Degassing a Decellularized Scaffold Enhances Wound Healing and Reduces Fibrosis during Tracheal Defect Reconstruction: A Preliminary Animal Study

Abstract

Few efforts have been made regarding the optimization of porcine small intestinal submucosa (SIS) to improve its biocompatibility. This study aims to evaluate the effect of SIS degassing on the promotion of cell attachment and wound healing. The degassed SIS was evaluated in vitro and in vivo, compared with the nondegassed SIS control. In the cell sheet reattachment model, the reattached cell sheet coverage was significantly higher in the degassed SIS group than in the nondegassed group. Cell sheet viability was also significantly higher in the SIS group than in the control group. In vivo studies showed that the tracheal defect repaired by the degassed SIS patch showed enhanced healing and reductions in fibrosis and luminal stenosis compared to the nondegassed SIS control group, with the thickness of the transplanted grafts in the degassed SIS group significantly lower than those in the control group (346.82 ± 28.02 µm vs. 771.29 ± 20.41 µm, p < 0.05). Degassing the SIS mesh significantly promoted cell sheet attachment and wound healing by reducing luminal fibrosis and stenosis compared to the nondegassed control SIS. The results suggest that the degassing processing might be a simple and effective way to improve the biocompatibility of SIS.

Keywords: degas; small intestinal submucosa; tissue engineering; tracheal patch model.

Figure 1

Flow diagram of the research design (A). Schematic diagram of the vacuum system (B).

Figure 2

Reattachment surface area analysis of the degassed SIS mesh. The area of reattached cell sheets in the degassed group was 34.57 ± 11.8%, which was significantly higher than that in the nontreated group (16.72 ± 3.8%, * p < 0.05).

Figure 3

Viability of the reattached cells in the degassed SIS mesh. The optical density (OD) measured by the ELISA reader in the degassed group was 0.363 ± 0.116, which was significantly higher than that in the nontreated group (0.228 ± 0.072, *** p < 0.001).

Figure 4

Histological comparison of the cell sheet reattached to the degassed SIS mesh. The cell sheet that reattached to the degassed SIS adhered to the surface very well. (A) No voids were observed between the two layers compare to the nondegassed control (B).

Figure 5

In vivo evaluation of the degassed SIS mesh in a trachea patch repair model: (A) H&E staining: the nondegassed control group showed dense fibrosis with high neovascularization (black arrow). (B) H&E staining: the DSA-treated experimental group showed significant epithelial regeneration (white arrow) and less fibrosis formation around the transplanted sheet (yellow arrow).

Figure 6

Image analysis of the measured thickness of the mucosal layer of each implanted graft. There was a significant reduction in the thickness of the transplanted degassed SIS mesh graft compared with and the nondegassed SIS mesh (346.82 ± 28.02 µm vs. 771.29 ± 20.41 µm, respectively; **** p < 0.0001).

Figure 7

Demonstrations of SIS degassing with blood. (A) Microscopically, the red blood cells become interlaced with the SIS interfiber spaces after degassing. (B) When blood was poured on the SIS surface, the red blood cells aggregated on only the SIS surface and did not penetrate the SIS matrix after immersion for more than 30 min.