Diagnosis Of Low-grade Squamous Intraepithelial Lesion Research Articles

Correlation Between hr-HPV Genotypes/Negative Cytology with Cervical Histology Findings

Abstract Introduction/Objective It is well established that the persistent of hr-HPV infection is necessary for the development of cervical dysplasia. Currently, routine cervical cancer screening include both high-risk human papilloma virus (hr- HPV) testing and cervical cytology tests. The guidelines for managing HPV-positive/cytology-negative results is cotesting at 1 year, and/or immediate colposcopy if HPV16 and/or HPV18 is positive. We analyzed the correlation between hr-HPV genotypes /negative cytology with cervical histology findings. Methods/Case Report Retrospective chart review was performed to identify all negative pap smears with positive hr-HPV and follow up colposcopy for patients who presented to our institution from January 2018 to December 2019. We calculated the distribution of HPV genotypes within low grade and high-grade cervical lesions among patients with HPV-positive/cytology-negative pap smears. Results (if a Case Study enter NA) 179 patients with HPV-positive/cytology-negative pap smears were included. The age range was 22-69 years old. Among these patients, 6.70% had low-grade squamous intraepithelial lesion (LGSIL) and 6.14% had high-grade squamous intraepithelial lesion (HSIL) on follow up biopsy. The detection rates of HPV genotypes among patients with LGSIL diagnosis were HPV16 (41.66%), HPV18 (16.66%) and other hr-HPV (41.66%). The detection rates of HPV genotypes among patients with HGSIL diagnosis were HPV16 (9.09), HPV18 (0%), other hr-HPV (72.72%), combined HPV16 + other hr-HPV (9.09), combined HPV18+ other hr-HPV (9.09%). Conclusion According to our data, patients with negative cytology and HGSIL on biopsy have higher association with other hr-HPV (not 16 or 18) than HPV 16 and 18. Patients with negative cytology and LGSIL on biopsy have higher association with HPV16 and other hr-HPV subtypes than HPV18. Thus, we recommend follow-up colposcopy even for patients with other hr-HPV infection.

Cervical Pre-cancers: Biopsy and Immunohistochemistry.

The existence of precursor lesions for invasive cervical cancer has been recognized for more than 50 years. Our understanding of the pathobiology and behavior of cervical cancer precursors has evolved considerably over the past five decades. Furthermore, the terminology used to classify pre-invasive lesions of the cervix has frequently changed. The realization that human papillomavirus (HPV) infections constitute a morphologic continuum has prompted efforts to include them within a single classification system, specifically the squamous intraepithelial lesions (SILs) which have now been embraced by the surgical pathologists. The reduced number of specific pathological categories has made clinical decision-making more straightforward. The generic criteria for SIL have two important histological parameters: Alterations in the density of superficial epithelial cells and superficial squamous atypia. The flat condyloma or cervical intraepithelial neoplasia (CIN) I is generally associated with intermediate and high-risk HPV types as against the low-risk viruses that cause exophytic/papillary growth patterns of condylomas. The diagnosis of low-grade SIL (LSIL) (flat and exophytic condylomas) requires first excluding benign mimics of LSIL and second to confirm the characteristic cytologic atypia. For high-grade SILs (HSILs), the extent and degree of atypia generally exceed the limits of that described in flat or exophytic condylomas (LSILs). Less maturation, abnormal cell differentiation, loss of cell polarity, and increased mitotic index with abnormal mitotic figures occupying increasing thickness of the epithelium define a lesion as CIN II or CIN III. Atypical immature metaplasia associated with inflammation and atrophy is a challenge in cervical biopsy interpretation. Careful attention to the growth pattern of the epithelium, the distribution of the atypia, nuclear spacing, and the degree of anisokaryosis and the presence of enlarged hyperchromatic nuclei help in differentiating a non-neoplastic from a neoplastic process. This chapter describes in depth the diagnostic difficulties in the interpretation of cervical biopsies. It also provides useful criteria in distinguishing benign mimics from true precancerous lesions and the role of biomarkers such as the p16ink4 and Ki-67 in the differential diagnosis of precursor lesions and the reactive and metaplastic epithelium.

The value of Ki67 for the diagnosis of LSIL and the problems of p16 in the diagnosis of HSIL

p16 and Ki67 are immunohistochemical markers related to cervical squamous intraepithelial lesions. p16 has been widely used to assist in the diagnosis of high-grade squamous intraepithelial lesions. However, a conclusion about the role of Ki67 in the diagnosis of squamous intraepithelial lesions has not been established. The aim of this study was to analyze the role of p16 and Ki67 immunohistochemical staining in assisting cervical squamous intraepithelial lesions. This study performed immunohistochemical staining for p16 and Ki67 on 1024 cervical biopsy specimens at our hospital to compare the differences between p16 and Ki67 in different cervical lesions using the chi-squared test and Fisher’s exact test. This study also evaluated the value of Ki67 for the diagnosis of low-grade squamous intraepithelial lesions (LSILs) using the receiver operating characteristic curve. The results indicated that Ki67 had high specificity and sensitivity in distinguishing LSIL from normal cervix. p16 was diffusely and strongly positive in some LSILs, and some problems were encountered in the interpretation of p16 staining. Therefore, we believe that Ki67 can be used as an immunohistochemical marker to help in the diagnosis of LSIL, to distinguish lesions that are difficult to morphologically determine and to avoid misdiagnosis. The practical application of p16 staining is still problematic. It may be necessary to find other auxiliary means to distinguish this small proportion of cervical lesions.

Risk factors modulating the course of low-grade squamous intraepithelial lesions in women of reproductive age

Background. Infection with oncogenic types of human papillomavirus (HPV) is the leading cause of cervical cancer and its immediate precursors – squamous intraepithelial lesions. However, the persistence of HPV may not be sufficient for the occurrence of malignant transformation, and there may be other exogenous or endogenous factors that, in combination with the virus, increase the risk of developing and progressing cervical neoplasia.Objective: identification of risk factors that modulate the course of low-grade squamous intraepithelial lesions (LSIL) in women of reproductive age.Materials and methods. In 110 women aged 18 to 45 years with a diagnosis of LSIL, we evaluated the impact of risk factors (sexual behavior, reproductive function, contraceptive methods, smoking, HPV infection) on the course of the process (recovery, persistence or progression) during the 12-month follow-up period. HPV testing was performed by real-time PCR.Results. The average age of the study participants was 31.55 ± 7.17 years. Our observations showed that frequent artificial abortions, the practice of risky sexual behavior (early sexual debut, frequent change of sexual partners) increased the risk of persistence and progression of mild cervical cytological abnormalities. The use of oral contraceptives, barrier contraception and smoking did not affect the outcome of the disease.The prevalence of HPV among women with LSIL was 70.91 % (78/110). The predominant types were HPV 16 (35.45 %), HPV 33 (12.73 %) and HPV 18 (12.73 %). HPV 16 was more common in the progression group (62.50 %) than in the regression and persistence groups (14.63 % and 43.40 %, p <0.05). Progression of neoplasias was more often observed in patients with signs of multiinfection of the cervical epithelium with HPV. Single HPV infection was higher in the group of recovered women.Conclusions. The study found that a large number of induced abortions, the practice of risky sexual behavior, the presence of HPV type 16 and multiple HPV infection are factors contributing to the persistence of LSIL and progression to HSIL. These data may be useful for the management of women with mild cervical cytological abnormalities.

Pap Smear miR-92a-5p and miR-155-5p as potential diagnostic biomarkers of squamous intraepithelial cervical cancer.

Background:one of the female-specific diseases with a high incidence and mortality is cervical cancer. The main cause of cervical cancer is infection with Human papilloma virus (HPV). Low-grade squamous intraepithelial lesions (LSIL) and High-grade squamous intraepithelial lesions (HSIL) usually is caused by an HPV infection. Considering the role of microRNAs (miRNAs) as diagnostic biomarkers for a variety of cancers, the aim of this study was to determine miR-92a-5p and miR-155-5p expression levels in LSIL and HSIL Pap Smear samples. Methods:After initial bioinformatic studies, A total of 75 samples (25 samples of patients with LSIL, 25 patients with HSIL and 25 healthy individuals) were subjected to RNA extraction and cDNA synthesis. The expressions levels of confirmed miRNAs in samples of patients with LSIL, HSIL and healthy individuals were evaluated by Real time PCR analysis. To demonstration the role of predicted miRNAs as novel biomarkers in diagnosis of LSIL and HSIL, ROC curve analysis was done. Results:Bioinformatics results showed that miR-92a-5p and miR-155-5p target the HPV E6 and E7 genes. The expression levels of these miRNAs were strikingly higher in Pap smear of patients with LSIL than in the healthy individuals (35.36, P = 0.001) (62.23, P = 0.001). Similarity, expression levels of miR-92a-5p and miR-155-5p were amazingly higher in patients with HSIL than in the healthy individuals (33.62, P= 0.001) (69.07, P= 0.001). Although, the levels of miR-92a-5p (0.95, P = 0. 85) and miR-155-5p (1.11, P = 0.84) exhibited no statistical differences between patients with LSIL and HSIL. Also, ROC curve analyses verified that miR-92a-5p and miR-155-5p are specific and sensitive and may serve as new biomarkers for the early detection of cervical cancer. Conclusion:These data suggest miR-92a-5p and miR-155-5p, which are upregulated in LSIL and HSIL, can be consider as predictive biomarkers for the prognosis of cervical cancer patients.

Prognostic value of microRNA assessment in cervical epithelial cells of patients with mild dysplasia

Background. Currently, there are no reliable markers for the prognosis of the low-grade squamous intraepithelial lesion (LSIL) of the cervical epithelium. Scientific literature provides with inconsistent recommendations regarding the management of the young patients with a cytological diagnosis of LSIL. The progression of cervical dysplasia and the development of cervical cancer are associated with characteristic alterations of the microRNA expression profile.Objective: to assess the prognostic value of microRNA in LSIL. Materials and methods. Samples (cytological smears) obtained from patients diagnosed with LSIL (n = 36), but with a different course of the disease over the next 6–12 months. Analysis of miRNA expression was carried out by the method of “two-tailed” reverse transcription and subsequent PCR.Results. The expression level of miR-126, miR-21, miR-1246, miR-182 was statistically significantly different in the compared groups, but the predictive value of the analysis of individual molecules was low (AUC <0.65). Calculation of the concentration ratios of the “reciprocal” pairs of microRNAs made it possible to obtain a more effective prognostic marker. ROC analysis of such ratios (miR-126/miR-182, miR-21/miR-182, miR-1246/miR-182) yielded AUC values: 0.82–0.89, sensitivity: 0.71–0.92; specificity: 0.86.Conclusions. Analysis of a panel of microRNA marker molecules in the material of the cervical epithelium and calculation of the concentration ratios of “reciprocal” pairs is a promising method for prognosis of LSIL course.

P16 Immunostaining Can Eliminate Approximately 20% Inaccurate Diagnosis of Low-Grade Squamous Intraepithelial Lesions (SIL) in Cervical Biopsies

Abstract Introduction/Objective Differentiating low-grade SIL from reactive changes on light microscopy has remained controversial. P16 immunostaining is widely accepted in high-grade SIL unlike in low-grade SIL. We focused on the use of p16 immunostaining in eliminating inaccurate diagnosis of low-grade SIL on light microscopy. Methods A retrospective study on 60 cervical biopsies results over a 2 year-period was conducted using 35 suspicious cases for low-grade SIL (bi-nucleation and some nuclear atypia) and 25 low suspicion cases (enlarged nuclei but nuclear atypia cannot differentiate from reactive changes). All cases were stained for p16, significant nuclear expression of p16 was determined as low- grade SIL. Results After p16 immunostaining, 30/35 (85%) cervical biopsies from the suspicious group were found to be truly positive for low-grade SIL while 5/35 (15%) biopsies negative for p16 immunostaining were regarded as reactive changes. In the low suspicious group, 7/25 (28%) were found to be positive for low-grade SIL and 18/25 (72%) biopsies were negative, favoring reactive changes. The overall percentage of all 60 biopsies that showed a changed diagnosis based on p16 immunostaining was 20% (5 were truly negative in the suspicious group and 7 were truly positive in the low suspicious group). Using a 2 x 2 table with p16 as a gold standard for all biopsies, sensitivity and specificity of light microscopy for low-grade SIL were only 81% and 78% respectively. Conclusion Our study showed that p16 immunostaining can also be utilized to eliminate inaccurate diagnosis of low- grade SIL up to 20% in cervical biopsies.

Use of AKR1C1 and TKTL1 in the Diagnosis of Low-grade Squamous Intraepithelial Lesions from Mexican Women.

To determine the differential protein profiles of cervical cancer cell lines in order to find potential targets that can be used as biomarkers in low-grade squamous intraepithelial lesions (LSIL) diagnosis. Proteomic analysis was performed on cervical cancer cell lines by 2D electrophoresis and liquid chromatography-mass spectrometry. Biomarker validation was performed in histological samples by immunofluorescence. Aldo-keto reductase C1 (AKR1C1) and transketolase-like 1 (TKTL1) proteins were selected as biomarkers and their expression was increased in samples with LSIL diagnosis. TKTL1 in combination with AKR1C1 increased sensitivity and specificity to 75% and 66%, respectively, with an area under curve of 0.76 in receiver operating characteristics curve analysis. AKR1C1 and TKTL1 showed potential as biomarkers for diagnosis of LSIL in Mexican women, with similar sensitivity and specificity to the biomarkers used in clinical trials for diagnosis of LSIL.

Potential influence of p16 immunohistochemical staining on the diagnosis of squamous cell lesions in cervical biopsy specimens: observation from cytologic-histologic correlation.

The p16 immunohistochemical (IHC) marker has been used increasingly as an adjunct to morphologic assessment of cervical biopsies in which the differential diagnoses include high-grade squamous intraepithelial lesion (HSIL) and its mimics. The objective of this study was to assess the potential influence of p16 IHC staining on the evaluation of cervical biopsy as observed through cytologic-histologic correlation (CHC). Cervical biopsy samples that had cytologic diagnoses of either low-grade squamous intraepithelial lesion (LSIL) or HSIL and also had histologic follow-up were retrieved from the department database. CHC and the use of p16 IHC from 2 periods (group 1, 2008; group 2, 2014-2016) were compared and analyzed. Histology on 452 samples from patients who had prior LSIL cytology in group 1 yielded 126 benign (27.9%), 272 LSIL (60.2%), and 54 HSIL (11.9%) diagnoses. By comparison, 491 samples from the patients in group 2 yielded 106 benign (21.6%), 277 LSIL (56.4%), and 108 HSIL (22.0%) diagnoses. The difference in CHC discrepancies between the 2 groups was significant (P = .0001), mainly because of the increased diagnosis of HSIL in group 2. Although p16 IHC was not applied to any sample from group 1, it was performed on 141 of 491 samples (28.7%) from group 2. Further follow-up of patients who had histologic HSIL revealed that residual HSIL was identified significantly more often in those who did not have p16 IHC applied in the preceding cervical biopsy than in those did (P = .0004). A similar comparison was performed between 113 patients from group 1 and 152 patients from group 2 who had a prior diagnosis of HSIL cytology, and the difference was statistically insignificant. The use of p16 IHC on cervical biopsies in patients who had a prior cytologic diagnosis of LSIL may lead to greater detection and upgrading of HSIL, thereby compounding the discrepancy in CHC.

Significance of bizarre cells in cervical screening liquid-based cytology: A prospective study of 15 cases.

The aim of this study was to assess the significance of bizarre cells (cells of squamous origin with a superficial squamous cell-type cytoplasm and characterised by multinucleation that produces bizarre nuclear shapes) in liquid-based cytology (LBC) Papanicoaou (pap) smears with clinical and histological follow-up correlation. Fifteen patients, all with LBC samples containing bizarre cells, were identified in routine ThinPrep® LBC workload. HPV testing was performed in each case using residual LBC material. Cytological-histological correlations were reviewed. All 15 LBC samples contained bizarre cells and tested positive for high-risk HPV types. Ten of the 15 cases were identified as atypical squamous cells - cannot exclude an HSIL (ASC-H) with secondary diagnosis of low-grade squamous intraepithelial lesion (LSIL), while five cases were identified as high-grade squamous intraepithelial lesion (HSIL), and a subsequent biopsy was recommended. Additionally, 13/15 cases underwent cone biopsy or hysterectomy within 1-11 months, of which 10 showed histologically confirmed HSIL end-points. LSIL was present in three cases. Bizarre cells were identified in the HSIL epithelium of five cone biopsies. Identification of bizarre cells in LBC is straightforward and may facilitate diagnosis. The cytology of bizarre cells is associated with HSIL in cone biopsies. We recommend assigning LBC samples containing bizarre cells as ASC-H with secondary diagnosis of LSIL.

Follow up in women with biopsy diagnosis of cervical low-grade squamous intraepithelial lesion (LSIL): how long should it be?

To analyse the regression rate and the risk of persistence or progression of the lesions in women with a histopathological diagnosis of cervical low-grade squamous intraepithelial lesion (LSIL). Retrospective cohort study of women with biopsy diagnosis of cervical LSIL, from January 2010 to December 2013. After the initial diagnosis of LSIL, all the women underwent scheduled follow-up examinations with cytology every 6months for 2years and an HPV test after 1year. At the 24-month follow-up, the regression of cervical LSIL was observed in 88.5% of the women. 10.8% of the women had a persistent lesion, while a progression towards cervical HSIL was reported in 0.7% of the women. The risk of persistence or progression of histological LSIL was higher in women with ASC-H or HSIL on the referral cytology and in tobacco users. In women with biopsy diagnosis of cervical LSIL, preceded by ASCUS or LSIL on cytology, a high rate of regression was observed and, in most of the cases, the regression occurred in the first year of follow-up. In women with cervical LSIL, preceded by ASC-H or HSIL on cytology, and in tobacco users, a higher risk of persistence and progression was observed. Thus, in these cases, repeated follow-up examinations, even with the HR-HPV test, are advisable.

HPV immunohistochemical testing and cervical dysplasia.

Background and aimHPV (Human Papilloma Virus) infection represents a necessary condition for cervical carcinogenesis. The purpose of this study was to evaluate the efficiency of HPV testing using an immunohistochemical staining kit with implications upon both diagnosis and treatment of cervical intraepithelial neoplasia (CIN).MethodsSeventy-nine patients and eighty-six controls were enrolled in the study. Each patient had completed a physical examination, gynecological examination with cervical sampling using a liquid-based cytology system and also colposcopy. The cervical samples were analyzed according to Bethesda terminology and HPV-HR immunohistochemical staining was performed. In all the patients with high-grade lesion a surgical excision procedure was performed followed by pathological examination of the specimen. The collected data were analyzed using statistical software.ResultsThe colposcopic examination has detected acetowhite modifications of the cervical epithelium in 47% of patients with ASC-US (Atypical squamous cells of undetermined significance) in 71% of patients with LSIL (Low grade squamous intraepithelial lesion) and in 100% of patients with HSIL ( High grade squamous intraepithelial lesion). The biopsy confirmed the diagnosis of LSIL in 27% of biopsy specimens in patients with ASC-US and in 79.99% of patients with LSIL respectively. In all patients with HSIL the diagnosis was CIN II or higher. The percentage of HPV-HR (Human Papilloma Virus – High Risk) positivity porportionaly increased with the severity of cytological diagnosis: 30% in ASC-US, 42.86% in LSIL and 75% in HSIL patients. The sensitivity of detection of HPV-HR was 50% with CI 95% [17.45;82.55] for ASC-US, 77.77% with CI 95% [51.91;92.62] for LSIL and 81.81% with CI 95% [58.99;94.00] for HSIL.ConclusionHPV testing can be an important screening tool for cervical dysplasia. The HPV testing targeting high risk types is indicated for ASC-US and LSIL triage. The present work sustains the idea of introducing HPV testing as a primary screening tool for cervical cancer.

Cytologic surveillance versus immediate colposcopy for women with a cervical smear diagnosis of low-grade squamous intraepithelial lesion in a poor setting in Nigeria.

BackgroundIdeally, all patients with abnormal Papanicolaou smear cytology results should undergo colposcopic examination of the cervix, but low-grade squamous intraepithelial lesions (LSILs) can also be followed up with further Papanicolaou smear surveillance. The aim of this study was to evaluate the outcome of cytologic surveillance versus immediate colposcopy in women with a cervical smear diagnosis of LSIL.MethodsThis was a prospective comparative study of 240 eligible consenting women consecutively grouped into cervical surveillance for 6 months versus immediate colposcopy at a ratio of 1:1. Free cervical smear cytology, colposcopy, and biopsy, as well as histology, were provided for all study participants.ResultsThe regression, persistence, and progression rates with 6 months of cytologic surveillance of LSIL were 46.1%, 43.4%, and 3.9%, respectively. The difference between the proportions of women who had an eventual histologic diagnosis of cervical intraepithelial neoplasia 2+ in both groups was not statistically significant (4.9% versus 8.7%; P=0.68). The default rates among women on cytologic surveillance and immediate colposcopy were 37% and 12.5%, respectively (P=0.0002).ConclusionAlthough the progression rate of LSIL is low, a high persistence rate and higher default rate from cytologic surveillance highlight the need to consider immediate referral for colposcopy, where available, for all women with a Papanicolaou smear diagnosis of LSIL in this environment.

Genotype distribution of human papillomavirus in women from the state of Bahia, Brazil

The aim of this study was to evaluate the human papillomavirus genotypes and the frequency of multiple human papillomavirus infections, as well as to assess the association between human papillomavirus genotype, cyto-histopathological abnormalities and age range. A retrospective cross-sectional study was carried out between June 2010 and October 2013 in Salvador, Bahia, Brazil. We analyzed 351 results of positive human papillomavirus genotyping performed using the PapilloCheck(r) test, designed to detect 24 human papillomavirus types. The cyto-histopathological abnormalities were classified as negative (negative cytology and histopathology), low-grade lesions (cytologic low-grade squamous intraepithelial lesion diagnosis or histopathologic cervical intraepithelial neoplasia 1 or vaginal intraepithelial neoplasia 1 diagnosis) and high-grade lesions (cytologic high-grade squamous intraepithelial lesion diagnosis or histopathologic cervical intraepithelial neoplasia 2+ or vaginal intraepithelial neoplasia 2+ diagnosis). The most frequently detected high risk human papillomavirus genotype was HPBV 16, with 18.5%, 95% confidence interval (95%CI) 14.6-23.0, followed by HPV 56 (14%; 95%CI 10.5-18.0) and HPV 39 (13.4%; 95%CI 9.5-16.8). HPV 18 (5.4%; 95%CI 3.3-8.3) was among the least frequent types. Among the low risk types, HPV 42 (15.7%; 95%CI 12.0-20.0), HPV 6 (11.4%; 95%CI 8.3-15.2) and HPV 44/55 (11.1%; 95%CI 8.0-14.9) were the most frequent, while HPV 11 (2.8%; 95%CI 1.4-5.2) was the least common. The proportion of HPV 16-positive women increased with severity of cyto-histopathological abnormalities: 13.8% (12/87) in low-grade lesion and 42.4% (14/33) in high-grade lesion. There was association between low- or high-grade cyto-histopathological lesion and the high risk genotypes, HPV16, HPV 52, HPV 73 and HPV 82, and the low risk type, HPV 43. Women under 30 years showed a significantly higher frequency of HPV 16 (22.2 versus 12.9%, p =0.01), HPV 42 (19.7 versus 10.9%, p=0.01) and HPV 45 (6.6 versus 1.4%, p=0.01), and multiple human papillomavirus infections (58.1 versus 47.4%, p=0.04). We observed variability of human papillomavirus genotype distribution in women from the state of Bahia. HPV 16 was the most frequently detected high risk human papillomavirus, as also reported for other geographic areas of Brazil and for the world in general. HPV 56 and HPV 39 were the second and the third most common genotypes, whereas HPV 18 was among the least frequent types. HPV 42, 6 and 44/55 were the most frequently detected low risk human papillomavirus, and HPV 11 was the least common.

Methylated ZNF582 gene as a marker for triage of women with Pap smear reporting low-grade squamous intraepithelial lesions — A Taiwanese Gynecologic Oncology Group (TGOG) study

ObjectiveOur previous work revealed that host genes ZNF582, PTPRR, PAX1, and SOX1 are highly methylated in cervical intraepithelial neoplasias grade 3 or worse (CIN3+). In this study, we used a standardized testing assay to evaluate the clinical efficacy of these biomarkers in the triage of cytological diagnoses of low-grade squamous intraepithelial lesions (LSILs), and compared the performance with human papillomavirus (HPV) testing. MethodsThis 2-year multicenter prospective study examined a population of 230 women from 12 medical centers who were diagnosed with LSILs on cervical cytology. Cervical scrapings were obtained prior to a colposcopy-directed biopsy for quantitative methylation analysis of ZNF582, PTPRR, PAX1, and SOX1, and HPV testing. Using logistic regression and receiver operating characteristic curve analyses, the abilities of methylated genes and HPV to predict CIN3+ were assessed. ResultsFifteen (6.5%) of the 230 women with a cytological diagnosis of LSIL were confirmed to have CIN3+ after a colposcopy-directed biopsy. Among the 4 methylated genes, ZNF582 was found to be the best biomarker for detecting CIN3+. The sensitivities for methylated ZNF582 and HPV testing were 73% and 80%, and the specificities were 71% and 28%, respectively. The odds ratio for predicting CIN3+ using methylated ZNF582 was 6.8 (95% confidence interval (CI) 2.1–22.1), which was much better than HPV testing (OR=1.6, 95% CI 0.4–5.8). ConclusionThis is the first study to show that ZNF582 methylation analysis of cervical swabs may be a promising choice in the positive triage of cytological diagnoses of LSILs.

HPV L1 capsid protein detection in high-risk human papillomavirus-positive cervical smears

Background: The L1 capsid protein is a viral nuclear protein that encapsidates the human papillomavirus (HPV) DNA to build new infectious particles. Previous studies in immune-competent patients have shown that detectable L1 protein in lowgrade squamous intraepithelial lesion (LSIL) smears is associated with remission in 60-75% of cases. Thus, the test can reduce interventions by approximately 75%. Objective: This study was performed to evaluate the use of HPV L1 capsid protein detection on cytology samples in a population with relatively high human immunodeficiency virus (HIV) prevalence and with known high-risk HPV (hrHPV) DNA results. Setting: Samples were obtained during a cervical cancer screening study at primary healthcare clinics in the Tshwane district, Gauteng. The HIV prevalence of the target group was estimated to be between 20% and 30%. Method: Conventional cervical cytology smears of 575 women were microscopically assessed and diagnosed. In addition, women were tested for the presence of HPV DNA on a cervical or vaginal sample. Immunocytochemical analysis was performed on morphologically abnormal smears and on a 52 control group smears reported to be negative for HPV DNA and morphological abnormalities. The detection of L1 capsid protein was carried out with the Cytoactiv® HPV L1 screening set, an antibody-based immunocytochemical stain. Results: A cytological diagnosis of LSIL was made in 19 women (3.3%), high-grade squamous intraepithelial lesions (HSILs) in 42 (7.5%) and malignancy in 1 (0.2%). Of the LSIL cases, 10 of 19 (52.6%) stained positive for the presence of the L1 protein, while only one of 42 HSIL (2.4%) cases stained positive. Three hundred and four cases (52.9%) tested positive for hrHPV DNA, including women with LSIL, HSIL and malignancy. All of the control cases stained negative for the L1 capsid protein. Conclusion: hrHPV was not a useful triage test for women with abnormal cellular morphology in this population. Promising results were reported for Cytoactiv® immunostaining as a triage test for patients with LSIL, but it added no value to samples known to be HPV-negative or reported to be morphologically negative. The immunostaining of smears reported to be HSIL or worse were almost universally negative, supporting the diagnostic accuracy of cytology and the expectation of persistence or progression of these lesions.

Verification of doubtful PAP smear results of women included in the screening program in the Podlaskie province

Verification of uncertain PAP-smear results in a group of women covered by the cervical screening program in the Podlaski province. The main aim of the study was to identify CIN (Cervical Intraepithelial Neoplasia) lesions present, with varying degrees of severity in women with cytological diagnosis of ASCUS (atypical squamous cells of undetermined significance), LSIL (low grade squamous intraepithelial lesion), and ASC-H (atypical squamous cells - cannot exclude high grade squamous intraepithelial lesion). The study evaluated 101 cervical smears taken from the vaginal part of the cervix in a group of screened women in the Podlaski province. Cytological evaluation was performed according the Bethesda System. We analyzed abnormal smears selected from a total of 7296 cytological examinations performed during 2012 at the University Center for Pathomorphological and Genetic - Molecular Diagnosis, Medical University in Białystok. The cytological results which were of interest to us included 19 cases with ASCUS, 59 with LSIL, and 23 with ASC-H, as well as with morphological features of the presence of Human Papilloma Virus (HPV). Staining was performed using CINtecPLUS test according to the manufacturer's instructions. CINtecPLUS is a immunocytochemical test based on specially designed monoclonal antibodies (E6H4TM) that let us identify protein p16ink4a within the cervical smear Additionally the diagnostic kit was provided with antibodies for diagnosing the presence of Ki-67 protein, a known marker of cell proliferation. The result was considered positive when staining of the nucleus and the cytoplasm appeared in red and brown, respectively. All abnormal results were eventually verified by histological examination of the tissue taken from cervical lesions by diagnostic-therapeutic procedure following colposcopic evaluation of cervical lesion topography In the group of cytological smears with ASCUS, the diagnosis was positive in 5 cases (26.3%), negative in 14 (73.7%). In the group with the diagnosis of LSIL, the cytology results were positive in 32 cases (54.2%), negative in 27 (45.8%). In the cytological diagnosis of ASC-H there were 20 positive (87%) and 3 negative (13%) results. Test CINtecPLUS could be a helpful tool in the final diagnosis of cervical abnormality in patients with the cytological diagnosis of ASCUS, LSIL and ASC-H. The combination of conventional cytological test and CINtecPLUS can help create a new procedure algorithm for cases with abnormal or ambiguous cytological screening results. It could be especially useful in a group of young women of childbearing age, when it is common to avoid a more radical treatment of cervical lesions.

Probiotics enhance the clearance of human papillomavirus-related cervical lesions

Probiotics have been proposed for a number of urogenital infectious conditions. In this study, we examine a possible effect on human papillomavirus (HPV)-related precancerous lesions in cervical cytology. We conducted a prospective controlled pilot study, in which 54 women with an HPV+low-grade squamous intraepithelial lesion diagnosis in their PAP smear were followed for 6 months. The intervention group consumed a daily probiotic drink during the study period; the control group received no treatment, according to common care policy. Outcome measures were the control PAP smear and HPV status after 6 months. Probiotic users had a twice as high chance of clearance of cytological abnormalities (60 vs. 31%, P=0.05). HPV was cleared in 19% of control patients versus 29% of probiotic users (P=0.41). This exploratory pilot study suggests that the probiotic studied promotes the clearance of HPV-related cytological abnormalities. If confirmed, this would represent an entirely new option to manage cervical cancer precursors.

Detection of abnormal cervical cytology in Papanicolaou smears

Background:Cervical cytology by Papanicolaou (Pap) smears is an effective means of screening for cervical premalignant and malignant conditions. Cervical intra-epithelial neoplasia (CIN) and cervical cancer remain important health problems for women worldwide.Aim:To study the role of Pap smear in detecting premalignant and malignant lesions of cervix; and to determine the prevalence of various lesions.Materials and Methods:This study is based on 300 patients who attended the out-patient Department of Obstetrics and Gynaecology. Pap smears were prepared from patients presenting with complaints like vaginal discharge, post-coital bleeding, inter-menstrual bleeding, dyspareunia, and pain lower abdomen. After fixation and staining, each smear was carefully examined.Results:Epithelial cell abnormalities were found in 5% smears, atypical squamous cells of undetermined significance (ASCUS) in 0.3%, squamous intraepithelial lesion (SIL) in 3.4% which includes low grade squamous intraepithelial lesion (LSIL) (2.7%) and high grade squamous intraepithelial lesion (HSIL) 0.7%. Invasive carcinoma was seen in 1.3% cases. Mean age of the patients with diagnosis of LSIL was 32.3 years and for HSIL, it was 40.5 years. The mean age of the patients with invasive carcinoma was 57 years.Conclusion:Premalignant and malignant lesions of cervix are not uncommon in our set up and can be diagnosed early by Pap smears.

Clinical Performance of a Human Papillomavirus Messenger RNA Test (Aptima HPV Assay) on Residual Material From Archived 3-Year-Old PreservCyt Samples With Low-Grade Squamous Intraepithelial Lesion

Human papillomavirus (HPV) testing is widely used in the triage of women with a borderline smear result but the efficiency of testing women with low-grade squamous intraepithelial lesion (LSIL) is less clear, mainly because of lack of specificity. New HPV tests are emerging, which detect E6/E7messenger RNA (mRNA), and preliminary data suggest that they might have a higher specificity. However, mRNA is less stable than DNA, thus posing a challenge to the preservation abilities of the cell-collecting medium. To evaluate the clinical performance of an HPV mRNA assay on 3-year-old archived liquid-based samples, all with a diagnosis of LSIL. The residual material from old archived PreservCyt samples from 442 women were tested with the Aptima HPV Assay, which detects E6/E7 mRNA from 14 high-risk HPV types. The samples had been stored at room temperature without any further handling. Follow-up was available for 405 women, 67 of whom had histologic confirmed cervical intraepithelial neoplasia (CIN) 2+ and 31 with CIN 3+. The sensitivity and specificity for the mRNA assay was 92.5% and 38.2%, respectively, for detecting CIN 2+, and 93.9% and 35.5%, respectively, for detecting CIN 3+. When evaluating separately the performance of the test for women younger than 30 years and for women 30 years or older, the sensitivity was found to be similar in the 2 groups, but the specificity was significantly lower for the younger women. Messenger RNA is well preserved in old archived PreservCyt samples. Triaging women with LSIL, using the Aptima HPV Assay, seems to be effective with a good sensitivity and a good specificity, especially for women 30 years or older.