[Porcine uterine decellularized extracellular matrix hydrogels and engineered microarrays regulate porcine...

The porcine uterine decellularized extracellular matrix (DECM) hydrogel represents a promising candidate for endometrial tissue engineering. However, conventional preparation methods face challenges in balancing decellularization efficiency and bioactive extracellular matrix (ECM) preservation. To investigate the regulatory mechanisms of the biochemical and biomechanical properties of the DECM hydrogel on porcine endometrial stromal cells (pESCs), while addressing the inherent conflict between decellularization efficiency and preserved ECM bioactivity in conventional processing techniques. In this study, we established an optimized protocol: combining freeze-thaw cycles with SDS/Triton X-100 decellularization, followed by orthogonal experimental determination of optimal pepsin digestion parameters (72 h, 4 ℃, 10 mg/mL). The refined protocol enhanced decellularization efficiency (P<0.001) while maintaining intact collagen architecture, glycosaminoglycans, growth factors, and porous microstructure, as validated through proteomic analysis and scanning electron microscopy. To address the inherent mechanical limitations of pure DECM hydrogels, we developed a composite hydrogel system, which was composed of 0.5% DECM and 5% methacrylated gelatin (GelMA). This composite hydrogel system demonstrated elastic modulus comparable to native endometrium (P<0.001), with improved degradation stability and optimized swelling ratio compared with single-component systems. Cellular investigations revealed that higher DECM content promoted porcine endometrial stromal cell migration and spheroid self-organization through activating the integrin α5β1-FAK signaling pathway. To overcome limitations in conventional 2D culture, we engineered DECM-GelMA composite microarrays with hemispherical/cylindrical configurations, successfully inducing directional cellular alignment via contact guidance effects. The present study achieved concurrent optimization of DECM preparation efficiency and bioactivity retention, while elucidating its regulatory mechanisms through integrin-mediated signaling and cytoskeletal reorganization. These findings provide new biomimetic strategies for designing functional endometrial scaffolds with improved histocompatibility and mechanobiological responsiveness.