The aim of the study was to determine optimal hydrolysis time for the Feulgen DNA staining of archival formalin fixed paraffin-embedded surgical samples, prepared as single cell suspensions for image ...cytometric measurements. The nuclear texture features along with the IOD (integrated optical density) of the tumor nuclei were analysed by an automated high resolution image cytometer as a function of duration of hydrolysis treatment (in 5 N HCl at room temperature). Tissue blocks of breast carcinoma, ovarian serous carcinoma, ovarian serous tumor of borderline malignancy and leiomyosarcoma were included in the study. IOD hydrolysis profiles showed plateau between 30 and 60 min in the breast carcinoma and leiomyosarcoma, and between 40 and 60 min in the ovarian serous carcinoma and ovarian serous tumor of borderline malignancy. Most of the nuclear texture features remained stable after 20 min of hydrolysis treatment. Our results indicate that the optimal hydrolysis time for IOD and for nuclear texture feature measurements, was between 40 and 60 min in the cell preparations from tissue blocks of three epithelial and one soft tissue tumor.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
BACKGROUND Oral lichen planus (OLP) is a chronic inflammatory disease of the oral mucosa with lowrisk of malignant transformation. At present there are no reliable tools to predict whichOLP will ...progress to cancer. Abnormal DNA content has been reported to be a valuablediagnostic marker of potential malignant transformation of oral white patches.Aims To analyze image cytometric DNA ploidy of OLP and assess chromatin changes reflectedin nuclear features, when OLP is compared to normal buccal mucosa.Materials and Twenty-eight patients with reticular form of OLP and 50 healthy control subjects weremethods included in the study. No malignant transformation of OLP was observed during a followupperiod of 25 years. Scrapings of buccal mucosa were suspended in transport medium.Monolayer filter cell preparations were Feulgen-Thionin stained. Image cytometric analysiswas performed by Cyto-Savant system. RESULTS All OLP in our study were diploid. Statistically significant differences were found betweennormal buccal mucosa and OLP in several nuclear texture features. CONCLUSIONS Our results indicate that diploidy predominates in OLP, confirming its benign nature.Minor chromatin changes of OLP were detected by IC, probably of reactive type
To analyze the diagnostic problems with fine needle aspiration biopsy in anaplastic thyroid carcinoma (ATC) and to describe the cytomorphologic characteristics in 113 cases.
A retrospective analysis ...of 113 fine needle aspirates and 67 surgical specimens from 113 patients with ATC admitted to the Institute of Oncology, Ljubljana, in 1972-1992.
In a series of 113 fine needle aspirates of ATC, 3 (2.7%) were inadequate, 3 (2.7%) suboptimal and 107 (94.7%) diagnostic of malignancy. On reexamination, 96/107 (89.7%) were diagnosed as ATC, 6 (5.6%) as differentiated thyroid carcinoma, and 5 (4.6%) as a malignant tumor not otherwise specified. As to the predominant cell population, fine needle aspirates showed three different cell patterns: (1) pleomorphic cell (43 cases), (2) round cell (33 cases), and (3) spindle cell pattern (7 cases). In the present retrospective analysis we identified three main reasons for inadequate or nonrepresentative fine needle aspiration biopsy sampling: (1) tumor regressive changes (necrosis, hemorrhage, leukocytic infiltration), (2) extensive tumor fibrosis, and (3) distinct differentiated and anaplastic patterns in the same tumor.
The major diagnostic problem with fine needle aspiration biopsy (FNAB) of ATC is related to sample quality. Cytomorphologic features of ATC are highly specific and easy to recognize. Due to the simple technique and high diagnostic accuracy, FNAB is the method of choice in patients with ATC.
To analyze image cytometric chromatin changes reflected in nuclear texture features and DNA ploidy of oral lichen planus in relation to the normal buccal mucosa and buccal mucosa expressing ...malignancy-associated changes in cancer patients.
Twenty-eight patients with the reticular form of oral lichen planus, with a follow-up period of 25 years, 50 healthy controls and 50 lung cancer patients were included in the study. Scrapings of buccal mucosa were suspended in transport medium. Monolayer filter preparations were Feulgen-thionin stained. Image cytometric analysis was performed by Cyto-Savant.
All oral lichen planus specimens in our study were diploid. In univariate analysis, differences between the normal buccal mucosa and oral lichen planus were found in several nuclear texture features, which gave an 80% correct classification rate in multivariate analysis. In the second part of the study, the classifier that recognizes malignancy-associated changes on the buccal mucosa of patients with lung cancer correctly recognized > 80% of oral lichen planus samples as normal buccal mucosa.
Our results indicate that chromatin changes in oral lichen planus exist compared to normal cells; however, the chromatin structure of the reticular form of oral lichen planus does not express malignancy-associated changes and is more similar to normal squamous cells.
To analyze the presence of malignancy associated changes (MACs) in normal buccal mucosa cells of lung and breast cancer patients and their relationship to tumor subtype, stage and size.
Buccal mucosa ...smears of 107 lung cancer and 100 breast cancer patients and corresponding healthy subjects were collected, stained by the DNA-specific Feulgen-thionin method and scanned using an automated high-resolution cytometer. Nuclear texture features of a minimum of 500 nuclei per slide were calculated, and statistical classifiers using Gaussian models of class-probability distribution were designed, trained and tested in 3 parts: (1) ability to separate cancer patient samples from controls, (2) cross-validation of classifiers for different cancer types, and (3) correlation of MAC expression with tumor subtype, stage and size.
Lung and breast cancer induce MACs in normal buccal mucosa cells. The classifiers based on the selected nuclear features correctly recognized >80% of lung and breast cancer cases. The results indicate that MAC detection is not dependent on the tumor subtype, stage or size.
The presence of MACs in buccal mucosa cells offers the potential for developing a new noninvasive cancer screening test.
To analyze the proportion of nontumor cells in fine needle aspirates of breast carcinoma and its influence on flow cytometric S-phase fraction (SPF) estimation.
We analyzed the proportion of nontumor ...cells in fine needle aspiration biopsy smears, performed flow cytometric analysis of DNA ploidy and SPF on freshly aspirated tumor material and analyzed histograms manually and automatically using Multi-Cycle AV software (Phoenix Flow Systems, San Diego, California, U.S.A.) for cell cycle analysis. We corrected SPF of diploid tumors for the dilution effect using an individually established percentage of nontumor cells (individual correction) and the mean proportion of nontumor cells in diploid tumors (factor correction).
The proportion of nontumor cells ranged from 0.5% to 76.6% (mean, 12.6; SD, 15.7) in 55 diploid tumors and from 0.5% to 53% (mean, 8.6; SD, 8.9) in 84 aneuploid tumors (p=0.178). In 14 of 139 (10%) samples, the proportion of nontumor cells exceeded 20%. The mean SPF values of diploid tumors without correction were 4.9% (manually) and 6.5% (automatically) and of aneuploid tumors, 9.5% and 11.0%, respectively. In univariate Cox survival analysis, noncorrected SPF was a significant prognostic factor in overall survival (p < 0.001). Neither individual nor factor correction of SPF significantly changed its prognostic value.
Fine needle aspirates contain low proportions of nontumor cells, having an insignificant dilution effect on SPF estimation. Most probably, SPF could be reliably estimated usingfreshly aspirated tumor material without any correction or adjustment.
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