Journal

Acta Cytologica

Papers (58)

Endocervical Cytology: Inter- and Intra-Observer Variability in Conventional Pap Smears

Introduction: Although the cytological diagnostic criteria for cervical squamous and glandular lesions are established by the Bethesda System for Reporting Cervical Cytology, the reproducibility of the diagnosis of these lesions has been shown to be variable in previous studies. At best, occasional good kappa (κ) values were reached both inter- and intra-observerly. Generally, consensus on high-grade lesions has been better compared to milder changes. Methods: Altogether, 167 conventional Pap smears from 50 patients with histologically confirmed endocervical adenocarcinomas (EAC) and adenocarcinomas in situ (AIS) and from 28 patients with histologically proven high-grade intraepithelial lesions were analyzed by four cytopathologists. Twenty of the smears were later re-evaluated by the same cytopathologists. κ-values between cytopathologists in the categories of squamous versus glandular, negative for intraepithelial lesion or malignancy (NILM), atypical, and preneoplastic/neoplastic were calculated. The diagnostic Pap smears of EAC and AIS with best and worst consensus between observers were then morphologically analyzed. Results: The reproducibility ranged from poor to substantial. The overall κ-values between the four cytopathologists were 0.412, 0.314, 0.272, and 0.082, respectively, in the categories of preneoplastic/neoplastic, squamous versus glandular, NILM, and atypical. Overall intra-observer κ-values were correspondingly 0.491, 0.616, 0.345, and 0.241. In the diagnostic smears of AIS and EAC, the nuclear size &#x3e;2 times the normal and nuclear pleomorphism were the commonest features associated with good diagnostic consensus and the lack of nuclear enlargement and degenerative changes were associated with poor consensus. Conclusions: The reproducibility of preneoplasia/neoplasia diagnoses was better than that of atypia and NILM both in the inter- and intra-observer part in this study. In the smears from AIS and EAC patients, general neoplasia-associated features were more common in samples with good agreement by the four cytopathologists of the neoplastic nature and the endocervical origin of the lesion.

Discriminant Analysis Using Gabor Filter Sets for Lobular Endocervical Glandular Hyperplasia: Numerical Interpretation of Nuclear Atypia by Gabor Filter Features

Introduction: Lobular endocervical glandular hyperplasia (LEGH) is a benign lesion; however, it is considered to be the origin of gastric-type adenocarcinoma in the uterine cervix, and early diagnosis is important. At Shinshu University Hospital, screening of LEGH cells is based on the difference in color tone of cytoplasmic mucin on Papanicolaou staining and detection of gastric mucin using HIK1083-labeled latex agglutination assay. However, it is sometimes difficult to distinguish LEGH cells with subtle nuclear atypia from endocervical (EC) cells. Methods: We calculated the Gabor filter features (mean signal value, standard deviation, skewness, kurtosis) from the nuclei of cytological specimens in EC cells (37 cases) and LEGH cells (33 cases) using microscopic images, and we performed statistical analysis and discriminant analysis by linear support vector machine (LSVM) using these features. A Gabor filter is a linear filter defined as a mathematical representation of the mammalian visual system. Gabor filters with three wavelengths and eight angles were used for analysis. Results: Gabor filter features in EC cells were higher than in LEGH cells, demonstrating that the gradient of LEGH cell nuclei was milder than that of EC cell nuclei. The accuracy calculated using all Gabor filters was 91.0% and the accuracy of four Gabor filters (λ = 2/3π and θ = 0°, 45°, 90°, 135°) was 88.9%. High accuracy with low computation costs was achieved by reducing the number of features used for LSVM. Conclusion: The application of a Gabor filter with convolutional processing resulted in the edges of LEGH cells being slightly rough and thick, whereas those of EC cells were fine and thin. Thus, it is thought that the frequency of abrupt gradients of pixels was higher in EC cells than in LEGH cells, and the gradient of chromatin distribution in LEGH cell nuclei was milder than that in EC cell nuclei. It was possible to evaluate nuclear findings of EC and LEGH cells objectively by quantifying morphological features of nuclei using Gabor filters. It was possible to differentiate EC cells from LEGH cells using LSVM using Gabor filter features.

p16INK4a/Ki67 Dual-Staining Immunocytochemistry to Refer Women Infected by High-Risk Human Papillomavirus for Colposcopy

Background: Cervical cancer is still a significant global health issue, especially in low- and middle-income countries. Human papillomavirus (HPV) infection is known to be the primary etiological factor in the development of cervical cancer. The identification of high-risk HPV (HR-HPV) infections in cervical cancer screening is becoming crucial for early detection and management of cervical pre-neoplasia. However, HPV testing requires triage markers since HR-HPV infections are widespread in the target population, of which only a small fraction requires detailed clinical workup by colposcopy. Summary: p16INK4a is a tumor suppressor protein that has emerged as a promising surrogate marker for HPV infection. In combination with a proliferation marker Ki67 in one diagnostic test kit referred to as CINtecPlus® or dual stain (DS), it revealed excellent sensitivity and specificity profiles to detect clinically relevant CIN2+ lesions among HPV-infected women in large screening cohorts. Key Message: This review examines the role of the cytology CINtecPlus® test (DS test) as a triage marker to identify patients with CIN2+ lesions among HPV+ tested women and summarizes the most recent recommendations for its clinical use.

The Founding Pioneer Cytotechnologists: The Women Who Assisted George N. Papanicolaou, MD, PhD, Develop the Pap Test for Cervical Cancer Prevention

Background: The earliest cytotechnologists are largely unknown. Summary: In 1943, the book “Diagnosis of Uterine Cancer by the Vaginal Smear” by Papanicolaou and Traut recognized several women who have largely faded from memory. While Mary Papanicolaou and Charlotte Street are familiar names, others like Alberta Kuder and Huldah Boerker, who inadvertently laid the groundwork for the field of cytotechnology, remain obscure. There were also women like Christine Rassias and Adele Reboul who did not receive recognition. Notably, Mrs. Lady Mary G. Papanicolaou, despite her significant contributions both in the lab and at home since 1914, was not acknowledged in her husband’s work until the publication of his Atlas in 1954. Key Message: These women set the benchmark for future cytotechnologists, unknowingly shaping the profession as we know it today.

Cytomorphological characteristics of cervical scrape from controlled and uncontrolled diabetes mellitus patients attending tertiary healthcare, Kenya

Background: Globally, the concurrence of diabetes mellitus and cervical cancer among women is compounded and increasing health burden. Furthermore, the incidence and prevalence of both diseases are increasing, particularly in Middle and Low-income countries. In Kenya, 19.7 % of new cancer cases among women is attributed to cervical cancer. While diabetes causes metabolic disorders and immune alterations that may promote persistent high-risk Human Papilloma virus infection, the magnitude of cervical cytomorphological abnormalities among diabetes women in Kenya remains unclear. Aim: To determine the cytomorphological characteristics of cervical scrape samples obtained from patients with controlled and uncontrolled diabetes mellitus who visited the Kitale County Hospital, Kenya. Methodology: This prospective, hospital-based analytical cross-sectional study was conducted between April 2023 and June 2024. A total of 156 patients with diabetes (n=156) between the ages of 18 and 65 years were enrolled. Alcohol fixed cervical smears were processed using Papanicolaou staining method. The findings were reported using the 2014 Bethesda system for reporting cervical cytology. Images were captured for cytomorphological abnormalities using X20 mobile camera. Rstudio 4.2.2 (2024-10-31 ucrt) software was used to analyze data. Mann-Whitney U test was used to compare distributions of laboratory characteristics between controlled and uncontrolled diabetes. Fisher’s exact test was performed to explore associations between diabetes control and cytomorphological abnormalities. Chi-square tests were performed to determine relationship between diabetes control and presence of cervical abnormalities. Logistic regression was performed to assess the relationship between glycemic levels and presence of cytomorphological abnormalities. Results: Out of 156 cases, cytomorphological abnormalities were reported in 63.5%. Negative for Intraepithelial lesion or Malignancy accounted for 84.0% while 16% were cervical intraepithelial lesions. Non-neoplastic abnormalities/infections were recorded in 48.1% of study participants. The study showed that there is statistically significant difference in the central tendency of age between controlled and uncontrolled diabetes Mellitus [95% CI= 2.0-10.0, p=0.001]. There was statistical significance association between reproductive status and presence of cervical infections [OR=2.18, 95% CI= 1.15 - 4.18, p=0.016]. There was no significant association between diabetes control and occurrence of cervical cytomorphological abnormalities [OR=1.47, 95% CI=0.76-2.86, p=0.704]. While a unit increase in HBA1c was associated with (1.12) times higher odds of cytomorphological abnormality, it was not statistically significant [OR=1.12, CI=-0.11-0.245, p=0.080]. Conclusion: The current study points both groups of patients with controlled and uncontrolled diabetes mellitus are at higher risk of cervical cytomorphological abnormalities. Further, glycemic control showed no significant association with occurrence of cervical cytomorphological abnormalities.

Image Quantification Analysis of Cytoplasmic Mucin and Interpretation of Mucin Color in Lobular Endocervical Glandular Hyperplasia

Introduction: Although the widespread use of screening tests and HPV vaccines for squamous cell carcinoma has led to early detection and treatment, effectiveness is limited for cervical adenocarcinoma. Lobular endocervical glandular hyperplasia (LEGH) corresponds to gastric metaplasia but is regarded as a pathological condition with subtle morphological abnormalities. LEGH is a benign lesion and a precursor to gastric-type adenocarcinoma. We herein developed an objective and quantitative method by applying an image analysis technique to overcome the difficulties associated with the differential diagnosis of LEGH in uterine cervical cytology. This approach is expected to enable the early detection and accurate diagnosis of LEGH. Methods: We extracted signal values for the nucleus and cytoplasm from microscopic images of cytological specimens of normal endocervical (EC) and LEGH cells. These values were then converted into CIELAB and sRGB values to create color distribution maps, and color unmixing techniques were applied to assess the spectral absorbance of each pigment. Results: The CIELAB signal values extracted from the nuclear images of LEGH cells exhibited lower values than those of EC cells. Furthermore, based on color distribution maps, the cytoplasm of EC cells exhibited shades from purple to pink, while LEGH cells showed a distribution toward yellow. Conclusions: This study reveals that, compared to EC cells, LEGH cells exhibit lower nuclear signal values and increased nuclear chromatin content. Thus, assessing the relative difference in cytoplasmic color tones between them may become an effective indicator for distinguishing between EC and LEGH cells.

Utility of p16INK4a Staining on Cell Blocks Prepared from Residual Liquid-Based Cervicovaginal Material

Introduction: Cervical cancer screening using Pap smears is affected by false-negative results. Liquid-based cytology (LBC) offers the technical advantage of preparing cell blocks from residual fluid to conduct ancillary tests on them. The p16INK4a gene product has been shown to be strongly overexpressed in dysplastic cervical epithelia and serves as surrogate marker for high-risk human papilloma virus infection. Materials and Methods: Microwave-processed cell blocks were prepared from residual material in vials after ThinPrep slide preparation, stained with hematoxylin and eosin and p16INK4a. Nuclear staining with or without cytoplasmic staining on p16 slides was considered positive. Four parameters were evaluated: percentage of positive cells, intensity of staining, number of positively stained cells in close contact, and full-thickness epithelial staining. We compared sensitivity and specificity of ThinPrep smears and p16-stained cell blocks in diagnosing invasive malignancy. Results: The intensity and percentage of p16-positive cells was found to increase with increasing grade of cervical abnormality. We found good concordance between ThinPrep smear and cell block diagnoses in cases which were negative for intraepithelial lesion or malignancy (97.6%), in low-grade squamous intraepithelial lesions (90%), high-grade squamous intraepithelial lesions (100%), and squamous cell carcinomas (93.5%). Of 16 discrepant cases, 9 were reported unsatisfactory on ThinPrep smears due to abundant necrosis or scant cellularity. All these turned out to have malignancies on follow-up and review of histology. The sensitivity of ThinPrep and p16-stained cell blocks in diagnosing invasive malignancy were 70.2% and 85.1%, respectively, while the specificity of both was 100%. Conclusions: Cell blocks prepared from residual fluid in LBC vials have the potential to reduce the rates of inadequacy and are feasible in routine practice. While the cost of p16 on cell blocks may be too prohibitive for use in routine cervical screening programs, if used judiciously in combination with clinical suspicion, a lot of valuable material which is usually discarded in the residual LBC vials can prove to be crucial in arriving at the correct diagnosis.

Pitfalls in Gynecological Cytology: Review of the Common and Less Frequent Entities in Pap Test

Background: Pitfalls in Pap test could be defined as false positive, false negative, or underdiagnosed results which can lead to unnecessary diagnostic procedures or delayed and inadequate treatment. It can be a consequence of misinterpretation of certain morphological entities which are described in this paper. Summary: The paper presents an overview of the morphological features and look-alikes of the common sources of pitfalls such as atrophy, repair, intrauterine device change, tubal metaplasia, hyperchromatic crowded groups, and radiation changes. Rare causes of pitfalls such as Arias-Stella changes, pemphigus, tumor diathesis per se, rare types of cervical cancer, including verrucous and papillary squamous cell cancer, gastric type, and endometrioid adenocarcinoma are also described. Key Messages: The awareness of pitfalls in cervical cytology is important for cytopathologists and clinicians to avoid future errors. Review of Pap tests with erroneous diagnosis is important for quality control in cytology laboratory, and it must be considered an educational- and experience-building procedure. Cytopathologist should not pull back in significant diagnoses, especially in human papillomavirus-negative cases.

PAX1 and SOX1 Gene Methylation as a Detection and Triage Method for Cervical Intraepithelial Neoplasia Diagnosis

Introduction: Methylation assays have demonstrated potential as dependable and high-precision approaches for identifying or triaging individuals with cervical cancer (CA) or cervical intraepithelial neoplasia (CIN). Our investigation aimed to assess the efficacy of the diagnosis and triage of the PAX1/SOX1 methylation panel in detecting CIN or CA. Methods: A total of 461 patients with abnormal high-risk human papillomavirus (hrHPV) or cytology test results were recruited for this study. Each patient underwent an assortment of assessments, comprising a cytology test, hrHPV test, colposcopy examination, and PAX1 and SOX1 methylation tests. Results: The extent of methylation of both genes demonstrates a positive correlation with the severity of CIN lesions and CA. To determine the correlation for patients with CIN2 or worse (CIN2+), the area under curve was 0.821 (95% CI: 0.782–0.853) for PAX1 and 0.800 (95% CI: 0.766–0.838) for SOX1, while for CIN3 or worse (CIN3+), 0.881 (95% CI: 0.839–0.908) for PAX1 and 0.867 (95% CI: 0.830–0.901) for SOX1. The PAX1/SOX1 methylation marker panel performed sensitivity and specificity of 77.16% and 91.67% for CIN2+, 84.76% and 90.50% for CIN3+, respectively. Regarding triaging hrHPV+ patients, the PAX1/SOX1 methylation test only referred 11.83% of the patients who are unnecessary for colonoscopy examination, which is comparatively lower than cytology, thereby signifying a promising triage strategy for hrHPV-positive women. Furthermore, we observed that the positive PAX1/SOX1 methylation test result for untreated CIN1 or fewer patients would result in a higher likelihood of progression upon a 24-month follow-up visit. Conclusion: The present investigation demonstrates that the PAX1/SOX1 methylation marker panel exhibits favorable diagnostic performance in CIN detection and holds the potential to be employed for individual CIN tests or hrHPV-positive triage.

Analysis of the Differences between Bethesda Groups according to Conventional Smear and Liquid-Based Cytology Methods in Cervicovaginal Cytology: A Single-Center Experience with 165,915 Cases

Introduction: Liquid-based cytology (LBC) has replaced conventional smear (CS) in the world. In this study, through a series with a large number of cases, we aimed to make a comparison and general evaluation in all groups, primarily epithelial abnormalities, according to LBC and CS methods. This study was carried out in a private pathology laboratory located in a metropolitan city, where cytological materials sent from many clinics were examined. Material and Methods: There were 165,915 cases whose smears were examined between 2012 and 2020, most of them conventional (131,224 CS, 34,691 LBC). Cases were evaluated on the basis of the Bethesda 2014 classification and divided into sub-diagnostic categories after they were divided into two main groups as “with epithelial abnormalities” and “without.” χ2 and Fischer’s precision statistical tests were conducted using SPSS 23.0 package. In the CS process, cervical samples were obtained using an endocervical brush and a spatula. Cells were directly spread onto the slides and promptly fixed in 95% ethanol, followed by staining with the standard Papanicolaou stain. For LBC ThinPrep, cervical specimens were gathered using a cervix brush. The brush was washed in a vial and discarded. Finally, cells were isolated through vacuum filtration and transferred to the slide using air pressure. Results: Squamous cell abnormalities (atypical squamous cells of undetermined significance [ASC-US], atypical squamous cells – cannot exclude high-grade squamous intraepithelial lesion [ASC-H], low-grade squamous intraepithelial lesion [LSIL], high-grade squamous intraepithelial lesion [HSIL], squamous cell carcinoma, atypical glandular cells of undetermined significance) were reported in 5,696 (3.43%) cases. ASC (ASC-US + ASC-H)/SIL ratio (1.36/2.04) was found to be 0.67 (recommended Bethesda ratio is &lt;3). ASC-US (p &lt; 0.001), ASC-H (p &lt; 0.001), and HSIL(p &lt; 0.001) detection rate of LBC was found to be significantly higher than CS. ASC-US (1.8/1.2), ASC-H (0.08/0.008), and HSIL (0.6/0.3) case ratios of LBC/CS were found to be significantly higher in LBC. LSIL (1.72/1.66) rate was similar. Conclusion: LBC is superior to CS in detecting epithelial lesions. In addition to being used as a screening method, it is clear that it makes a great contribution to reducing cervical carcinomas due to HPV typing. Definitive comments regarding comparison of methods on reactive changes and microorganism detection are challenging. Preanalytical factors might account for these situations.

Accuracy and Reproducibility of Cytology Triage in a HPV-Based Primary Screening Setting: A Revision of 384 Pap Tests

Introduction: After the transition toward the HPV-based screening protocol, which has led to an increase in sensitivity, and in order to bring the specificity back to acceptable values, cytology underwent a change of approach, becoming a triage test. For these reasons, in the Tuscany region (after the recommendations of the GISCi document), it was decided to reduce, as much as possible, the use of ASC-US category in cytology triage, classifying these morphological cases as negative for intraepithelial lesion or malignancies (NILM) or LSIL, basing on the grade of nuclear atypia. So, in Italy, in a cytology triage context (HPV primary screening), a modified Bethesda system (TBS) is currently used. The aim of this study was to evaluate the performance of the review activity of 384 cytology triage cases and of the cervical cancer screening indicators (sensitivity and specificity for CIN2+ lesions) using the TBS 2014 or the modified TBS. Materials and Methods: 384 HPV positive cases at one-year recall (192 with a cytology result of NILM both at baseline and at one-year recall; 192 with a cytology result of NILM at baseline but abnormal at one-year recall), all with a histologically confirmed result (128 CIN2+, 256 ≤ CIN1), were selected, and their baseline Pap tests were reviewed in blind mode by 5 expert cytologists. Results: The cytological results of NILM were confirmed for 92.5% and 83.8% of cases using TBS 2014 or modified TBS, respectively. 20/128 CIN2+ cases could have been reported at the baseline cytology triage, causing an anticipatory effect and an improvement in sensitivity of the screening protocol at baseline (+15.6%). Using TBS 2014, the number of false positives more than tripled with respect to the modified TBS 2014, with a significant increase in unnecessary colposcopies (+11.4%). Conclusion: This work demonstrated that a greater expertise of cytologists, acquired during the following 3 years of experience with cytological triage, and a strong IQC system could lead to the identification of a significant number of lesions reported to baseline rather than at one-year recall (diagnostic anticipation).

False-Positive Atypical Endocervical Cells in Conventional Pap Smears: Cyto-Histological Correlation and Analysis

Introduction: Endocervical glandular atypia is relatively rarely diagnosed by Pap smears. A significant proportion of follow-up histological samples show no premalignant or malignant lesions. The observed cytomorphological findings in premalignant glandular lesions overlap with histologically proven reactive lesions. Methods: A total of 45 conventional Pap smears diagnosed as atypical endocervical cells, not otherwise specified (AEC, NOS) with human papillomavirus (HPV) status available were blindly evaluated in a search for 38 cytomorphological features representing background, architectural, cellular, and nuclear features. Of the cases, 30 represented histologically proven benign changes, and 15 represented histologically proven adenocarcinoma in situ (AIS) or endocervical adenocarcinoma (EAC) cases. The benign biopsies were re-evaluated, and the associations of the cytomorphological features or combinations of them with specific histological features and entities were statistically examined. Results: The most frequent histological findings in the benign group were squamous metaplasia, inflammation, tubal metaplasia, and microglandular hyperplasia. The statistical analysis revealed cytological features associated with squamous metaplastic changes, inflammation, and microglandular hyperplasia. Unfortunately, no cytomorphological feature was sufficiently specific to confidently leave the lesion without follow-up and histological correlation. Degeneration and nuclear crowding were the most salient features that distinguished the instances of glandular atypia with benign follow-up histology from those with histologically proven AIS or EAC (26.7 vs. 60.0%, p = 0.030, and 50.0 vs. 86.7%, p = 0.017). Conclusion: Additional methods besides cytomorphology are required to reliably distinguish smears with AEC, NOS harbouring only benign histological changes from those exhibiting endocervical glandular malignancy.

Histopathologic Follow-Up of Women with Inadequate ThinPrep™ Cervical Pap Tests and Positive High-Risk Human Papillomavirus Test Results

Introduction: We found only a few studies that had performed high-risk human papillomavirus (hrHPV) analyses of inadequate ThinPrep™ Papanicolaou (Pap) tests. Therefore, this study aimed to analyze unsatisfactory ThinPrep Pap tests using hrHPV tests. The colposcopic biopsy results of cases with an unsatisfactory ThinPrep Pap test and positive hrHPV results were revealed. Methods: Between January 1, 2018, and October 31, 2022, 965 (3.7%) of 25,958 liquid-based cytology specimens were evaluated as unsatisfactory. Ninety-five (9.8%) of 965 patients were positive for hrHPV. The colposcopic evaluation was performed in 28 (29.4%) of 95 patients, in whom 23 tests were adequate. Results: Twenty-three colposcopy biopsy results showed that 17 (73.9%) of 23 patients had benign biopsy results. High-grade squamous intraepithelial lesions were observed in three (13%) of the 23 patients, and low-grade squamous intraepithelial lesions were observed in two (8.6%) of the 23 patients. One of the 23 (4.3%) patients had keratinized squamous cell carcinoma of the cervix diagnosed histologically, although no tumor was visible upon gynecologic examination. Conclusion: For the management of unsatisfactory Pap tests, The American Society for Colposcopy and Cervical Pathology (ASCCP) recommends repeat cytology within 2–4 months. Evaluation of such patients using hrHPV tests may triage those with squamous intraepithelial lesions, even invasive cervical cancer. More studies with a larger number of cases are needed to analyze the hrHPV status and biopsy follow-up of cases with unsatisfactory cytology.

Cytology of Benign Peritoneal Fluids and Pelvic Washing Specimens: Diagnostic Cytomorphologic Features and Pitfalls

Background: Pelvic washing and peritoneal fluid cytology specimens are used to detect peritoneal spread of malignancies. In most cases, identification of malignancy in these specimens is straightforward, but benign processes may occasionally mimic neoplasia and cause diagnostic difficulty. Summary: In this article, we perform a focused review of common benign entities encountered in pelvic washing and peritoneal fluid specimens during routine practice which may cause difficulty and discuss helpful features for avoiding diagnostic pitfalls. Key Messages: Application of strict cytomorphologic criteria, along with judicious use of ancillary studies and correlation with clinical, intraoperative, radiologic, and other pathologic findings, can help resolve most problematic cases.

Clinical Usefulness of Endometrial Cytology in Determining the Therapeutic Effect of Fertility Preserving Therapy

Introduction: The significance of endometrial cytology in determining the therapeutic efficacy of medroxyprogesterone acetate (MPA) therapy is unclear. This study aimed to evaluate the clinical usefulness of endometrial cytology during MPA therapy. Methods: Overall, 77 patients who underwent dilatation and curettage (D&amp;C) to evaluate the therapeutic efficacy of MPA therapy at our hospital between January 2018 and December 2019 were retrospectively analyzed. The results of D&amp;C, cytological evaluation, and other clinicopathological factors were analyzed based on the patients’ medical records. Results: The sensitivity and specificity of cytology were 61% and 92%, respectively, with D&amp;C being the gold standard for diagnosis in 142 D&amp;C/cytological examinations. Among patients with no residual disease on D&amp;C, 5 (4%) had suspicious or positive cytology. Although MPA therapy was terminated in 3 of these patients, only 1 patient had early recurrence, and the frequency of recurrence was similar to that of patients who showed negative results in both D&amp;C and cytology. Discussion/Conclusion: The sensitivity of endometrial cytology in determining the therapeutic effect of MPA therapy is low, and we confirmed that the omission of D&amp;C is unacceptable. Our findings also suggested that the addition of cytological evaluation to D&amp;C during MPA therapy had a low clinical significance.

Influence of Estradiol-Producing Ovarian Tumors on the Maturation Index of Cervical Cytology in Postmenopausal Women

Introduction: The aim of the study was to evaluate the influence of estradiol-producing ovarian tumors, including surface epithelial-stromal tumors, on the cervical cytology of postmenopausal women. Methods: This case-controlled study included 160 postmenopausal women who underwent a gynecological surgery between January 2009 and December 2016. The relationship between serum estradiol levels and the maturation index of cervical cytology was examined. Patients with ovarian tumors and a high estradiol level (≥28 pg/mL) constituted the estradiol-producing ovarian tumor group (30 women, including 23 with surface epithelial-stromal tumors). The maturation index of this group was compared with that of the control group (130 women with normal estradiol levels [&#x3c;28 pg/mL] with either ovarian tumors or uterine tumors). Results: For all patients, the serum estradiol levels were significantly correlated with the maturation index (p &#x3c; 0.001, r = 0.65). The maturation index of the estradiol-producing ovarian tumor group was significantly higher than that of the control group (0.67 ± 0.21 vs. 0.075 ± 0.16, p &#x3c; 0.001). The area under the receiver operating characteristic curve for the maturation index was 0.94. The best maturation index cut-off level for estradiol-producing ovarian tumors was 0.20. Using this cut-off, the sensitivity and specificity were 94% and 82%, respectively. Conclusion: Estradiol-producing ovarian tumors influence cervical epithelial maturation in postmenopausal women. An increased maturation index may trigger the early detection of asymptomatic ovarian tumors.

HPV Screening Test for the Detection of Precancerous Cervical Lesions and Cervical Cancer in Israeli Women

Objectives: This work is aimed to summarize the first year of the high-risk human papillomavirus (hrHPV) screening test and compare it to the cytology screening test, regarding positivity rates and premalignant lesions diagnosed in the Israeli population. A specific consideration is for the age group 25–30 that is not considered mandatory for the HPV primary screening testing. Methods: A retrospective study was performed in women who were screened for prevention of cervical cancer in Maccabi HealthCare HMO from March 2017 to March 2019. Screening methods included hrHPV typing for types 16, 18, and the other 12 hrHPV types and the PAP LBC test. Results: A total of 115,807 cervical samples were tested for HPV presence and 91% (105,225) were found negative for hrHPV. The other 9% (10,582) were positive for one or more of the 14 hrHPV types tested, and 37% (3,916) of them showed abnormal PAP LBC results. In the age group of 25–30, 3,104 (17.5%) women were found positive for hrHPV (825 had hrHPV types 16 and/or 18), of which 42% (1,293) of them showed abnormal PAP LBC results. During the hrHPV versus PAP LBC screening era, 258 more women were diagnosed with precancerous cervical lesions (CIN2/3), 70% increased detection versus cytology screening. Conclusions: The hrHPV screening test is currently the best method for the detection of precancerous cervical lesions and cervical cancer, and it is better started at age 25.

The Application of DNA Ploidy Analysis in Large-Scale Population Screening for Cervical Cancer

Objective: The objective of this study was to evaluate the application of DNA ploidy analysis in large-scale population screening for cervical cancer. Methods: From March 2016 to March 2019, eligible subjects were enrolled and recommended to undergo DNA ploidy analysis, the ThinPrep cytology test (TCT), and high-risk human papillomavirus (hrHPV) detection concurrently. Patients with positive results were recommended for colposcopy, and biopsy diagnosis was regarded as the “gold standard.” We compared the test efficiencies of the 3 methods and compared the efficiency and accuracy of the TCT in our hospital and the “2-cancer screening” project in Hubei Province during the same period. Results: Among 20,574 women, the positive rates of DNA ploidy analysis, cytology, and hrHPV testing were 4.01%, 4.71%, and 16.28%, respectively. The sensitivities of these methods for screening for grade 2+ cervical intraepithelial neoplasia were 0.70, 0.68, and 0.96, and their specificities were 0.79, 0.82, and 0.45, respectively. On comparing DNA ploidy analysis with the TCT, there was no significant difference in the sensitivity, specificity, positive predictive value, negative predictive value, and missed diagnosis rate. In opportunistic screening and the 2-cancer screening project, the positive rates of cytology were 4.71% and 2.87%, respectively. And the efficiency and accuracy of the TCT in opportunistic screening were higher than in the 2-cancer screening project. Conclusion: Therefore, DNA ploidy analysis, which is of low-cost and does not depend on cytopathologists, can replace cytology and be applied in large-scale population screening for cervical cancer.

Back from the Future: Rational Accountabilities for Cytopathology in Pap Test Cervical Cancer Screening Arising from Covid19

Utility of Fallopian Tube Brush Cytology and Cell Blocks as a Screening Tool for Epithelial Ovarian Cancer in Patients Undergoing Gynaecological Surgeries for Benign and Malignant Indications: Attempt at a Classification System

Introduction: Fallopian tube (FT) cytology is an evolving and as yet not well-established field. Through this study, we aimed to establish the utility of FT brush cytology by stratification into cytological diagnostic categories. Methods: Cytological specimens were collected using an endobrush from the fimbrial end of the tubes at the time of gynaecological surgeries, and LBC preparation (liquid-based cytology slides prepared by SurePath technique) and cell blocks were prepared. Smears were stratified into unsatisfactory/non-diagnostic (ND), benign, atypical, suspicious of malignancy (SOM), and malignant. Correlation with histopathology was done, and the risk of malignancy (ROM) was calculated for each category. Negative predictive value (NPV) and positive predictive value (PPV) were calculated. Diagnostic accuracy was calculated. Results: A total of 392 tubal cytology specimens of 225 patients were collected. 8.2% (n = 32) of the specimens were unsatisfactory/ND, 87% (n = 343) were benign, 2.6% (n = 10) were atypical, 0.8% (n = 3) were SOM, and 1% (n = 4) were malignant. All the cases in the SOM and malignant categories were serous carcinomas on histopathology. Of the ten atypical cases, all were non-malignant on histopathology: two were serous tubal intraepithelial lesions and negative for serous tubal intraepithelial carcinoma (STIC), four showed salpingitis, and four showed normal histology. ROM for ND, benign, and atypical categories was 0%. ROM for the malignant category, as well as the SOM category, was 100%. NPV for the benign category, as well as the benign and atypical categories, was 100%. PPV for the malignant category, as well as the malignant and SOM categories, was 100%. Cell blocks were prepared for all cases, and the grey zone categories of atypical and SOM were reduced from 13 to 8. The diagnostic accuracy was 91.3% without and 99.4% with consideration of the ND category. Conclusion: FT brush cytology shows excellent concordance with the follow-up histopathology in all categories, barring the ND category. Excellent concordance with histopathology was seen in cases of the benign category, which comprised the majority of the samples (87.5%). Although excellent concordance was also seen in the other categories with the final histopathology, the number of samples in these categories was less for a definite conclusion. Cell block preparation, though useful, especially in the grey zone categories, did not offer statistically significant results. Another important finding was that not even a single case of incidental STIC was found. This finding raises questions on the accepted current routine practice of preventive salpingectomy for all in the correct setting.

Prognostic Value of Human Papillomavirus Genotyping before and after Cervical Conization for Squamous Intraepithelial Lesion

Introduction: We previously reported that preoperative human papillomavirus (HPV) type 16/18 positivity and postoperative high-risk (HR)-HPV test positivity are associated with abnormal postoperative cytology. In this study, we further examined whether preoperative and postoperative HR-HPV genotyping could predict cytological abnormalities and the risk of additional surgery. Methods: Patients who underwent cervical conization at our hospital between July 2009 and June 2018 were enrolled. HPV genotyping was performed preoperatively for all patients with HPV-positive. The association among preoperative and postoperative HR-HPV genotyping results, the cumulative risk of cytological abnormalities, and additional surgery were evaluated. The endpoint approach was used to investigate the cumulative incidence of additional surgery owing to cytological abnormalities, such as low-grade squamous intraepithelial lesion positivity and recurrence 2 years after cervical conization. Results: Positive and negative histological margins were observed in 21 and 287 of 308 cases, respectively. The cumulative incidence of abnormal cytology and additional surgery was significantly higher in margin-positive cases than that in margin-negative cases. Examination established according to the margin status demonstrated that the postoperative HR-HPV-positive group had a significantly worse prognosis than the HR-HPV-negative group. Additionally, 32 cases, wherein the same genotype detected before and after surgery, demonstrated significantly unfavorable outcomes. Fifteen patients with persistent HPV 16/18 had the worst prognosis than the other types. Conclusion: Preoperative and postoperative HR-HPV genotype tests were used to predict the cumulative incidence of abnormal cytology and additional surgery. Particularly, patients with suspected persistent HPV type 16/18 infection are at a high recurrence risk.

Cytological Appearances of Ovarian Seromucinous Borderline Tumor in Ascites: Presentation of 2 Cases

Background: Seromucinous borderline tumor (SMBT) is a rare neoplasm which was newly defined in the 2014 WHO classification. Although the clinical and histopathological characteristics of SMBT have been well described, its cytological characteristics have not. We recently experienced 2 cases of SMBT which were defined by cytological findings of ascites. Case Presentation: Case 1 was a 65-year-old Japanese woman. Preoperative imaging studies revealed abundant ascites and a cystic tumor, with a solid component measuring 13 cm on the left ovary. Radical surgery was performed during the intraoperative diagnosis of ovarian borderline tumor, made by histological examinations of frozen tumor sections. The cytological smears of preoperatively and intraoperatively collected ascites showed many atypical cells resembling reactive mesothelial cells. Alcian-blue staining of cell block sections revealed intracytoplasmic mucin, and the results of immunohistochemistry were consistent with SMBT. The final pathological diagnosis of tumor was SMBT. Case 2 was a 28-year-old Japanese woman. Preoperative imaging studies revealed a small amount of ascites and cystic tumors with solid components in the bilateral ovaries. She initially underwent fertility preservation surgery. SMBT was suspected by cytological examination of smears of intraoperatively collected ascites and the findings of cell block. She underwent additional radical surgery based on a final pathological diagnosis of SMBT. Conclusion: In our experience, the tumor cells of SMBT in ascites mimicked reactive mesothelial cells. The nuclear atypia of SMBTs was intermediate between that of reactive mesothelial cells and serous carcinoma. The immunohistochemistry and mucin staining using cell block were very helpful for facilitating the cytodiagnosis of SMBT.

Evaluation of Elafin Immunohistochemical Expression as Marker of Cervical Cancer Severity

Introduction: The main risk factor for the development of cervical cancer (CC) is persistent infection by human papillomavirus (HPV) oncogenic types. In order to persist, HPV exhibits a plethora of immune evasion mechanisms. PI3/Elafin (Peptidase Inhibitor 3) is an endogenous serine protease inhibitor involved in epithelial protection against pathogens. PI3/Elafin’s role in CC is still poorly understood. Materials and Methods: In the present study, we addressed PI3/Elafin protein detection in 123 CC samples by immunohistochemistry and mRNA expression in several datasets available at Gene Expression Omnibus and The Cancer Genome Atlas platforms. Results: We observed that PI3/Elafin is consistently downregulated in CC samples when compared to normal tissue. Most of PI3/Elafin-positive samples exhibited this protein at the plasma membrane. Besides, high PI3/Elafin expression at the cellular membrane was more frequent in in situ stages I + II than in invasive cervical tumor stages III + IV. This indicates that PI3/Elafin expression is gradually lost during the CC progression. Of note, advanced stages of CC were more frequently associated with a more intense PI3/Elafin reaction in the nuclei and cytoplasm. Conclusion: Our results suggest that PI3/Elafin levels and subcellular localization may be used as a biomarker for CC severity.

Conventional Direct Smear Yields Diagnostic Indicators of Gastric-Type Mucinous Carcinoma Compared with Cytomorphological Features Identified by Liquid-Based Cervical Cytology

Introduction: Gastric-type mucinous carcinoma (GAS) of the uterine cervix is an adenocarcinoma subtype with a gastric phenotype that poses diagnostic pitfalls in cervical screening cytology because of its blunt morphologic atypia and the limited utility of human papillomavirus testing and ancillary immunochemical staining. Despite the recent widespread uptake of liquid-based cytology (LBC) systems, the cytomorphological features of GAS in LBC samples and the differential features between GAS and usual-type endocervical adenocarcinoma (UEA) remain unclear. Methods: Eight GAS cases, all of which were surgically treated following histological confirmation, were examined. Direct Papanicolaou-stained smears and LBC samples were reviewed and compared with 10 UEA cases as controls. Featured cytomorphological findings were as follows: background (mucinous, inflammatory, or necrotic), cell crowding (size of neoplastic cell clusters), cytoplasm (golden mucin and cell border), and nuclei (nuclear chromatin and nucleoli). Results: Of 18 adenocarcinomas, 16 were detected against a non-mucinous background in LBC samples, most of which were accompanied by mild to moderate inflammation. Clusters comprising &#x3e;300 neoplastic cells were identified in both GAS and UEA in conventional smears (CSs), while no LBC samples harboured clusters as large as these. Cell borders of GAS were more distinct than those of UEA in CSs (p &#x3c; 0.001), although fewer populations of neoplastic clusters revealed distinct cell borders in both GAS and UEA in LBC samples. Three of 8 and 2 of 8 GAS cases had golden mucin in CSs and in LBC samples, respectively, which was not detected in UEA at all. Nucleoli against fine nuclear chromatin were more pronounced in GAS than in UEA on CS (p = 0.03), although the difference between GAS and UEA was not apparent in LBC samples. Discussion/Conclusion: This study demonstrated that the diagnostic clues to detect GAS using the conventional approach, namely distinct cell borders and prominent nucleoli, are not useful for excluding UEA in LBC samples. Conventional cervical smears may indicate a diagnosis of GAS; however, specific high-risk HPV detection approaches, such as HPV test or immunocytochemical p16/Ki-67 dual staining, are desirable to differentiate GAS from UEA in the setting of LBC with ambiguous cytomorphological features.

Correlation of High-Risk HPV Genotypes with Pap Test Findings: A Retrospective Study in Eastern Province, Saudi Arabia

Introduction: High-risk human papillomavirus (HR HPV) is found to be responsible for 4.5% of cancer in general, primarily cervical cancer. We aim here to highlight the prevalence and genotypes of HR HPV and correlate its association with Pap tests’ results, which are still not well known in the Eastern Province of Saudi Arabia. Methods: Over 7 years (2013–2019), the results of 164 Saudi women coinvestigated for HR HPV along with Pap tests were collected from the archive of King Fahd University Hospital. Only women who had atypical squamous cells of undetermined significance (ASCUS) on the Pap test and those at elevated risk of infection were cotested for HR HPV; otherwise, the Pap test was the only screening modality for cervical cancer. Data were organized and statistically analyzed using IBM SPSS v26. Results: Out of 164 Saudi women, 14.5% (n = 24/164) showed positive results for HR HPV (8 patients had HPV16 and 2 had both HPV16 and HPV18/45, while the remaining 14 had other HR HPV); among them, 41.5% (n = 10/24) had an abnormal Pap test (5 ASCUS and 5 LSIL), while 58.5% (n = 14/24) had a negative Pap test. On the other hand, 21% (n = 35/164) of patients, in the study, had an abnormal Pap test (24 ASCUS, 8 low-grade squamous intraepithelial lesion [LSIL], and 3 atypical glandular cell [AGC]). In 80% (n = 19/24) of ASCUS cases, HR HPV was not detected; however, 20% (n = 5/24) were positive for other HR HPV. Concerning LSIL cases, 62.5% (n = 5/8) were positive for HR HPV (1 case showed HPV16 and HPV18/45, 2 cases showed HPV16, and 2 cases showed other HR HPV), while in the remaining 37.5% (n = 3/8) LSIL cases, HR HPV was negative; similarly, all AGC cases were negative for HR HPV. Statistical analysis showed a significant correlation between HPV status and Pap test findings (p value &#x3c;0.001). Discussion/Conclusion: HR HPV frequency and genotype distribution, in this study, might reflect a different regional infection pattern. The high association of HR HPV with negative cytology emphasizes the need to add the HR HPV test to screening modalities of cervix cancer.

PIK3CA Mutations and Their Impact on Survival Outcomes of Patients with Cervical Cancer: A Systematic Review

Introduction: Several studies have implicated the PIK3/AKT pathway in the pathophysiology of cancer progression as its activation seems to be aberrant in several forms of cancer. The purpose of the present systematic review is to evaluate the impact of PIK3CA mutations on survival outcomes of patients with cervical cancer. Methods: We used the Medline (1966–2020), Scopus (2004–2020), ClinicalTrials.gov (2008–2020), EMBASE (1980–2020), Cochrane Central Register of Controlled Trials (CENTRAL) (1999–2020), and Google Scholar (2004–2020) databases in our primary search along with the reference lists of electronically retrieved full-text papers. Statistical meta-analysis was performed with the RevMan 5.3 software. Results: Overall, 12 articles were included in the present study that comprised 2,196 women with cervical cancer. Of those, 3 studies did not report significant differences in survival outcomes among patients with mutated versus wild-type PIK3CA tumors, 5 studies reported decreased survival outcomes, and 3 studies revealed increased survival rates. The meta-analysis revealed that patients with the mutated PIK3CA genotypes had worse overall survival compared to patients with wild-type PIK3CA (HR 2.31; 95% CI: 1.51, 3.55; 95% PI: 0.54, 9.96; data from 3 studies) and the same was observed in the case of DFS rates (HR 1.82; 95% CI: 1.47, 2.25; 95% PI: 1.29, 2.56; data from 4 studies). Conclusion: Current evidence concerning the impact of PIK3CA mutations on survival outcomes of patients with cervical cancer is inconclusive, although the majority of included studies support a potential negative effect, primarily among those with squamous cell carcinoma tumors.

Downregulation of IL-2 and IL-23 in Cervical Biopsies of Cervical Intraepithelial Lesions: A Cross-Sectional Study

Introduction: Persistent infection with high-risk human papillomavirus (HPV) types is associated with high-grade intraepithelial lesions (HSILs) and invasive cervical cancer. The host immune response plays a key role in whether HPV clears or persists. Most studies on local immune response to HPV collect cervical mucus in order to quantify secreted cytokines; however, cells located inside the tissue can release different cytokines associated with HPV infection. Objective: This study compared the cytokine levels in cervical biopsy specimens of women with abnormal colposcopic findings according to the histopathological results: low-grade intraepithelial lesion (LSIL), HSIL, and no intraepithelial lesion (NSIL). Methods: A cross-sectional study enrolling 141 cervical biopsy specimens examined the cytokine profile for interleukin (IL-) 2, IL-4, IL-10, IL-12, IL-17, and IL-23 and interferon-γ, using the Luminex assay/ELISA. Differences in cytokine levels among the cervical lesion groups were assessed using the Kruskal-Wallis test. Results: The 141 specimens included 90 HSILs, 22 LSILs, and 29 NSILs. IL-2 levels were significantly higher in NSIL samples than in LSIL or in HSIL samples (p = 0.0001) and IL-23 levels were significantly higher in NSIL than in HSIL samples (p = 0.003). Conclusions: Our study shows that in samples from the lesion site point, 2 important pro-inflammatory cytokines, IL-2 and IL-23, are downregulated in HPV lesions.

Self-Sampling as a Plausible Alternative to Screen Cervical Cancer Precursor Lesions in a Population with Low Adherence to Screening: A Systematic Review

Background: Cervical cancer is the fourth most common cancer in the world. A raised incidence and mortality parallel a low participation rate in screening, namely screening in poor countries and among specific populations of developed countries. Cervical or vaginal self-sampling may increase adherence to screening due to its low costs and elevated sensibility and specificity. Our main goal was to compare self-sampling with physician sampling and to evaluate the participation rate of women formerly non-adherent to cervical cancer screening. Study Design: We identified relevant studies from PubMed. Studies were eligible for inclusion if they fulfilled the following criteria: women aged between 16 and 80 years, poor and developed countries that use self-sampling, women with a low participation rate in screening and studies published since 2013. After fully reading the articles, data were extracted to an Excel sheet to display all relevant information in an organized manner. Results: We identified 18 studies, which altogether enrolled 22,118 women. Self-sampling and physician sampling are quite similar regarding the HPV detection rate. Women have a low participation rate mainly because of limited access to health services, religious and culture beliefs, and lack of time due to several demanding tasks in daily life. Self-sampling shows a significant increase in acceptability and preference compared to physician sampling. Conclusion: Self-sampling is a reliable method to involve women in opportunistic or organized screening programs for cervical cancer prevention.

DNA Recovery Using Ethanol-Based Liquid Medium from FTA Card-Stored Samples for HPV Detection

Introduction: Alternative methods of dry storage and transportation may be a viable alternative to the use of liquid storage medium for cervical samples, especially for screening programs in places with few resources. Objective: The objective of this study is to verify the viability and efficacy of human papillomavirus DNA (HPV-DNA) detection in cervical cell samples collected and stored on a Flinders Technology Associates (FTA) card (Whatman Indicating FTA® Elute Micro Card) and subsequently recovered in ethanol-based liquid medium and to compare the results to those obtained using samples stored directly in ethanol-based liquid medium. Study Design: Thirty-four women submitted to ETZ (excision of the transformation zone of the cervix) were included in this study. Before ETZ, 2 samples of exfoliated cervical cells were collected from each woman by a doctor and stored in ethanol-based liquid medium and on an FTA card. DNA recovery from FTA samples was performed using ethanol-based liquid medium. Detection of HPV-DNA in the samples was performed using the Cobas® 4800 HPV Test Platform. Results and Conclusions: The HPV-DNA detection positivity rates were 70.6% for the samples collected directly in liquid medium and 64.7% for the samples stored on the FTA card, with high detection accuracy in the DNA samples recovered from the FTA card (area under the curve = 0.958; 95% confidence interval = 0.890–1.000). The concordance between the results obtained using the 2 storage media was 94.1% (Kappa = 0.866). These preliminary results suggest that collection of cervical material on an FTA card may be an alternative to storage in liquid medium since the liquid medium has some limitations. In addition, DNA recovery from the card using ethanol-based liquid medium streamlines the workflow in the laboratory and reduces the cost associated with reagents, thereby facilitating access to the HPV test in places with few resources and potentially improving cervical cancer screening.

Seborrheic Keratosis-Like Lesion of the Cervix: First Report of the Cytological Features of a Low-Risk HPV 42-Associated Lesion

Introduction: Seborrheic keratosis-like lesion of the cervix and vagina is a rare lesion and shows similar morphology to vulvar seborrheic keratosis; 3 of the 7 previously reported cases were associated with low-risk human papillomavirus (HPV) type 42. We report a case of seborrheic keratosis-like lesion of the cervix and provide the first description of the cytological features of this lesion. Case Presentation: A woman in her late forties presented with postcoital bleeding. She had a cervical screening test following which she underwent cervical biopsy, endocervical and endometrial curettage, large loop excision of the transformation zone of the cervix, and hysterectomy. Results: The liquid-based cytology preparation showed cohesive groups of mildly atypical squamoid cells with a spindle cell morphology, mildly increased nuclear to cytoplasmic ratio, prominent nucleoli, and occasional nuclear grooves. No koilocytes were identified. Molecular genotyping revealed positivity for HPV type 42. Discussion/Conclusion: This represents the first description of the cytological features of a seborrheic keratosis-like lesion of the cervix, which are distinctive and unusual. Whilst the mild squamous atypia raised the possibility of a low-grade squamous intraepithelial lesion, no koilocytes were identified. The association in our case with a low-risk HPV type, HPV 42, provides further evidence for a role of this HPV type in the pathogenesis of these lesions.

Diagnostic Performance of the Newly Developed WellPrep® Liquid-Based Cytology System and Its Comparison with SurePathTM in Cervical Squamous Lesions

Introduction: WellPrep® (WP), a fully automated, one-step liquid-based cytology (LBC) platform using an all-in-one closed chamber, has recently been developed as a next-generation LBC technology. This study aimed to evaluate the diagnostic performance and cytomorphologic features of WP regarding cervical cytology and also to compare WP with the SurePathTM (SP), one of the most widely used LBC systems used worldwide. Methods: Cervicovaginal samples were taken from 212 females who enrolled in the study, and each sample was split and subsequently used for WP and SP LBC. Following the exclusion of seven cases with insufficient quality, a total of 205 cases were used for subsequent analysis. Among them, 75 (36.6%) received histologic follow-up. All cases were interpreted according to the Bethesda System, while three experienced pathologists evaluated their cytomorphologic features. Results: The diagnostic concordance rate between the two LBC technologies was 84.4% (kappa = 0.776). Furthermore, the diagnostic concordance rates between SP and histology and between WP and histology were 73.3% (kappa = 0.516) and 70.7% (kappa = 0.497), respectively. The two LBC methods showed comparable sensitivity, specificity, and area under the curve (AUC) for histologic HSIL+ (SP: sensitivity 82.8%, specificity 84.8%, and AUC 0.838; WP: sensitivity 79.3%, specificity 87.0%, and AUC 0.831). No significant difference was found regarding the sensitivity, specificity, and AUC between SP and WP (p = 0.586, p = 0.670, and p = 0.924, respectively). In terms of cytomorphologic features, WP revealed more often than SP the presence of coarse chromatin (p = 0.031) and mitoses (p = 0.008) but less commonly perinuclear clearing (p = 0.001). Conclusion: This is the first study demonstrating that WP has a comparable performance to SP. In conclusion, WP may be an alternative LBC technology for cervical cancer screening.

High Prevalence of Cervical High-Grade Lesions and High-Risk Human Papillomavirus Infections in Women Living with HIV: A Case for Prioritizing Cervical Screening in This Vulnerable Group

Introduction: Women living with HIV (WLHIV) are at an increased risk of developing cervical precancerous lesions and cervical human papillomavirus (HPV) infection. This study aimed at evaluating the prevalence of cervical lesions and high-risk HPV (HR-HPV) infection in WLHIV in comparison to the HIV-negative women undergoing opportunistic screening. In addition, these findings among WLHIV were correlated with the clinic-demographic factors. Methods: A cross-sectional study was conducted among WLHIVs at a tertiary hospital and linked antiretroviral therapy (ART) center, while HIV-negative women were recruited from the health promotion clinic at our institute. With informed consent, a semi-structured questionnaire was filled on demographic and epidemiological parameters. Conventional cervical smears and samples for HPV DNA detection by HC2 high-risk HPV DNA test were collected in all participants. Cervical smears were reported using the Bethesda system 2014. Appropriate statistical analysis was performed for bivariate and multivariate logistic regression analysis for comparison between WLHIV and HIV-negative women and for correlation of abnormal cervical cytology and HR-HPV infection among WLHIVs. Results: The clinic-demographic characteristics of WLHIVs and HIV-negative women were similar. On cytology, the prevalence of cervical cytological abnormalities were significantly higher (p < 0.001) among WLHIVs (14.1%) compared to HIV-negative women (3.1%). High-grade lesions were seen in 3.7% of WLHIVs, while no high-grade lesions were detected in HIV-negative women. Cervical HR-HPV infection was also significantly higher (p < 0.001) in WLHIVs (28.9%) than HIV-negative women (9.3%). Cervical precancerous lesions in WLHIVs showed positive association with current sexually transmitted infection (STI), multiple sexual partners, tobacco use, and CD4 count less than 200/µL, while cervical HPV was positively associated with current STI, tobacco use, CD4 count less than 200/µL and negatively with ART intake. On multivariate logistic regression, cervical cytological abnormalities showed a significant association with multiple sexual partners (p < 0.001), while cervical HR-HPV infection was positively associated with current STI (p = 0.01), nadir CD4 count <200/µL (p = 0.004), abnormal cervical cytology (p = 0.002) and negatively with ART intake (p = 0.03). Conclusion: Women living with HIV have a significantly higher prevalence of cervical precancerous lesions and HR-HPV infection compared to the general population. Considering the lack of an organized population-based cervical cancer screening program in many low-resource countries like ours, specific focus on screening this highly vulnerable population to reduce the morbidity and mortality due to cervical cancer is imperative.

Effects of Menstrual Cycle on Various Morphologies of High-Grade Squamous Intraepithelial Lesions in SurePath™ Liquid-Based Cervical Cytology

Introduction: The morphology of high-grade squamous intraepithelial lesion (HSIL) on Papanicolaou (Pap) smears widely varied, including syncytial aggregates, sheets, and scattered single cells, and no particular cellular pattern is consistently observed. Therefore, this study aimed to determine whether the menstrual cycle affects the cellular pattern of HSILs, an effort to avoid false negatives due to the oversight of scattered small single HSIL cells in the cytological triage of human papillomavirus-positive women. Methods: A total of 147 HSIL samples of liquid-based cytology (LBC) in patients with cervical intraepithelial neoplasia grade 2 or 3 were obtained, and then, the relationship between cellular patterns, such as single-cell-like and syncytial aggregates, and menstrual cycles classified into six phases was analyzed. If a syncytial aggregate was present, the number of cells constituting the aggregate was visually counted under the microscope. Results: HSILs in scattered single cells and small sheets of &#x3c;6 on LBC samples accounted for 43% (23/54) during the late proliferative phase of the menstrual cycle. A moderately strong statistically significant association was observed between cellular patterns and menstrual cycles (χ2 [3] = 9.423, p &#x3c; 0.05) (Cramer’s V = 0.253). The value of adjusted residuals showed a statistically significant increased proportion of single-cell-like patterns during the late proliferative phase (p &#x3c; 0.01). Conclusions: The present study demonstrated that HSIL cells in Pap smears in the late proliferation phase have a high frequency of single-cell-like patterns. In human papillomavirus-positive Pap smears with a clean background and predominantly superficial cells, careful microscopic observation by targeting single HSIL cells can potentially reduce false negatives.

A SYSTEMATIC REVIEW AND META-ANALYSIS ON SEDIMENT CYTOLOGY: AN ADJUNCT TO EARLY DIAGNOSIS

Introduction: Sediment cytology, initially applied in urinary diagnostics, has expanded to formalin-fixed biopsy specimens, evaluating exfoliated cells suspended in formalin fixative. Though traditionally discarded, this sediment contains diagnostic material, especially in friable or malignant tissues. It offers a rapid, cost-effective, preliminary diagnostic tool when histopathology is delayed, such as in bone lesions requiring decalcification. This systematic review aimed to comprehensively assess the diagnostic accuracy, applicability, and limitations of sediment cytology across diverse anatomical sites. Methods: This systematic review followed PRISMA guidelines, and an electronic search was conducted across PubMed, Scopus, and Google Scholar databases until January 1, 2025, without year restrictions. Keywords included “Sediment cytology”, “oral neoplasms”, “bone lesions”, and related terms. Articles in English reporting sediment cytology on formalin-fixed biopsies with cytological-histological correlation were included. Results: Nine eligible studies were identified, involving various lesions including OSCC, OPMDs, bone lesions, GIT, cervical and ovarian neoplasms. Pooled sensitivity and specificity were 73.3% and 92.1% respectively. Subgroup analysis revealed excellent concordance for OSCC and bone malignancies. Sensitivity was lower in benign lesions and OPMDs, attributed to poor cellularity and exfoliation. Methodological heterogeneity was noted, though overall bias remained low. Conclusion: Sediment cytology shows valuable diagnostic potential as a preliminary adjunct to histopathology, particularly for malignancies, warranting further standardized multicenter studies.

Comparative Diagnostic Utility of Squash, Scrape, and Imprint cytology in Intraoperative Diagnosis of Ovarian Tumours

Introduction: Intraoperative cytology in ovarian tumours involves collecting cell samples from the ovarian sample sent during surgery and quickly examining them for diagnostic information. Frozen section provides rapid diagnosis to guide intraoperative patient management. The indications of frozen section are identification of tissue, evaluation of margins, and identification of lymph nodes metastasis. Materials and Methods: Intraoperative tissue from clinico-radiologically suspected ovarian tumour for frozen section taken and processed in Department of Pathology and Laboratory Medicine. Squash smear, scrape smear, and imprint smear were made. Three stains rapid May-Grünwald Giemsa, rapid papanicolaou (Pap), and rapid hematoxylin and eosin with expected turnaround time of <15 min were done. Intraoperative cytological smear (squash, scrape, and imprint smear) were correlated with frozen section and histopathology slide. Final assessment of intraoperative cytological smears for diagnostic accuracy was done using statistical study. The aim of this study was to evaluate comparative diagnostic utility of squash smear, scrape smear, and imprint cytology with frozen section in intraoperative ovarian tumour is the aim of study. Results: Sensitivity, specificity, and diagnostic accuracy for frozen and cytology were: sensitivity of frozen section, squash cytology, and scrape cytology was 91.67% in all three, whereas sensitivity of imprint was 87.5%. Specificity of frozen section, imprint cytology, squash cytology, and scrape cytology was 96.77%, 93.55%, 90.32%, and 90.32%, respectively, and accuracy was 94.55%, 90.91%, 90.91%, and 90.91%, respectively. Conclusion: Imprint, squash, and scrape cytology have similar sensitivity and specificity compared to frozen section in identifying the nature of lesion and can be an alternative to frozen section in resource stricken setting.

Role of Pap Smear Cervical Cytology in the Diagnosis of Extrauterine Malignancies: Largest Study of 104 Cases from Tertiary Care Cancer Centre in India

Introduction: Extrauterine malignancies in cervical samples are rarely seen. It is important to differentiate these cells from those of primary uterine malignancies to determine appropriate line of further investigations and management. Literature on these lesions is limited largely restricted to case reports. The aim of the present study was to study the spectrum and cytomorphological features of extrauterine malignancies in cervical Pap smears. Materials and Methods: It is a retrospective and descriptive study conducted in Department of Cytopathology from January 2019 to July 2023. All cases of extrauterine malignancies with available cytology material were included in this study. All cases of primary uterine malignancies, i.e., uterine corpus or cervix confirmed by clinical, radiological, and histopathological examination were excluded. Results: 104 out of 11,674 cytology Pap smears were those of extrauterine malignancy. Diagnosis of extrauterine malignancy was given in 47.1% cases, 30.9% were reported as positive for malignancy without giving the possibility of an extrauterine origin, and 22.0% were reported as atypical glandular cells only. In 56 cases where Pap smear was the first investigation which led to the diagnosis. Most common extrauterine malignancy was adenocarcinoma principally from ovarian, colorectal, and vaginal origin. Other epithelial malignancies noted were urothelial carcinoma and invasive breast carcinoma. Among non-epithelial malignancies, we reported vaginal mucosal melanoma, cutaneous melanoma, acute leukaemia, and anaplastic large cell lymphoma. Conclusion: High index of suspicion, presence of squamous and glandular elements with no atypical features, and occasional clusters of cells with marked atypia and usually no necrosis in the background are helpful cytomorphological clues to raise suspicion for extrauterine malignancy. Correlation with serology, radiology, and immunocytochemistry can help in reaching final diagnosis.

Clinical Management of Atypical Endometrial Cells of Undetermined Significance Followed by Negative Cytology

Introduction: In Japan, endometrial cytology is widely performed to evaluate the status of the endometrium in women with suspected endometrial cancer. A new classification system for endometrial cytology has recently been used: the Yokohama system, based on a descriptive reporting format. This study aimed to clarify the triage for patients with atypical endometrial cells of undetermined significance (ATEC-US) when followed by negative endometrial cytology. Methods: We enrolled patients diagnosed with ATEC-US at the Cancer Institute Hospital between January 2016 and December 2017, based on the following inclusion criteria: (1) ATEC-US diagnosed by office endometrial cytology, with or without office endometrial biopsy; (2) follow-up endometrial cytology was performed 3–6 months after initial sampling, with a negative result for malignancy; and (3) no prior history of conservative treatment with progestin for endometrial cancer or atypical endometrial hyperplasia (ATEC-A). Among eligible patients, we analyzed those later diagnosed by endometrial biopsy with ATEC-A or carcinoma. Results: Among 187 patients, 65 met the inclusion criteria. Forty-two patients (64.6%) were observed for more than 24 months. Two patients (3.1%) developed ATEC-A during a median observation time of 26.5 months; the times to diagnosis were 32 months and 22 months. Discussion/Conclusion: No patient developed ATEC-A or worse within 1 year. For patients with ATEC-US, if negative cytology is obtained at the next examination, a close follow-up is not necessary.

A Diagnostic Approach to Endometrial Cytology by Means of Liquid-Based Preparations

The adoption of endometrial cytology as a diagnostic procedure has been hampered in the past by difficulties arising in interpreting the cellular findings due to a number of factors (such as excess blood, cellular overlapping, and the complex physiology of endometrium). Recently, the use of liquid-based cytology (LBC), with its ability to remove blood and mucus and to distribute cells uniformly in a thin layer on the slide, has provided an opportunity to reevaluate the role of endometrial cytology. LBC samples are easier to screen compared to conventional ones, due to a smaller screening area and an excellent quality of cell preparations. LBC by using peculiar cytoarchitectural features is a useful tool in the cellular diagnosis and follow-up of abnormalities, which, however, remains complementary to histopathology and to the emerging molecular diagnostic cytopathology. This review discusses these various entities and takes into consideration the ancillary techniques that may be useful in the diagnostic procedure. Herein, we also summarize the process and rationale by which updates were made to the standardized terminology in 2018 and outline the contents of the new Bethesda-style classification (the Yokohama system) for the endometrial cytology.

Artificial Intelligence in Gynecologic Cytology

Background: Cervical cancer is the fourth most common cancer in women globally with highest incidence and mortality identified in less developed and medically underserved areas in the world. The diminishing cytology workforce, unavailability of expert consultation, and the high volume of Pap tests needing manual screening are the main reasons for exploring innovative solutions to help mitigate the negative effects resulting from the dearth of timely cervical cancer screening in certain population groups. Summary: Developments in whole slide imaging and artificial intelligence (AI) have enabled the emergence of new computer-assisted systems that have the potential for transforming traditional cytopathology practice. However, AI-based systems are relatively new with limited published data on their validation and clinical utility in clinical practice. Our article aims to increase awareness of the availability of such systems, explores the history and development of AI-assisted screening platforms for Pap tests, compares the performance characteristics of various systems, elaborates on technical challenges associated with conducting clinical trials employing this technology, and discusses considerations around deploying such systems in routine cytopathology practice. Key Message: Revolutionary AI-based systems are being developed and utilized in cytopathology practice to screen Pap tests. Some of these systems have good performance characteristics and provide opportunities to combat various issues such as workload and standardization faced by cytology laboratories globally. However, judicious review of these systems using evidence-based studies is imperative to promote widespread adoption and maintain high-quality standards for patient safety.

Critical Evaluation of Pap Test Adherence to Routine Screening in Amazonas State, Brazil

Introduction: This study critically evaluates adherence to Pap test screening practices in cytology-based cervical cancer screening in the state of Amazonas over a 10-year period. Materials and Methods: A retrospective analysis was conducted of the results of cytological screening examinations (Pap test) in Amazonas State from 2013 to 2023. For this purpose, Brazilian public databases Cervical Cancer Information System (SISCOLO) and Cancer Information System (SISCAN) (from the Department of Information and IT of the Unified Health System [DATASUS]) were consulted. Results: There was a decrease in the number of Pap tests performed during the period from 2019 to 2021, likely related to the COVID-19 pandemic. This was followed by a subsequent increase in the post-pandemic period. Notably, in municipalities with fewer than 10,000 annual Pap examinations there was a decrease in the average number of tests when comparing the years 2016–2018 to 2013–2015, and an even greater decrease during the pandemic. Conclusions: There is considerable variation in utilization of the cytological Pap test across different municipalities. This lack of uniformity throughout the state likely compromises the capacity to detect early stage cervical intraepithelial lesions.

Practical Approach to the Evaluation of Malignant Peritoneal Fluids in the Setting of Gynecologic Neoplasms

Background: Evaluation of peritoneal fluid cytology, either from ascitic fluids or as a result of peritoneal washings, is a fundamental aspect in the evaluation of women presenting with clinically concerning or histologically confirmed gynecologic neoplasms. Summary: Ascitic fluid samples are often the initial and only source of diagnostic material in women presenting with gynecologic malignancies, and important therapeutic decisions will result from the information provided in the cytology report. On the other hand, cytologic evaluation of peritoneal washing specimens obtained during surgical excision of a presumed gynecologic neoplasm provides crucial information to the clinical team regarding tumor staging, often with significant therapeutic implications. While recognition of high-grade tumors in either of these samples is generally straightforward, low-grade tumors and unusual neoplasms can prove to be more difficult to recognize, differentiate from benign mimics, and correctly diagnose, particularly in low-cellularity specimens. Even with high-grade tumors, a mere diagnosis of “positive for malignancy” in diagnostic ascitic fluid specimens might not suffice to guide clinical management, and the use of ancillary techniques to further and more definitively characterize the lesional cells is required. Key Messages: This review will focus on the clinically relevant issues surrounding interpretation of peritoneal fluid cytology specimens in the setting of gynecologic neoplasms, making emphasis on the salient cytomorphologic and immunocytochemical features of the various neoplastic processes, in an attempt to provide a practical yet effective guide on how to best evaluate, diagnose, and report these samples.

Cytomorphologic Features of Gastric-Type Endocervical Adenocarcinoma in Liquid-Based Preparations

Objective: Gastric-type endocervical adenocarcinoma (GAS) is a recently described, uncommon, and aggressive tumor with distinct morphologic features and HPV-independent etiology. Data on GAS in liquid-based cytology (LBC) Papanicolaou (Pap) test preparations from a North American patient population are scant. We systematically assessed the cytomorphologic characteristics of GAS in LBC from patients in Ontario and examined if glandular cell nuclear area could represent a readily assessable feature which may aid in GAS detection. Study Design: Pap test slides preceding the diagnosis of GAS were retrieved locally or requested from outside laboratories. A structured review of 15 cytomorphologic features was performed using the available LBC Pap test slides of GAS and a set of usual-type endocervical adenocarcinomas (UEA). Morphometry of the glandular cell nuclear area was performed, and normalized values were compared to UEA and benign endocervical cells. Results: At least 1 Pap test (5 ThinPrep®, 11 SurePath®, and 1 direct smear) was available for 14 patients. Original LBC Pap test diagnoses were negative for intraepithelial lesion or malignancy (NILM) (7), adenocarcinoma/carcinoma (6), atypical glandular cells (2), and adenocarcinoma in situ (1). Review detected abnormal glandular cells in 6/7 NILM cases. Honeycomb-like sheets, nuclear enlargement, and microvesicular cytoplasm were the single most common architectural, nuclear, and cytoplasmic features, respectively. Microvesicular cytoplasm (100 vs. 17%), honeycomb-like sheets (87 vs. 8%), prominent nucleoli (93 vs. 25%), and anisonucleosis (93 vs. 50%) were most discriminatory for GAS versus UEA, respectively. Yellow mucin, intranuclear cytoplasmic pseudoinclusions, and goblet/Paneth-like cells were uncommon, but unique for GAS. Glandular cell nuclear area normalized to neutrophils was found to be significantly increased in GAS compared to benign endocervical cells. Conclusions: GAS is under-recognized and may mimic reactive endocervical cells. Awareness of the tumor type and its cytomorphology is critical for early detection. Identification of glandular cells with uniform nuclear enlargement in conjunction with any of the other cytologic features may help avoid false-negative Pap results. Neutrophils may serve as convenient size reference and visual aid.

Nationwide Rereview of Normal Cervical Cytologies before High-Grade Cervical Lesions or before Invasive Cervical Cancer

Sweden has experienced an unexpected &#x3e;30% increase in cervical cancer incidence among women with normal cytological screening results. We therefore performed a nationwide assessment of false-negative cytology before invasive cervical cancer. The Swedish national cervical screening registry identified 2,150 normal cytologies taken up to 10 years before 903 cases of invasive cervical cancer. The 27 cytological laboratories in Sweden were asked to rereview the slides, and all of them completed the rereview. One thousand nine hundred fifteen slides were retrieved and reviewed. Abnormalities were found in 30% of the slides, and the proportion of slides that had a changed diagnosis on rereview increased on average by 3.9% per sampling year during 2001–2016 (p &#x3c; 0.03). We also asked for rereview of normal smears taken up to 42 months before a histopathologically diagnosed high-grade squamous intraepithelial lesion (HSIL) or adenocarcinoma in situ (AIS). 19/27 laboratories responded, and out of 6,101 normal smears taken before HSIL/AIS, 5,918 were retrieved and rereviewed. The diagnosis was changed in 25% of cases. In summary, we found an increasing time trend of false-negative smears taken before invasive cervical cancer. This indicates a decreased protection of normal cytology in the screening program supporting earlier findings that this is the main reason behind the recent Swedish increase in cervical cancer. We suggest that optimal cervical cancer control may be promoted by routine nationally coordinated rereview of negative smears before high-grade cervical lesions or invasive cervical cancer.

Comparison of ThinPrep Integrated Imager-Assisted Screening versus Manual Screening of ThinPrep Liquid-Based Cytology Specimens

Objective: The aim of this study was to find out whether ThinPrep Integrated Imager (Hologic Inc.) screening is non-inferior to manual screening in the detection of cervical lesion. Study Design: For a total of 4,011 ThinPrep Pap test specimens stained by ThinPrep staining, manual screening (Manual arm) and ThinPrep Integrated Imager screening (Imager arm) were performed so as not to be screened by the same cytotechnologist, and the sensitivity and specificity in the detection of cervical lesion were compared using McNemar’s test. Results: The sensitivity to detect CIN1 or more squamous cell abnormalities or glandular abnormalities was 91.67% (= 374/408, 95% confidence interval [CI]: 88.44–94.08%) for the Manual arm and 92.40% (= 377/408, 95% CI: 89.28–94.70%) for the Imager arm, and the specificity was 88.87% (= 3,113/3,503, 95% CI: 87.77–89.88%) for the Manual arm and 89.55% (= 3,137/3,503, 95% CI: 88.48–90.54%) for the Imager arm. The differences in sensitivity and in specificity, respectively, were 0.74% (95% CI: −3.14–4.61%, McNemar’s test, p = 0.8041) and 0.69% (95% CI: −0.13–1.50%, McNemar’s test, p = 0.1125). About the equality of sensitivity and specificity between the 2 methods, 95% CIs of the difference between sensitivity and specificity are in the clinical equivalence range of ±5%, so the Imager arm is non-inferior to the Manual arm. Conclusion: The Imager arm was confirmed to have an equivalent and non-inferior capacity in the detection of cervical lesions compared with the Manual arm, suggesting that its practical application in cervical cytology tests is highly possible.

Not Enough Cells: How Insufficient Cytological Specimens Are Mirrored by Reporting Systems – Journey from the Bethesda to the WHO Reporting Systems

Background: Pap classes have been replaced by organ-specific reporting systems in recent decades; however, part of the cytological specimens is insufficient. The present review summarizes how different organ-specific systems define the insufficient category: both quantitative and qualitative criteria are used. In addition, the sample volume may be evaluated in certain specimens. Summary: The reasons for an insufficient sample may vary and depend either on the lesion itself or the sampling procedure. Key Messages: The management recommendations for insufficient specimens improve communication between cytopathologists and treating physicians.

Human Papillomavirus Primary Screening and Collaboration in Quality Assurance Work between Laboratories

Background: Cervical screening is changing to the use of human papillomavirus (HPV) testing as the primary screening test. It is essential that the well-established and critically important systems for quality assurance based on laboratory audits of seemingly negative samples taken before HSIL and cervical cancer are maintained. They provide a means of verifying if the actual screening is effective for the intended purpose. Together with international proficiency panels, audits provide a simple and unambiguous way to evaluate if the screening is adequate. Detailed knowledge of how these systems work and how they are dependent on the genotyping of HPV, biobanking, and screening registries is vital to cytologists and pathologists involved in quality assurance work and follow-up of cervical lesions and cervical cancer. Interpretation and communication of outcome and results are equally important for successful quality assurance work and should ideally be done together with expertise in HPV. Summary: The internationally defined procedures for laboratory audit, similar to those used for cytology, require sensitivities before HSIL of >95% and before invasive cervical cancer of >90%. If also results on blinded proficiency panels and international criteria for analytic sensitivity, specificity, and reproducibility are achieved, the HPV screening test can be said to be adequate. Key Messages: Performance of HPV screening tests in a cervical screening program includes similar laboratory audits as hitherto used for cytology. Similarly, technical proficiency of a laboratory is established using blinded proficiency panels with defined contents of virus. Detailed knowledge of quality assurance work is necessary for cytologists and pathologists. Communication of outcome and results depends on collaboration between laboratories.

Outer Cutoff Value for the Box-Counting Method for Fractal Analysis of the Nucleus Using Kirsch Edge Detection

Objective: The complexity of chromatin (i.e., irregular geometry and distribution) is one of the important factors considered in the cytological diagnosis of cancer. Fractal analysis with Kirsch edge detection is a known technique to detect irregular geometry and distribution in an image. We examined the outer cutoff value for the box-counting (BC) method for fractal analysis of the complexity of chromatin using Kirsch edge detection. Materials: The following images were used for the analysis: (1) image of the nucleus for Kirsch edge detection measuring 97 × 122 pix (10.7 × 13.4 μm) with a Feret diameter of chromatin mesh (n = 50) measuring 17.3 ± 1.8 pix (1.9 ± 0.5 μm) and chromatin network distance (n = 50) measuring 4.4 ± 1.6 pix (0.49 ± 0.18 μm), and (2) sample images for Kirsch edge detection with varying diameters (10.4, 15.9, and 18.1 μm) and network width of 0.4 μm. Methods: Three types of bias that can affect the outcomes of fractal analysis in cytological diagnosis were defined. (1) Nuclear position bias: images of 9 different positions generated by shifting the original position of the nucleus in the middle of a 256 × 256 pix (28.1 μm) square frame in 8 compass directions. (2) Nuclear rotation bias: images of 8 different rotations obtained by rotating the original position of the nucleus in 45° increments (0°, 45°, 90°, 135°, 180°, 225°, 270°, and 315°). (3) Nuclear size bias: images of varying size (diameter: 190 pix [10.4 μm], 290 pix [15.9 μm], and 330 pix [18.1 μm]) with the same mesh pattern (network width: 8 pix [0.4 μm]) within a 512 × 512 pix square. Different outer cutoff values for the BC method (256, 128, 64, 32, 16, and 8 pix) were applied for each bias to assess the fractal dimension and to compare the coefficient of variation (CV). Results: The BC method with the outer cutoff value of 32 pix resulted in the least variation of fractal dimension. Specifically, with the cutoff value of 32 pix, the CV of nuclear position bias, nuclear rotation bias, and nuclear size bias were &#x3c;1% (0.1, 0.4, and 0.3%, respectively), with no significant difference between the position and rotation bias (p = 0.19). Our study suggests that the BC method with the outer cutoff value of 32 pix is suitable for the analysis of the complexity of chromatin with chromatin mesh.

Personalized Medicine and Cervical Screening: Development of Individualized Quantitative Risk Assessments for Cervical Adenocarcinoma and Adenocarcinoma in situ

Introduction: Cervical screening has decreased the incidence of cervical carcinoma around the world primarily by preventing cervical squamous carcinoma, with significantly less measurable protective benefits in prevention of cervical adenocarcinoma. In this study, we apply Bayesian modeling of cervical clinical, screening, and biopsy data from a large integrated health system to explore the feasibility of calculating personalized risk assessments on screened system patients for subsequent histopathologic diagnoses of invasive cervical adenocarcinoma (AdCa) or cervical adenocarcinoma in situ (AIS). Materials and Methods: Diagnoses of cervical AIS or AdCa rendered between 2005 and 2018 were identified in our large health system database with 1,053,713 cytology results, 354,843 high-risk (hr) human papillomavirus (HPV) test results, and 99,012 cervical histopathologic results. Using our continuously updated Bayesian cervical cancer screening model which includes clinical data, cervical screening results, and cervical biopsy results, we projected quantitative estimates of patients’ 5-year cumulative risk for cervical AIS or AdCa. Results: 161 patients were identified with AIS (ages 17–75, mean 37 years), and 99 patients had diagnoses of cervical AdCa (ages 26–91, mean 48 years). Quantitative Bayesian 5-year cumulative risk projections for diagnoses of cervical AdCa or AIS in patients with different cervical screening test and biopsy histories were determined. The highest patient risk projections for subsequent cervical AdCa and/or AIS histopathologic diagnoses were associated with prior cervical screening test results of HPV-positive atypical glandular cells. Prior squamous cytologic abnormalities were associated with lower risk estimates. Prior histopathologic diagnoses of squamous abnormalities also influenced quantitative risk. A prior histopathologic diagnosis of AIS was associated with a very low risk of subsequent AdCa, consistent with effective excisional treatment. AdCa risk was greatest in women aged 30–65 years with prior CIN3 biopsy results, whereas AIS risk was greatest in women &#x3c;30. Conclusion: Prevention of cervical AdCa in screened patients remains a major challenge for cervical screening. Individualized risk projections for cervical glandular neoplasia reflecting patient age, prior cervical screening test results, and prior cervical biopsy history are feasible using Bayesian modeling of health system data.

Characteristic Cytological Findings of Lobular Endocervical Glandular Hyperplasia Associated with Adenocarcinoma of the Uterine Cervix

Objective: To investigate the cytological findings of lobular endocervical glandular hyperplasia (LEGH) associated with adenocarcinoma and to clarify its characteristics and the coexisting adenocarcinoma using histochemistry and immunohistochemistry. Methods: Eighteen surgical cases of LEGH of the uterine cervix were retrospectively reviewed and classified into 3 groups: pure (pure type), atypical (atypical type), and LEGH with adenocarcinoma (mixed type). The mixed type is defined as LEGH or atypical LEGH with in situ or invasive adenocarcinoma. Cytological findings of conventional endocervical smear specimens (Papanicolaou stain) were analyzed. Histochemistry (periodic acid-Schiff reaction) and immunohistochemistry (M-GGMC-1, Muc-6 glycoprotein, and Ki-67) were performed using tissue specimens. Results: Cytologically, the pure type (7 cases) is characterized by glandular cell clusters that tended to form monolayered sheets with uniformly small nuclei and contain golden-yellowish mucin, whereas atypical (5 cases) and mixed (6 cases) types are characterized by glandular cell clusters similar to those of the pure type, but with complex glandular structures and mucin localization on the surface of glandular cell clusters. Ki-67 labeling index was significantly higher in atypical and mixed types than that in the pure type. Gastric-type mucinous carcinoma (MC-G) was observed in 2 out of 6 cases with mixed type. Conclusions: LEGH is found to be associated with adenocarcinoma types other than MC-G. Complex glandular structures or mucin localization on the surface of glandular cell clusters may be useful cytological findings to detect atypical and mixed types of LEGH.

Computational Cytology: Lessons Learned from Pap Test Computer-Assisted Screening

Background: In the face of rapid technological advances in computational cytology including artificial intelligence (AI), optimization of its application to clinical practice would benefit from reflection on the lessons learned from the decades-long journey in the development of computer-assisted Pap test screening. Summary: The initial driving force for automated screening in cytology was the overwhelming number of Pap tests requiring manual screening, leading to workflow backlogs and incorrect diagnoses. Several companies invested resources to address these concerns utilizing different specimen processing techniques and imaging systems. However, not all companies were commercially prosperous. Successful implementation of this new technology required viable use cases, improved clinical outcomes, and an acceptable means of integration into the daily workflow of cytopathology laboratories. Several factors including supply and demand, Food and Drug Administration (FDA) oversight, reimbursement, overcoming learning curves and workflow changes associated with the adoption of new technology, and cytologist apprehension, played a significant role in either promoting or preventing the widespread adoption of automated screening technologies. Key Messages: Any change in health care, particularly those involving new technology that impacts clinical workflow, is bound to have its successes and failures. However, perseverance through learning curves, optimizing workflow processes, improvements in diagnostic accuracy, and regulatory and financial approval can facilitate widespread adoption of these technologies. Given their history with successfully implementing automated Pap test screening, cytologists are uniquely positioned to not only help with the development of AI technology for other areas of pathology, but also to guide how they are utilized, regulated, and managed.

Comparison between Conventional Cytology and Liquid-Based Cytology in the Tertiary Brazilian Navy Hospital in Rio de Janeiro

Introduction: Cervical cancer screening is an important tool in public health. Liquid-based cytology (LBC) has been performed at the studied hospital for 7 years. The present study compares the performance of 2 LBC techniques with conventional cytology. Objective: Our objective is to verify the sensitivity for the detection of neoplastic and preneoplastic epithelial atypia, as well as the positive predictive value of the 3 methodologies. Methods: We analyzed retrospectively 24,529 cases and evaluated the conventional cytology, ThinPrep®, and BD SurePath® performance categorizing the results according to the Bethesda system. We also compared the level of unsatisfactory samples, the presence of elements from the squamocolumnar junction, and the detection of pathogenic microorganisms. Results: ThinPrep® (1.43%) showed superior sensitivity over BD SurePath® (0.91%) and conventional cytology (0.71%) in terms of the detection of high-grade lesions; however, in terms of squamous atypia as a whole (ASC-US+), BD SurePath® (6.44%) proved to be more sensitive than conventional cytology (5.28%) and ThinPrep® (3.73%). Conclusions: The results show the advantage of implementing LBC in routine screening for cervical lesions. In this study, BD SurePath® achieved the overall best performance considering the studied variables.

HPV Genotype Prevalence and Success of Vaccination to Prevent Cervical Cancer

Background: Nearly 500,000 new cases of cervical cancer are estimated annually worldwide. Three vaccines are currently licensed to prevent cervical cancer. The success of vaccination depends mainly on the prevalence of HPV genotypes, and many cases of HPV infection have been diagnosed after vaccination. Our aim was to search for HPV genotyping in cervical samples to verify the proportion of women that remain susceptible to infection even after vaccination. Methods: 21,017 liquid-based cervical (LBC) specimens were received for cytology and HPV detection from 2015 to 2018. Before slide preparations for cytology, a 1,000-μL aliquot was taken from the LBC fixative and subjected to automated DNA extraction and multiplex PCR followed by capillary electrophoresis to detect and classify HPV. Results: HPV was detected in 895 (4.3%) specimens. The most prevalent genotype was HPV-16, followed by HPV-58 and HPV-66. A total of 258 (28.8%) cases were positive for high-risk (HR)-HPV types (66, 59, 39, 56, 30, 35, 53, 51, 68, 82, and 70) that are not covered by the HPV vaccines. Conclusion: A significant proportion of HPV types detected in cytological specimens are representative of HR-HPV not covered by the available vaccines. The health system should be aware of the considerable percentage of women who are not being immunized and will continue to need cervical cancer screening.

Role of Ancillary Techniques in Gynecologic Cytopathology Specimens

The association between high-risk genotypes of human papillomavirus (hr-HPV) and cervical cancer is well established. As hr-HPV testing is rapidly becoming a part of routine cervical cancer screening, either in conjunction with cytology or as primary testing, the management of hr-HPV-positive women has to be tailored in a way that increases the detection of cervical abnormalities while decreasing unnecessary colposcopic biopsies or other invasive procedures. In this review, we discuss the overall utility and strategies of hr-HPV testing, as well as the advantages and limitations of potential triage strategies for hr-HPV-positive women, including HPV genotyping, p16/Ki-67 dual staining, and methylation assays.

Utility of Pipet Curet Cytology and Biopsy as a Diagnostic Method for Endometrial Endometrioid Carcinoma

Introduction: We aimed to determine the utility of Pipet Curet cytology (PCC) and Pipet Curet biopsy (PCB) for diagnosing uterine endometrial endometrioid carcinoma (EEC). Methods: We divided 77 patients with EEC into two groups per Federation of Gynecology and Obstetrics (FIGO) grades: G1 (n = 44) and G2/3 (n = 33) and compared the diagnostic sensitivity of PCC, PCB, and PCC and PCB combined, retrospectively. Next, we investigated any diagnostic discordance between PCC-based and PCB-based diagnoses per FIGO grade group. Results: The diagnostic sensitivity of PCC, PCB, and the two modalities combined was significantly higher for G2/3 EECs than for G1 EECs (72.7% vs. 45.5%, p = 0.0209; 84.8% vs. 63.6%, p = 0.0434; and 93.9% vs. 65.9%, p = 0.0046, respectively), likely due to more friable cancer cells in higher grade EEC cases. Among our 77 EEC patients, there were 4 patients (5.19%) with PCC-based concordant but PCB-based discordant results against EEC, in the G2/3 group predominantly. Diagnostic sensitivity of all cases increased from 72.7% (56/77) by PCB alone to 77.9% (60/77) by use of both modalities combined. Conclusion: Cytologic evaluation can reduce the number of false-negative histologic diagnoses. By providing complementary information, the two modalities combined from the Pipet Curet procedure would be valuable as a diagnostic method for EEC.

Can Endometrial Cytology Identify Patients Who Would Benefit from Immunotherapy?

Introduction: Patients with polymerase epsilon (POLE) mutation (POLEmut) subtype, MMR-deficient (MMR-d) subtype as classified by The Cancer Genome Atlas (TCGA), and a high tumor mutation burden (TMB-high) potentially benefit from immunotherapy. However, characteristics of the cytological morphology within these populations remain unknown. Methods: DNA extracted from formalin-fixed paraffin-embedded tissues was subjected to next-generation sequencing analysis. Genomic mutations related to gynecological cancers, TMB, and microsatellite instability were analyzed and were placed in four TCGA classification types. The following morphological cytological investigations were conducted on endometrial cancer using a liquid-based preparation method, prior to the commencement of initial treatment: (i) cytological backgrounds; (ii) differences between each count of neutrophils and lymphocytes as described below. Results: Insignificant differences in the cytological background patterns of TCGA groups and TMB status were found. Although there was no significant difference in neutrophil count (p = 0.955) in the TCGA groups, POLEmut and MMR-d had significantly higher lymphocyte counts than no specific molecular profile (NSMP) (p = 0.019 and 0.037, respectively); furthermore, p53mut also tended to be significant (p = 0.064). Lymphocyte counts in TMB-high were also significantly greater than TMB-low (p = 0.002). POLEmut showed a positive correlation between TMB levels and lymphocyte counts. For predicting patients with POLEmut plus MMR-d, lymphocyte counts demonstrated a superior diagnostic accuracy of area under the curve (AUC) (0.70, 95% CI: 0.57–0.84), with a cutoff value of 26 high-power field. Conclusion: Lymphocyte count using liquid-based cytology for patients with endometrial cancer may predict POLEmut plus MMR-d of TCGA groups and TMB-high in those who can benefit from immunotherapy.

Characteristics of Pelvic Cytology in Patients with Gynecologic Malignancies: Comparison of Pre and Post-Neoadjuvant Specimens

Introduction: The presence of tumor cells in pelvic cytology (PC) specimens can portend a worse outcome for patients undergoing gynecologic surgery. Primary debulking surgery (PDS) was a mainstay for most of these tumors; however, recent advances have triaged selected patients to neoadjuvant chemotherapy (NACT) with interval debulking surgery (IDS). Reduction in tumor cellularity and histologic alterations has been noted in these cases; however, similar cytologic characterization has not been performed. Materials and Methods: PC was searched to find those in NACT patients. Additional PDS were included as controls. Cases were scored for cellularity of malignant cells and background components were described, and when available, pretreatment and posttreatment specimens from the same patients were compared. Results: In all, 19 specimens from 16 patients were found, 6 (32%) of which were paired PTS and IDS from the same patient. Only 6/19 (32%) were from IDS, the remainder PTS. A majority (15/19; 79%) of specimens were malignant; all negative cases were PTS. Few (4/16; 24%) were endometrial primaries; the remainder were pelvic high-grade serous carcinoma. No difference in tumor cell morphology or inflammatory component was noted between the 2 groups, though in 3/3 paired specimens from PDS and IDS, the cellularity of malignant cells decreased in the IDS specimens. Discussion/Conclusion: No identifiable trend was noted regarding cellularity of specimens in the pre compared to the post-neoadjuvant setting. A trend toward reduced cellularity was noted in individual patients, but no alteration in background cells or tumor morphology was noted.

Morphometrical Differences among Endometrial Endometrioid Carcinoma Grade 1, Grade 3, and Serous Carcinoma in Endometrial Liquid-Based Cytology Preparations

Introduction: In Japan, the direct smearing preparation (conventional preparation) has been widely used for cytological examination of the endometrium. Problems with the conventional preparation can be dissolved by liquid-based cytology (LBC) preparation. The Yokohama System is a method for reporting endometrial cytology, but the system lumps cancers together and does not distinguish between histological types. The objective of this study was to clarify morphometrical differences among endometrial endometrioid carcinoma grade 1 (G1), grade 3 (G3), and serous carcinoma (Serous) by image analysis of endometrial LBC. Methods: Using Papanicolaou smears prepared by LBC after sampling with a brush from 32 G1, 16 G3, and 16 Serous patients, image analysis was performed concerning the following 11 items: (1) number of layers of cluster, (2) area of cluster, (3) perimeter of cluster, (4) roundness of cluster, (5) complexity of cluster, (6) area of nucleus, (7) perimeter of nucleus, (8) roundness of nucleus, (9) complexity of nucleus, (10) area of nucleolus, and (11) nucleolus/nucleus (N/N) ratio. The data were statistically compared among G1, G3, and Serous. Results: Significant differences were observed in the number of layers of cluster (G1&#x3c;G3&#x3c;Serous), area of cluster (G1&#x3e;G3&#x3c;Serous), area of nucleus (G1&#x3c;G3&#x3c;Serous), area of nucleolus (G1&#x3c;G3&#x3c;Serous), roundness of cluster (G1&#x3e;G3, G1&#x3e;Serous), complexity of cluster (G1&#x3c;G3, G1&#x3c;Serous), roundness of nucleus (G1&#x3c;Serous), complexity of nucleus (G1&#x3e;Serous), and N/N ratio (G1&#x3e;G3, G3&#x3c;Serous). Conclusion: The number of layers and area of cluster, complexity of cluster, area and complexity of nucleus, area of nucleolus, and N/N ratio were suggested to be useful for the differentiation of histological types among G1, G3, and Serous. Particularly, differences in the number of layers of cluster, size of nucleus, and size of nucleolus among histological types were large and are considered to be potentially effective points of differential diagnosis.

Morphological Differences between Liquid-Based Cytology and Conventional Preparation in Endometrial Endometrioid Carcinoma Grade 1 and Grade 3, and the Differentiation of Grades in Each Method

Introduction: Direct smearing preparation (conventional preparation [CP]) has been widely used for endometrial cytology in Japan. In CP, sampling and screening errors are problematic. In liquid-based cytology preparation (LBC), the problems of CP can be solved. But there is a problem that cytological findings of LBC are different from those of CP. The purpose of this study was to evaluate the differences of morphological findings of endometrial cytology between LBC and CP, and the usefulness of the endometrial LBC to differentiate endometrioid carcinoma grade 1 (G1) from grade 3 (G3). Methods: Thirteen cases of endometrioid carcinoma G1, and 5 cases of G3 collected by the Softcyte device and prepared by LBC and CP (split specimen) were used. We focused on the following items: (1) the number of clusters per cm2, (2) the number of layers of clusters, (3) area of clusters, (4) perimeter of clusters, (5) roundness of clusters, (6) complexity of clusters, (7) area of nucleus, (8) perimeter of nucleus, (9) roundness of nucleus, (10) complexity of nucleus, (11) area of nucleolus, and (12) nucleolus-nucleus ratio (N/N). Results: Compared with CP, the number of clusters and layers of the clusters in LBC were significantly larger in G1. The area and perimeters of the clusters and the nucleus were significant smaller, and the N/N ratio was greater in LBC than that in CP in both G1 and G3. Regarding morphological differences between G1 and G3 in LBC and CP, the number of layers was significantly larger in G1 than in G3 in LBC and CP. The area of the clusters in LBC was significantly larger in G1 than in G3. The area and perimeters of the nucleus in CP and the area of the nucleolus and N/N ratio in LBC and CP were significantly smaller in G1 than in G3. Conclusion: In the endometrial cytology, it became clear that the cell image was different between LBC and CP and between G1 and G3. By microscopic examination understanding the characteristics of the cell image in LBC, endometrial LBC could be useful to diagnose endometrial carcinoma.

Publisher

S. Karger AG

ISSN

0001-5547