HFTMSCs alleviate uterine fibrosis and restore fertility in intrauterine adhesion models

BACKGROUND: Endometrial intrauterine adhesions (IUA) arise due to comprised regenerative capacity of endometrium and aberrant fibrotic processes. These adhesions significantly affect fertility by impeding embryo implantation, sustaining pregnancy, and facilitating parturition. While human fallopian tube-derived MSCs (HFTMSCs), as a newly identified MSCs population, exhibit substantial potential for endometrial regeneration and fibrosis resolution in IUA treatment. METHODS: HFTMSCs were isolated by collagenase digestion and characterized through multilineage differentiation induction (adiopogenic/osteogenic), surface marker analysis, gene expression profiling, and immunofluorescence staining. The safety profile of HFTMSCs was assessed via subcutaneous tumorigenicity assays in nude mice. An IUA mouse model was established in C57BL/6 J mice via mechanical endometrial curettage. Mice were then randomly allocated into five groups: Sham-operated controls, IUA model, HFTMSCs monotherapy, Estradiol (E2) monotherapy, and HFTMSCs + E2 combination therapy. For in vivo tracking, HFTMSCs were labeled with the fluorescent dye CM-DiI. HFTMSCs retention and distribution were assessed using in vivo imaging and immunofluorescence. Uterine functional recovery was evaluated by embryo implantation rate measurement, with endometrial morphological structure examined via histological analyses. Moreover, Transcriptomic analysis (RNA sequencing) was performed on uterine tissues to elucidate the molecular mechanisms of repair. RESULTS: The CM-DiI labeling experiment confirmed that HFTMSCs can migrate directionally to sites of uterine injury. Histological analysis showed that the combination of HFTMSCs and Estradiol (E2) treatment significantly increased endometrial thickness (P < 0.001), improved glandular number (P < 0.001), reduced fibrosis levels (P < 0.001), and enhanced both angiogenesis in the uterine myometrium and endometrial regeneration on days 14 and 28, respectively. Western blot analysis showed that on day 14, LIF/αvβ3 expression in the IUA group was significantly downregulated compared to the Sham group (P < 0.001). Notably, combined treatment with HFTMSCs and E2 markedly upregulated the expression of these markers (P < 0.001), whereas individual treatments with either HFTMSCs or E2 demonstrated limited effects. By 28 day, LIF/αvβ3 expression in the IUA group remained significantly suppressed compared to the Sham group (P < 0.01), while other treatment groups exhibited varying degrees of marker upregulation. RNA sequencing analysis revealed that HFTMSCs treatment reversed IUA-induced transcriptional changes, modulating key pathways involved in fibrosis (e.g., TGF-β, PI3K-Akt) and promoting those associated with tissue regeneration. Importantly, subcutaneous tumorigenicity assays confirmed the safety of HFTMSCs, with no tumor formation observed throughout the study period. Furthermore, functional fertility assessment demonstrated that the combined HFTMSCs and E2 therapy significantly restored reproductive capability in IUA mice, as evidenced by substantially improved embryo implantation rates and pregnancy rates compared to the IUA model group. CONCLUSIONS: This study demonstrates that combined HFTMSCs and E2 therapy significantly enhances endometrial regeneration in IUA mice, restoring both structural integrity (evidenced by increased endometrial thickness, glandular number, and vascularization with reduced fibrosis) and functional competence, as indicated by recovered fertility. This dual restoration demonstrates synergistic efficacy surpassing monotherapies.