To assess histologic changes in the peritoneal neovagina following robotic-assisted peritoneal flap gender-affirming vaginoplasty (RPGAV) to better guide shared decision-making regarding vaginoplasty techniques. RPGAV uses pedicled flaps of pelvic peritoneum to form the proximal neovagina. Early reports of peritoneal vaginoplasty suggested potential for "self-lubrication"; however, long-term maintenance of fluid production remains uncertain.

Five patients were recruited between August 2024 and March 2025. Patients included were at least 12 months post-RPGAV and had elected for vulvar revision surgery. Patients with a diagnosis of neovaginal stenosis or extensive granulation tissue were excluded. Biopsies were obtained from the peritoneal portion of the neovagina during revision surgery. Primary pathology review was blinded other than biopsy location: “vagina”. Unblinded secondary pathology review was performed to identify diagnostic pitfalls.

All 5 biopsies showed stratified squamous epithelium and no residual mesothelial cells. This demonstrated metaplasia from peritoneal mesothelium to stratified squamous epithelium. Sixty percent of the biopsies showed histologic features mimicking low-grade condylomatous dysplasia. One case required p16 immunostaining to exclude moderate to high-grade dysplasia. Diffuse strong superficial hypergranulosis was a key element to exclude condyloma.

We observed a process of metaplasia: a transition from peritoneal mesothelium to stratified squamous epithelium. While the peritoneal neovagina may produce fluid initially, it is unlikely to be maintained. There was also evidence of atypia, with histologic features mimicking low-grade condylomatous dysplasia; in the context of vaginoplasty, these findings likely represent chronic reactive changes. These histologic changes may have implications for cancer screening and tailored surveillance strategies.