Electron microscopy renders the diagnostic capabilities of cytopathology more precise: an approach to everyday practice

Cytology is a powerful diagnostic tool but to make definitive diagnoses, the use of ancillary techniques is imperative. By combining immunohistochemistry (IHC) and electron microscopy (EM), cytologic diagnoses can be as precise as those of surgical pathology. In the authors' daily practice of cytopathology they use all ancillary techniques available to them: histochemistry, IHC, EM, flow cytometry, and molecular pathology. IHC is frequently used as an ancillary technique in their daily practice but EM is many times their technique of choice. By the use of EM the authors can make specific final diagnoses, make the diagnosis more definitive, narrow the differential diagnosis, or determine the origin of a neoplasm with unknown primary site. Specimens obtained by fine-needle aspiration as well as all body fluids are suitable for EM. The limiting factor is to obtain the appropriate material with the diagnostic cells for ultrastructural examination. The common diagnostic dilemmas in the everyday practice of cytology are the following: mesothelioma vs. adenocarcinoma, neuroendocrine differentiation or not, the distinction of melanoma from adenocarcinoma and sarcoma, hepatocellular carcinoma vs. adenocarcinoma, and the origin of adenocarcinomas of unknown primary. The authors discuss how they approach these diagnostic problems in their everyday practice and how they incorporate EM in solving them.     

Electron Microscopy in the Diagnosis of Percutaneous Fine Needle Aspiration Specimens

Five years experience in the application of electron microscopy to fine needle aspiration biopsy specimens is reviewed. In an initial evaluation, 200 consecutive unselected specimens were examined; 89 proved diagnostic and, in a third of these, electron microscopy gave additional information that was often essential to diagnosis. Negative specimens resulted almost entirely from failure to obtain an adequate amount of material. Results were improved by the adoption of a preparation technique involving concentration of the cells in bovine serum albumin and by the inspection of a rapidly stained smear at the time of the aspiration procedure, with a further needle pass for electron microscopy being performed if necessary. Despite the small size of specimens, adequate examination was usually possible and electron microscopy has proved of value in the diagnosis of tumor samples acquired by fine needle aspiration in the same way as has been established with the larger sized specimens obtained by conventional biopsy and surgical resection.     

Electron Microscopy in the Diagnosis of Percutaneous Fine Needle Aspiration Specimens

Five years experience in the application of electron microscopy to fine needle aspiration biopsy specimens is reviewed. In an initial evaluation, 200 consecutive unselected specimens were examined; 89 proved diagnostic and, in a third of these, electron microscopy gave additional information that was often essential to diagnosis. Negative specimens resulted almost entirely from failure to obtain an adequate amount of material. Results were improved by the adoption of a preparation technique involving concentration of the cells in bovine serum albumin and by the inspection of a rapidly stained smear at the time of the aspiration procedure, with a further needle pass for electron microscopy being performed if necessary. Despite the small size of specimens, adequate examination was usually possible and electron microscopy has proved of value in the diagnosis of tumor samples acquired by fine needle aspiration in the same way as has been established with the larger sized specimens obtained by conventional biopsy and surgical resection.     

Cytopathology of insular carcinoma of the thyroid.

Four pure insular carcinomas (IC) and one IC with focal anaplastic carcinoma (AC) of the thyroid with cytologic evaluation by fine-needle aspiration (FNA) were reviewed. The needle aspirates from the four pure ICs revealed abundant monomorphic follicular cells present singly, in small, loose aggregates, and in cohesive trabecular and acinar clusters. Tumor cells showed fragile, ill-defined, granular cytoplasm and oval nuclei with conspicuous or inconspicuous nucleoli. The case of IC with focal AC yielded, in addition to the follicular cells as seen in the FNA of the 4 cases of pure IC, large pleomorphic malignant cells with prominent nucleoli that were characteristic for an AC, giant-cell type. No intact insulae of tumor cells were identified in any of the 5 cases. Thus, a thyroid IC may be suspected if abundant cohesive and dyshesive monomorphic follicular cells are present in the tumor FNA. However, a firm diagnosis of thyroid IC can only be made by histologic examination of the excised tumor.     

The Utility of Electron Microscopy in Cytopathology

Background: The use of ultrastructural analysis in the diagnostic work-up of histologic specimens has been well studied but less is known about the utility of electron microscopy (EM) in cytopathology. Design: 149,006 non-gynecologic cytology cases at the Massachusetts General Hospital between the years 1993 and 2006 were searched to identify those in which material had been submitted for EM. Cytologic and EM diagnoses were correlated with available histologic diagnoses. The results were put into one of three categories: confirmatory, diagnostic, or insufficient material for diagnosis (IMFD). Results: Material for EM was obtained from 178 cytology cases that included 131 fine-needle aspirates (FNA) and 47 exfoliative specimens. EM provided additional diagnostic information beyond that offered by cytologic examination alone in 32% of cases, and in 48% of cases EM confirmed the cytologic findings. Insufficient material and discrepant results were noted for EM evaluation in 19% of cases and in 1% cases respectively. EM was most useful when applied to FNAs for subclassifying tumors as epithelial or mesenchymal (45.6%), for the diagnosis of non-neoplastic processes (15.7%) such as alveolar proteinosis and amyloidosis, and for the identification of microorganisms (12.2%). In our study, although EM was infrequently applied to exfoliative specimens to distinguish mesothelioma from adenocarcinoma, it proved to be very useful in this setting. Conclusion: When adequate material is obtained, EM can contribute significantly to the evaluation of both FNA and exfoliative cytology cases, including the diagnosis and subclassification of epithelial and mesenchymal tumors, non-neoplastic processes, and the identification of microorganisms.     

Guidelines of the Papanicolaou Society of Cytopathology for fine-needle aspiration procedure and reporting

This guideline document was developed by the Standards of Practice Task Force of the Papanicolaou Society of Cytopathology, based on extensive literature reviews and the personal practical experience of task force members. The draft guidelines were then subjected to expert review. The task force made revisions to the drafts based on the responses received from the consultant members, who are recognized experts in fine-needle aspiration biopsy. Diagn. Cytopathol. 1997;17:239–247. © 1997 Wiley-Liss, Inc.     

Cytopathology of follicular dendritic cell tumors

Follicular dendritic cell tumors are rarely described entities, arising from antigen-presenting immune accessory cells, found within B-lymphocyte follicles in nodal and extranodal sites. We report two cases, one associated with Castleman's disease, in whom fine-needle aspiration biopsy was performed followed by surgical biopsy. The diagnosis was confirmed using immunoreactivity with CD21 and CD35 antibodies and by ultrastructural demonstration of interdigitating cell processes with desmosomes. Both the cytologic and histologic findings are presented, and the differential diagnoses are discussed. Awareness of this entity and recognition of the pathologic features may lead to a presumptive diagnosis which must be confirmed using immunohistochemistry and/or electron microscopy. Diagn. Cytopathol. 17:138–142, 1997. © 1997 Wiley-Liss, Inc.     

Diagnostic significance of signet ring cells in fine-needle aspirates of the breast

Fine-needle aspiration (FNA) is a reliable and cost-effective procedure in the evaluation and management of breast lesions. One diagnostic dilemma that may sometimes arise is the finding of signet ring cells. The isolated finding of such cells in aspirate smears may be particularly problematic in cases of low cellularity or those with otherwise benign features. Although it is generally held that such cells are almost exclusively associated with carcinoma (particularly the lobular subtype), their significance in FNA smears has never been systematically evaluated. To establish their diagnostic utility, we evaluated aspirate smears from 150 cases of histologically proven benign (77) and malignant (73) breast lesions for the presence of signet ring cells, defined as those containing a prominent intracytoplasmic vacuole with nuclear displacement. Signet ring cells were identified in 71% of malignant cases (75% of ductal carcinomas and 71% of lobular carcinomas), mostly as single cells or within small, loosely cohesive tissue fragments. Such cells were also present in 6% of histologically proven benign lesions, most commonly within large tissue fragments. Many of these cells were proven to be vacuolated myoepithelial cells, based on histologic correlation and immunostaining results using anti-muscle-specific actin. On the basis of these findings, we conclude that (1) the presence of signet ring cells within small loose tissue fragments or as single cells in FNA smears should prompt close clinical follow-up (including repeat FNA and perhaps surgical biopsy), regardless of smear cellularity, (2) the presence of signet ring cells in cases of adenocarcinoma does not predict a particular tumor subtype, and (3) rare benign breast lesions may contain signet ring cells, particularly within large tissue fragments, and do not, in isolation, warrant surgical biopsy to exclude malignancy. Diagn. Cytopathol. 16:117–121, 1997. © 1997 Wiley-Liss, Inc.     

Fine-needle aspiration cytology of chondromyxoid fibroma: a case report

The cytologic presentation of a case of chondromyxoid fibroma studied by fine-needle aspiration in a 17-yr-old black male is described. The cytologic features of chondromyxoid fibroma are presented, and the differential diagnosis of intraosseous cartilaginous neoplasms is reviewed, emphasizing the cytomorphologic differentiation of chondromyxoid fibroma from chondrosarcoma, enchondroma, and chondroblastoma.     

Fine-needle aspiration cytology of pleuropulmonary blastoma: Case report and review of the literature

This report describes the fine-needle aspiration (FNA) cytology of a case of pleuropulmonary blastoma in a 3-yr-9-mo-old male. Pleuropulmonary blastoma is considered by most authors to be distinct from pulmonary blastoma and is a rare malignant tumor of the intrathoracic cavity. FNA smears were cellular with numerous small ovoid to spindled cells with oval to elliptical nuclei exhibiting finely granular chromatin and inconspicuous nucleoli. The cytoplasm was scant and cosinophilic with indistinct borders. Focal chondroid material and blastema-like cells were noted. The differential diagnosis suggested by the cytologic findings included rhabdomysosarcoma, teratoma, neuroblastoma, malignant mesenchymoma, pleuropulmonary blastoma, and metastatic tumor. To our knowledge, this is the first report of the cytology of this tumor. Diagn. Cytopathol. 16:336–340, 1997. © 1997 Wiley-Liss, Inc.     

The use of electron microscopy to refine diagnoses in the daily practice of cytopathology

Conceived as a screening tool, cytology is a field that since the 1980s has become more diagnostic in its scope. The advent of the fine-needle aspiration biopsy (FNAB) is responsible for cytology's new place in pathology. In the everyday practice of cytopathology, about 85-90% of the nongynecologic cases can be diagnosed with the use of routine stains (i.e., Papanicolaou and Diff Quik). The other 10-15% of the cases require the use of ancillary diagnostic techniques for a precise diagnosis. Immunohistochemistry helps solve approximately 50% of these cases, and the other half of these challenging cases are best approached and diagnosed by using electron microscopy (EM). In their practice, the authors obtain cytologic samples for EM routinely in difficult cases. Unfortunately, a percentage of these cases collected for ultrastructural evaluation do not have enough cells after processing, and others only have a few diagnostic cells available. In the cases in which at least a handful of cells are available, EM is almost invariably helpful in one way or another, either making a definitive diagnosis or refining the diagnosis. A sampling of FNAB cases from the authors' everyday practice is prevented to illustrate the use of EM in the practice of cytopathology. The cases have been selected from among the most common diagnostic challenges to highlight the important role that ultrastructural evaluation plays in a busy cytology practice. In our practice ultrastructural evaluation is a piece of the puzzle, which, along with the clinical history, clinical impression, light microscopic/cytologic features, and other ancillary techniques (IHC, flow cytometry, and molecular pathology), help compile an accurate diagnosis. Many times EM is the most important component of the diagnostic algorithm.     

New Techniques: The use of Fresh-Frozen Tissue in Diagnostic Transmission Electron Microscopy

We have examined the effects of freezing on fresh pathologic human tissue specimens with regard to the fine structural preservation of various organelles that can be of diagnostic importance. Specimens included in this study were frozen either in the cryostat (-20°C) or by quenching in isopentane cooled by liquid nitrogen (-115°C). The results indicate that fresh-frozen tissue that is subsequently fixed and processed for transmission electron microscopy (TEM) provides ultrastructural information that is very nearly equal to that of conventionally processed specimens. Additionally, we performed identical freezing experiments using animal (rat) tissue in order to ensure that the results we obtained using a rather limited number of human specimens were valid. The outcome of these animal experiments thoroughly supports our observations using human pathologic tissue. We believe an appreciation of the fact that fresh-frozen tissue is suitable for TEM examination may become a very valuable resource in the surgical pathology laboratory.     

Fine-needle aspiration cytology of mammary fibromatosis: Report of two cases

The purpose of this study is to evaluate cytologically two cases of mammary fibromatosis (MF). Prior to FNAC, clinical and mammographic suspicion of carcinoma and fibroadenoma were present. In both cases cytology disclosed the presence of numerous spindle cells admixed with epithelial cells. In the first case, carcinoma was excluded and the diagnosis of “spindle-cell proliferative lesion” was established. The second case was erroneously diagnosed as “cellular fibroadenoma” due to the presence of monolayered ductal epithelial groups and stromal tissue. In both cases local excision of the lesion was recommended. Although in a strict sense fibromatosis is a pure stromal lesion, the frequent presence of epithelial groups in the smears should raise a differential diagnosis with other more frequent mixed (epithelial and stromal) lesions such as fibroadenoma, cystosarcoma phyllodes, and metaplastic carcinoma. Due to the fact that clinically and mamographically MF is frequently confused with malignancy, preoperative recognition is essential since in many cases it would avoid unnecessary radical surgery. In this sense cytology offers very important preoperative information. Diagn. Cytopathol. 1997;17:363–368. © 1997 Wiley-Liss, Inc.     

Fine-needle aspiration cytology of renal angiomyolipoma: Report of a case with immunocytochemical and electron microscopic findings

A case is presented in which the diagnosis of renal angiomyolipoma was made by computed tomography (CT)-guided, fine-needle aspiration cytology and the examination of a cell block. The tumor was characterized by epithelioid smooth muscle cells, blood vessels and fat in the cytologic material. The cytodiagnosis was further substantiated by positive staining of the epithelioid muscle cells for markers of muscle-specific actin, HMB45, and trace positivity for vimentin but not with S100 protein, desmin, or cytokeratin. Further studies for the recently described crystalloids were also performed by using the periodic-acid Schiff stain with and without diastase treatment and electron microscopy on the cytologic material. However, no such crystalloids were found. A preoperative cytologic diagnosis of renal angiomyolipoma was considered to be of value due to the difference in management between angiomyolipoma and a renal carcinoma, both of which can present as a renal mass on imaging. Diagn. Cytopathol. 1998;18:297–300. © 1998 Wiley-Liss, Inc.     

Poorly-differentiated adenoid cystic carcinoma: Cytologic appearance in fine-needle aspirates of distant metastases

Adenoid cystic carcinoma (ACC) is a primary salivary-gland neoplasm which typically yields characteristic cytomorphology upon fine-needle aspiration (FNA). We report on the FNA findings of a case of ACC metastatic to the liver which demonstrated a predominantly solid, poorly-differentiated pattern, an unusual but well-recognized subtype associated with a poor clinical outcome. The FNA findings in 7 additional cases of ACC metastatic to distant sites were also reviewed, with 4 cases displaying a prominent poorly-differentiated component. These findings suggest that, although not commonly recognized in salivary-gland FNAs, the poorly-differentiated pattern of ACC does occur in metastatic deposits and should be recognized as such, thereby preventing a needless search for a second primary malignancy. Diagn Cytopathol 1996;15: 296–300. © 1996 Wiley-Liss, Inc.     

Fine-needle aspiration of basaloid squamous carcinoma: A case report with review of differential diagnostic considerations

Basaloid squamous carcinoma is a distinct variant of squamous carcinoma with a particularly poor prognosis. To our knowledge, there are only two papers in the cytopathology literature which describe this entity. We report the fine-needle aspiration findings of an additional case of metastatic basaloid squamous carcinoma in a cervical lymph node and compare its cytomorphologic features to those observed on touch imprints of the subsequent surgical specimen. Smears of the aspirate showed a mixed lymphoid background with interspersed cohesive clusters of small cells roughly 3 times the size of small mature lymphocytes. Some cells were angulated and others exhibited irregular nuclear contours. The cells were generally hyperchromatic with evenly staining dense chromatin or irregularly distributed coarse chromatin. Focally there was evidence of nuclear molding. On Diff-Quik staining, irregular globules of magenta-stained extracellular dense material were noted within or adherent to the periphery of some clusters or as somewhat linear formations with small epithelial cells clinging to the edges. Abundant mitotic figures and clumps of necrotic tumor were more apparent on touch preps of the subsequent surgical specimen. The differential diagnosis by fine-needle aspiration includes adenoid cystic carcinoma, basal-cell adenocarcinoma, adenosquamous carcinoma, and small-cell carcinoma. If a fine-needle aspirate of a cervical lymph node shows the features described above and the primary tumor is unknown, suggesting the possibility of metastatic basaloid squamous carcinoma may aid clinicians in the search for a primary site, as basaloid squamous carcinoma occurs most frequently at the base of the tongue, hypopharynx, and supraglottic larynx. Diagn. Cytopathol. 1998;19:210–215. Published 1998 Wiley-Liss, Inc.

1 This article is a US Government work and, as such, is in the public domain in the United States of America.

     

Extracranial metastasis of glioblastoma multiforme diagnosed by fine-needle aspiration: A report of two cases and a review of the literature

Extracranial metastasis of cerebral glioblastoma is rarely seen. Craniotomy and diversionary shunt are widely accepted causes of dissemination. Prognosis is poor but new therapeutic modalities may improve the survival and lessen the patient's symptoms. It is also important to diagnose extracranial metastasis because of possible response to treatment and fine-needle aspiration cytology can then be helpful. Two cases of extracranial metastases of glioblastoma multiforme diagnosed by fine-needle aspiration cytology are reported and a review of the literature is presented. Diagn Cytopathol 1996;15:60–65. © 1996 Wiley-Liss, Inc.     

Fine-needle aspiration cytology of soft-tissue calcinosis presenting as an enlarging mass in the chest wall

Calcinosis cutis is an uncommon condition, and calcific deposits in patients with end-stage renal disease are now somewhat rare, due to improvement in management. Since calcific deposits may clinically resemble a tumor, it is feasible to investigate them by fine-needle aspiration cytology (FNAC). A 52-yr-old male with a history of end-stage renal failure presented with a 2-cm mass in the left chest wall. Within 6 mo the mass enlarged to nearly four times its original size. In view of clinical suspicion of malignancy, FNAC was performed. Cytopreparations showed abundant calcium, indicative of soft-tissue calcinosis. The case is interesting, since the known history of end-stage renal failure enabled the FNAC diagnosis of soft-tissue calcinosis. Diagn. Cytopathol. 1998;19:465–467. © 1998 Wiley-Liss, Inc.