Inter-observer reproducibility of endometrial cytology by the Osaki Study Group method: utilising the Becton Dickinson SurePath™ liquid-based cytology.

OBJECTIVE: The purpose of the present study was to evaluate the reproducibility of the cytological diagnosis of endometrial lesions by the Osaki Study Group (OSG) method of new cytological diagnostic criteria using BD SurePath™ (SP)-liquid-based cytology (LBC). METHODS: This cytological classification using the OSG method consists of six categories: (i) normal endometrium (NE), (ii) endometrial glandular and stromal breakdown (EGBD), (iii) atypical endometrial cells, cannot exclude atypical endometrial hyperplasia or more (ATEC-A), (iv) adenocarcinoma including atypical endometrial hyperplasia or malignant tumour (Malignancy), (v) endometrial hyperplasia without atypia (EH) and (vi) atypical endometrial cells of undetermined significance (ATEC-US). For this study, a total 244 endometrial samplings were classified by two academic cytopathologists as follows: 147 NE cases , 36 EGBD cases , 47 Malignant cases, eight ATEC-A cases, two EH cases and four ATEC-US cases. To confirm the reproducibility of the diagnosis and to study the inter- and intra-observer agreement further, a second review round followed at 3-month intervals, which included three additional cytopathologists. RESULTS: The inter-observer agreement of NE classes improved progressively from 'good to fair' to 'excellent', with values increasing from 0.70 to 0.81. Both EGBD and Malignancy classes improved progressively from 'good to fair' to 'excellent', with values increasing from 0.62-0.63 to 0.84-0.95, respectively. The overall intra-observer agreement between the first and the second rounds was 'good to fair' to 'excellent', with values changing from 0.79 to 0.85. All kappa improvements were significant (P < 0.0001). CONCLUSION: In this study, it seemed that the use of the OSG method as the new diagnostic criteria for SP-LBC preparation, may be a valid method to improve the precision (reproducibility) of endometrial cytology.

Body cavity fluid can induce epithelial and mesothelial differentiation from CD34 positive peripheral blood stem cells in vitro.

OBJECTIVE: Primary culture of CD34 positive stem cells collected from human peripheral blood was performed with and without supplementation with concentrated ascitic fluid; morphological and immunocytochemical pictures of cultured cells were taken chronologically and compared. METHODS: CD34-positive stem cells collected from peripheral blood were cultured for 1, 24 and 48 hours. Concentrated ascitic fluid was added to the plates for the 24-and 48-hour cultures. For immunocytochemical studies, CD34, AE1/AE3, Ber-Ep4 (EA), EMA, EGFR, CD31, CA125 and D2-40 monoclonal antibodies were used. RESULTS: After culture, small round cells with naked nuclei began to enlarge and to exhibit various changes in the cytoplasm and nucleus. Supplementation with concentrated body cavity fluid enhanced these changes. CD34-positive cells with small round cell features were detected 1 hour after culture and these had no epithelial or mesothelial markers. After 24 hours, CD34-positive cells had disappeared and cells weakly positive for EGFR, EMA, CA125 and D2-40 were detected. Cells with strong and moderate positive reactions for EGFR, AE1/AE3, EA, EMA, D2-40 and CA125 were detected after 48 hours. Supplementation with concentrated body cavity fluid increased the intensity and number of positive cells for these markers compared with the control group. The positive reaction, not only for the epithelial markers such as EGFR and AE1/AE3, but also for mesothelial markers such as CA125 and D2-40, was found to be increased in small numbers of cells in direct proportion to the duration of the primary culture of the peripheral blood cells. CD31, characteristically expressed in endothelial cells, was negative in the cultured cells. CONCLUSION: Supplementation of peripheral blood CD34-positive stem cells with body cavity fluid in vitro enhanced their differentiation toward cells of an epithelial or mesothelial phenotype, concomitant with loss of immunoreactivity for CD34. It is assumed that the routine cytological observation of cells obtained from body cavity fluid might cause possible cytomorphological and immunophenotypical changes due to the action of the growth factors contained in the body cavity fluid.

EMMPRIN (CD147) expression and differentiation of papillary thyroid carcinoma: implications for immunocytochemistry in FNA cytology.

OBJECTIVE: Extracellular matrix metalloproteinase inducer (EMMPRIN) and its induced matrix metalloproteinases (MMPs) play a crucial role in tumour progression, invasion and metastasis. EMMPRIN expression has been demonstrated in several tumours, but its expression profile in thyroid cancer remains unclear. METHODS: We evaluated the expression profile of EMMPRIN at various stages of differentiation of thyroid carcinoma, including 20 cases of well-differentiated papillary carcinoma (WDPC), 15 cases of papillary carcinoma with a poorly differentiated carcinoma component (PC/PDC) and four cases with an undifferentiated carcinoma (UDC) component, using paraffin-embedded sections for immunohistochemical stains. Also, we used 32 fine needle aspiration cytology and imprint smears from the same cases for immunocytochemical stains. The staining results were evaluated with a scoring system. RESULTS: Immunohistochemical staining showed that EMMPRIN expression was absent or weak in almost all WDPC specimens, whereas it was moderate or strong in PDC and UDC components. In tumours that showed a gradual morphological transformation from WDPC to PDC components, the expression of EMMPRIN was progressively stronger from the areas of WDPC to those of PDC. WDPC, PC/PDC and UDC had expression scores of 4.9, 45.0 and 245.7, respectively. Results of immunocytochemical staining showed almost the same staining profile as those of immunohistochemical staining. The cytological atypia of EMMPRIN-positive cells was greater than that of negative cells. CONCLUSION: These results indicated that EMMPRIN expression correlates significantly with the degree of dedifferentiation of thyroid carcinoma. This study demonstrates the feasibility of expression of EMMPRIN using fine needle aspiration samples. Therefore, immunocytochemical analysis of EMMPRIN may be a novel aid to evaluate the differentiation of thyroid carcinoma.

Utility of liquid-based cytology in endometrial pathology: diagnosis of endometrial carcinoma.

OBJECTIVE: The purpose of this study was to examine the utility of SurePath-liquid-based cytology (LBC) compared to conventional cytological preparations (CCP) in the identification of endometrial carcinoma. METHODS: During a 13-month period, direct endometrial samples were collected from 120 patients using the Uterobrush. The material comprised 30 cases each of endometrial carcinoma, proliferative endometrium, secretory endometrium and atrophic endometrium. The following points were investigated:(i) the frequency of cell clumps in endometrial carcinoma; (ii) the area of cell nuclei; (iii) overlapping nuclei. RESULTS: (i) Comparison of the frequency of cell clumps with irregular protrusion pattern and papillo-tubular pattern showed no statistically significant difference in either type of cell clump between CCP and LBC. (ii) Comparison of the nuclear area of cells showed a sequential decrease from endometrial carcinoma to secretory endometrium, to proliferative endometrium and to atrophic endometrium, which was significant in CCP and LBC. (iii) Nuclear area was significantly lower with LBC compared with CCP in endometrial carcinoma, secretory endometrium and proliferative endometrium but not atrophic endometrium. (iv) Comparison of the degree of overlapping nuclei showed a sequential decrease from endometrial carcinoma to proliferative endometrium, to secretory endometrium and to atrophic endometrium, which was significant in both CCP and LBC. (v) Comparison of the degree of overlapping nuclei between CCP and LBC showed no significant difference for normal types of endometrium, but LBC had significantly higher values (P < 0.0001) in endometrial carcinoma than in CCP. CONCLUSIONS: The results of this study revealed that applying diagnostic criteria used in CCP to LBC was easy to achieve, because LBC had excellent cytoarchitectural preservation and cells were well presented. Although we have not examined all cytological features of malignancy and have not considered atypical hyperplasia, we believe that this method may be a useful tool in the diagnosis of endometrial cytology.

New diagnostic reporting format for endometrial cytology based on cytoarchitectural criteria.

OBJECTIVE: The aim of this study was to develop a new reporting format for endometrial cytology that would standardize the diagnostic criteria and the terminology used for reporting. METHODS: In previous studies, cytoarchitectural criteria were found to be useful for the cytological assessment of endometrial lesions. To apply these criteria, an appropriate cytological specimen is imperative. In this article, the requirements of an adequate endometrial cytological specimen for the new diagnostic criteria are first discussed. Then, the diagnostic criteria, standardized on a combination of conventional and cytoarchitectural criteria, are presented. Third, terminology that could be used, not only for reporting the histopathological diagnosis, but also for providing better guidance for the gynaecologist to determine further clinical action, is introduced. The proposed reporting format was investigated using endometrial cytology of 58 cases that were cytologically underestimated or overestimated compared to the histopathological diagnosis made on the subsequent endometrial biopsy or surgical specimens. RESULTS: Of the 58 cases, 12 were reassessed as being unsatisfactory for evaluation. Among the remaining 46 cases, 25 of the 27 cases, which had been underestimated and subsequently diagnosed as having endometrial carcinoma or a precursor stage on histopathological examination,were reassessed as recommended for endometrial biopsy. On the other hand, 19 cases overestimated by cytology were all reassessed as not requiring biopsy. CONCLUSIONS: The reporting format for endometrial cytology proposed in this article may improve diagnostic accuracy and reduce the number of patients managed inappropriately.

Fine needle aspiration cytology: a survey of current European practice.

Fine needle aspiration cytology (FNAC) is practised widely throughout Europe. The majority of countries have dedicated cytopathologists as well as histopathologists practicing cytology. Despite this, FNAC is performed mostly by clinicians and radiologists except in the larger centres with dedicated staff with a special interest in cytopathology. The advent of One-Stop diagnostic services and image-guided procedures are prompting further development of FNAC clinics where cytopathologists take their own samples, issue reports in the same clinical session and take extra material for ancillary tests to complete the diagnosis. The volume of FNAC work varies accordingly; in dedicated centres FNAC represents up to 80% of the workload whilst, in the majority of countries, it represents one quarter or less. Hence, the rate of inadequate FNAC varies widely, depending on the local sampling policies and the organ, but does not exceed 25% in any of the countries. The most sampled organs are breast and thyroid, followed by lymph nodes. Most countries have dedicated training in cytopathology for pathology trainees, the duration varying between 6 months and 2 years of the total training time. This discussion, focusing on European practices, highlights the heterogeneity of FNAC activity but also its success in many centres where it is practiced to a high standard, particularly in breast, thyroid and lymph node pathology. The relatively high rate of inadequate material in some centres reflects local policies and calls for greater uniformity of FNAC practice, particularly specimen sampling. To achieve this, the future direction should concentrate on specialist training, to include performing as well as interpreting FNAC, as part of the curriculum. Current emphasis on web-based training may not provide first hand experience of the FNAC procedure and should be supplemented by attending FNAC clinics and developing the technique to its full potential.

Application of the cell block method, utilizing mount quick mounting medium.

Our objective was to determine the applicability of cell transfer and cell block methods using Mount Quick (Daido Sangyo, Saitama, Japan) mounting medium (MQ) for hematoxylin-eosin (H&E) and immunohistochemical staining of several limited amounts of biological materials in slide preparations. The materials investigated were histopathologically confirmed malignant mesotheliomas (pleural effusions) and malignant lymphomas, a malignant melanoma, and an amelanotic melanoma in sealed slides. Monoclonal antibodies against carcinoembryonic antigen (CEA), epithelial membrane antigen (EMA), cancer antigen 125 (CA-125), vimentin, thrombomodulin (TM), cytokeratin, UCHL-1, L-26, melanoma-specific antigen (HMB45), and S-100 protein (S-100) were applied in the investigation. The malignant mesotheliomas were found to be positive for EMA, cytokeratin, vimentin, TM, and CA-125, and negative for CEA, with no differences being observed in findings from direct contact preparations. Using T-cell-type malignant lymphomas for immunohistochemistry, UCHL-1 positivity and L-26 negativity were clearly demonstrated. The malignant melanoma and amelanotic melanoma materials stained strongly for HMB45 and S-100. Cell transfer employing MQ is a suitable approach for immunohistochemical investigations of limited materials. In addition, cell blocks derived from MQ-embedded smears can be used for both H&E and immunohistochemical staining. Diagn. Cytopathol. 2000;22:117-119.

Cytopathology of pregnancy-induced cell patterns in cervicovaginal smears.

Considerable interest has been devoted to cytology in pregnancy, especially the morphologic changes that may cause problems in differential diagnosis. It is surprising that the published discussion of the cytologic appearance of smears from pregnant women has been so limited. This review emphasizes that retained trophoblastic tissue may be a source of highly atypical appearing cells in the cervicovaginal cytology obtained under various clinical conditions. Distinguishing between Arias-Stella cells and cells of glandular abnormalities can be problematic, since the morphologic characterization of the former is poor. This review also emphasizes that a full awareness of the morphology of pregnancy as well as of the patient's clinical history are needed for greater precision in diagnosing cell patterns as pregnancy-related and not malignant.

Association of mast cells with Warthin's tumor in fine needle aspirates of the salivary gland.

OBJECTIVE: To determine the significance of the presence of mast cells in Warthin's tumor by evaluating the occurrence of these cells in cellular and immunohistochemical preparations. STUDY DESIGN: Specimens derived from five cases of FNAC were examined. A total of four slides from five cases were prepared from each: two air-dried smears were stained with May-Grünwald-Giemsa (MGG) stain and two with Hansel's stain. The other two were alcohol fixed and stained using the Papanicolaou method. The smears were evaluated for the presence of mast cells, especially associated with oxyphilic cells. In order to investigate the location of mast cells, we also counted those cells by means of immunohistochemistry using anti-mast cell monoclonal antibody AA1. RESULTS: The Hanselstained cellular sample from Warthin's tumor contained numerous mast cells, associated mainly with large, oxyphilic cell sheets. The number of AA1-positive cells (mast cells) stained with immunohistochemistry was greater in epithelial component than in lymphoid stroma. CONCLUSION: Mast cells in a salivary gland aspirate might be indicative of Warthin's tumor; therefore, MGG-stained slides offer the advantage of ease of preparation, particularly when the typical cytologic features are not present.

Fine-needle aspiration cytology of malignant hemangiopericytoma of the salivary gland: A case report.

A 79-yr-old woman presented with a 5-yr history of swelling of the left cheek. The fine-needle aspiration (FNA) smear showed a spindle-cell neoplasm with capillaries and benign endothelial cells. The spindle cells possessed pleomorphic, hyperchromatic elongated nuclei and a moderate amount of ill-defined cytoplasm. They also showed papillary arcades surrounded and encased by relatively small ovoid to short spindle cells. Subsequent surgical excision confirmed the presence of malignant hemangiopericytoma (HP). Immunohistochemical studies on the histologic section using vimentin were strongly positive, consistent with HP. To the best of our knowledge, this is the second published report of FNA cellular features of malignant HP of the salivary gland. Besides delineating the FNA cellular features of HP of the salivary gland, the present case illustrates the value of using immunohistochemical approaches. Diagn. Cytopathol. 1999;21:398-401.

Scrape cytology of oral pemphigus. Report of a case with immunocytochemistry and light, scanning electron and transmission electron microscopy.

BACKGROUND: Pemphigus vulgaris is a disseminated disease of the skin and mucous membranes characterized by recurrent vesicular and bullous lesions due to the autoantigen belonging to the cadherin type of cell adhesion molecules. The presence of acantholysis associated with immunoglobulins in the intercellular spaces and on the cell membrane are diagnostic features. However, the appearance of smears from the oral cavity by scanning (SEM) and transmission electron microscopic (TEM) study as well as immunocytochemistry of cadherin does not appear to have been previously reported. CASE: A 67-year-old female developed erosion on her gingiva with severe pain. On oral examination, there were ulcerations on the palate, and the Nikolsky sign was positive. The characteristic cytologic findings from oral scrapes were high cellularity, a bloody background and a predominant cell population consisting of polygonal basal and parabasal cells with pronounced nucleoli. Also present were degenerative cell changes: e.g., cytoplasmic vacuoles and a homogeneous nuclear appearance. Immunocytochemical staining for IgG and cadherin gave a positive reaction in the intercellular spaces and on the cell membranes. The surface of cells in pemphigus vulgaris by SEM showed somewhat irregularly distributed microridges, and TEM revealed desmosomal attachments, degenerated tonofilaments with pronounced nucleoli and heterochromatin. As a result of cytodiagnosis, additional appropriate specimens were obtained at the time of the scraping for confirmatory immunocytochemistry for cadherin, SEM and TEM studies. CONCLUSION: The results demonstrate that a precise diagnosis of pemphigus vulgaris can be rendered on cellular material and cadherin immunocytochemistry obtained by scrape from the oral mucosa.

Polymerase chain reaction amplification of Trichomonas vaginalis DNA from Papanicolaou-stained smears.

DNA from Papanicolaou-stained smears was successfully amplified using the polymerase chain reaction (PCR) to investigate if it could be used for retrospective genome studies, such as for the detection of Trichomonas vaginalis. DNA was isolated from 20 archival Papanicolaou-stained smears (19 cervical and 1 seminal fluid samples) and purified by treatment with proteinase K, phenol/chloroform, and ethanol. The method of recovering DNA from the Papanicolaou-stained smear is very important in the successful amplification of Trichomonas DNA in archival cytological smears. We used a specific sequence and successfully amplified DNA in cytological smears of T. vaginalis using PCR. We conclude that the diagnostic accuracy of the detection of T. vaginalis in cervical smears is enhanced by using a combination of a Papanicolaou-stained smear and PCR.

Coexistence of Pemphigus vulgaris and herpes simplex virus infection in oral mucosa diagnosed by cytology, immunohistochemistry, and polymerase chain reaction.

A case of Pemphigus vulgaris concurrent with hepres simplex virus (HSV) infection in a 53-yr-old female is described, in which the diagnosis was based on oral scraping cytology. Two populations of abnormal cells were identified in the oral smear. One abnormal cell population was characterized by the presence of numerous single cells and sheets and smaller aggregates of loosely cohesive epithelial cells that appeared to have only a few points of intercellular attachment. A second population of abnormal cells showed characteristic signs of HSV infections such as ground-glass nuclear appearance and multinucleation. Subsequently, diagnosis of HSV infections based on polymerase chain reaction was applied to identify the specific DNA for HSV type 1 in the Papanicolaou specimens. To our knowledge, this is the first case in which the coexistence of Pemphigus vulgaris and HSV infection in the oral mucosa was established by cytologic diagnosis. This is discussed in view of our recent experience with this unusual oral lesion.