Computerized morphometric image analysis of cytologic nuclear parameters in breast cancer

The value of nuclear morphometry in the preoperative fine needle aspiration (FNA) cytologic diagnosis of mammary lesions was investigated and correlated with the lymph node status of the patients. The subjects consisted of four groups of patients: 49 with invasive ductal carcinomas (18 with no positive nodes, 16 with one to three positive nodes and 15 with four or more positive nodes) and 14 patients with benign lesions. The FNA specimens were smeared onto slides and stained by the May-Grünwald-Giemsa technique. The area, perimeter and maximum diameter of 100 randomly chosen nuclei were both measured with the IBAS image analysis system and semiquantitatively estimated with an eyepiece micrometer. For all three parameters, significant differences were found between benign and malignant lesions. The mean nuclear perimeter allowed the morphometric discrimination between all four groups with statistical significance; nuclear area and maximum diameter did not discriminate patients with invasive carcinoma and one to three positive nodes from those with no positive nodes or more than three positive nodes. Morphometry proved to be far superior to eyepiece measurements with respect to accuracy and reproducibility of the results. The results suggest that nuclear perimeter can be used as an additional parameter not only for the FNA cytologic diagnosis of breast cancer, but also for the estimation of patients' prognosis.     

Quantification of angiogenesis by a computerized image analysis system in renal cell carcinoma

OBJECTIVE: To ascertain whether tumor angiogenesis quantitated by a computerized image analysis system correlates with clinical outcome in renal cell carcinoma. STUDY DESIGN: Microvessels were immunohistochemically labeled with antibodies to CD34 in sections from 62 cases of renal cell carcinoma. Computerized image analysis was used to evaluate the mean microvessel count (MMC) and mean percentage microvessel area (MPMA). RESULTS: MMC ranged from 19.3 to 315.0, while MPMA was 0.6-17.9%. There was a highly significant correlation between MMC and MPMA (r = .867, P < .01). Although MMC and MPMA decreased with increasing nuclear grade and TNM stage, this difference failed to achieve statistical significance. No statistically significant differences in survival were found for MMC or MPMA. CONCLUSION: Our results indicate that computerized image analysis can evaluate accurately tumor angiogenesis, but tumor angiogenesis in renal cell carcinoma does not provide significant prognostic information in renal cell carcinoma.     

Use of Feulgen image analysis densitometry to study the effect of genome size on nuclear size in polyploid sturgeons

Feulgen image analysis densitometry (FIA) and image cytometry were used to study the relationship between the DNA content (pgDNA nucleus^?1) and nuclear area (μm²) in blood smears of evolutionary tetraploid (4n) sterlet (Acipenser ruthenus) and stellate sturgeon (A. stellatus); evolutionary octaploid (8n) Siberian sturgeon (A. baerii) and Russian sturgeon (A. gueldenstaedtii); hexaploid (6n) and decaploid (10n) fish found within A. baerii stock; and A. baerii and A. gueldenstaedtii exhibiting dodecaploidy (12n). Standards used for FIA were blood smears of chicken (Gallus gallus domesticus; 2.5 pgDNA nucleus^?1) and diploid and induced triploid tench, Tinca tinca (2.04 and 3.1 pgDNA nucleus^?1, respectively). All ploidy levels were first verified by means of flow cytometry. Species of the same ploidy level, however differing in their DNA content, exhibited a similar mean erythrocyte nuclear area, as could be demonstrated on A. ruthenus and A. stellatus (19.27 and 19.79 μm²_, respectively) with a respective mean DNA content of 3.72 and 4.68 pgDNA nucleus^?1 and the same relationship was found for evolutionary octaploid (8n) A. baerii and A. gueldenstaedtii (29.87 and 30.09 μm², respectively) with respective mean DNA content 8.29 and 7.87 pgDNA nucleus^?1. The 0.19–0.32 pgDNA increments in DNA content of erythrocytes thus had no effect on their nuclear area. With increasing ploidy level, the DNA concentration (pgDNA per μm² of erythrocyte nuclear area) was found not to increase linearly. The DNA in erythrocyte nuclei appeared to be more and more densely packed with an increase of the ploidy level (r = 0.98; R² = 0.95).     

Intranuclear localization of proliferating cell nuclear antigen during the cell cycle in renal cell carcinoma

OBJECTIVE: To investigate, with laser scanning cytometry (LSC), proliferating cell nuclear antigen (PCNA) expression during the cell cycle in renal cell carcinoma. STUDY DESIGN: DNA ploidy and intracellular localization of PCNA in renal cell carcinoma were determined using LSC and immunohistochemistry. The subjects were nine patients who had received surgery for renal cell carcinoma. After DNA ploidy analysis, the glass slides were restained by immunohistochemistry of PCNA. LSC allowed direct observation of PCNA localization during the cell cycle because we could obtain immunohistochemical staining of PCNA as a function of cell cycle phase for individual cells. RESULTS: PCNA was not demonstrated in the nuclei of G0/G1 cells. PCNA expression increased from the S phase of the cell cycle. PCNA rapidly degraded at the end of the G2 phase. In the late G2 and M phase, PCNA was not detected in almost any nucleus. CONCLUSION: LSC allows morphologic observation of the intracellular distribution of PCNA during the cell cycle in renal cell carcinoma.     

The nuclear DNA content of vulvar skin using Feulgen microspectrophotometry and "four-quarter" analysis

The nuclear DNA contents in subbasal (nonreplicating) cells of 161 vulvar biopsies from 31 patients were studied by Feulgen staining, integrated scanning microspectrophotometry (densitometry) and subsequent "four-quarter" analysis of the measurements. The results showed a constancy of the nuclear DNA content in the nonreplicating cells, regardless of the patient or the biopsy site.     

Standardization of the Feulgen reaction for absorption DNA image cytometry: a review.

The present paper gives a review of the current potentials and problems of a standardized Feulgen reaction for absorption DNA image cytometry. The cytochemical basis of the Feulgen reaction is described in the first part of this review. Subsequently, several preparatory factors which influence the performance of the Feulgen reaction, such as fixation, acid hydrolysis, composition of the Feulgen reagent and, in histology, embedding, are discussed in more detail. Some user-oriented recommendations for a standard Feulgen technique are given.     

Some aspects of the Feulgen reaction in situ

Summary Cytological observations combined with studies on absorption spectra of Feulgen stained normal and lipid — extractet HeLa and ehrlich-Lettré mouse ascites cells were performed after fixation of the cells as well in neutral formaldehyde as in Serra fixative. The effects of formaldehyde treatment of the stained cells to substitute all the free amino groups of DNA bond pararosaniline molecules, were also studied. The results obtained by using DNA samples containing 2% protein and relatively free from protein, led to the conclusion that after acid hydrolysis for a short period purines in DNA become splitted and these released aldehydes react with one or two amino groups of pararosaniline, a triphenylmethane dye (according to the arrangement of purines and pyrimidines in the helices). Some protein molecules also take part in the reaction and substitute some of the free amino groups of DNA bound pararosaniline. Peulgen stained cells fixed in Serra fixative show an absorption maximum at 546–550 m. Under appropriate conditions, as in cells fixed in formaldehyde, other substances e.g. phospholipids and lipoproteins interfere with the reaction by substituting most of the free amino groups of DNA bound pararosaniline molecules. It has been argued that in histochemical reactions monosubstituted pararosaniline molecules should be coloured and further substitution of free amino groups of pararosaniline, bound in DNA helices, does not change the intensity of the colour, but gives a shift in the wavelength of the absorption spectra.It has been suggested that the differential response of the nucleoli to the Feulgen-reaction, depending on whether the cells were fixed in formaldehyde or in Serra fixative, may be due to the formation of a protecting shield around the finely distributed intranucleolar chromatin strands, when formaldehyde is being used. After this fixation lipoproteins and other lipids, present in a relatively high percentage and closely associated with the intranucleolar chromatin strands, are especially well preserved.Evidences have been put foreward in support of the amino alkylsulfonic acid theory of Rumpf (1935) and Hörmann et al. (1958) whereas the amino sulfinic acid theory to explain the Schiffs reaction (Wieland and Scheuing, 1921) was shown not to be in agreement with our results.On leave from the Department of Botany, Calcutta University, 35, Ballygunge Circular Road, Calcutta-19, India; on a fellowship from the German Academic Exchange Service.     

Classification of lung carcinoma by means of digital nuclear image analysis

An investigation was performed of the maximum discriminating efficiency for each subgroup of digital nuclear image features and of the overall classification of nuclei from three types of human lung carcinomas in histologic sections: adenocarcinoma, small-cell carcinoma and squamous-cell carcinoma. The results indicate that, for each subgroup of features, the nuclei of the small-cell carcinomas are generally "correctly" classified in a higher percentage (80% to 100%) than are the nuclei of the adenocarcinomas (46% to 74%) and squamous-cell carcinomas (29% to 68%). The discriminant analysis for the overall classification selected features from most of the subgroups, suggesting that it is useful to perform nuclear image analysis with many subgroups having different properties. The overall classifications for the nuclei of the adenocarcinomas, small-cell carcinomas and squamous-cell carcinomas were, respectively, 81.4%, 93.2% and 74.7%. Before this technique can be applied to histopathologic diagnosis, a larger number of unselected lung carcinomas must be evaluated.     

Computer aided morphometric analysis of oral leukoplakia and oral squamous cell carcinoma

We compared the changes in the cells in the basal layer of normal mucosa, oral leukoplakia with dysplasia and different grades of oral squamous cell carcinoma (OSCC) using computer aided image analysis of tissue sections. We investigated three morphometric parameters: nuclear area (NA), cell area (CA) and their ratio (NA:CA). NA and NA:CA ratio showed a statistically significant increase from dysplasia to increasing grades of OSCC. Nuclear size was useful for differentiating normal tissue, potentially malignant leukoplakia and OSCC.     

Ultrastructural and morphometric analysis of the Paneth cell reaction to administration of cholera toxin]

Experiments were made on 40 immature guinea--pigs which were divided into two groups. 20 animals received intraduodenal injections of cholera toxin and 20 others were injected normal saline (groups I and II, respectively). Group I animals exhibited a rapid fall in one--cell secretion of XS recorded on hour 6-12 after the injection of cholera toxin. Inhibition of functional activity was associated with defected synthesis of granules in the cells of Paneth.     

肾透明细胞癌预后相关miRNAs的生物信息学分析

目的寻找可作为肾透明细胞癌（ccRCC）生物标志物的miRNA,以及ccRCC与正常组织间miRNA差异表达情况。 方法利用TCGA数据库下载ccRCC中miRNA表达数据,分析肿瘤与正常组织间差异表达miRNA。使用Kaplan-Meier曲线对患者进行生存分析,筛选出表达情况与临床预后相关的miRNA。通过生物信息学对miRNA的靶基因进行预测,然后运用FunRich软件和ClueGO对靶基因进行GO和KEGG富集分析。 结果通过TCGA数据库分析发现,ccRCC较正常组织差异表达miRNA共54个,其中上调33个,下调21个。通过生存分析发现hsa-miR-21和hsa-miR-155与患者预后相关,P≤0.05。进一步通过Perl软件在Targetscan、miRDB、miRTarBase、miRPath这四个数据库中预测miRNA靶基因并将结果取交集,共发现129个靶基因。GO和KEGG分析结果表明,这些靶基因主要与转录因子活性、信号转导以及FoxO、TNF等信号通路密切相关。 结论通过生物信息学分析发现了ccRCC与正常组织的差异表达miRNA;其中hsa-miR-21和hsa-miR-155与患者总体生存率相关,并通过调控靶基因参与相关的信号通路进而影响ccRCC的发生发展进程,提示hsa-miR-21和hsa-miR-155可能是ccRCC潜在的生物标志物。