(207) Exploring the Extent of Endodermal Tissue Exhibiting Neuroproliferation: An Immunohistochemical Analysis of the Vestibule and the Hymen in Patients With Neuroproliferative Vestibulodynia

Abstract Introduction Understanding the extent of the endodermal region affected by neuroproliferative vestibulodynia (NPV) is critical when performing complete vestibulectomy. Anatomy textbooks have classically described the upper two-thirds of the vagina as mesoderm and the lower one-third as endoderm. Complete vestibulectomy involves excision of endodermal tissue distal to the mesodermal vagina with the lateral border being Hart’s line and medial border being hymen. However, the extent to which NPV, an endodermal field disease, affects hymenal tissue has not been previously investigated. Objective Our aim was to determine, for the first time, in a series of patients with NPV, if hymenal tissue demonstrated similar neuroproliferative findings to vestibular tissue, based on immunohistochemical (IHC) studies. These findings would help us better determine the extent of endodermal tissue with neuroproliferation causing vestibular pain in patients with NPV. Methods Medical records from patients who underwent a hymenal biopsy between 4/1/24-04/30/25 were reviewed. Biopsies were obtained in-office or during complete vestibulectomy which included additional specimens from 1:00-11:00 (posterior) vestibule and 12:00 (anterior) vestibule (Figure 1). Tissue was processed by pathology, slides stained with CD117, (consistent with mast cells) and PGP 9.5 (consistent with nerves), and stained slides digitally photographed using a LCD35 CMOS color camera attached to an Olympus CX43 microscope at 100× magnification. A minimum of 2 photomicrographs were taken of representative regions including the epithelial basement membrane and adjacent sub-epithelium, for each stained slide. Density of nerves and mast cells per high power field were assessed using Fiji version 1.53. NPV was confirmed using Bornstein’s criteria of >8 mast cells positively immunostained for CD117 in a field viewed at 100x magnification and high density of positively immuno-stained cells with neuronal marker. Comparisons between hymen and the other locations were performed using a one-way ANOVA for repeated measures with Geisser-Greenhouse correction. Post-hoc pairwise comparisons were done using Tukey’s test. Results Biopsies were obtained in office (n=16) and intraoperatively (n=14) from 19 patients with NPV. Mean age was 33 years (range 18–61) at the time of the procedure. All patients met diagnostic criteria for NPV. There were no statistically significant differences between the density of mast cells and nerves of the hymen and anterior and posterior vestibule (Table 1). Conclusions Our data demonstrate for the first time that the endodermal field affected by NPV includes the hymen in addition to the anterior and posterior vestibule. While the lateral border of the vestibule is Hart’s line, the medial border of the endodermal tissue affected by NPV is, in fact, the junction between endodermal and mesodermal vagina which is proximal to the hymen. Thus, the tissue removed during vestibulectomy should include the hymen and extend to the vaginal transition between endoderm and mesoderm. Disclosure No