Cells In Cytology Specimens Research Articles

Artificial intelligence algorithm accurately assesses oestrogen receptor immunohistochemistry in metastatic breast cancer cytology specimens: A pilot study.

The Visiopharm artificial intelligence (AI) algorithm for oestrogen receptor (ER) immunohistochemistry (IHC) in whole slide images (WSIs) has been successfully validated in surgical pathology. This study aimed to assess its efficacy in cytology specimens. The study cohort comprised 105 consecutive cytology specimens with metastatic breast carcinoma. ER IHC WSIs were seamlessly integrated into the Visiopharm platform from the Image Management System (IMS) during our routine digital workflow, and an AI algorithm was employed for analysis. ER AI scores were compared with pathologists' manual consensus scores. Optimization steps were implemented and evaluated to reduce discordance. The overall concordance between pathologists' scores and AI scores was excellent (99/105, 94.3%). Six cases exhibited discordant results, including two false-negative (FN) cases due to abundant histiocytes incorrectly counted as negatively stained tumour cells by AI, two FN cases owing to weak staining, and two false-positive (FP) cases where pigmented macrophages were erroneously counted as positively stained tumour cells by AI. The Pearson correlation coefficient of ER-positive percentages between pathologists' and AI scores was 0.8483. Optimization steps, such as lowering the cut-off threshold and additional training using higher input magnification, significantly improved accuracy. The automated ER AI algorithm demonstrated excellent concordance with pathologists' assessments and accurately differentiated ER-positive from ER-negative metastatic breast carcinoma cytology cases. However, precision in identifying tumour cells in cytology specimens requires further enhancement.

A multivalent activatable aptamer probe with ultralow background signal and high sensitivity for diagnosis of lung adenocarcinoma

Lung adenocarcinoma is the most common subtype of lung cancer. Developing a simple, rapid, and sensitive method for the detection of lung adenocarcinoma is helpful to reduce the clinical workload and facilitate clinical treatment. To detect lung adenocarcinoma cells, this study designed a multivalent activatable aptamer probe (multi-AP) using S1 aptamer that can specifically recognize A549 cells (a lung adenocarcinoma cell line). Firstly, the monovalent activatable aptamer probe (mono-AP) was designed by strand-displacement strategy and fluorescent resonance energy transfer (FRET). The multi-AP was constructed by mono-AP assembled on the nucleic acid scaffold obtained by hybridization chain reaction (HCR). The presence of the target cell activated the multi-AP, the complementary strand was released, and the fluorescence detection signal was reported based on the FRET. The multi-AP has good specificity and sensitivity for detecting A549 in different matrices and greatly reduces the background signal with the lowest detection limit of 6 cells/200 μL. Furthermore, the multi-AP could directly detect the lung adenocarcinoma cells in cytology specimens, indicating a good potential of the multi-AP in clinical cytology specimens for rapid diagnosis, such as rapid on-site cytological evaluation. Besides, the multi-AP was further modified with biotin (bio-multi-AP) and used to detect clinical cytology cell blocks, implying the value of bio-multi-AP in immunocytochemistry for accurate diagnosis of lung adenocarcinoma.

A deep learning system to diagnose the malignant potential of urothelial carcinoma cells in cytology specimens.

Although deep learning algorithms for clinical cytology have recently been developed, their application to practical assistance systems has not been achieved. In addition, whether deep learning systems (DLSs) can perform diagnoses that cannot be performed by pathologists has not been fully evaluated. The authors initially obtained low-power field cytology images from archived Papanicolaou-stained urinary cytology glass slides from 232 patients. To aid in the development of a diagnosis support system that could identify suspicious atypical cells, the images were divided into high-power field panel image sets for training and testing of the 16-layer Visual Geometry Group convolutional neural network. The DLS was trained using linked information pertaining to whether urothelial carcinoma (UC) in the corresponding histology specimen was invasive or noninvasive, or high-grade or low-grade, followed by an evaluation of whether the DLS could diagnose these characteristics. The DLS achieved excellent performance (eg, an area under the curve [AUC] of 0.9890; F1 score, 0.9002) when trained on high-power field images of malignant and benign cases. The DLS could diagnose whether the lesions were invasive UC (AUC, 0.8628; F1 score, 0.8239) or high-grade UC (AUC, 0.8661; F1 score, 0.8218). Gradient-weighted class activation mapping of these images indicated that the diagnoses were based on the color of tumor cell nuclei. The DLS could accurately screen UC cells and determine the malignant potential of tumors more accurately than classical cytology. The use of a DLS during cytopathology screening could help urologists plan therapeutic strategies, which, in turn, may be beneficial for patients.

Proportion of mast cells in normal canine hepatic cytologic specimens: comparison of 2 staining methods

Although the presence of mast cells is considered a normal finding in canine hepatic tissue, the normal proportion of these cells in cytologic specimens from normal livers was not previously established. The objective of this study was to determine the normal proportion of hepatic mast cells in cytologic specimens from normal canine liver by the use and comparison of the May-Grünwald-Giemsa and the Toluidine Blue staining methods. The cytologic specimens obtained from fine-needle aspirates of livers of 13 healthy dogs were investigated by counting of hepatocytes and mast cells per high-power field (HPF, x40 objective). The ratio of mast cells/100 hepatocytes was then compared between the 2 staining methods. A median of 0.17 (range: 0-0.33) mast cells/100 hepatocytes and of 0.47 (range: 0-0.86) mast cells/100 hepatocytes was obtained with May-Grünwald-Giemsa and Toluidine Blue stains, respectively. Toluidine Blue stain detects more hepatic mast cells than May-Grunwald-Giemsa in normal canine livers. With both stains, < 1% mast cells were detected in normal canine liver samples.

Cytologic subtyping of lung adenocarcinoma by using the proposed International Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society (IASLC/ATS/ERS) adenocarcinoma classification

The significance of histologic subtyping of surgically resected lung adenocarcinoma (ADC) was recently proposed by the International Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society (IASLC/ATS/ERS) classification. Approximately 70% of lung cancer patients present with advanced disease, and small biopsies or cytology specimens are frequently the only available diagnostic material. It is uncertain whether proposed morphologic subtyping of ADC can be applied to small specimens. The objective of this study was to assess the applicability of morphologic subtyping of ADC on cytologic specimens. Consecutive, newly diagnosed primary lung ADC specimens from patients with matched surgical resection and cytology specimens (n = 66) were selected for the study. The dominant morphologic pattern was determined according to the IASLC/ATS/ERS classification. The number and percentage of malignant cells in cytology specimens were also evaluated. Concordant subtyping of ADC between the dominant pattern on resection and cytology specimens was observed in 26 cases (40%), and was discordant in 32 cases (48%). Concordance increased in specimens that had >200 cells and when correlating with the primary or secondary histologic pattern. The acinar pattern was the most common in concordant cases, whereas discordant cases had a predominantly solid pattern. Application of the IASLC/ATS/ERS ADC classification to cytologic specimens is challenging and depends on the sufficient cellularity of cytologic preparations. The identification of solid and micropapillary patterns is prognostically important but may be unreliable and difficult on cytology specimens. Future studies are needed to establish reproducible cytologic criteria for the precise subtyping of lung ADC on small specimens.

Cytologic study of normal canine testis

It has been shown that cytology has a role in the diagnosis of canine infertility, but there are few reports dealing with normal cytologic features of the testis. This is in contrast with human medicine, in which the normal testicular cytology has been thoroughly described and assessed both by qualitative and quantitative approaches. This study was performed to provide a description of the normal cells in cytologic specimens of the testis and to establish ratios between cells that quantify testicular function. Six male dogs ( Canis lupus familiaris) with no testicular pathology, aged 3 to 8 yr, were subjected to elected orchiectomy and included in this study. For each testis, six impression smears were obtained, air dried, and stained with a rapid Romanowsky-type stain. Specimens presented low blood contamination and a heterogeneous population of isolated or loosely packed cells, with high pleomorphism and anisokaryosis; naked nuclei, smeared chromatin, and multinucleation were frequent. The morphologic features of the normal testicular cells from spermatogonia to spermatozoa, including Sertoli and Leydig cells, were described. Cells were classified according to slightly modified criteria taken from human cytology. Simultaneously, 500 cells were counted per slide to determine the sperm index (SI) and Sertoli cell index (SEI), which were 26.6 ± 3.8 and 4.2 ± 0.8, respectively. In general, the cells encountered in the dog resemble those of men, although some particularities exist, such as the much larger size of the Sertoli cells, the highly vacuolated appearance of Leydig cells, or the lower SI and SEI in dogs. The descriptions and indexes defined here may assist in the interpretation of a testicular cytology.

Architectural patterns in cytology: correlation with histology

Cellular arrangements are highly relevant in the evaluation of histologic sections, but cytologic specimens are not thought to accurately reflect the same patterns. Only occasionally do cytologic descriptions comprise, apart from cytoplasmic and nuclear features, architectural patterns. Architectural arrangement of cells in cytologic specimens, however, may mirror the corresponding arrangement of cells in histologic specimens provided the cytologic preparations are collected and prepared appropriately. Recognition and understanding of the different types of cell arrangements that may be seen in cytologic specimens can provide important information for obtaining a definitive diagnosis, particularly for neoplastic lesions. The goal of this review is to describe the most common architectural patterns observed in cytologic specimens and to draw correlations with similar histologic arrangements.

Use of mesothelin as a marker for mesothelial cells in cytologic specimens

Immunocytochemistry is often employed for the distinction between mesothelial cells and adenocarcinoma. Mesothelin has recently been reported to be expressed in reactive mesothelial cells and epithelioid mesotheliomas. The objective of this study is to determine the utility of mesothelin as marker for mesothelial cells in cytologic preparations. Thirty cell blocks were retrieved from the archives and immunostained with monoclonal antibody directed against mesothelin and calretinin. Heat-induced epitope retrieval technique was employed, and the immunostaining was accomplished using an automated stainer. These tissue blocks were from 35 patients (17 females and 18 males) with a median age of 64 years. Nine were benign effusions, 11 mesotheliomas, and 18 metastatic adenocarcinomas. The presence of any immunoreactivity, irrespective of level of intensity or percentage of cells, was considered positive for mesothelin expression. Follow up included correlation with pathology materials obtained at surgery and review of medical records. Mesothelin staining was positive in 7/9 benign cases, 8/11 mesotheliomas, and 8/18 adenocarcinomas. The difference of mesothelin expression between mesothelial cells and adenocarcinoma was statistically significant. For calretinin, all cases, except 2 malignant mesotheliomas and 3 adenocarcinomas, showed positive staining with calretinin. As a marker for mesothelial cells, the sensitivity and specificity of mesothelin were 73% and 55%, respectively, and the sensitivity and specificity of calretinin were 95% and 86%, respectively. Therefore, mesothelin is not a sensitive or a specific marker for mesothelial cells in cytologic specimens when compared with calretinin.

Detection of malignancy in cytology specimens using spectral–spatial analysis

Despite low sensitivity (around 60%), cytomorphologic examination of urine specimens represents the standard procedure in the diagnosis and follow-up of bladder cancer. Although color is information-rich, morphologic diagnoses are rendered almost exclusively on the basis of spatial information. We hypothesized that quantitative assessment of color (more precisely, of spectral properties) using liquid crystal-based spectral fractionation, combined with genetic algorithm-based spatial analysis, can improve the accuracy of traditional cytologic examination. Images of various cytological specimens were collected every 10?nm from 400 to 700?nm to create an image stack. The resulting data sets were analyzed using the Los Alamos-developed GENetic Imagery Exploitation (GENIE) package, a hybrid genetic algorithm that segments (classifies) images using automatically ‘learned' spatio-spectral features. In an evolutionary fashion, GENIE generates a series of algorithms or ‘chromosomes', keeping the one with best fitness with respect to a user-defined training set. First, we tested the system to determine if it could recognize malignant cells using artificial cytology specimens constructed to completely avoid the requirement for human interpretation. GENIE was able to differentiate malignant from benign cells and to estimate their relative proportions in controlled mixtures. We then tested the system on routine cytology specimens. When targeted to detect malignant urothelial cells in cytology specimens, GENIE showed a combined sensitivity and specificity of 85 and 95%, in samples drawn from two separate institutions over a span of 4 years. When trained on cases initially diagnosed as ‘atypical' but with unequivocal follow-up by biopsy, surgical specimen or cytology, GENIE showed efficiency superior to the cytopathologist with respect to predicting the follow-up result in a cohort of 85 cases. We believe that, in future, this type of methodology could be used as an ancillary test in cytopathology, in a manner analogous to immunostaining, in those situations when a definitive diagnosis cannot be rendered based solely on the morphology.

Comparison of immunomarkers for the identification of adrenocortical cells in cytology specimens

We studied the immunoreactivity of three antibodies--A103, calretinin, and inhibin alpha in destained Papanicolaou (Pap) smears and cell-blocks of 40 fine-needle aspiration biopsy cases of adrenocortical lesions (35 cases of hyperplasia/adenoma and 5 cases of carcinoma). Five cases of carcinoma (4) and melanoma (1) metastases to the adrenal gland and five cases of renal-cell carcinoma were also included for comparison. In benign adrenocortical lesions, A103 staining was noted in 82% of the destained Pap smears and in 92% of cell-blocks. In malignant adrenocortical lesions, A103 staining was noted in 50% of the destained Pap smears and in 80% of cell-blocks. In comparison, calretinin staining was noted in 6% and 50% of destained smears and in 78% and 60% of the cell-blocks of benign and malignant adrenocortical lesions. Inhibin alpha was not positive in any of the smears and showed the lowest level of positivity in the cell-block sections, namely in 11% of the benign lesions and 25% of the malignant lesions. The sensitivity of A103 was 90% on cell-blocks and 74% on smears, that of calretinin 75% on cell-blocks and 11% on smears, and that of inhibin alpha, 13% on cell-blocks alone. The specificity of A103 was lower than the other two makers, 90% vs. 100% because of positivity in metastatic melanoma in the adrenal gland. Our data show A103 to be the immunomarker with the highest sensitivity for identifying cells of adrenocortical origin in destained Pap's smears and cell-block sections with, however, a lower specificity when compared with calretinin and inhibin alpha. Calretinin is comparable in sensitivity with A103 on cell-block sections alone and not on smears. The results of this study suggest that if metastatic melanoma in adrenal gland is not a consideration then A103 is the marker of choice for identifying cells of adrenocortical origin in the limited material available for diagnostic purposes in cytology specimens.

Novel tyramide-based tyrosinase assay for the detection of melanoma cells in cytological preparations.

Fine-needle aspiration (FNA) has been used as a fast, minimally invasive, and reliable method for the evaluation of enlarged lymph nodes. However, there are some cases where the definitive diagnosis cannot be elicited with morphology alone, especially cases without a known primary lesion. Although immunocytochemical studies may be helpful in some situations, they are often complicated by nonspecific staining. Recently, a novel tyramide-based tyrosinase assay was developed. Since melanocytes, both benign and malignant, produce tyrosinase, we postulated that this assay could be useful as an in situ biochemical diagnostic test. We modified the Perkin Elmer TSA assay, a commercial assay based on tyramide, a tyrosine analog that is a substrate for tyrosinase, for use on air-dried cytological preparations. We validated the assay on cell lines, then tested a small series of melanoma and nonmelanoma cytology specimens. The YUGEN8 melanoma cell line was used to optimize the assay and it showed abundant reaction product, while HeLa cells served as a negative control. All melanoma cytology specimens were positive and all nonmelanoma specimens were negative. These results suggest that this simple, fast, and inexpensive assay is a sensitive and specific method for detection of melanoma cells in cytology specimens. This method may be a useful ancillary procedure for the resolution of challenging melanoma cases.

Diagnostic classification of urothelial cells in urine cytology specimens using exclusively spectral information.

Although cytologic evaluation of urine specimens is a standard procedure in the diagnosis and follow-up of bladder carcinoma, its sensitivity and specificity are low. Cytopathologic diagnoses are driven primarily by spatial relations or morphology. Although color enhances the pathologist's perception of the specimen, spectral information plays a minimal role in diagnostic processes. Recently, methods have been developed to capture and analyze spectral information from clinical specimens. In the current study, the authors determined the classification value of spectral information by testing its ability to discriminate between malignant and benign urothelial cells in cytology specimens. Multiple images of benign urothelial cells (n = 39) and urothelial carcinoma cells (n = 35) were collected at serial wavelengths using a liquid crystal tunable optical filter and composited into a mosaic using ENVI (Environment for Visualizing Images) software. Through minimum noise fractionation and principal component analysis, the spectral information in the mosaic was compressed into a 29-dimensional scatter plot. The data generated were analyzed using visual and spectral end member extraction on both the original data set and a second independent data set (test set). One area of spectral clustering in the scatter plot segmented with carcinoma cells exclusively (100% specific), but was not present in every cell (approximately 50%), which may indicate that these spectral profiles are present in a subpopulation of malignant cells or at specific points of their cell cycle. Using ENVI algorithms, the authors found that a particular classification spectrum (end member 9) and its closest relatives identified malignant cell clusters, with a sensitivity and specificity that reached 82% and 81%, respectively. To validate this mechanism in a test set, a second mosaic comprised of 15 benign and 15 malignant clusters was analyzed using end member 9, resulting in a combined sensitivity and specificity of 73%. The results of the current study demonstrate that spectral information, in the complete absence of morphologic or spatial information, allows discrimination of benign and malignant urothelial cells in routine urine cytology specimens. The authors believe that this novel technology, combined with spatial analysis, has the potential to serve as an ancillary test for improved detection of bladder carcinoma.

Ultrastructure of Pleural Mesothelioma and Pulmonary Adenocarcinoma in Malignant Effusions as Compared with Reactive Mesothelial Cells

To determine the ultrastructural features of diffuse malignant pleural mesothelioma cells in cytologic specimens from pleural effusions. We retrospectively studied 35 pleural effusions: 12 diffuse malignant pleural mesotheliomas (8 epithelial type, 4 biphasic type), 12 pulmonary adenocarcinomas and 11 cases of reactive mesothelial cells. In the cytoplasm, reactive and malignant mesothelial cells had more-abundant intermediate filaments (P < .05, P < .01) and fewer free ribosomes (P < .001, P < .001) than adenocarcinoma cells. Reactive mesothelial cells had fewer mitochondria than mesothelioma cells (P < .05). Mesothelioma cells had longer, thinner microvilli on the cell surfaces (P < .001); length/diameter ratios of microvilli were 19.1 +/- 7.0 (mesothelioma) vs. 9.1 +/- 2.2 (adenocarcinoma) and 9.2 +/- 2.4 (mesothelial cells). Giant intercellular junctions (desmosomes or desmosomelike structures > 1 micron in length) were found in eight cases of mesothelioma. Core filaments or rootlets in microvilli were present in two cases of adenocarcinoma. Because cytologic specimens from pleural effusions were easy to obtain, we think ultrastructural cytology is useful in distinguishing mesothelioma from adenocarcinoma and benign effusions.

Detection of apoptotic cells in cytology specimens: an application of TdT-mediated dUTP-biotin nick end labeling to cell smears.

We applied TdT-mediated deoxyuridine triphosphate (dUTP)-biotin nick end labeling (TUNEL) to cytologic smears in order to detect the cells undergoing apoptosis. These smears were obtained by scraping the cut surface of 9 cases of carcinoma, including renal-cell carcinoma (3 cases), esophageal squamous-cell carcinoma (3 cases), and gastric adenocarcinoma (3 cases), and were fixed and prepared by different methods. The results were also compared with those of tissue sections. TUNEL in smears was generally associated with higher background nuclear stain than in tissue sections. Smears that were fixed in 4% or 8% paraformaldehyde or absolute methanol exhibited results comparable with those of tissue sections, with minimum background in all cases examined. There were no significant differences in TUNEL labeling index among tissue sections and smears fixed in 4% or 8% paraformaldehyde or in absolute methanol. Smears treated in Carnoy's fixative (3:1 methanol:acetic acid) and air-dried smears demonstrated a higher background. TUNEL positivity could not be detected in slides decolorized from May-Grünwald-Giemsa stain. Markedly high background, which may occur as a result of artifactural DNA breaks, was also observed in slides decolorized from Papanicolaou stain, in which TUNEL-positive cells could be evaluated only in 3/8 cases. Application of the TUNEL method to cytology specimens has disadvantages or limitations compared to its application to histological sections, but the method is considered the most suitable one for detecting cells undergoing apoptosis in cytology materials.

Detection of apoptotic cells in cytology specimens: An application of TdT‐mediated dUTP‐biotin nick end labeling to cell smears

Detection of apoptotic cells in cytology specimens: An application of TdT‐mediated dUTP‐biotin nick end labeling to cell smears

Keratin and carcinoembryonic antigen in exfoliated mesothelial and malignant cells: an immunoperoxidase study.

Immunoperoxidase technics were used to identify keratin and carcinoembryonic antigen (CEA) in exfoliated cells of fine-needle aspirates and body cavity fluids. Staining was evaluated in cytocentrifuge preparations from 27 malignant and 30 benign cytologic specimens. Most reactive mesothelial cell preparations were strongly positive for keratin and negative or only weakly positive for CEA. Diffuse, peripheral, and perinuclear concentration of staining for keratin was noted in exfoliated reactive mesothelial cells. Positive staining for keratin, predominantly diffuse, was noted in exfoliated cells from 56% of the adenocarcinomas. Sixty-nine per cent of adenocarcinoma preparations were strongly positive for CEA. These findings suggest that keratin proteins are not restricted to squamous cells and that keratin staining does not permit distinction between adenocarcinoma and mesothelial cells in cytologic specimens. Staining for CEA and keratin was compared in cytocentrifuge preparations and histologic sections of 12 adenocarcinomas and 7 lymphomas. In some adenocarcinomas, staining was detected only in cytologic preparations. Possible explanations for these differences are discussed. Variable staining for keratin was observed among exfoliated reactive mesothelial cells, possibly identifying different mesothelial cell populations. All reactive and neoplastic lymphoid cells were negative for keratin and CEA in cytologic and histologic preparations. Immunoperoxidase technics can be applied to rehydrated Papanicolaou-fixed and Papanicolaou-stained cytologic preparations with excellent preservation of cytologic detail.