Survey of medical training in cytopathology carried out by the journal Cytopathology.

This report of the Editorial Advisory Board of Cytopathology gives the results of a survey of medical practitioners in cytopathology, which aimed to find out their views on the current situation in undergraduate and postgraduate training in their institutions and countries. The results show that training in cytopathology and histopathology are largely carried out at postgraduate level and tend to be organized nationally rather than locally. Histopathology was regarded as essential for training in cytopathology by 89.5% of respondents and was mandatory according to 83.1%. Mandatory cytopathology sections of histopathology were reported by 67.3% and specific examinations in cytopathology by 55.4%. The main deficiencies in training were due to its variability; there were insufficient numbers of pathologists interested in cytology and a consequent lack of training to a high level of competence. Pathologists without specific training in cytopathology signed out cytology reports according to 54.7% of responses, more often in centres where training was 3-6 months or less duration. Although 92.2% of respondents thought that specialist cytology should not be reported by pathologists without experience in general cytopathology, that practice was reported by 30.9%, more often in centres with small workloads. The survey report recommends that 6-12 months should be dedicated to cytopathology during histopathology training, with optional additional training for those wanting to carry out independent practice in cytopathology. Formal accreditation should be mandatory for independent practice in cytopathology. When necessary, temporary placements to centres of good practice should be available for trainees intending to practise independently in cytopathology. There should be adequate numbers of pathologists trained in cytopathology to a high level of competence; some of their time could be released by training cytotechnologists and trainee pathologists to prescreen cytology slides and assess adequacy of fine-needle aspiration samples when immediate diagnosis was not required. The survey demonstrated a clear need for European and international guidelines for training in cytopathology.

Prognostic impact of HER2/neu protein in urothelial bladder cancer. Survival analysis of 80 cases and an overview of almost 20 years' research.

PURPOSE: This study was conducted to evaluate the quantitative assessment of HER2/neu immunohistochemical expression in urothelial bladder cancer in order to determine its prognostic significance. MATERIALS AND METHODS: Archival tumor tissue from 80 patients with primary urothelial carcinoma were analysed for HER2/neu immunohistochemical expression. A highly reproducible standardized procedure on a Bond-X automated slide stainer was used. RESULTS: HER2 protein was overexpressed in 41 of 80 patients (51.25%), demonstrating an increase in the expression rate corresponding to progressively advanced tumor stage (p=0.032) and tumor grade (p=0.0001). Kaplan-Meier analyses showed that positive membranous expression of HER2/neu was not associated with an increased probability of tumor recurrence (p=0.362). In contrast, HER2 scores correlated strongly with specific survival probability (p=0.002) and overall survival (p=0.025). Multivariate analysis revealed that only stage was an independent predictor of specific survival (p=0.016). HER2 expression was an independent predictor of specific survival with borderline statistical significance (p=0.08). CONCLUSION: HER2 overexpression represents a prognostic factor for adverse disease outcome.

The role of breast FNAC in diagnosis and clinical management: a survey of current practice.

Most participating countries have now adopted a triple assessment approach, i.e. clinical,imaging and pathology, to breast diagnosis, with FNAC as the first-line pathological investigation in both screening and symptomatic populations, with the exception of microcalcifications. Pathologists specialized in cytopathology are best qualified to collect and interpret FNAC samples, but this is not always possible or practical. Radiologists involved in breast imaging should ensure that they have the necessary skills to carry out FNAC under all forms of image guidance. Best results are achieved by a combination of both techniques, as shown in the image-guided FNAC in the presence of the cytopathologist. The majority of European countries use similar reporting systems for breast FNAC (C1-C5), in keeping with European Guidelines for Quality Assurance in Breast Cancer Screening and Diagnosis, although some still prefer descriptive reporting only. When triple assessment is concordant, final treatment may proceed on the basis of FNAC, without a tissue biopsy. ER and PR assessment can be done safely on FNAC material. However, not all institutions may have expertise in doing this. HER-2 protein expression on direct cytological preparations is insufficiently reliable for clinical use, although its use for FISH is possible, if expertise is available. The majority of participants practise a degree of one-stop diagnosis with a cytopathologist present in the out-patient clinic. Formal recognition of the importance of the time spent outside the laboratory, both for cytopathologist and cytotechnologist, is necessary in order to ensure appropriate resourcing. The use of core biopsy (CB) has increased, although not always for evidence-based reasons. CB and FNAC are not mutually exclusive. FNAC should be used in diagnosis of benign, symptomatic lesions and CB in microcalcifications, suspicious FNAC findings and malignancies where radiology cannot guarantee stromal invasion.

Extracellular regulated kinase-2 immunoreactivity increases in parallel with cervical intraepithelial neoplasia grade in cervical neoplasia.

The cell cycle control system includes cyclins, cyclin-dependent kinases (CDK), and their inhibitors (CDK1). Extracellular regulated kinase (ERK1/2) (p44 and p42 mitogen-activated protein kinases [MAPKs]) is a component of the MAPK pathway, which is associated with cyclin D1 and CDK. It is a critical signaling system for the induction of cell proliferation, differentiation, and cell survival. The aim of this study was to investigate the usefulness of ERK2 expression as a marker of biological aggressiveness complementary to cervical intraepithelial neoplasia (CIN) grade as well as to compare its expression in preinvasive lesions with that in invasive carcinoma. Paraffin-embedded sections of 146 CIN lesions (32 CIN I, 49 CIN II, and 43 CIN III) and 22 invasive cervical carcinomas (13 squamous and 9 adenocarcinomas) were used for the standard immunohistochemical procedure with the application of the ERK2 monoclonal antibody. ERK2 staining displayed a cytoplasmic and nuclear pattern. The staining intensity was gradually increased according to the severity of the dysplastic lesions; ERK2 immunoreactivity was significantly increased in high-grade dysplastic lesions (CIN II and CIN III) and invasive carcinomas by comparison to low-grade dysplastic lesions (CIN I) (P < 0.001). When high-grade lesions were separately assessed, the differences between each one of them and CIN I retained their statistical significance: CIN II versus CIN I (P < 0.001) and CIN III versus CIN I (P < 0.001). In conclusion, our study found a direct relationship between the increasing grade of the dysplastic cervical lesions and the intensity of ERK2 staining, thus implying a role of ERK2 as an early event in cervical carcinogenesis.

Altered expression of adhesion molecules in inflammatory cervical smears.

OBJECTIVE: The aim of the present study was to evaluate the expression of pan-cadherin and beta-catenin in cervical smears with various types of infectious agents. PATIENTS AND METHODS: Cervical smears obtained from 53 women, aged 21-65 years, with a diagnosis of specific inflammation were examined in our study. Eighteen subjects were infected by Candida albicans, 18 by Gardnerella vaginalis, nine by Bacteroides spp. and eight by Chlamydia trachomatis. All infectious agents found in the smears were at the same time confirmed by the microbiological laboratory methods. We performed a biotin-streptavidin-peroxidase immunocytochemical method using anti-beta-catenin (Clone 12F7) and anti-pan-cadherin (pan, polyclonal) antibodies. RESULTS: Aberrant expression of pan-cadherin was found in the cytoplasmic membrane of glandular, metaplastic, superficial and intermediate squamous cells in all types of infections. With regard to beta-catenin, this was expressed in majority (90%) of glandular and metaplastic cells in all types of infections and in a small proportion (15%) of superficial and intermediate squamous cells in infections caused by C. albicans and G. vaginalis. CONCLUSION: Our data show that infectious agents may cause alterations in the expression and distribution of these adhesive molecules, which can be recognized in cervical smears. Additional studies in larger sets of patients should help clarify this issue further.

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.

Epidermal growth factor receptor and proliferating cell nuclear antigen expression in urine ThinPrep specimens.

OBJECTIVE: To evaluate proliferating cell nuclear antigen (PCNA) and epidermal growth factor receptor (EGFR) expression in urine ThinPrep (TP) specimens, to compare these findings with clinical and histological features and to determine whether these immunomarkers are predictive of clinical stage. PATIENTS AND METHODS: The TP processed urine samples and the corresponding tissue sections from 42 patients with newly diagnosed bladder cancer (18 non-muscle invasive and 24 muscle invasive) were included in our study. Urine was collected for cytological evaluation before transurethral resection. Tumour grade and clinical stage were assessed from the transurethral resection specimens. The EGFR and PCNA expression was obtained by an automated immunostainer. RESULTS: There was a remarkable concordance in the expression of both antibodies in TP smears and tissue sections. No significant association was detected for any of the immunomarkers examined with regard to tumour grade. The EGFR expression as well as grade of malignancy were significantly associated with stage of disease (P = 0.0001). PCNA was not found to be a significant predictor of stage (P = 0.210). CONCLUSION: Our data suggest that the evaluation of grade of malignancy and EGFR immunopositivity can be considered as reliable predictors of disease stage in urine TP specimens.

BSCC, Bethesda or other? Terminology in cervical cytology European panel discussion.

The European panel agreed that reproducibility and translatability of terminology in cervical cytology were essential, arguing well for harmonization of reporting systems. The majority at this meeting use a modification of the Bethesda system (BS). Local modifications involved reporting subcategories within high grade and low grade lesions, which would not alter the overall translatability of their systems both with each other and BS. The majority agree that low grade lesions with and without koilocytosis should be managed similarly as should high grade lesions (moderate dysplasia/CIN2 or worse). Those systems linking moderate dysplasia with mild rather than severe dysplasia would need to define moderate dysplasia as such, if their results were to be translatable, which would be preferable to their using a different definition of low grade and high grade lesions. Translation between systems might anyway be facilitated by reporting moderate dysplasia as a subcategory within high grade, which was favoured by most of those present. Therefore, there is no need for exact agreement of terminology if broad principles are agreed. This useful discussion adds weight to the British Society for Clinical Cytology recommendation that the new classification should be adopted by the UK National Health Service Cervical Screening Programme. If the new classification is adopted, the UK would join the European consensus opinion on terminology.

Tumour-stroma interactions in carcinogenesis: basic aspects and perspectives.

In contrast to the conventional notion regarding tumour development as a cell autonomous process in which the major participants were the cancer cells, increasing evidence attributes important role in the stromal components, namely fibroblasts, and view the tumour as a heterogenous mixture of different cell types. These different types of cells, being cancer cells, fibroblasts, endothelial cells, and others, interact reciprocally and play an almost equally important role in the manifestation of certain aspects of the malignant phenotype. The elucidation of the mechanistic base of such interactions, besides the contribution to understand fundamental aspects of tumour cell biology, promises important applications in diagnosis, prognosis and therapy of the disease.

Immunohistochemical detection of cell cycle regulators, Fhit protein and apoptotic cells in parathyroid lesions.

OBJECTIVE: The pathological distinction between parathyroid neoplasms and hyperplasias remains difficult. Changes in cell cycle control may lead to clonal proliferation and precede tumorigenesis. The parathyroid adenoma 1 oncogene, subsequently identified as the gene encoding cyclin D1, has been shown to be important to parathyroid tumour development. In addition to cell proliferation, the mechanisms of parathyroid cell turnover include apoptosis. The tumour-suppressor activity of the fragile histidine triad gene (FHIT) is linked to its proapoptotic function and cell cycle control. We attempted to evaluate the cellular proliferative kinetics and apoptotic function of the parathyroid glands in patients with non-familial hyperparathyroidism (HPT). DESIGN: TIssue specimens were taken from 40 patients with primary HPT (17 adenomas, two carcinomas and 21 primary hyperplasias) and from 30 patients with secondary HPT. Normal glands served as controls. METHODS: In a standard immunohistochemical procedure, monoclonal antibodies to Ki-67 antigen and single-stranded DNA were applied to detect cycling and apoptotic cells respectively; polyclonal antibodies to cyclin D1 and Fhit protein were used. Immunostaining was estimated by image analysis and statistical analysis was subsequently performed. RESULTS: Significantly higher proliferative and apoptotic indexes were detected in the diseased glands in comparison with normal controls. In neoplastic and secondarily hyperplastic glands, apoptotic indexes were higher than in primarily hyperplastic glands; the difference between neoplastic and primarily hyperplastic glands was statistically significant (P=0.034). Cyclin D1 was overexpressed in a considerable proportion of tumours (68.4%). A reduction of Fhit protein immunoreactivity was selectively noticed in carcinomas. CONCLUSIONS: In primary hyperplasia, the remarkable proliferation of parathyroid glands may be due to the reduction of the apoptotic process. FHIT gene abnormalities are worthy of investigation in parathyroid carcinogenesis.

Papillary serous carcinoma of peritoneum: case study and review of the literature on the differential diagnosis of malignant peritoneal tumors.

The distinction between malignant mesothelioma and other malignant neoplasms diffusely involving the peritoneum is important for proper patient treatment. The extra-ovarian peritoneal serous papillary carcinoma is a rare, primary, multicentric peritoneal tumor that is morphologically identical to ovarian serous carcinoma of equivalent grade, but can spare or minimally involve the ovaries. We report such a tumor in a 65-year-old female who had abdominal swelling, ascites with positive cytology and a high grade of nuclear atypia in malignant cells as well as elevated serum CA125. Exploratory laparotomy findings of intrabdominal carcinomatosis were not accompanied by any evident primary site; so the diagnosis of a primary papillary serous neoplasia of the peritoneum was strongly considered. Since the amount of residual disease may be an important prognostic determining factor in primary papillary serous carcinoma of the peritoneum, the patient was debulked to no macroscopic disease and was then given platin-based chemotherapy. The tumor's differential diagnosis from malignant mesothelioma was based, apart from morphologic criteria, on the tumor's immunoreactivity to MOC-31, Ber-EP4 and TAG-72, as well as on the lack of immunostaining for keratin 5/6 and calretinin. Differential diagnosis from ovarian cancer was possible only after the pathological examination of the surgically resected ovaries; the tumor showed minimal superficial invasion of the ovarian cortex.

Significance of nuclear morphometry as a diagnostic tool in fine-needle aspirates of the liver.

OBJECTIVE: Classification of hepatic tumours and tumour-like conditions can sometimes be difficult to establish by light microscopy. Our aim was to determine the value of computerized interactive nuclear morphometry in the preoperative prediction of primary or metastatic malignancy as opposed to non-malignant lesions, in fine-needle aspirates (FNA) of hepatic lesions. METHODS: Alcohol-fixed, Papanicolaou-stained FNA smears of 99 histologically proven hepatocellular carcinomas (HCCs), metastatic neoplasms and benign lesions were measured by computerized image analysis with regard to nuclear major axis, minor axis, perimeter, area, shape factor and hyperchromasia. Data were coded, entered into a computerized database, and statistically analysed with SPSS programs. RESULTS: Tukey HSD test for multiple comparisons, assessed for all features except hyperchromasia, showed that the mean values of morphometric features of nonmalignant cells were significantly different from those of malignant cells, either primary or metastatic, whereas differences between morphometric characteristics of HCCs and metastatic neoplasms were insignificant. Kruskal-Wallis analysis of variance revealed that hyperchromasia varied among the three groups of samples proportionally to the other nuclear features. The mean differences of all evaluated nuclear variables except minor axis were significant between grades II & III and grade IV HCCs. Morphometric mean values of well-differentiated HCCs differed significantly in comparison with those of non-malignant lesions; however, some degree of overlap was observed in the ranges of minor axis and hyperchromasia mean values. CONCLUSIONS: The three most important cytological criteria of nuclear malignancy (hyperchromasia, enlargement and anisonucleosis), when quantified by morphometry, may be helpful in the differential diagnosis between non-cancerous liver lesions and HCCs (even those of high differentiation), since all the morphometric data showed pronounced differences between malignant and benign groups. Morphometry may also be used as a complementary tool in the cytological grading of HCCs.

Relationship of epidermal growth factor receptor (EGFR), proliferating cell nuclear antigen (PCNA) and vimentin expression and various prognostic factors in breast cancer patients.

Cytologic specimens (FNA) from 42 primary invasive ductal breast carcinomas and 22 matched specimens of cancer tissue were tested for EGFR status, PCNA index and vimentin expression by immunocytochemical staining, using an Extravidin-Biotin method, and their relationship with various prognostic factors was investigated. EGFR positivity, high PC10 score and vimentin positivity were significantly correlated with high histologic grade. The coordinate expression of EGFR, PCNA and vimentin was significantly associated with ER-negative breast carcinomas. A positive trend was observed between high proliferating tumours and EGFR expression. EGFR status and PCNA index were not correlated with axillary lymph node involvement, tumour size, age and menopausal status. Vimentin was preferentially expressed in tumours, with lymph node metastases. Co-expression of EGFR, PCNA and vimentin was determined in most cases. These data suggest that EGFR status, PCNA index and vimentin expression may be important for the prediction of biologically aggressive tumours.

Detection of human papillomavirus DNA in nongenital seborrhoeic keratoses.

The histological similarities of seborrhoeic keratoses and common warts led to the investigation of the possible occurrence of human papillomavirus DNA (HPV-DNA) in a large number of nongenital seborrhoeic keratoses using the in situ hybridization technique. All specimens derived from normal skin (n = 173) were negative for the applied HPV-DNA probe, whereas the HPV genome was detected in 34 of 173 seborrhoeic keratosis specimens (19.65%). Of 34 HPV-positive specimens, 15 contained types 6/11 and 14 types 31/33/35, and 5 showed no positive reaction to the applied types. These results suggest that a considerable percentage of nongenital seborrhoeic keratoses may be related to an HPV infection.

C-jun oncogene expression in nonsmall cell lung-cancer.

Primary lung cancer specimens of the non-small cell (NSC) types were examined for expression of the c-jun gene at the protein level by immunocytochemistry using the polyclonal antibody c-jun 890. The results obtained revealed that the c-jun p39 protein was expressed in 82% (18/22) of squamous cell carcinomas, 77% (10/15) of adenocarcinomas and 40% (2/5) of large cell anaplastic carcinomas, but no staining was seen in the non-cancer cases. Positive immunostaining was also seen in 16.7% (3/18) of the nonneoplastic bronchial cells of positively stained squamous cell carcinomas and in 40% (4/10) of adenocarcinomas. We suggest that elevated expression df c-jun p39 protein which was seen in a total of 75% (30/40) of the NSCLC, plays a role in lung cancer.

p53 expression in cytologic specimens from benign and malignant breast lesions.

This study was undertaken to determine the expression of p53 gene in cytologic specimens from benign and malignant breast lesions. To detect p53 an immunocytochemical assay with p53 (pAb421) monoclonal antibody was used. Abnormalities in p53 expression were found in 19 out of 40 Fine Needle Aspiration (FNA) smears with infiltrating ductal breast carcinomas. Benign epithelial breast cells obtained from fibroadenomas, fibrocystic disease and smears from nipple discharge reacted negatively for p53 in 38 out of 39 cases. Moderate positive reaction, confined to a few clusters of epithelial cells, was observed in one smear of fibroadenoma with cellularity. The results recorded in this study show that no significant association was found between p53 staining and stage of disease, tumor size or nodal status and that the immunocytochemical assay represents a simple method for the detection of p53 associated proteins in breast lesions.

Immunocytochemical study of RAS oncoprotein in cytologic specimens of primary lung tumours.

We have examined the distribution of ras p21 oncoprotein expression in cytologic specimens from 73 primary bronchial carcinomas using an immunocytochemical analysis. The cytologic preparations studied represent the two major groups of histological types of lung cancer: Small Cell Lung Carcinoma (SCLC) and Non-Small Cell Lung Carcinoma (NSCLC) (squamous cell carcinoma and adenocarcinoma). The differential expression of ras p21 oncoprotein correlated with histological classification and was found in 30% of 23 small cell lesions, 61% of 28 squamous cell lung carcinomas and 32% of 22 adenocarcinomas. The ras p21 oncoprotein was commonly expressed in NSCLC cases (48%) as compared to SCLC cases (30%).