Ultrastructure of pleural mesothelioma and pulmonary adenocarcinoma in malignant effusions as compared with reactive mesothelial cells.

OBJECTIVE: To determine the ultrastructural features of diffuse malignant pleural mesothelioma cells in cytologic specimens from pleural effusions. STUDY DESIGN: 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. RESULTS: 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. CONCLUSION: Because cytologic specimens from pleural effusions were easy to obtain, we think ultrastructural cytology is useful in distinguishing mesothelioma from adenocarcinoma and benign effusions.     

Cytologic presentation of malignant mesothelioma in pleural effusions

1,601 pleural effusions were found to be malignant between 1976 and 1987. Among these were 26 (1.6% of the malignant effusions) mesothelioma. Only 2 cases showed pronounced cytologic features that made a definite diagnosis possible on cytologic criteria alone. In 20 cases diagnosis of mesothelioma was strongly suggested by the patient's history and cytology of the effusion was compatible with mesothelioma. In the other 4 cases special examinations (histo- and immunohistochemistry, electron microscopy) led to the final diagnosis. The cytologic features of mesothelioma and other examination techniques, needed to resolve the differential diagnosis of mesothelioma versus other neoplasm in pleural effusions, are discussed.     

Positive effusion cytology as the initial presentation of malignancy

During a period of four years (1981 to 1984), 641 ascitic, 860 pleural and 47 pericardial fluid specimens were examined cytologically. Of these, 154 ascitic samples, 174 pleural specimens and 10 pericardial effusions, obtained, respectively, from 108, 133 and 7 patients, were found to contain malignant cells. In 7 patients, ascites, and in 18 cases, pleural effusions were the first indication of cancer. None of the positive pericardial fluids was the initial presentation of malignancy. The cytologic findings and follow-up data on these 25 patients are the subject of this study. The most common type of neoplasm in these effusions was adenocarcinoma (86% of the ascitic and 78% of the pleural fluids). Most of the malignant neoplasms in ascitic fluids were derived from ovarian tumors (5 of 7) while those in pleural effusions came mainly from lung tumors (12 of 18). Mammary carcinoma, which was the most common malignant tumor found in cases of pleural effusions, did not present initially with an effusion in any of our cases. The cytologic diagnosis was confirmed in all cases by either biopsy or strong clinical evidence. The prognosis in patients who initially presented with an effusion was poor. All of the patients with an adequate follow-up died within 29 months in cases of ascites and within 19 months in cases of pleural effusions.     

Fluorescence microscopy of mycobacteria in pleural effusions

The diagnostic advantage of fluorescence microscopy (FM) of Papanicolaou-stained cytological specimens obtained by bronchoscopy has been described previously. This study was designed to evaluate the method's diagnostic benefit in cytological preparations of pleural effusions in cases of active pulmonary tuberculosis. In contrast to bronchial material there is no advantage in cytological evaluation of pleural effusions by FM.     

DNA flow cytometry of pleural effusions. Comparison with pathology for the diagnosis of malignancy

A study was undertaken to determine the potential value of DNA flow cytometry (FCM) for the diagnosis of malignancy in pleural effusions and to compare its results with those of traditional cytomorphology. Forty-one pleural fluids from 37 patients were evaluated by DNA FCM and routine cytologic techniques. Of the 41 pleural fluids, 29 (70.7%) demonstrated an abnormal DNA content by FCM. Two of the 29 pleural fluids had been originally diagnosed as benign by cytology. Cytologic review of these two cases showed no abnormal cells; therefore, there were no false-negative cases based on cytology. In 12 (29.3%) of the 41 fluids, DNA FCM and cytologic evaluation both indicated benign processes. Our preliminary observations indicate that FCM is an accurate and reliable technique that may be of aid in the diagnosis of malignant effusions. The technique may prove to be of special value in the differential diagnosis of reactive mesothelial cells versus malignant mesothelioma as well as for following patients who receive chemotherapy for malignant pleural effusions. DNA FCM may also complement cytomorphologic diagnoses in other serous, exfoliative or aspiration material.     

The distinction of mesothelioma from adenocarcinoma in malignant effusions by electron microscopy

To determine the usefulness of the electron microscopic (EM) differential diagnosis between malignant mesothelioma and metastatic adenocarcinoma in cytologic specimens of serous fluids, we undertook a prospective study of 17 pleural and peritoneal effusions from 14 patients. In the nine effusions identified as malignant by routine cytologic examination, EM correctly diagnosed three mesotheliomas and six adenocarcinomas. EM resolved the differential diagnosis of mesothelioma versus adenocarcinoma in three cases in which routine cytologic examination could not. As with tissue specimens, EM cannot be used to diagnose the malignancy of cytologic specimens; it can, however, reliably identify the origin of cells diagnosed as malignant by routine cytologic examination. We conclude that, when EM is used to evaluate cytologically malignant effusions, it can accurately distinguish mesothelioma from adenocarcinoma. This technique will be diagnostically useful in selected cases and may be helpful in avoiding more invasive procedures as well as delays in diagnosis and therapy.     

Oncofetal antigens as tumor markers in the cytologic diagnosis of effusions

To test the value of oncofetal antigens in the cytologic diagnosis of effusions, immunoperoxidase staining with antisera to carcinoembryonic antigen (CEA), alpha fetoprotein (AFP), pregnancy-specific beta 1-glycoprotein (SP1) and placental alkaline phosphatase (PLAP) was carried out on pleural and peritoneal fluids from 72 cases. Sections of formalin-fixed, paraffin-embedded cell blocks were used in most cases; cytocentrifuge preparations were used in some. Reactions were negative with all antisera in 23 of 24 nonmalignant effusions as well as in all 7 cases of malignant mesothelioma and 4 cases of malignant lymphoma. In 24 of 36 confirmed carcinomatous effusions, staining was positive with one or more antisera, including anti-CEA positivity in 23 of the 24 cases. In 5 of the 24 cases with positive staining, a confident diagnosis of malignancy had not been made on routine cytologic preparations. Immunoperoxidase staining for CEA appears to be of supportive value in the cytologic diagnosis of malignancy in effusions.     

Survival of patients with malignancy-associated effusions

For a better understanding of the prognosis after the onset of a malignancy-associated effusion in patients known or subsequently shown to have cancer, survival time was compared with the findings and the date of the first cytologic diagnosis of an effusion. The number of patients studied was 254; 171 had a pleural and 83 a peritoneal effusion. The average survival time was 25.5 weeks, which was about equal for both sites of effusions. After two years, only 6% of all patients were alive. When the cytologic diagnosis of the effusion was "malignant," only 4% survived after two years; when the cytologic diagnosis was "suspicious for malignancy" and "nonmalignant," these figures were 5% and 7%, respectively. This indicates that a cytologic diagnosis of benign or nonmalignant is not a good indicator of a better prognosis in cancer patients for whom benign causes of the effusion have been excluded. There appeared to be a prognostic relationship between the length of the interval from the initial diagnosis of cancer to the time of examination of the first sample of the effusion: a longer interval was correlated with a better survival. When survival time was viewed in relation to therapy, patients whose pleural effusions were only treated by aspiration were found to have a particularly short average survival (13.9 weeks).     

Macrophage size determinations in the diagnosis of tuberculous effusions

This study investigated the usefulness of macrophage size determinations in lymphocyte-rich pleural effusions to improve the cytologic diagnosis of tuberculous pleurisy. The size of pleural macrophages was analyzed by quantitative morphometric planimetry in 18 effusions due to tuberculosis, 21 effusions following radiotherapy for malignant disease and 10 effusions due to congestive heart failure. Macrophages were identified and clearly separated from mesothelial cells by latex phagocytosis and immunostaining with the monoclonal antibody My4 (CD14). The mean macrophage area (+/- standard deviation) in tuberculous effusions (92 +/- 14 sq micron) was significantly smaller than in postradiation (141 +/- 28 sq micron) and heart-failure effusions (154 +/- 22 sq micron) (P less than .0001). There was also a smaller ratio of mesothelial cells in tuberculous effusions (0.5 +/- 0.9%) in comparison with effusions following radiotherapy (4 +/- 5%) or congestive heart failure (10 +/- 12%). In summary, this study demonstrated some cytomorphologic parameters that may be helpful in the differential diagnosis of tuberculous effusions.     

Cytologic finding of chyloascites in lymphangioleiomyomatosis. A case report

BACKGROUND: Lymphangioleiomyomatosis is a rare disease, histologically characterized by an abnormal proliferation of smooth muscle around the lymphatics. Lung is the most common site of involvement, and patients usually present with dyspnea, chest pain, and cough. Chylous pleural effusion and ascites occasionally appear during the course of the disease. There are only a few reports on the cytologic findings in this disease. To our knowledge, the cytologic findings of chylous pleural effusion and chyloascites have not been reported before. CASE: A 23-year-old female presented with chylothorax, chyloascites and a retroperitoneal mass. Cytologic examination of chylous pleural effusion and chyloascites revealed numerous cohesive and thick clusters of cells with a high nuclear/cytoplasmic ratio, oval nuclei and slightly increased chromatin content. Mitosis and necrosis were not observed. Exploratory laparotomy and transbronchial lung biopsy were performed, and the histologic diagnosis was lymphangioleiomyomatosis involving the retroperitoneal lymph nodes, uterine fundus and lungs. Immunohistochemistry showed that the characteristic clusters in chylous fluids were positive for alpha-smooth muscle actin. CONCLUSION: A diagnosis of lymphangioleiomyomatosis is possible from cytologic findings of effusions with the aid of clinical findings.     

Diagnostic accuracy of toluidine blue-stained wet films in effusion cytology

OBJECTIVE: To evaluate the usefulness of toluidine blue-stained wet films in the preliminary cytologic evaluation of serous effusions by means of specificity, sensitivity, and positive and negative predictive value. STUDY DESIGN: One hundred seventy-six samples consisting of 122 pleural and pericardial effusions and 54 peritoneal effusions from 160 patients were included in the study. A toluidine blue-stained wet film of each sample was evaluated, and diagnoses were compared with the diagnoses by conventional smears and cell blocks on the same sample. RESULTS: The sensitivity of wet films was 69%, 68% in pleural and pericardial effusions and peritoneal effusions, respectively, and the specificity of wet films was 93%, 92% in pleural and pericardial effusions and peritoneal effusions, respectively. Positive predictive values of smears alone were 78% and 75%; negative predictive values of smears alone were 96% and 95% in pleural and pericardial effusions and peritoneal effusions, respectively. CONCLUSION: Preliminary cytologic evaluation of serous effusions with toluidine blue-stained wet films is simple and economical. It provides the opportunity to plan additional procedures for the samples.     

Mycobacterium kansasii infection diagnosed by pleural fluid cytology: a case report

BACKGROUND: Identification of disseminated nontuberculous Mycobacterium infection is a challenge, especially when it occurs in patients without a known cause of immunosuppression. Acid-fast organisms in the pleural fluid are rare and easily missed, especially when they occur in patients without a clinical suspicion of infection. The classical cytologic picture of tuberculous pleural fluid with lymphocytosis and paucity of mesothelial cells is not seen. CASE: A 57-year-old man presented with chronic neutrophilia of unknown etiology together with chest pain and bilateral pleural effusions. Pleural fluid cytology revealed organisms seen in the cytoplasm of numerous macrophages and neutrophils, creating a "negative image" on Diff-Quik smears. Acid-fast stains demonstrated intracellular acid-fast bacilli consistent with mycobacteria. Microbiologic studies with DNA probe technology resulted in identification of the mycobacterial organism as Mycobacterium kansasii. CONCLUSION: Nontuberculous Mycobacterium should be included in the differential diagnosis in patients with inflammatory, exudative pleural effusions.     

Fine needle aspiration cytology of malignant mesothelioma

The results of fine needle aspiration (FNA) cytology in 19 cases of malignant mesothelioma are presented. Adequate material for a diagnosis of malignancy was obtained in 17 cases, and in 8 cases a specific diagnosis of mesothelioma could be made. In four other cases, the findings were either consistent with or suggestive of mesothelioma; in four, accurate distinction from other neoplasms was not possible, and in two cases, adenocarcinoma was suggested. The spectrum of cytologic findings ranged from neoplasms of purely epithelial appearance through more pleomorphic biphasic neoplasms to anaplastic tumors. A combination of epithelial-like cell clusters, pavement-like sheets of epithelial cells with well-defined cell borders and prominent cell separation, dispersed angular cells with dense cytoplasm and some spindle-cell forms was the most specific cytologic pattern for mesothelioma. In four neoplasms, ultrastructural examination of aspirated material provided the additional evidence for a definitive diagnosis. The identification of hyaluronic acid within intracytoplasmic vacuoles, either in smears or in cell blocks, confirmed the diagnosis in three tumors. Only in one case, with a strong clinical background suggesting mesothelioma, was the cytologic preparation sufficient for diagnosis without ancillary diagnostic methods. FNA is of particular value in the diagnosis of pleural mesothelioma in patients who do not present with a pleural effusion. Obtaining material for cell block preparations, cytochemistry or ultrastructural study is generally necessary for definitive tumor typing.     

Potential use of loss of heterozygosity in pleural effusions of breast cancer metastases using the microsatellite marker of the 16q22.1 region of the CDH1 gene

OBJECTIVE: To evaluate the presence of allelic loss in 16q22.1, including the locus of E-cadherin, in pleural effusions in breast cancer patients. STUDY DESIGN: Molecular analysis of DNA was performed using a DNA extraction kit (NucleoSpin, Macherey-Nagel, Germany). Loss of heterozygosity (LOH) in primary tumors and pleural effusions was analyzed using a microsatellite marker of the CDH1 gene, D16S265, described in previous studies. LOH was evaluated by radioactive polymerase chain reaction assay in 17 samples of pleural effusions and breast tissues (primary tumors and nonneoplastic adjacent tissue) from breast cancer patients: 7 positive for neoplastic cells, 6 suspected and 4 cases without evidence of neoplastic cells in the effusions. RESULTS: Thirteen cases (76%) were informative. LOH was detected in 5 cases (38.5%). In 3 of them LOH was detected only in the cytologic sample, and in 2 of them LOH was detected in the primary tumor and cytologic sample. CONCLUSION: Results show that LOH in the CDH1 gene can identify tumor cells in pleural effusions when morphologic analysis is difficult.     

Bronchial cytology in pleural mesothelioma. A report of 3 positive cases, including 1 diagnosed initially on bronchial brushings

OBJECTIVE: To evaluate retrospectively the value of bronchial aspiration cytology in patients with histologically proven pleural mesothelioma, reappraising positive smears in light of conventional microscopic features and, when feasible, immunocytochemical investigations. STUDY DESIGN: In 3 cases of mesothelioma with bronchial brushings positive for malignant cells, the cytologic features were correlated with the histologic findings. RESULTS: Salient microscopic features included scant to moderate cellularity arranged in micropapillary clusters, morular aggregates with scalloped borders and isolated malignant cells. Intercellular clear spaces or windows suggesting a brush border on cell membranes were also noted. In cases with available material, immunocytochemistry was positive for keratins, epithelial membrane antigen and calretinin and negative for carcinoembryonic antigen. All the cases were histologically confirmed epithelial mesotheliomas. CONCLUSION: In rare cases, pleural mesothelioma cells are shed within the airway lumina and can be detected in bronchoscopic cytology specimens. Cytologic features seem comparable to their analogues in pleural effusions. Although no single criterion appears diagnostic, their combined documentation could ensure correct interpretation, especially if supported by a limited immunocytochemical panel.     

Microfilariae of Wuchereria bancrofti in cytologic smears

Microfilariae of Wuchereria bancrofti were observed in cytologic material in 35 cases. The material included cervicovaginal smears (17 cases), effusions (14), urine (2), bronchial washings (1) and ovarian cyst fluid (1). The initial diagnosis was made from the cytologic smear in all cases; none had clinical filariasis. Symptomatic vaginal bleeding in 9 of the 17 cases with microfilaria-positive cervicovaginal smears was reflected in the large numbers of red blood cells found in the smear. Blood eosinophilia was present in 11 of 19 cases investigated. Eosinophils were seen in the smears in 20 cases. In the majority of the cases of effusions with microfilariae the effusions were malignant. Significant adherence of inflammatory cells and macrophages to microfilariae was present in 7 of the 35 cases. The significance of these findings is discussed.     

Immunohistochemical study of lysozyme, alpha 1-anti-chymotrypsin, tissue polypeptide antigen, keratin and carcinoembryonic antigen in effusion sediments

The cellular sediments of 42 malignant and 16 benign effusions (58 cases) were studied using the immunoperoxidase technique. Serial sections of formalin-fixed, paraffin-embedded residual sediments of effusions, sent for routine cytologic examination, were studied by commercially available polyclonal antisera against lysozyme, alpha 1-anti-trypsin, alpha 1-anti-chymotrypsin, tissue polypeptide antigen (TPA), a wide-spectrum anti-keratin, carcinoembryonic antigen (CEA) and, in single cases, thyroglobulin and prostate-specific antigen. A final definite diagnosis from histologic study of biopsy or autopsy specimens was known in all cases. All carcinomas, the mesotheliomas and the reactive mesothelial cells showed a positive reaction for TPA and, partly, the wide-spectrum keratin. Lysozyme could be demonstrated in the cells of the one proven malignant fibrous histiocytoma; all malignant epithelial cells were negative. Alpha 1-anti-chymotrypsin and alpha 1-anti-trypsin showed similar reactions: they were often positive in carcinoma cells of the breast, the bronchial system and the pancreas, in contrast to a mostly negative reaction in carcinomas of the stomach and ovary. CEA showed considerable differences; it was always negative in benign and malignant mesothelial proliferations but mostly positive in carcinomas of the stomach, pancreas and bronchial system. It was only positive in less than 20% of the carcinomas of the breast and always negative in the proven malignant effusions of primary carcinomas of the ovary and prostate. Studying a combination of several tumor markers is possible in serial paraffin-embedded sections and may be a valuable criterion in the cytologic diagnosis of effusions.     

The cytologic diagnosis of malignant neoplasms in pleural and peritoneal effusions

This study presents data on 3,011 pleural and peritoneal effusion specimens that were examined over a three-year period (1982 to 1984). Totals of 812 (44%) of 1,846 pleural and 423 (36%) of 1,165 peritoneal specimens were positive for malignant cells. While 535 patients had malignant pleural effusions, 254 patients had malignant peritoneal effusions, and 57 had both malignant pleural and peritoneal effusions. The most common primary neoplasms causing malignant pleural effusions were carcinomas of breast (24%) and lung (19%) and lymphoreticular neoplasms (16%). The most common primary neoplasms causing malignant peritoneal effusions were carcinomas of ovary (32%) and breast (13%) and lymphoreticular neoplasms (7%). There was an average interval of more than 30 months between the histologic diagnosis of the primary neoplasm and the diagnosis of malignant effusions in patients with carcinoma of breast, lymphoreticular neoplasm and malignant melanoma. The average time until death following the diagnosis of a malignant effusions was five months or less, except for patients with carcinoma of the breast and carcinoma of the ovary. One hundred twenty-five patients (15%) presented with malignant effusions caused by neoplasms of unknown primary sites. The most common primary neoplasms that were later diagnosed were, in decreasing order of frequency, carcinoma of the ovary, carcinoma of the lung and lymphoreticular neoplasms.     

Differential proteome profiling of pleural effusions from lung cancer and benign inflammatory disease patients

The pleural effusion proteome has been found containing information that directly reflects pathophysiological status and represents a potential diagnostic value for pulmonary diseases. However, the variability in protein composition between malignant and benign effusions is not well understood. Herein, we investigated the changes of proteins in pleural effusions from lung adenocarcinoma and benign inflammatory disease (pneumonia and tuberculosis) patients by two-dimensional difference gel electrophoresis (2D-DIGE). Twenty-eight protein spots displayed significantly different expression levels were positively identified by MALDI-TOF-MS representing 16 unique proteins. Five identified protein candidates were further validated and analyzed in effusions, sera or tissues. Among them, hemopexin, fibrinogen gamma and transthyretin (TTR) were up-regulated in cancer samples. The effusion concentration of serum amyloid P component (SAP) was significantly lower in lung cancer patients than in benign inflammatory patients, but no differences were found in sera samples. Moreover, a Jumonji C (JmjC)-domain-containing protein, JMJD5, was observed to be down-regulated in malignant effusions, lung cancer tissues and cancer cells. These results shed light on the altered pleural effusion proteins as a useful and important complement to plasma or other routine clinical tests for pulmonary disease diagnosis.     

Relation between cell composition of pleural effusions in patients with pulmonary carcinomas and their clinical courses.

1) Cancer cells of the effusions were decreased by chemotherapy, but this did not always indicate a good clinical course. 2) When the clinical condition was fairly good, the percentage of lymphocytes predominated in the pleural effusion. 3) When the clinical conditions became rather critical, there was an increase in the number of macrophage. 4) The cytologic findings were observed prior to the time the clinical symptoms became manifest. 5) The percentage of lymphocytes and macrophages in pleural effusions was not related to the percentage of lymphocytes and monocytes in peripheral blood.