Aristotelis Loufopoulos

Human papillomavirus DNA and mRNA positivity of the anal canal in women with lower genital tract HPV lesions: Predictors and clinical implications

OBJECTIVE Women with HPV related pathology of the lower genital tract are at higher risk for AIN and anal cancer than the general population. A strategy to identify anal disease in these women has not been formulated. The aim of this study is to examine the feasibility of HPV related biomarker testing on anal smears, to identify the risk factors for anal HPV positivity and to provide information of the clinical implications of anal HPV infection in this population. METHODS In women referred for colposcopy because of HPV related pathology of the lower genital tract (cervical cancer, CIN, VIN, warts) a detailed questionnaire, an anal smear and a cervical smear were taken. On each sample morphological cytology, flow cytometric evaluation of E6&7 mRNA, and HPV DNA detection and typing were performed. Women with a positive anal result were referred for high resolution anoscopy. RESULTS So far 235 women have been included (mean age 34.3). HPV DNA, high-risk HPV DNA, high-risk mRNA was detected in 45%, 31% and 8% of the anal smears and in 56%, 39% and 25% of the cervical smears respectively. Absolute or partial concordance of the types between the cervix and the anus was seen in 74%. Positivity for mRNA was significantly lower in the anus than the cervix (8% vs 25%). Logistic regression analysis revealed risk factors for the presence of anal HPV DNA (>3 lifetime sexual partners and presence of cervical HPV DNA), hr HPV DNA (presence of cervical hr HPV DNA), and hr mRNA (presence of cervical hr mRNA). Twelve months after LLETZ 53% of women were cervical HPV negative, but 25% of those were still HPV positive in the anus. CONCLUSIONS HPV infection of the anus is common in this group and is interlinked with the cervical infection. Anal HPV E6&7 mRNA expression is less common than in the cervix. Possible clinical implications of anal infection could be the development of AIN and recurrence of CIN after treatment due to cervical reinfection from the anal reservoir. The use of HPV biomarkers is feasible in anal smears, although especially DNA testing as triage method for referral to anoscopy is probably inappropriate due to high positivity rate.

The evaluation of p16INK4a immunoexpression/immunostaining and human papillomavirus DNA test in cervical liquid-based cytological samples.

Aim: To evaluate the role of p16INK4a immunoexpression and human papillomavirus (HPV) DNA test for the detection of dyskaryotic cells in high-risk women. Materials and Methods: This work was a retrospective diagnostic study conducted in the University Hospital of Thessaloniki from January to December 2008. The subjects were women with current or previous HPV infection and current or previous cervical intraepithelial lesion (with or without treatment) or clinical warts. All liquid-based cytological samples were tested for P16INKa and HPV DNA test. The accuracy parameters used for the outcome included sensitivity, specificity, and positive predictive value. Results: A total of 226 women were included; the mean age was 29 years. Expression of p16INK4a was detected in the cytological samples of 13% of the negative cases, 44% of the cases of atypical squamous cells of undetermined significance, 46% of the cases of low-grade squamous intraepithelial lesion, and 78% of the cases of high-grade squamous intraepithelial lesion. A total of 91 women tested positive for high-risk HPV infection, and 54 of those had p16INK4a-positive staining reaction cells. The concordance between the 2 tests, HPV DNA and p16, was 59% regarding infection-positive cases. Diffuse strong parabasal p16INK4a immunostaining (nuclear score >2) was observed in 17 cases of the abnormal cytological findings (atypical squamous cells of undetermined significance, 2 cases; low-grade squamous intraepithelial lesion, 8 cases; high-grade squamous intraepithelial lesion, 7 cases). Colposcopy-directed biopsies were used as the criterion standard for the detection of cervical intraepithelial neoplasia in 91 women. The sensitivity of p16INK4a was 95% and the specificity was 92%, whereas the sensitivity of high-risk HPV was 100% and the specificity was 78%. The positive predictive value of p16INK4a was 71%, whereas that of HPV DNA was 44%. Conclusion: The findings suggest that p16INK4a immunostaining can improve the accuracy of cytological examination and HPV DNA test and may be particularly useful in the triage of low-grade lesions.

Histology Verification Demonstrates That Biospectroscopy Analysis of Cervical Cytology Identifies Underlying Disease More Accurately than Conventional Screening: Removing the Confounder of Discordance

Background Subjective visual assessment of cervical cytology is flawed, and this can manifest itself by inter- and intra-observer variability resulting ultimately in the degree of discordance in the grading categorisation of samples in screening vs. representative histology. Biospectroscopy methods have been suggested as sensor-based tools that can deliver objective assessments of cytology. However, studies to date have been apparently flawed by a corresponding lack of diagnostic efficiency when samples have previously been classed using cytology screening. This raises the question as to whether categorisation of cervical cytology based on imperfect conventional screening reduces the diagnostic accuracy of biospectroscopy approaches; are these latter methods more accurate and diagnose underlying disease? The purpose of this study was to compare the objective accuracy of infrared (IR) spectroscopy of cervical cytology samples using conventional cytology vs. histology-based categorisation. Methods Within a typical clinical setting, a total of n = 322 liquid-based cytology samples were collected immediately before biopsy. Of these, it was possible to acquire subsequent histology for n = 154. Cytology samples were categorised according to conventional screening methods and subsequently interrogated employing attenuated total reflection Fourier-transform IR (ATR-FTIR) spectroscopy. IR spectra were pre-processed and analysed using linear discriminant analysis. Dunn’s test was applied to identify the differences in spectra. Within the diagnostic categories, histology allowed us to determine the comparative efficiency of conventional screening vs. biospectroscopy to correctly identify either true atypia or underlying disease. Results Conventional cytology-based screening results in poor sensitivity and specificity. IR spectra derived from cervical cytology do not appear to discriminate in a diagnostic fashion when categories were based on conventional screening. Scores plots of IR spectra exhibit marked crossover of spectral points between different cytological categories. Although, significant differences between spectral bands in different categories are noted, crossover samples point to the potential for poor specificity and hampers the development of biospectroscopy as a diagnostic tool. However, when histology-based categories are used to conduct analyses, the scores plot of IR spectra exhibit markedly better segregation. Conclusions Histology demonstrates that ATR-FTIR spectroscopy of liquid-based cytology identifies the presence of underlying atypia or disease missed in conventional cytology screening. This study points to an urgent need for a future biospectroscopy study where categories are based on such histology. It will allow for the validation of this approach as a screening tool.

Laparoscopic management of unicornuate uterus with non-communicating rudimentary horn (three cases)

This article describes the laparoscopic management of unicornuate uterus with a non-communicating rudimentary horn in three women who presented with lower abdominal pain and dysmenorrhoea. The patients were managed with laparoscopic removal of the rudimentary horn along with the ipsilateral salpinx. No complications were noted intraoperatively and the patients remain asymptomatic.

Postparacentesis bilateral massive vulvar edema in a patient with severe ovarian hyperstimulation syndrome

OBJECTIVE To report a case of bilateral massive vulvar edema following lower abdominal paracentesis in a patient with ovarian hyperstimulation syndrome. DESIGN Case report. SETTING University teaching hospital. PATIENT(S) A 32-year-old woman with primary infertility. INTERVENTION(S) The patient underwent ovarian stimulation with leuprolide acetate, highly purified FSH, and hCG. Because of the development of severe ovarian hyperstimulation syndrome, bilateral paracentesis through the lower abdominal quadrants was performed. MAIN OUTCOME MEASURE(S) Treatment of ovarian hyperstimulation syndrome. RESULT(S) Development of bilateral massive vulvar edema 24 hours after lower abdominal paracentesis. CONCLUSION This case report suggests that lower abdominal paracentesis could be the cause of vulvar edema development in ovarian hyperstimulation syndrome, probably due to a fistulous tract created between the peritoneal cavity and the subcutaneous tissues.

Molecular epidemiology of HPV infection using a clinical array methodology in 2952 women in Greece

The molecular epidemiology of human papillomavirus (HPV) infection in a sample of Greek women (n = 2952, mean age 42.2 ± 13.3 years) was examined. HPV prevalence was 50.7% (95% confidence interval, 48.8-52.6). The most frequent HPV types were HPV 53, 51 and 66 (10.2%, 9.4% and 9.3%, respectively). HPV positivity was associated with age, age of sexual debut, number of sexual partners and duration of sexual relationship, while marriage or multiparity protected against infection (all p <0.001). Follow-up of this cohort will assist in predicting the effect of vaccination with the new HPV vaccines on future screening with HPV-based tests for cervical cancer.

Personalised management of women with cervical abnormalities using a clinical decision support scoring system

OBJECTIVES To develop a clinical decision support scoring system (DSSS) based on artificial neural networks (ANN) for personalised management of women with cervical abnormalities. METHODS We recruited women with cervical abnormalities and healthy controls that attended for opportunistic screening between 2006 and 2014 in 3 University Hospitals. We prospectively collected detailed patient characteristics, the colposcopic impression and performed a series of biomarkers using a liquid-based cytology sample. These included HPV DNA typing, E6&E7 mRNA by NASBA or flow cytometry and p16INK4a immunostaining. We used ANNs to combine the cytology and biomarker results and develop a clinical DSSS with the aim to improve the diagnostic accuracy of tests and quantify the individuals risk for different histological diagnoses. We used histology as the gold standard. RESULTS We analysed data from 2267 women that had complete or partial dataset of clinical and molecular data during their initial or followup visits (N=3565). Accuracy parameters (sensitivity, specificity, positive and negative predictive values) were assessed for the cytological result and/or HPV status and for the DSSS. The ANN predicted with higher accuracy the chances of high-grade (CIN2+), low grade (HPV/CIN1) and normal histology than cytology with or without HPV test. The sensitivity for prediction of CIN2 or worse was 93.0%, specificity 99.2% with high positive (93.3%) and negative (99.2%) predictive values. CONCLUSIONS The DSSS based on an ANN of multilayer perceptron (MLP) type, can predict with the highest accuracy the histological diagnosis in women with abnormalities at cytology when compared with the use of tests alone. A user-friendly software based on this technology could be used to guide clinician decision making towards a more personalised care.

Expression of HPV-related biomarkers and grade of cervical intraepithelial lesion at treatment.

New human papillomavirus (HPV)‐related biomarkers may allow better identification of clinically significant lesions that warrant excision and, conversely, identification of the false positive cases that have been overreported by traditional techniques. The aim of this study was to investigate how the expression of several HPV‐related biomarkers correlates to the severity of the lesion at treatment.

Transition to ISO 15189: 2012 for Cytopathology Laboratories Part 1: General and Management Requirements

Nowadays, due to the latest advances in Laboratory Medicine, diagnostic medical laboratories with their highly qualified personnel and state-of-the-art analytical equipment, have completely changed the way modern healthcare is offered. In order to maintain or even increase the already high quality level of the provided testing services, diagnostic laboratories including the cytopathology ones, need to design and apply a Quality Management System QMS in agreement with the requirements of the ISO 15189 International Standard. The authors present their experience on the implementation of such a QMS in cytopathology laboratories and highlight the most important general and management parameters that should be taken into consideration when moving from ISO 15189:2007 to the latest ISO 15189:2012. In addition, useful recommendations and suggestions that could make the transition to the latest Standard easier are included. Finally, possible issues and potential adverse events associated with the laboratorys implementation of the ISO 15189:2012 are also described.

Transition to ISO 15189: 2012 for Cytopathology Laboratories Part 2: Technical Requirements

Modern diagnostic medical laboratories personnel should be an ideal combination of highly qualified scientists and technologists who use innovative analytical equipment to provide high quality diagnostic services to mankind. A well-designed and a well-implemented Quality Management System QMS according to the ISO 15189 requirements is regarded as a prerequisite that will ensure that a cytopathology laboratory is capable enough to provide precise and credible results. The authors present their knowledge on the implementation of such a QMS in cytopathology laboratories and emphasize the most crucial technical parameters that should be considered when advancing from ISO 15189:2007 to the latest ISO 15189:2012. Furthermore, helpful advice and pointers that could accommodate relative transition are included. Finally, potential issues related to the laboratorys implementation of ISO 15189:2012 and a mobile technology application for better personnel management are also depicted.