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In vivo magnetic resonance imaging and semiautomated image analysis extend the brain phenotype for cdf/cdf mice.

Magnetic resonance imaging and computer image analysis in human clinical studies effectively identify abnormal neuroanatomy in disease populations. As more mouse models of neurological disorders are discovered, such an approach may prove useful for translational studies. Here, we demonstrate the effectiveness of a similar strategy for mouse neuroscience studies by phenotyping mice with the cerebellar deficient folia (cdf) mutation. Using in vivo multiple-mouse magnetic resonance imaging for increased throughput, we imaged groups of cdf mutant, heterozygous, and wild-type mice and made an atlas-based segmentation of the structures in 15 individual brains. We then performed computer automated volume measurements on the structures. We found a reduced cerebellar volume in the cdf mutants, which was expected, but we also found a new phenotype in the inferior colliculus and the olfactory bulbs. Subsequent local histology revealed additional cytoarchitectural abnormalities in the olfactory bulbs. This demonstrates the utility of anatomical magnetic resonance imaging and semiautomated image analysis for detecting abnormal neuroarchitecture in mutant mice.

Animals↗

[Quantitative determination with image analysis of immunohistochemically identified steroid hormone receptors in breast carcinoma].

Oestrogen receptors were demonstrated immunohistochemically in 120 breast cancer cases; in 90 cases, progesterone receptors were additionally assessed. Immunohistochemical staining was evaluated by the so-called "immuno-reactive score" and quantified densitometrically by a method of computer-assisted image analysis. The measuring method allowed an objective evaluation of the portion of receptor-positive cells, the mean optical density and the distribution pattern of receptor concentrations within receptor-positive nuclei. Image analysis was compared with the semiquantitative evaluation by IRS. In 29 out of 58 weakly stained sections, which were scored as receptor-negative, receptor-positive cells could be demonstrated by image analysis because of the definition of an objective cut-off level for specific staining. Furthermore, three different patterns of distribution of receptor concentration could be distinguished: Type 0: receptor-negative carcinomas Type 1: homogeneous receptor expression within receptor-positive cells Type 2: heterogeneous receptor expression with high receptor concentrations in single cells. Type 2 patterns were almost exclusively found in postmenopausal patients, a dependence of expression pattern from grading was not demonstrated. These distribution patterns were found in the same manner in the expression analysis of progesterone receptor. With immunohistochemical staining only 45% of ER-negative carcinomas expressed PR, but 74% of ER-positive carcinomas. When ER was expressed heterogeneously (Type 2) PR was expressed in the same way in 62% of the cases. Beneath the exact determination of the portion of receptor-positive cells the analysis of the pattern of distribution is only possible by image analysis.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

[Comparative study on image analysis and manual counting of immunohistochemistry].

With the use of computer image analysis (IA) and manual counting, the immunohistochemistry (IHC) results on PS2, ER and c-erB-2 of breast cancer which represent positive expression of cytoplasm, cell nucleus and cytomembrane are analyzed and compared. Then the advantages, disadvantages and repetition of the two methods are discussed and the correlation between image analysis and manual counting is estimated. The results of this study indicate that IA has better repetition in estimation of positive expression of cytoplasm, cell nucleus and cytomembrane, whereas manual counting has better repetition only in the positive expression of cell nucleus and cytomembrane, and the two methods have positive correlation in estimation of IHC results. In addition, some problems in image processing are discussed in this paper.

Breast Neoplasms↗

Image analysis for the evaluation of p53 expression in human cancers.

We compared the results of a subjective microscopic evaluation and an interactive image analysis of p53 expression in different human tumours. Expression of p53 was immunocytochemically detected by using the monoclonal antibody PAb 1801, and the evaluation was made blindly by two observers. Image analysis was performed on the same sections using DISCOVERY. The image was segmented into objects and background by interactive thresholding. Several morphological and densitometric features were selected in order to remove artefacts, lymphocytes, stromal cells and overlapping nuclei from the counting. The study was performed on series of 30 cases for different tumour types: breast, colon, oral cavity, ovary and lung cancer. Spearman's correlation coefficient ranged from 0.72 to 0.95 (P = 0.0001). Image analysis can therefore be considered as a quick and alternative approach to microscopic evaluation of nuclear immuno-determinations.

Antibodies, Monoclonal↗

An automatic analysis method for in situ hybridization using high-resolution image analysis.

Specific mRNA for alpha 1 (I) collagen has been detected on a cellular level by in situ hybridization using radioactively labelled alpha 1 (I) antisense RNA probes. We here present an automatic, quantitative, evaluation technique for the determination of grain densities using the high-resolution image analysis system IPS KONTRON. The reliability and objectivity of this method were evaluated by comparing the values of grains per cell obtained by conventional and automatic techniques following in situ hybridization with alpha 1 (I) collagen DNA probes in various specimens. The correlation coefficients between conventional and automatic analysis of grain densities were 0.97 for fibroblasts embedded into a three-dimensional collagen gel, 0.94 for dermal fibroblasts in skin obtained from a patient with progressive systemic scleroderma, and 0.90 for fibroblasts in a 2-week-old scar. All correlation coefficients were significant on the P less than 0.0001 level. This new analysis technique therefore allows a more rapid and reliable quantitative evaluation of in situ hybridization and may also be helpful in differentiating between various cell populations characterized by different biosynthetic capacities.

Autoradiography↗

Quantitation of nucleolar organizer regions by image analysis in glottic squamous cell carcinoma.

OBJECTIVE: To apply computer image analysis as a quantitative method for analyzing interphase nucleolar organizer regions (NORs) to determine whether this proliferative marker provides useful prognostic information in glottic squamous cell carcinoma. DESIGN: Retrospective testing of biopsy samples and resected tissue. SETTING: Nova Scotia Regional Cancer Centre and regional hospitals in Nova Scotia and Prince Edward Island, Canada. PATIENTS: Patients with primary glottic cancer presenting to the cancer centre between 1984 and 1991. INTERVENTIONS: Semiautomated image analysis was used to measure the nuclear area and the NOR area in formalin-fixed paraffin-embedded tumour samples. OUTCOME MEASURES: Mean nuclear area, mean NOR area and NOR percentage of nuclear area, calculated as the mean NOR area divided by the mean nuclear area, expressed as a percentage. RESULTS: Of 154 cases, 90 samples were received; however, 8 paraffin blocks were exhausted and 29 samples stained poorly due to extent of fixation. Analysis of the remaining 53 cases, all primary squamous cell carcinomas, showed no statistically significant association between, on one hand, mean NOR area or NOR percentage of nuclear area and, on the other hand, tumour grade, tumour stage, tumour recurrence or disease-related death. CONCLUSIONS: This study does not demonstrate a prognostic value of NOR measurement as a proliferative marker in primary glottic squamous carcinoma. However, given the small number of cases in this study, further research should be conducted using a larger number of cases from one centre and comparing NOR measures with other markers of cell proliferation.

Biomarkers, Tumor↗

Value of morphometric nuclear image analysis using the Feulgen reaction in renal cell carcinoma.

OBJECTIVE: To evaluate retrospectively the ability of morphometric nuclear image analysis to predict survival in patients with renal cell carcinoma. STUDY DESIGN: The subjects were 40 patients with previously untreated renal cell carcinoma. Pathologic stage was determined using Robson's stage system. Nuclear grade was assigned according to the criteria of Fuhrman et al. We used the Feulgen staining technique, which has been widely used for the histochemical assessment of nuclear DNA content. A minimum of 300 nuclei were analyzed from each subject. Five variables in morphometric nuclear image analysis were measured: nuclear area, nuclear perimeter, nuclear ellipticity, nuclear regularity and DNA content. Cox's proportional hazard model was applied to identify prognostic usefulness with respect to survival time. RESULTS: All nuclear morphometric variables but nuclear regularity correlated with tumor grade. According to univariate survival analyses, Robson stage and nuclear ellipticity revealed a prognosis on survival with statistical significance. After adjustments for age and sex, nuclear ellipticity remained the only significant prognostic factor related to survival (P < .01). The survival rates were relatively high for patients with nuclear ellipticity > 773 as compared to those with nuclear ellipticity < 773 (P < .05). CONCLUSION: These findings indicate that morphometric nuclear image analysis using the Feulgen reaction is a reliable and efficient technique and that nuclear ellipticity is the most discriminating morphometric variable for predicting the prognosis of renal cell carcinoma patients.

Adult↗

[A longitudinal research on palate growth in the normal mixed dentition by three-dimensional image analysis].

OBJECTIVE: To investigate the trait of the palate growth and to provide a method of three-dimensional image analysis on revealing the entire variation of palate. METHODS: A longitudinal study on the palate growth of 30 subjects (male 8, female 22) with normal occlusion during mixed dentition was carried out by three-dimensional image analysis. The observation period was from 7 to 10 years of age. RESULTS: During growth, the palate vaults in 16 cases (53%) descend for 1-5 mm, whereas, the palate vaults in 14 cases (47%) ascend for 2.6 mm at most. CONCLUSION: The variations of palate growth can be investigated by three-dimensional image analysis. During growth, the palate vault may descend or ascend, which shows the individual differences of oronasal function.

Adolescent↗

Genome size determination in peronosporales (Oomycota) by Feulgen image analysis.

Genome size was determined, by nuclear Feulgen staining and image analysis, in 46 accessions of 31 species of Peronosporales (Oomycota), including important plant pathogens such as Bremia lactucae, Plasmopara viticola, Pseudoperonospora cubensis, and Pseudoperonospora humuli. The 1C DNA contents ranged from 0.046 (45. 6 Mb) to 0.163 pg (159.9 Mb). This is 0.041- to 0.144-fold that of Glycine max (soybean, 1C = 1.134 pg), which was used as an internal standard for genome size determination. The linearity of Feulgen absorbance photometry method over this range was demonstrated by calibration of Aspergillus species (1C = 31-38 Mb) against Glycine, which revealed differences of less than 6% compared to the published CHEF data. The low coefficients of variation (usually between 5 and 10%), repeatability of the results, and compatibility with CHEF data prove the resolution power of Feulgen image analysis. The applicability and limitations of Feulgen photometry are discussed in relation to other methods of genome size determination (CHEF gel electrophoresis, reassociation kinetics, genomic reconstruction) that have been previously applied to Oomycota.

Aspergillus↗

Application of flow cytometry and automated image analysis to the study of prostate cancer.

Flow cytometry and image analysis are complementary quantitative cytologic techniques that have demonstrated utility in the assessment and analysis of prostate cancer and other urologic and nonurologic tumors. This review is intended to assess the current state of the art and to project future directions and applications of these modalities in the pathologic assessment of prostate cancer. Special attention is directed toward the topics of prostate cancer detection and diagnosis, determination of patient prognosis, and the monitoring of patient response to therapy. We recommend that 1) flow cytometry and image analysis be used to determine pathologic parameters that will help in predicting poor patient prognosis and 2) quantitative cytologic determinations of DNA content be included in clinical trials so that their ultimate role in monitoring patient response to therapy can be determined. This knowledge will allow the development of protocols designed to test the value of earlier institution of multimodal therapy in high-risk populations.

Flow Cytometry↗

Focal spot motion of linear accelerators and its effect on portal image analysis.

The focal spot of a linear accelerator is often considered to have a fully stable position. In practice, however, the beam control loop of a linear accelerator needs to stabilize after the beam is turned on. As a result, some motion of the focal spot might occur during the start-up phase of irradiation. When acquiring portal images, this motion will affect the projected position of anatomy and field edges, especially when low exposures are used. In this paper, the motion of the focal spot and the effect of this motion on portal image analysis are quantified. A slightly tilted narrow slit phantom was placed at the isocenter of several linear accelerators and images were acquired (3.5 frames per second) by means of an amorphous silicon flat panel imager positioned approximately 0.7 m below the isocenter. The motion of the focal spot was determined by converting the tilted slit images to subpixel accurate line spread functions. The error in portal image analysis due to focal spot motionwas estimated by a subtraction of the relative displacement of the projected slit from the relative displacement of the field edges. It was found that the motion of the focal spot depends on the control system and design of the accelerator. The shift of the focal spot at the start of irradiation ranges between 0.05-0.7 mm in the gun-target (GT) direction. In the left-right (AB) direction the shift is generally smaller. The resulting error in portal image analysis due to focal spotmotion ranges between 0.05-1.1 mm for a dose corresponding to two monitor units (MUs). For 20 MUs, the effect of the focal spot motion reduces to 0.01-0.3 mm. The error in portal image analysis due to focal spot motion can be reduced by reducing the applied dose rate.

Artifacts↗

Quantitation of chromatin patterns by image analysis as a predictive tool in chemopreventive trials with vitamin A.

Nuclear textures of oral mucosal cells were quantitated by image analysis, and their suitability as markers in a chemopreventive trial explored. Subjects were chewers of tobacco-containing betel quids with well established oral leukoplakias. Treatment consisted of a weekly oral administration of vitamin A (200,000 IU/week) for six months. Leukoplakias regressed in 57.1% of the 21 trial participants. The original leukoplakias did not redevelop within four months after termination of treatment. For image analysis, biopsies were taken from leukoplakias of five chewers before administration of vitamin A, at the end of the administration, and four months after termination of treatment. Sections of paraffin-embedded biopsies were stained with the Feulgen reaction and submitted to quantitative image analysis of two parameters: variance of intensity and entropy. Both these parameters were significantly reduced in all five trial participants as a result of the six-month vitamin A treatment. During the post-treatment period, nuclei with condensed chromatin, as measured by the variance of intensity, reappeared in the mucosa of four of the five chewers examined, although no leukoplakias were detectable on visual examination of the oral cavity. The results indicate that the quantitation of nuclear textures in a small subpopulation of a chemopreventive trial could conceivably be a simple marker with a predictive value.

Biomarkers, Tumor↗

The myocardial fibrosis in patients with dilated cardiomyopathy. The application of image analysis in the myocardial biopsies.

BACKGROUND: The aim of our study is to investigate whether myocardial fibrosis measured by image analysis may be considered as an important and accurate index of dilated cardiomyopathy and its prognosis. METHODS: The study group consisted of 24 patients with dilated cardiomyopathy, which was diagnosed by echocardiography, radionuclide ventriculography, cardiac catheterization, and left ventricular endomyocardial biopsy. The patients' overall disability was conventionally expressed with the criteria for functional capacity. Using image analysis, the percentage of fibrosis in a total of 35 myocardial biopsies was accurately measured, followed by a study comparing the percentage of myocardial fibrosis and the clinical parameters (left ventricular ejection fraction and overall functional capacity), showing the degree of each patient's heart failure. RESULTS: A correlation was found among fibrosis, left ventricular ejection fraction, and overall functional capacity. The cases with small values of fibrosis (< 10%) have big values of ejection fraction and belong to Class I of overall functional capacity. The cases with big values of fibrosis (> 10%) belong to Classes III and IV of overall functional capacity and have small values of ejection fraction. The results of the comparative study were graphically presented and considered significant. CONCLUSION: Myocardial fibrosis measured by image analysis might be considered an important prognostic index of dilated cardiomyopathy.

Adult↗

Image analysis quantification of peptide-immunoreactive nerves in the skin of patients with Raynaud's phenomenon and systemic sclerosis.

Image analysis quantification was used to assess the results of immunocytochemistry for a neuronal marker and neuropeptides in digital skin biopsies from Raynaud's phenomenon (RP) and systemic sclerosis (SS) patients, to verify the possibility of a selective quantitative abnormality of immunoreactive nerves. The field area of specific immunostaining and nerve counts were evaluated on coded specimens, and the data compared by statistical analysis. Nerves immunoreactive for protein gene product 9.5 (PGP), a marker for neuronal elements, were decreased significantly in epidermal and subepidermal layers of digital skin in RP patients (P less than 0.0001). This change was paralleled by a decrease of calcitonin gene-related peptide (CGRP) immunoreactive nerves in the epidermis and around capillaries in the dermal papillae (P = 0.005). In the skin of RP patients, these changes were readily demonstrated by image analysis, although they were not always apparent on visual screening. In digital skin of SS patients, there was a generalized and very significant decrease of PGP, CGRP, and VIP immunoreactivities in all areas (P less than 0.0001). These results demonstrate that neuropeptide-containing nerves are involved in the digital pathology of RP and SS, and that image analysis quantification is an accurate and sensitive method for assessing morphological changes in pathological samples.

Adult↗

Development of a new radiographic scoring system using digital image analysis.

A scoring system using computerised analysis of digital stored images of knee radiographs has been developed. Measurement is based on the assessment of joint space size. It is sensitive, rapid, and reproducible. Plain radiographs are positioned on an acetate grid; a computer generated grid is superimposed on a digital image of the radiograph viewed on a closed circuit television monitor and the joint space measured automatically. Area and distance have been assessed; area measurements are more reproducible. Application of microcomputer based digital image analysis to radiological scoring systems is an important step in understanding the nature and progression of arthritis.

Arthritis↗

Computed tomographic image analysis of ears with otosclerosis.

To evaluate the usefulness of quantitative analysis of computed tomographic images in diagnosing stapedial otosclerosis, high-resolution temporal bone computed tomography (CT) of 32 ears with otosclerosis and 19 control ears was used. The images on the section containing the oval window were digitized using a scanner, and the ratio of the mean gray scale value of 5 areas in the optic capsule including the area anterior to the oval window to that of the cortical bone was calculated. The mean value of the ratio of the area anterior to the oval window was significantly lower in the otosclerosis group (0.85 +/- 0.10) than in the control group (1.03 +/- 0.03; Student t test: t = 7.94, p < 0.0001). In 30 of 32 (93.7%) otosclerotic ears, the values were lower than the normal range calculated in the control group. This method of quantitative CT image analysis comparing the area anterior to the oval window was considered useful in diagnosing stapedial otosclerosis.

Adolescent↗

Protecting spermatogenesis from damage induced by doxorubicin using the luteinizing hormone-releasing hormone agonist leuprorelin: an image analysis study of a rat experimental model.

BACKGROUND: This study was performed to investigate the protective effect of a luteinizing hormone-releasing hormone (LHRH) agonist, leuprorelin, against spermatogenetic damage caused by doxorubicin in rats. METHODS: Sprague-Dawley rats were divided into 4 groups: (1) a control group, (2) a group given LHRH agonist (subcutaneous injections, total dose 9 mg/kg), (3) a group given doxorubicin (intraperitoneal injections, total dose 7.5 mg/kg), and (4) a group given both LHRH agonist (subcutaneous injections, total dose 9 mg/ kg) and doxorubicin (intraperitoneal injections, total dose 7.5 mg/kg). Evaluations were made by measuring body and testicular weights, determining Johnsen's score, and conducting DNA image analysis consisting of DNA content measurement (%1C, %2C, and %4C) by image cytometry. RESULTS: In the group given doxorubicin, the testicular weight was 1.47 +/- 0.24 mg, Johnsen's score was 4.4 +/- 1.2, and image analysis revealed %1C: 33.8 +/- 9.2, %2C: 43.9 +/- 16.3, and %4C: 5.0 +/- 4.4. In the group given both LHRH agonist and doxorubicin, the testicular weight was 1.32 + 0.23, Johnsen's score was 5.90 + 1.6, and image analysis revealed %1C: 46.9 +/- 15.0, %2C: 28.4 +/- 13.3, and %4C: 8.8 +/- 3.5. CONCLUSIONS: The significant prophylactic effect (P < 0.05) of the LHRH agonist against doxorubicin-induced spermatogenetic damage was demonstrated by Johnsen's score and image analysis (%1C, %2C, and %4C).

Animals↗

Fully automated TV-image analysis of the cell-cycle: comparison of the PLM method with determinations of the percentage and the DNA content of labelled cells.

A cell-cycle analysis based on a fully automated TV-image scanning system is proposed to replace the laborious PLM method. To compare the efficiency of the two procedures, cell-cycle parameters were assessed in Ehrlich (diploid and hyperdiploid), L-1210, and JB-1 mouse ascites tumours and in rat jejunal crypts. The percentages of labelled mitoses (PLM) were counted visually on Feulgen-stained autoradiographs obtained at various times after a single 3H-thymidine pulse. The fraction of labelled cells (P) and the DNA ratio of labelled and unlabelled cells were measured by TV-image analysis in the same slides and plotted against time. Within practical limits, TV-image analysis using the P-curve gives the same results as the PLM method. Using the P-curve has the important advantage that its first part, beginning at the time of 3H-thymidine injection and ending at the first maximum, furnishes more information about the cell cycle than the corresponding part of the PLM curve. It can be used to compute tG2M tS and the ratio of the growth faction index to the cell-cycle time (IP/tC) whereas the first part of the PLM-curve reveals only the length of the S-phase (tS). The IP/tC ratio is a readily accessible measure of growth and increases when the cells divide more frequently. Cell death rates may be neglected since the ratio is determined within less than the duration of one cell cycle. Moreover, the data from the first part of the P curve indicate whether there is a large non-growth fraction. If the non-growth fraction is small, i.e. if IP approximately 1, the P curve need only be measured until the first maximum is reached so that fewer samples and animals are required. If the non-growth fraction is large or unknown, the cell-cycle parameters are calculated by reference to the position and size not only of the first minimum and the first maximum, but also of the second minimum of the P curve.

Animals↗