Introduction and Hypothesis <p>The treatment of pelvic organ prolapse (POP) presents significant challenges. It is important to explore safer and more effective treatment modalities. Recombinant humanized collagen (rhCol) is a promising biomaterial with excellent biocompatibility and pro-regenerative properties. Therefore, this study aims to evaluate the potential applications of rhCol in POP treatment.</p> Methods <p>Vaginal wall tissues were collected from three non-POP and five POP patients to analyze extracellular matrix (ECM) changes via histological staining. Primary fibroblasts isolated from POP vaginal tissues were treated with rhCol III. Cell proliferation, migration, senescence, and ECM synthesis were assessed. A simulated birth injury (SBI) rat model was used to evaluate ECM remodeling following rhCol injection into the vaginal wall. Additionally, the angiogenic potential of rhCol III was examined in vivo and in vitro.</p> Results <p>POP patient tissues and fibroblasts exhibited lower expression levels of type I and III collagen compared to non-POP samples. At a 1&#xa0;mg/ml concentration, rhCol III promoted fibroblast proliferation and migration, reduced cellular senescence, and enhanced ECM synthesis. In the vaginal wall, the expression of COL1A1 and COL3A1 in the rhCol group was significantly higher than that in the SBI group, with a marked increase in the levels of CD31, CD34, and VEGFA. Furthermore, rhCol III improved the proliferation, migration, and tubule formation capacities of HUVECs.</p> Conclusions <p>rhCol III may promote ECM remodeling in an injured vaginal wall by restoring fibroblast function and stimulating angiogenesis, offering a novel biomaterial-based strategy for POP treatment.</p>