of pH levels, good injectability, and sustained release of PPy nanozymes and PFD. PPy nanozymes demonstrated superior RONS scavenging capacity in vitro and in vivo. The gel exhibited excellent biocompatibility in vitro and in vivo. Oral administration of the gel modulated the gut microbiota, increasing the abundance of beneficial bacteria like *Akkermansia*. In both prophylactic and delayed treatment models of DSS-induced colitis, the PPy/PFD@Inulin gel significantly improved body weight, colon length, and reduced inflammatory markers (MPO, TNF-α, IL-1β, IL-6). The gel also enhanced intestinal barrier function by upregulating ZO-1 and occludin expression. In a chronic colitis model, the gel effectively reduced intestinal fibrosis by inhibiting fibroblast proliferation and attenuating the TGF-β/Smad signaling pathway. RNA sequencing revealed that the gel modulated gene expression related to inflammatory and immune responses.

The findings demonstrate that the PPy/PFD@Inulin gel effectively addresses multiple aspects of IBD and its fibrotic complication. The sustained release of PPy nanozymes and PFD, enabled by the inulin hydrogel, contributes to its therapeutic efficacy. The combination of RONS scavenging, gut microbiota modulation, and antifibrotic effects makes this a promising therapeutic approach. The results from RNA sequencing support the observed improvements in inflammation and immune response. The study's strength lies in its multi-pronged approach targeting various mechanisms involved in IBD and fibrosis. The use of mouse models provides valuable in vivo data, although further research in larger animal models and clinical trials is warranted.

This study successfully developed a ternary PPy/PFD@Inulin gel with excellent biocompatibility and therapeutic efficacy in treating IBD and its fibrotic complication. The gel's multi-functional properties, ease of preparation, and potential for scalability make it a promising therapeutic modality. Future research should focus on optimizing the drug release profile and exploring functional modifications to enhance targeting specificity. Clinical trials are needed to evaluate the gel's safety and efficacy in human patients.

The study used mouse models, and the results may not fully translate to humans. The sample sizes in some experiments were relatively small. Long-term effects of the gel were not extensively evaluated. Further research is necessary to investigate the optimal dosage and duration of treatment for different patient populations.