Human papillomavirus‐related cellular changes measured by cytometric analysis of DNA ploidy and chromatin texture

Abstract

Background: Image cytometry has provided two highly sensitive markers for the identification of the malignant potential of squamous lesions. Aneuploidy and chromatin texture have been investigated as quantitative measures of nuclear damage in premalignant lesions and carcinoma. Real-time PCR methods have evolved to yield highly specific measurements of mRNA expression in very sparse cellular samples.Methods: Human papillomavirus (HPV) 16 and 18 E7 mRNA expression was measured using quantitative RT-PCR. DNA index and chromatin measures were taken from image cytology samples. The chromatin features, through discriminant analysis, were aggregated into a score, and both measurements were related to mRNA expression.Results: mRNA level and DNA index show an increasing trend over increasing histological grades. However, DNA index and chromatin score were not correlated to mRNA levels in these samples. Chromatin score differed by mRNA type found with HPV 18 infected samples having a higher score than those with HPV 16. Samples infected with HPV 16 and HPV 18 had even higher chromatin scores.Conclusions: DNA index and chromatin score were not directly correlated with mRNA levels. However both mRNA and DNA index were related to histological grade, and chromatin score was associated with HPV type. Therefore, DNA index and mRNA levels could be independent predictors of cervical dysplasia, and chromatin score could be related to the viral integration process in cells infected with HPV 18 or dual infections. q 2007 Clinical Cytometry Society

Comparing the Performance of Hybrid Capture II and Polymerase Chain Reaction (PCR) for the Identification of Cervical Dysplasia in the Screening and Diagnostic Settings

“…DNA specimens were placed in 250 μL PBS with 0.02% sodium azide immediately after cytologic sampling and were stored at −80 °C until extracted. Specimen extraction was performed approximately one month after collection 19. Biopsies were taken from colposcopicly abnormal and normal sites for histopathologic confirmation of disease, as described elsewhere 18,19.…”

Optical Technologies and Molecular Imaging for Cervical Neoplasia: A Program Project Update

“…Over the course of this project, we have undertaken multiple pilot and Phase I/II studies. Through Phase II trials of quantitative cytology, we saw 1850 patients at five clinical sites, with this work confirming the feasibility of obtaining quick results by this technique (68, 70). Data from a Phase II trial that evaluated >3500 cervical biopsies using quantitative pathology methodologies helped hone automated components of this technique and identify cell-level changes associated with different neoplastic stages (66–67, 71–73).…”

“…All of the cytologic and histopathologic specimens needed for clinical and quantitative assessment have been collected and managed by this core. This core has also developed imaging systems for evaluating morphometric and architectural features on tissue cross-sections, with phenotypic scores calculated by this method correlating well with pathology classifications and HPV status (48, 66–68, 70, 73, 120–121). Algorithms based on this approach have had particular utility in characterizing the underlying biology of cervical disease.…”

Human Papillomavirus and Related Diseases - From Bench to Bedside - A Clinical Perspective

“…The distinction of florid reactive changes, immature metaplastic patterns, and atrophic changes from HPVinduced alterations may cause difficulties. Attempts have been made to redefine the traditional criteria for lesion diagnosis, while other efforts aimed at the adoption of new, more objective methods, which might support the former (Bollmann et al, 2005;Cho et al, 2005;Guillaud et al, 2005;Prasad et al, 1994;Salvia et al, 2004;Scheurer et al, 2007). However, studies attempting to correlate HPV presence and replication to certain cytohistologic alterations are becoming less frequent and/or fruitful.…”