Evaluation of HPV Infection and Smoking Status Impacts on Cell Proliferation in Epithelial Layers of Cervical Neoplasia

Abstract

Accurate cervical intra-epithelial neoplasia (CIN) lesion grading is needed for effective patient management. We applied computer-assisted scanning and analytic approaches to immuno-stained CIN lesion sections to more accurately delineate disease states and decipher cell proliferation impacts from HPV and smoking within individual epithelial layers. A patient cohort undergoing cervical screening was identified (n = 196) and biopsies of varying disease grades and with intact basement membranes and epithelial layers were obtained (n = 261). Specimens were sectioned, stained (Mib1), and scanned using a high-resolution imaging system. We achieved semi-automated delineation of proliferation status and epithelial cell layers using Otsu segmentation, manual image review, Voronoi tessellation, and immuno-staining. Data were interrogated against known status for HPV infection, smoking, and disease grade. We observed increased cell proliferation and decreased epithelial thickness with increased disease grade (when analyzing the epithelium at full thickness). Analysis within individual cell layers showed a ≥50% increase in cell proliferation for CIN2 vs. CIN1 lesions in higher epithelial layers (with minimal differences seen in basal/parabasal layers). Higher rates of proliferation for HPV-positive vs. -negative cases were seen in epithelial layers beyond the basal/parabasal layers in normal and CIN1 tissues. Comparing smokers vs. non-smokers, we observed increased cell proliferation in parabasal (low and high grade lesions) and basal layers (high grade only). In sum, we report CIN grade-specific differences in cell proliferation within individual epithelial layers. We also show HPV and smoking impacts on cell layer-specific proliferation. Our findings yield insight into CIN progression biology and demonstrate that rigorous, semi-automated imaging of histopathological specimens may be applied to improve disease grading accuracy.

Diffuse optical microscopy for quantification of depth-dependent epithelial backscattering in the cervix

“…In HSIL, this lack of differentiation is often accompanied by a reduction in epithelial thickness. 48 Sample histology images of benign and dysplastic areas for both columnar and squamous epithelium are presented in the bottom part of Fig. 10.…”

Limits of Applicability of the Voronoi Tessellation Determined by Centers of Cell Nuclei to Epithelium Morphology

“…One of the first attempts to use CMVT in a clinically relevant situation was to estimate cell areas and perimeters in primary lung carcinoma (Kayser and Stute, 1989). More accurate and complex procedures developed over time include one using CMVT to characterize a number of morphological measures of cell shapes in different cancers with poor and good prognosis (Sudbø et al, 2000) Recently, CMVT became the foundation of an automatic analysis routine and is today used for the analysis of biopsies to distinguish cervical inter-epithelial neoplasia from normal tissues (Guillaud et al, 2014) (Sheikhzadeh et al, 2015). …”

Quantification of large scale DNA organization for predicting prostate cancer recurrence

“…Voronoi Original Article tessellation is a partition of the ROI such that each geometric center of a nucleus is associated with a convex polygon; each point of this polygon is closer to the center of the nucleus than to any other nuclei in the ROI (37,40,41). Voronoi Original Article tessellation is a partition of the ROI such that each geometric center of a nucleus is associated with a convex polygon; each point of this polygon is closer to the center of the nucleus than to any other nuclei in the ROI (37,40,41).…”

“…These tools-which define "cellular sociology"-can be used in conjunction with nuclear GOALS scores to examine tissue characteristics in unprecedented detail and enable us to determine which cells are in a specific layer of the epithelium (Layer 1 cells touch the basement membrane, Layer 2 cells touch Layer 1 cells, etc.). Moreover, they allow us to characterize GOALS behavior of cells in a specific layer (1) individually, (2) as a whole layer, (3) as separate cells according to some other criteria (e.g., cells in a specific duct or adjacent to a specific type of cell as defined by morphology or IHC results) (35,36), or (4) for differential cell sociology behavior based on patient characteristics like HPV or smoking status (37).…”

“…The locations of the geometric centers of the nuclei were used to create a Voronoi tessellation of the ROIs. Voronoi Original Article tessellation is a partition of the ROI such that each geometric center of a nucleus is associated with a convex polygon; each point of this polygon is closer to the center of the nucleus than to any other nuclei in the ROI (37,40,41). Characteristics assigned to the cell nuclei based on this Voronoi tessellation was used to identify possibly biologically meaningful cell subsets.…”