An injectable nanofiber hydrogel composite loaded exosome is effective in a Crohn’s disease perianal fistula model

Introduction: Perianal fistulas represent an aggressive and disabling complication of Crohn’s disease (CD). Despite the introduction of advanced anti-inflammatory biologic and small molecule therapeutics, novel surgical approaches as well as multidisciplinary treatment approaches, the outcome is dismal with less than 50% durable fistula resolution. Adipose Derived Stem Cell (ADSCs) therapy has been met with variable success. In some studies, ADSC based therapy has been successful, but the recent Phase 3 ADMIRE-CD II study for complex Crohn’s Perianal Fistulas (CPF) in 2023 did not meet its primary endpoint of combined remission at 24 weeks. The failure of this trial may be due, at least in part, to the migration of ADSCs away from the site of injection. The beneficial effects of ADSCs are therefore temporally limited. One modality to prevent the migration of ADSC is the utilization of scaffolds or other biomaterials to ensure a durable presence of ADSC close to fistula tracks. Exosomes, as a major component of paracrine signaling responsible for beneficial effects of ADSC, have recently been proposed as an acellular and equally efficacious replacement for ADSCs. In addition, compared to ADSCs, exosomes display more stable membrane structures, and lower immunogenicity. An ideal fistula repair modality would include ADSC-derived exosomes delivered with a biomaterial that would retain the exosomes in the fistula track. In this study, we reported an injectable, biodegraded nanofiber-hydrogel composite (NHC) loaded with ADSC-derived exosomes (exo-NHC) utilized for CD fistula therapy.

Materials and

Methods: NHC was formulated as described by us previously. ADSCs-derived exosomes were collected from passage 4-6 rat ADSCs cell culture medium via ultracentrifugation. A rat model of Crohn’s disease perianal fistula was established as we described before. In brief, TNBS was given via enema twice a week for one week to induce proctitis. Then, a 14-gauge needle was used to create 2 perianal fistulas (at 3 and 9 o’clock of anal, respectively). Vascular loop setons were then inserted into fistula tract, and TNBS was instilled into the fistula tracts as well as anus twice a week for a total of one month to maintain chronic inflammation. Rats were then randomized to receive one of the following 4 treatments: (1) standard fistula surgery (brushing of fistula tract followed by closure of internal orifice); (2) standard fistula surgery plus umbilical cord derived tissue grafts (Stravix) placed into fistula tract, (3) standard fistula surgery plus NHC placed into fistula tract, (4) standard fistula surgery plus NHC+ADSCs placed into fistula tract and (5) standard fistula surgery plus (exo-NHC) placed into fistula tract. At day 20 after treatment, animals were subjected to MRI scanning to assess the healing of fistula. Animals were then euthanized and tissue was collected for further analysis.

Results, Conclusions, and Discussions: MRI images showed that exo-NHC treatment group reach the best fistula healing when compared with to the other 4 treatment approaches. 4 out 5 fistulas achieve completing healing, and 1/5 displayed 90% healing at 20 days. Mechanistic studies showed that exo-NHC was able to inhibit inflammation within fistula. In addition, exosomes promoted tissue healing or regeneration via polarization of macrophages and promotion of new vessel formation.

Conclusions: This study demonstrated that exo-NHC platform is a promising approach for perianal fistula treatment in Crohn’s disease.

Acknowledgements (Optional): This work was financially supported by the Helmsley Foundation, and NIH.