Pelvic organ prolapse (POP) is a common cause for diminished quality of life worldwide and may recur years after successful surgical repair. Microstructural connective tissue aberrations are suggested to cause mechanical failure of pelvic organ support over time.
The aim of this study was to explore the potential of electrically stimulated (ES) patient derived fibroblasts to restore connective tissue microarchitecture in a POP murine model.

Vaginal wall fibroblasts from 3 consenting human females with POP were electrically stimulated (ES), cultured, and injected into a murine POP model of multiparous female lysyl oxidase like-1 (LOXL1) knockout (KO) mice demonstrating POP. This experimental group was named PKO-ES. Controls included:
•	prolapsed multiparous LOXL1 KO mice treated with non-stimulated fibroblasts (PKO-NS);
•	prolapsed multiparous LOXL1 KO mice treated with vehicle injections (PKO-V);
•	non-prolapsed multiparous LOXL1 KO mice treated with vehicle injections (NPKO); and
•	nulliparous, age-matched wild type (WT) mice. 
Images of mouse vaginal tissues, sectioned and stained using Masson’s trichrome for collagen, were systematically processed for analysis and connective tissue quantified using ImageJ® software to measure collagen area as a percentage of total tissue and calculate the collagen alignment index. We performed aggregate and per patient ANOVA with Tukey-Kramer post-hoc testing for between-group differences.

78 mice were studied, 30 were PKO-ES, 30 PKO-NS, 5 PKO-V, 6 NPKO, and 7 WT.

In aggregate, percent collagen did not differ significantly between PKO-ES (22.2 ± 19.2%) and PKO-NS (19.7 ± 16.7%, p=0.98). Both demonstrated a decrease in percent collagen compared with PKO-V controls (58.6 ± 28.4%, p= 0.0005 & 0.0002, respectively), and their collagen area values resembled those of NPKO mice (26.2 ± 9.9% p non-significant) and WT mice (21.6 ± 6.0%, p non-significant).

When tested per patient, PKO-ES mice injected with fibroblasts from a human patient with stage III POP had significantly higher collagen area then their PKO-NS counterparts (42.8 ± 19.2% vs 13.6 ± 4.1%, p= 0.02), a difference not seen with fibroblasts from patients with minimal (stage I) or no POP.

PKO-ES mice had a lower collagen alignment index (0.14 ± 0.03) compared with PKO-NS (0.17 ± 0.02, p=0.0004). Collagen alignment index was similar between PKO-ES and the controls (PKO-V: 0.15 ± 0.01, p=0.9032, NPKO: 0.13 ± 0.003, p=0.5670, WT: 0.12 ± 0.02, p=0.0805.), however it was significantly increased in PKO-NS compared with NPKO (p=0.0014) and WT (p<0.0001). These differences were more pronounced when testing the mice treated with fibroblasts derived from a patient with stage III POP.

Vaginal wall fibroblasts derived from human subjects with varying degrees of POP, with and without electrical stimulation, had varying effects on the microscopic architecture of collagen fibrils.
Electrically stimulated fibroblasts from a subject with significant prolapse seemed to have the greatest effect. Total collagen (as measured by percent of tissue area) was increased, and alignment index was decreased - resembling more the alignment index of collagen in non-prolapsed KO and WT mice than that of prolapsed KO mice treated with non-stimulated fibroblasts or vehicle.

Human derived, electrically stimulated vaginal wall fibroblasts had significant effects on pelvic collagen quantity and alignment in a prolapsed LOXL1 KO mouse model.

Chi, N. et al. Distinctive structure, composition and biomechanics of collagen fibrils in vaginal wall connective tissues associated with pelvic organ prolapse. Acta Biomaterialia 152, 335–344 (2022).

Continence 12S (2024) 101498
DOI: 10.1016/j.cont.2024.101498