Adjusting interphase FISH results in epithelial tissue sections to whole cell complement

OBJECTIVE: To derive an equation to compensate for the discrepancies between whole cell preparations and tissue sections for more accurate enumeration of fluorescence in situ hybridization (FISH) signals per cell. STUDY DESIGN: Mean centromere signal counts in touch preparations and corresponding 4-6-micron sections of paraffin-embedded tissue were calculated. Mean widest nuclear diameters were also determined from the tissue sections. The observed data were analyzed to define the volumetric relationships between tissue sections and whole cell preparations. RESULTS: Analysis of results from six lung specimens yielded an equation that approximates whole versus sectioned nuclear volume and permits accurate quantification of mean FISH signal count in histologic sections, as follows: [formula: see text].     

Immunohistochemical and immunocytochemical findings associated with Marek’s disease virus in naturally infected laying hens

We compared immunohistochemical (IHC) staining of tissue sections of liver, kidney, spleen, lung, proventriculus, sciatic nerve, bursa of Fabricius, brain, heart, intestine and skin; immunocytochemical (ICC) staining of peripheral blood samples and touch preparations of liver, spleen and kidney of laying hens naturally infected with Marek’s disease (MD) virus. We used one hundred and fifty 5-17-week-old commercial hens. IHC and ICC staining were performed using polymer-based techniques. IHC staining exhibited mostly free immunopositive reactions in tumor cells and in the cytoplasm of the parenchymal cells of liver, kidney, spleen and bursa of Fabricius. In the sciatic nerve, severe reactions were observed in the cytoplasm of plasma and MD cells in the lymphoproliferative areas. Pronounced staining was found in the lymphoid cells in the medulla of intrafollicular regions in the bursa of Fabricius. Although immunostaining was observed in the liver and spleen touch preparations, there was no staining in the kidneys and peripheral blood cell samples. The presence of virus in the tissue and peripheral blood samples and in touch preparations was compared immunohistochemically and immunocytochemically. IHC and ICC techniques were helpful for diagnosis of MD. Peripheral blood samples are inappropriate for field conditions and natural infections.     

Cytodiagnosis of Campylobacter pylori in Papanicolaou-stained imprints of gastric biopsy specimens.

The use of Papanicolaou-stained touch preparations of gastric antral biopsies for the identification of Campylobacter pylori was examined using specimens obtained from 63 consecutive patients with endoscopic evidence of antral gastritis, with the results compared to routine histologic examination and Warthin-Starry silver staining. Organisms were readily identifiable in the Papanicolaou-stained imprints of the gastric mucus. The sensitivity in detecting organisms was 92.5% for the Warthin-Starry-stained sections, 71.4% for the Papanicolaou-stained imprints and 100% for both techniques combined. False-negative imprints were attributed to poor smears and/or the submission of duodenal tissue rather than antral biopsies. Properly performed touch preparations stained by the Papanicolaou method are a cost-effective adjunct to Warthin-Starry-stained section for improving the sensitivity of gastric biopsies for the diagnosis of C pylori.     

Pathogenesis of Sarcocystis falcatula (Apicomplexa: Sarcocystidae) in the budgerigar (Melopsittacus undulatus). III. Pathologic and quantitative parasitologic analysis of extrapulmonary disease

Forty-four budgerigars (Melopsittacus undulatus) were administered sporocysts of Sarcocystis falcatula orally and were examined at necropsy intervals from less than 12 hr to 168 days. Tissue were examined by touch preparations (of organ cut surfaces), light microscopy, and electron microscopy. Meront and cyst burdens were determined in various organs and correlated with duration of infection, inoculum, and the meront or cyst burdens of other organs. Host inflammatory tissue reactions were quantitated and correlated with meront and cyst burdens. Quantitation of meronts was more accurate in tissue sections than in touch preparations, but quantitation of merozoites was better in touch preparations. More than 97% of meronts were found in capillary, venular, and venous endothelial cells. Cysts were found only in cardiac and skeletal myocytes. Merogony began in the lamina propria of the small intestine less than 12 hr postinoculation (PI). Meronts were in liver and lung by the second day PI and in other organs by 3-7 days PI. Mean meront burdens were highest in lung (33 meronts/mm2), lower in liver and kidney (1-3 meronts/mm2), and infrequent in other organs (less than 0.9 meronts/mm2). Cysts were first seen in cardiac myocytes 7 days PI. They developed through the metrocyte stage and then degenerated, rarely reaching maturity. Cysts were first noted in skeletal muscle at 8 days PI. In leg, upper esophagus, and tongue, cysts matured between 44 and 77 days PI. In pectoral muscles, the majority of cysts degenerated during the late metrocyte and early intermediate stages (28-42 days PI). In addition to a previously reported and often fatal acute interstitial pneumonitis, S. falcatula-infected budgerigars also sustained a chronic active hepatitis, interstitial myocarditis, myositis, nephritis, splenitis, and encephalitis. These lesions weakly correlated with meront burdens in most sites during early infection (up to 50 days PI).     

A method of assessing the adequacy of trucut biopsy specimens obtained with a 19-gauge trucut core biopsy needle

OBJECTIVE: To develop a simple and accurate method of evaluating trucut biopsy (TCB) specimen. STUDY DESIGN: We performed 40 ex vivo TCBs of surgical specimens under direct visual guidance with the Quick-Core needle (Wilson-Cook, Winston-Salem, North Carolina, U.S.A). Touch preparations were made using 4 methods. Slides were stained with a modified Wright-Giemsa stain, and biopsies were processed for histology. Slides were blindly classified as malignant, suspicious, atypical, benign or inadequate. The ease of the 4 touch preparation methods was compared. RESULTS: There were 20 TCBs of benign tissue and 20 of malignant tissue. Cytologic interpretation could be made in 34 of 40 cases and was correct in 25 of 34 cases. Touching the biopsy to a glass slide was significantly easier than the 3 other methods tested. CONCLUSION: Touch preparations can be made easily with TCB samples and can provide information about specimen adequacy and diagnosis.     

Supraspinal input is dispensable to generate glycine‐mediated locomotive behaviors in the zebrafish embryo

The anatomy of the developing zebrafish spinal cord is relatively simple but, despite this simplicity, it generates a sequence of three patterns of locomotive behaviors. The first behavior exhibited is spontaneous movement, then touch‐evoked coiling, and finally swimming. Previous studies in zebrafish have suggested that spontaneous movements occur independent of supraspinal input and do not require chemical neurotransmission, while touch‐evoked coiling and swimming depend on glycinergic neurotransmission as well as supraspinal input. In contrast, studies in other vertebrate preparations have shown that spontaneous movement requires glycine and other neurotransmitters and that later behaviors do not require supraspinal input. Here, we use lesion analysis combined with high‐speed kinematic analysis to re‐examine the role of glycine and supraspinal input in each of the three behaviors. We find that, similar to other vertebrate preparations, supraspinal input is not essential for spontaneous movement, touch‐evoked coiling, or swimming behavior. Moreover, we find that blockade of glycinergic neurotransmission decreases the rate of spontaneous movement and impairs touch‐evoked coiling and swimming, suggesting that glycinergic neurotransmission plays critical yet distinct roles for individual patterns of locomotive behaviors. © 2006 Wiley Periodicals, Inc. J Neurobiol, 2006     

Network interactions among sensory neurons in the leech

Interactions among mechanosensory neurons, sensitive to touch, pressure and nociceptive stimuli in the leech nervous system were studied in isolated ganglia and in body-wall preparations. Pairs of touch-pressure, touch-nociceptive and pressure-nociceptive neurons were tested by suprathreshold stimulation of one neuron while recording the response of the other, in both directions. Pressure and nociceptive stimulation evoked depolarizing and hyperpolarizing responses in touch cells, mediated by interneurons. The relative expression of these responses depended on the stimulus duration. One or two pressure cell spikes produced, predominantly, a depolarization of the touch cells, and increasing number of spikes evoked a hyperpolarization. Nociceptive cells produced primarily the hyperpolarization of touch cells at any stimulus duration. When touch cells were induced to fire by injection of positive current into the soma, stimulation of pressure cells inhibited touch cell activity. However, when touch cells were induced to fire by peripheral stimulation, pressure cell activation failed to inhibit touch cell firing. The results suggest that excitation of pressure and nociceptive cells would not limit the responses of touch cells to peripheral stimuli, but would inhibit the firing of touch cells evoked by their central connectivity network.     

Fine needle aspiration cytology of clear cell sarcoma. Report of a case with immunocytochemical, immunohistochemical and ultrastructural studies.

Cytologic findings of clear cell sarcoma obtained by fine needle aspiration (FNA) of a tumor are described. The tumor probably originated in the retroperitoneal tissue, and the diagnosis was confirmed histologically by open biopsy. Percutaneous needle aspirates of the intraabdominal tumor and touch preparations obtained from the open biopsy specimen revealed numerous atypical cells with an extremely hyperchromatic nucleus, prominent nucleoli and clear cytoplasm. The cytoplasm was rich in glycogen. The immunocytochemical technique demonstrated S-100 protein and neuron-specific enolase in the cytoplasm, both of which were exhibited also in the histologic specimen. Clear cell sarcoma is a rare tumor of soft tissue, and to our knowledge, detailed cytologic appearances of this tumor obtained by FNA have not been reported. In addition, the present tumor was unique in location. It is possible to diagnose clear cell sarcoma accurately on an FNA cytologic specimen if the periodic acid-Schiff stain and immunocytochemical technique are utilized in addition to the routine Papanicolaou method.     

Touch imprint cytology in biopsy of the infertile testis.

OBJECTIVE: To identify different cell types in the testis by using touch imprint cytology and to compare the cytologic findings to the histopathologic diagnosis in infertile men. STUDY DESIGN: This prospective study used touch imprint preparations and included 20 infertile men. The biopsy material obtained was stained with toluidine blue, May-Grünwald-Giemsa stain and Papanicolaou stain. The cytologic results for oligospermic, normospermic and azospermic men were compared to the specific histopathologic diagnosis. The proportion of spermatogenic versus Sertoli cells was calculated. The scores were compared between three groups based on the results of the histologic biopsy: normal spermatogenesis, hypospermatogenesis and incomplete spermatogenic arrest. RESULTS: The mean ratio of the spermatogenic cells versus Sertoli cells was statistically significantly different in the three groups (P < .01). The mean ratio of spermatogenic cells to Sertoli cells was higher in cases with normal spermatogenesis than in cases with hypospermatogenesis and incomplete spermatogenic arrest, revealing a statistical difference (P<.01). This ratio was not statistically significantly different between the hypospermatogenesis and incomplete spermatogenic arrest groups. CONCLUSION: A cytologic demonstration of germinal cells by using touch imprint preparations may be a guide for histologic diagnosis.     

Relationship of ploidy and chromatin condensation in the rat liver,moreover a comparison of the nuclear texture in sections and touch preparations

Summary In touch preparations and tissue sections from normal rat hepatocytes and -amanitine- and actinomycind-poisoned liver cells the extent of chromatin condensation relatively to the degree of ploidy and the chromatin distribution were studied by means of computer aided cytophotometry. It could be found, that the relationship of the condensed and decondensed chromatin is independent of the degree of ploidy and first of all dependent upon the intoxicant. Therefore, the extent of chromatin condensation can be utilized also in sections as additional parameter for automated cell screening.Supported by the Deutsche Forschungsgemeinschaft, Sonderforschungsbereich 105     

Keratin-positive reticulum cells in fine needle aspirates and touch imprints of hyperplastic lymph nodes. A possible pitfall in the immunocytochemical diagnosis of metastatic carcinoma.

Fine needle aspirates and touch imprints of 36 hyperplastic (reactive) lymph nodes were tested for the presence of keratin and desmin. Keratin-positive cells with morphologic characteristics corresponding to extrafollicular (fibroblastic) reticulum cells were found in 18% of the fine needle aspirates and 42% of the touch imprints. The number of keratin-positive reticulum cells varied from 1 to greater than 30 per slide. Desmin-positive cells with similar morphology were found in 23% of fine needle aspirates and 37% of touch imprints, and the number of such cells per slide ranged from 2 to greater than 70. The relatively frequent occurrence of keratin-positive reticulum cells in these preparations from hyperplastic lymph nodes should be taken into account if keratin antibodies are used to search for carcinoma micrometastases.     

Diagnostic cytologic features of an epididymal melanotic neuroectodermal tumor of infancy present in scrotal fluid: a case report

BACKGROUND: The cytologic features of melanotic neuroectodermal tumor of infancy (MNTI) have been described in rare cases only, and these reports have been based solely on direct smears obtained from aspirated tumor material or from touch preparations of resected tumors. CASE: We report, to our knowledge, the first documented case of an epididymal MNTI diagnosed from aspirated hydrocele fluid processed using the ThinPrep system (Cytyc Corp., Boxborough, Massachusetts, U.S.A.). CONCLUSION: Based on a 15 year retrospective review of scrotal fluids submitted for cytologic evaluation to our laboratory, MNTI represented the sole neoplasm that was represented in the fluid and could be specifically diagnosed. Compared to the direct smear evaluation in our case and other published reports, the ThinPrep method enables the recognition of both neuroblast-like cells and larger melanin-containing epithelial cells in scrotal fluid, thereby aiding in the specific diagnosis of MNTI and minimizing the risk of misdiagnosing this tumor as a more aggressive neoplasm.     

Amelanotic malignant melanoma arising in an ovarian cystic teratoma: a case report

BACKGROUND: Malignant melanoma arising in a cystic teratoma is extremely rare. We report the clinicopathologic and cytopathologic features of an amelanotic malignant melanoma arising in an ovarian cystic teratoma. CASE: A 55-year-old woman presented with an asymptomatic right ovarian mass, showing features of cystic teratoma according to preoperative computed tomography and magnetic resonance imaging. The resected teratoma was suspected to include a nonepithelial malignancy in a touch preparation from a solid component. The tumor showed immunoreactivity for Melan-A, S-100 and HMB-45 in the absence of melanin granules, which established the diagnosis of amelanotic malignant melanoma arising in an ovarian cystic teratoma. CONCLUSION: Cytopathologic findings from touch preparations and immunohistochemical staining are useful for the diagnosis of amelanotic malignant melanoma arising in an ovarian cystic teratoma.     

Chondrosarcoma of the trachea. A case report and literature review

The cytologic presentation of a case of chondrosarcoma of the trachea in a 72-year-old man is described. A mass detected on routine chest roentgenogram and defined by CT scan was used to make a touch imprint smear during partial tumor resection. The cytologic findings included round or polygonal cells with occasional binucleation, round hyperchromatic nuclei and prominent nucleoli, present in an amorphous pink-violet or light-blue background containing fragments of chondroid tissue. The histopathology was interpreted as a low-grade chondrosarcoma. Cartilaginous tumors of the trachea should be considered in the differential diagnosis of upper airway obstruction.     

A gastrointestinal-specific monoclonal antibody that may be of clinical value in cytologic material

A new monoclonal antibody (MAb), 29-10, produced by immunization of mice with cells from the SW 1116 colorectal carcinoma cell line, detected an antigen present in cytologic touch imprints of surgically resected normal and neoplastic gastrointestinal (GI) tissue, including specimens from the stomach and the colon. These imprints were fixed in 95% ethanol and stained with the avidin-biotin immunoperoxidase technique. In tested cases, 22 (100%) of 22 imprints from GI adenocarcinomas and from normal GI tissue, as well as 13 (56.6%) of 23 imprints from colonic polyps, stained positively while no staining was demonstrable in imprints from other tissues. In histologic sections, only 4 (23%) of 17 colonic adenocarcinomas and 3 (11.5%) of 26 polyps stained positively. The staining ability of MAb 29-10 was compared to that of MAb 19-9, another colorectal antibody, and was found to be markedly superior for binding of the antigen in cytologic preparations. This tissue-specific antibody may be useful in identifying malignant cells of metastatic carcinoma as to their GI tract origin.     

Cytology of lumpectomy specimens.

Residual microscopic disease after lumpectomy may cause significant local recurrence. This study evaluated the use of touch preparation cytology to assess lumpectomy margins. For 90 specimens, the findings on Diff-Quik-stained touch preparations (rendered intraoperatively within 15 minutes after the lumpectomy) were correlated with the gross findings, frozen-section results and, later, the findings in the permanent histologic sections. Three specimens were cytologically unsatisfactory, 68 yielded benign findings, and 19 were suspicious or diagnostic for malignancy. The margins showed tumor involvement in 5 lumpectomy samples by gross examination, in 13 by frozen-section evaluation and in 17 by the study of permanent sections. Touch preparation cytology was putatively falsely positive in two cases while frozen-section evaluation was falsely negative in four cases. Cytology had a sensitivity of 100%, a specificity of 97.1% and a diagnostic accuracy of 97.7%. These results demonstrate that touch preparation cytology rapidly and reliably evaluates lumpectomy margins and can overcome some sampling errors and artifacts related to frozen-section analysis. Touch preparation cytology is now being used to complement frozen-section evaluation of lumpectomy margins as part of a protocol aimed at reducing the local recurrence rate.     

Qualitative and quantitative analysis of cytologic assessment of astrocytoma, oligodendroglioma and oligoastrocytoma

OBJECTIVE: To evaluate the usefulness of intraoperative cytology for differential diagnoses of astrocytoma, oligodendroglioma and oligoastrocytoma. STUDY DESIGN: Qualitative analysis of cytologic features of the 3 brain tumors was conducted using intraoperative touch or squash preparations that were stained with the Papanicolaou method, targeting the cellular density, cytoplasmic and nuclear profiles and blood vessel morphology. In addition, we attempted a computer-assisted image analysis of tumor cell nuclei and compared the results with qualitative observations. RESULTS: Astrocytomas were characterized by many fibrillary cytoplasmic processes and large, irregular nuclei. Oligodendrogliomas were characterized by small, round nuclei and a fine, delicate capillary network. In both tumors of a higher grade, anaplastic large nuclei and proliferating endothelial cells were noted. Oligoastrocytomas showed combined cytologic profiles of astrocytomas and oligodendrogliomas. Quantitative studies suggested that nuclei of oligodendroglial tumors were significantly rounder than those of astrocytomas. In general, the quantitative results were consistent with the qualitative observations. CONCLUSION; Cytologic evaluation using touch or squash preparations is of great help for intraoperative differential diagnosis of astrocytoma, oligodendroglioma and oligoastrocytoma. Cytologic as well as histologic assessment should be conducted at the intraoperative diagnosis of these tumors.     

Nuclear grooves in cytologic preparations. A study of the utility of this feature in the diagnosis of papillary carcinoma

The sensitivity and specificity of nuclear grooves and inclusions for papillary carcinoma was investigated in 32 touch preparations and 69 fine needle aspiration cytologic preparations of the thyroid. Ultrastructurally, these grooves and inclusions are cytoplasmic invaginations into the nucleus. Overall, 100% of the papillary carcinomas contained nuclear grooves while only 70% contained inclusions. Grooves, however, could be seen in 70% of nonpapillary neoplasms and in 56% of nonneoplastic conditions of the thyroid, albeit generally fewer in number and often not as distinct. Inclusions were present in 13% of nonpapillary neoplasms and were absent in nonneoplastic conditions. Some nuclei on cytologic preparations contain lines that are probably artifacts of chromatin alignment and do not represent true nuclear grooves. Since such lines may be indistinguishable from true grooves, grooves should be used cautiously and in conjunction with other criteria in the diagnosis of papillary carcinoma.     

Inhibition of the reversible deactivation of chicken adipose tissue hormone-sensitive lipase by heat-stable proteins from adipose tissue and skeletal muscle.

The reversible deactivation of chicken adipose tissue hormone-sensitive lipase is catalyzed by a lipase phosphatase. Heat-stable protein preparations from rat epididymal fat pads, chicken adipose tissue, and rabbit skeletal muscle inhibited lipase phosphatase activity. Phosphatase inhibitor preparations from rat adipose tissue did not inhibit the protein kinase-catalyzed activation of hormone-sensitive lipase, whereas inhibitor preparations from rabbit skeletal muscle were contaminated with protein kinase inhibitor.     

The role of conformational changes in the functioning of brain Na,K-ATPase]

In the experiments with enzyme preparations of Na,K-ATPase from normal brain tissue (NBT) and tumorous brain tissue (TBT) the following data were established: 1) the cooperativity of Na,K-ATPase with Na+ from NBT is temperature-dependent, the Hill coefficient (nH) at 37, 27.0-30.5 and 20-22 degrees C being 1.80 +/- 0.07, 1.30 +/- 0.09 and 1.10 +/- 0.08, respectively; the cooperativity of Na+ with Na,K-ATPase from TBT was absent; 2) the cooperativity for ouabain (nH-1.30 +/- 0.05) was revealed only in the case of Na-pump from TBT; 3) the protective effect of ATP against the inhibitory action of pCMB is temperature-dependent and differs significantly in enzyme preparations from NBT and TBT; 4) the parameters of the temperature inactivation of enzyme preparations at 45-52 degrees C, especially the change of entropy (delta S*) were different in the case of NBT and TBT; 5) a peptide fraction isolated from sheep brain differently inhibited the Na,K-ATPase from NBT and TBT. In conclusion, these data demonstrate that there are significant differences in functioning of Na,K-ATPase from NBT and TBT, and that besides lipid-protein interactions the local domenic conformational changes in the enzyme molecule may play a definite role in these differences.