Cytoplasmic localization of endothelial constitutive nitric oxide synthase in endometrial carcinomas

Background: Production of nitric oxide by nitric oxide synthase (NOS) has been implicated in numerous physiologic and pathophysiologic processes including mutagenesis. This study was designed to examine the expression of the endothelial constitutive isoform of NOS (ecNOS) in endometrial carcinomas. Methods: Fifty endometrial carcinomas (42 endometrioid, 4 serous papillary, 2 clear cell, and 2 adenosquamous carcinomas) and 21 normal endometrial gland tissue specimens (5 cases of proliferative, 5 early secretory, 5 mid-secretory, and 5 late secretory and 1 menstrual phase endometrium), previously formalin fixed and paraffin embedded, were immunostained using a commercially available anti-ecNOS monoclonal antibody. Localization of ecNOS staining to the plasma membrane, cytoplasm and nuclei was graded with respect to overall staining intensity (0-3+ scale) and frequency (percentage of immunoreactive cells). Results: Relatively little staining for ecNOS was localized to the plasma membrane in either normal or neoplastic tissues. Normal and hyperplastic endometrial glands demonstrated moderate cytoplasmic and weak nuclear staining in a small percentage of cells. While ecNOS expression was most prominent in epithelial cells, weak expression was also rarely noted in endometrial stroma, blood vessel walls, and endothelium. We found a broad range of ecNOS expression in endometrial carcinomas, predominantly localized to the cytoplasm and nuclei. No statistically significant difference in ecNOS staining frequency or intensity was found between different histologic subtypes of endometrial carcinomas. No apparent correlation was found between ecNOS expression and tumor stage, grade, extension to the lower uterine segment or cervix, nodal or distant metastases, recurrence, or final patient status among patients with endometrioid adenocarcinomas. Endometrioid tumors invading more than 1/2 of myometrial thickness (n = 18) had significantly higher cytoplasmic staining intensity than those tumors limited to the inner 1/2 of myometrium (n = 27; 2.0 vs. 1.3, p < 0.04). Furthermore, a trend toward shorter disease-free survival was noted with increased staining intensity and decreased staining frequency. Conclusions: Cytoplasmic and nuclear expression of ecNOS, which is primarily limited to the glandular elements of normal endometrium, is also found to be expressed in endometrial carcinoma. Increased ecNOS staining intensity and decreased frequency tends to correlate with decreased disease-free survival. Lastly, increased cytoplasmic ecNOS staining intensity correlates with increased myometrial invasion.