Detection of pathological aortic tissues by infrared multispectral imaging and chemometrics.

Processing of multispectral images is becoming an important issue, especially in terms of data mining for disease diagnosis. We report here an original image analysis procedure developed in order to compare 42 infrared multispectral images acquired on human ascending aortic healthy and pathological tissues. Each image contained about 2500 infrared absorption spectra, each composed of 1641 variables (wavenumbers). To process this large data set, we have restricted the spectral window used to the 1800-950 cm(-1) spectral range and selected 100 spectra from the aortic media, which is the most altered part of the aortic tissue in aneurysms. Prior to this selection, a spectral quality test was performed to eliminate 'bad' spectra. Our data set was first subjected to a discriminant analysis, which allowed separation of aortic tissues in two groups corresponding respectively to normal and aneurysmal states. Then a K-means analysis, based on 20 groups, allowed reconstruction of infrared images using false-colours and discriminated between pathological and healthy tissues. These results demonstrate the usefulness of such data processing methods for the analysis and comparison of a set of spectral images.

In-vitro analysis of normal and aneurismal human ascending aortic tissues using FT-IR microspectroscopy.

FTIR microspectroscopy has shown to be a proven tool in the investigation of many tissue types. We have used this spectroscopic approach to analyse structural differences between normal and aneurismal aortic tissues and also aortas from patients with congenital anomalies like aortic bicuspid valves. Spectral analysis showed important variations in amide I and II regions, related to changes in alpha-helix and beta-sheet secondary structure of proteins that seem to be correlated to structural modifications of collagen and elastin. These proteins are the major constituents of the aortic wall associated to smooth muscular cells. The amide regions have thus been identified as a marker of structural modifications related to these proteins whose modifications can be associated to a given aortic pathological situation. Both univariate (total absorbance image and band ratio) and multivariate (principal components analysis) analyses of the spectral information contained in the infrared images have been performed. Differences between tissues have been identified by these two approaches and allowed to separate each group of aortic tissues. However, with univariate band ratio analysis, the pathological group was found to be composed of samples from aneurismal aortas associated or not with an aortic bicuspid valve. In contrast, PCA was able to separate these two types of aortic pathologies. For other groups, PCA and band ratio analysis can differentiate between normal, aneurismal, and none dilated aortas from patients with a bicuspid aortic valve.

[Dilatation of Virchow-Robin perivascular spaces (types III cerebral lacunae): radio-clinical correlations]. / Dilatation des espaces périvasculaires de Virchow-Robin (Lacunes de type III): corrélations radio-cliniques.

BACKGROUND AND PURPOSE: Virchow-Robin spaces are pia-lined extensions of the subarachnoid space surrounding the path of brain vessels. When enlarged, such dilated perivascular spaces are often seen as foci of cerebrospinal fluid signal on MRI or CT scan. These foci are found in patients with miscellaneous clinical status. It is necessary to determine the radiological significance and clinical associations, if any, in such patients in order to give them the appropriate treatment. METHODS: We describe the clinical and radiological findings of five patients and review the literature on perivascular Virchow-Robin spaces. RESULTS: The mechanisms of dilated Virchow-Robin spaces are still not well understood. Such dilated perivascular spaces are found in two locations: typically in the high-convexity white matter of healthy elderly subjects, or surrounding the lenticulostriate vessels as they enter the basal ganglia. On MR images, they may be confused with lacunar infarcts. Most of the patients present with no symptoms: small dilatations located in the high convexity actually represent an anatomic variant, also called "état criblé". Sometimes, giant dilatations, or Poirier's type IIIb "expanding lacunae", found in the basal ganglia and midbrain may result in symptomatic hydrocephalus needing appropriate treatment. For other miscellaneous symptoms as headache, generalized epilepsy, dysmorphy, macrocephaly, there is no reliable correlation with enlarged perivascular spaces seen on MR images. CONCLUSIONS: The real symptomatic dilated perivascular spaces need appropriate and quick treatment. Most of the other patients present with no symptoms and will remain asymptomatic.

[Systemic lupus erythematosus presenting as severe alveolar hypoventilation and the shrinking lung syndrome]. / Hypoventilation alvéolaire sévère révélant un shrinking lung syndrome lupique.

INTRODUCTION: The shrinking lung syndrome (SLS) is a rare complication of systemic lupus erythematosus. CASE REPORT: A 69-year-old man presented with exertional dyspnoea, muscle weakness, and weight loss of 15kg in 6months. Pulmonary function tests revealed a restrictive lung disorder, with a dramatic decrease in maximal inspiratory pressure (17% of theoretical value), and alveolar hypoventilation (pH 7.43; PaCO2 55mmHg). A thoracic CT-scan showed bilateral diaphragmatic elevation. The creatinine phophokinase level was increased at 280U/L. Progress was marked by a rapidly increasing respiratory acidosis (pH 7.24, PaCO2 109mmHg) requiring invasive ventilation. Auto-immune studies revealed positive anti-nuclear antibodies (1/800) and positive anti-native DNA antibody at 45U/L. Treatment with systemic corticosteroids led to an initial improvement but it was not possible to discontinue mechanical ventilation. The outcome was fatal. Autopsy did not reveal any other cause and a diagnosis of the SLS associated with lupus was confirmed. CONCLUSION: The interesting features of this case report consist of: 1) the presentation of the SLS as an alveolar hypoventilation with a fatal outcome, 2) the presentation of systemic lupus as SLS.

Recurrent deletion of the short arm of chromosome 3 in human renal cell carcinoma: shift of the c-raf 1 locus.

A cytogenetic study performed on 6 human renal cell carcinomas after short-term culture on extracellular matrix with serum-free medium showed aneuploidy without structural changes in 2 tumors and a rearrangement of the short arm of chromosome 3 in 4 tumors, including deletions and a translocation involving the 3p14 and 3p21 bands. Chromosomal in situ hybridization with a c-raf 1 probe demonstrated that in 2 renal cancers with del3(p14 or 21) the cellular oncogene had shifted from 3p25 to 3p14 as a result of an interstitial deletion.

[Nano-biocaptures for research and diagnostics in inflammation diseases and cancer]. / Nano-biocapteurs pour la recherche et les diagnostics des maladies inflammatoires et du cancer.

As part of the ongoing search for ways to decrease the mortality of different pathological conditions related to cancer and inflammatory diseases, nanotechnologies currently under evaluation offer potentially attractive tools for innovative methodologies for early diagnosis, new bioimaging techniques and therapeutic strategies. Nano-tools can be employed for various functions, such as the detection of lesions at very early stages of disease development, extremely precise anatomical localization, or evaluation of the efficacy of medications specifically targeted against cells and pathological tissues. We have synthesized homogeneous CdSe/ZnS (core/shell) highly fluorescent nanocrystals (NC) detectable as individual nanoparticules with a routine fluorescent microscope. These NC are at least 10-fold brighter than the best organic fluorophores and at least 1000-fold more stable against photobleaching than AlexaFluor, for example. When conjugated with proteins, DNA or with drugs, NCs may be excited with the light of any wavelength from UV through visible spectral region providing a range of fluorescence colors depending on their diameter. These properties provide excellent perspectives for high through-put multiplexing and long-term tracking of labeled precursors for days or even weeks. We present here NC applications for ultrasensitive detection of p-glycoprotein, cytokeratins, LCA, Ki67, etc. both on the cellular level and in pathological human surgical specimens.

Quantitative immunolocalization of a P29 protein (GRA7), a new antigen of toxoplasma gondii.

Ultrastructural localization of a P29 protein of Toxoplasma gondii was examined on thin sections by an immunogold technique using a P29 antigen-specific monoclonal antibody (5-241-178). Immunolocalization of the P29 protein in extracellular tachyzoites demonstrated that this antigen was present in the dense granules. Thus, we have identified this P29 antigen as the seventh protein (GRA7) to be localized to the dense granules of T. gondii. P29 immunolocalization in intracellular tachyzoites demonstrated association of this antigen with the parasite membrane complex, tubular elements of the intravacuolar network, and with the parasitophorous vacuolar membrane. Our immunolabeling data suggest trafficking of the P29 (GRA7) antigen from the dense granule via the intravacuolar network to the parasitophorous vacuolar membrane on invasion of the tachyzoite into the host cell. (J Histochem Cytochem 46:1411-1421, 1998)

Epstein-Barr virus-associated primary malignant lymphomas of the pleural cavity occurring in longstanding pleural chronic inflammation.

Primary pleural non-Hodgkin's lymphomas (NHLs) have been described mostly in Japan and occasionally in western countries. We report three cases occurring in nonimmunocompromised patients demonstrating close similarities to the Japanese cases. These lymphomas occurred more than 20 years after thoracoplasty for the treatment of pleural or pulmonary tuberculosis. They were all high grade lymphomas of B phenotype (two immunoblastic with plasmocytoid differentiation and one immunoblastic with anaplastic large cells). In situ hybridization (ISH) detected small nuclear RNAs encoded by Epstein-Barr virus (EBERs) in most lymphomatous cells in all cases. This suggests a link between primary pleural lymphoma and the longstanding chronic inflammation, which could favor a clonal evolution of Epstein-Barr virus (EBV)-infected B cells through the production of growth factors or immunosuppressive cytokines produced by inflammatory cells.

Immunohistochemical study of thrombospondin and its receptors alpha root of beta 3 and CD36 in normal thyroid and in thyroid tumours.

AIM: To describe the pattern of distribution of thrombospondin (TSP1) and its receptors, alpha root of beta 3 integrin and CD36, in normal human thyroid tissue and to compare their expression in different benign and malignant thyroid conditions. METHODS: Immunohistochemistry was used to study TSP1 and its receptors in 40 surgical thyroidectomy specimens (normal parenchyma, 7; follicular adenoma, 4; multinodular goitre, 13; papillary carcinoma, 6; follicular carcinoma, 8; anaplastic carcinoma, 2). RESULTS: In the normal thyroid parenchyma, there was weak expression of TSP1 limited to the vessels with no staining of the extracellular matrix. In goitres, the expression of TSP1 was more pronounced in areas of fibrosis, with staining of alpha root of beta 3 on thyrocytes located in the vicinity. In thyroid adenomas, expression of TSP1 was slightly enhanced compared with normal tissue, located in the basement membrane of vessels. In papillary carcinomas, TSP1 was abundant in the desmoplastic stroma with a cytoplasmic distribution of alpha root of beta 3 integrin in thyrocytes. In follicular carcinomas, TSP1 was less abundant in the extracellular matrix, limited to the vessels of the stroma with a weaker expression of alpha root of beta 3 on thyrocytes than in papillary carcinomas. In anaplastic carcinomas, TSP1 was only present in the numerous capillaries of the stroma, with a marked positivity for alpha root of beta 3 in one case. No immunostaining of thyrocytes is observed with CD36. CONCLUSIONS: These results suggest the importance of the interaction between alpha root of beta 3 integrin and TSP1 during remodelling of the matrix in fibrous goitres with areas of early sclerosis comparable with wound healing. In papillary carcinomas, the overexpression of TSP1 restricted to the stroma suggests protective effects against tumour progression.

Subcellular localization and quantitative analysis of Toxoplasma gondii target antigens of specific immunoglobulins G, M, A, and E.

The target antigens of specific immunoglobulins G, M, A, and E from patients with acquired acute toxoplasmosis were determined using immunocytochemistry. The relative repartition of these antigens in four cellular compartments of Toxoplasma (membrane complex, apical area, rhoptries, and dense granules) was quantitatively evaluated. Rhoptry antigens mainly react positively with IgA. Membrane, submembrane area (membrane complex), and rhoptry antigens are immunodominant for IgA and IgM. Apical area antigens are recognized by IgM two times more than IgG and IgA. IgE recognized only rhoptry antigens. The localization of pathogenetically antigenic components and their identification by the immune system appeared to be of importance for selection of immunodominant or recombinant antigens. Such localization would improve laboratory diagnosis of serious congenital toxoplasmosis or in immunocompromised patients with toxoplasmic complications after cyst reactivation.

Subcellular calcium localization in Toxoplasma gondii by electron microscopy and by X-ray and electron energy loss spectroscopies.

The localization of calcium in Toxoplasma gondii tachyzoites was studied at the ultrastructural level, with a cytochemical pyroantimonate precipitation method (PA) and controlled by EGTA chelating and EDX and EELS microanalyses. Appropriate conditions for material preparation, fixation and embedding, were defined. The proportion of precipitates that were either free or inside vacuoles and their distribution inside Toxoplasma appeared to be PA dose-dependent. Precipitation mainly occurred in the anterior pole of the Epon-embedded tachyzoites. EDX and EELS analyses showed that out of 30 PA precipitates inside tachyzoites, 78% contained Ca. In Melamine sections, 96% of the tachyzoites had intracellular precipitates and the membrane complex was stained; 25% of the tachyzoites inside host cells contained PA-Ca precipitates, but most of them were retained in the reticular network of the parasitophorous vacuole. Melamine embedding appeared to improve the preservation of calcium pyroantimonate precipitates.

Effects of gallium chloride oral administration on transplanted C3HBA mammary adenocarcinoma: Ga, Mg, Ca and Fe concentration and anatomopathological characteristics.

To determine the influence of the length of the treatment on the anatomopathological and biochemical intratumor changes induced by gallium, we treated C3H BA mammary adenocarcinoma-bearing C3H/HeJ mice with gallium chloride daily, for a period of either 21 or 42 days. In both cases the same dose of 200 mg/kg/24h was administered. An increase of collagen fibrosis in treated tumors as opposed to controls was only noted after 42 days, as well as a significant decrease of the intratumor magnesium and calcium concentrations that could be responsible for a reduction in the metabolic activities of the malignant cells. Remarkable intratumor gallium concentrations (38.4 +/- 30.3 nmol/g after 21 days of treatment; 13.4 +/- 7.3 nmol/g after 42 days where the necrosis is much more important) are obtained after this oral administration. There is no renal toxicity and a higher tumor/kidney concentration ratio is obtained than after acute parenteral administration. The effect of gallium may be different according to the mode of administration: it may be more cytotoxic after parenteral administration, while after oral administration it may act as a better metabolic regulator with a more selective tumor uptake and fewer side effects.

Contribution of quantitative cytology to the cytological diagnosis of thyroid neoplasms.

In a previous study we determined that the fine needle aspiration (FNA) biopsy of thyroid produces 16% of false-negative cases (FNC). In order to determine the value of quantitative cytology (QC) as a tool for predicting the FNC result, thirty-seven cases, 18 of which were operated (histologically benign: 13; malignant: 3; atypical adenoma; 2) were examined for quantitative morphologic characteristics. The smears were stained by the Feulgen method and nuclear parameters of morphometry, densitometry and texture were computed by a cell image processor. The system was first taught to recognize the benign and malignant cells from 3 histologically benign and malignant lesions. Thereafter, prospective cases were submitted to decisional analysis. For 10 histologically benign cases, the benign cell rate (bcr) ranged from 65% to 99.4 (95% confidence interval). Among the patients with a cytologically benign lesion (and an unknown histological diagnosis), 2 had a bcr less than 65% and so were not to be regarded as benign. The follow-up of these patients will show whether they represent FNC and whether QC can be of predictive value in assessing the FNC.

Epidermal growth factor receptors in breast cancer: comparison of radioligand and immunocytochemical assays.

Epidermal growth factor receptors (EGFR) are part of second generation biological factors that clinicians caring for breast cancer patients wish to evaluate for their prognostic value. This aim requires the standardization of methods: the radioligand assay (RLA) for the quantification of EGF binding sites was performed on membrane pellets from 261 breast cancer samples (ligand binding and hydroxylapatite separation as recommended by the EORTC Receptor Study Group); the immunocytochemical assay (ICA) for the staining of EGFR antigenic sites was performed on fine needle aspiration (FNA) cytology or touch imprints from 97 surgical specimens. The percentage of EGFR positivity by RLA (specific binding higher than 1% of total radioactivity) and the EGFR positive rate by ICA (more than 5% of stained cells) were respectively 43% and 38%. For 61 cases assayed on the same patient both methods revealed a concordance of 85%. Our results show that both methods are complementary and give quantitative data and information on tumor heterogeneity when they are performed in parallel. The next step of this study will be to determine the prognostic value of EGFR in these subpopulations of tumors for the adjustment of adjuvant treatment.

Cytogenetic changes in 67 cranial and spinal meningiomas: relation to histopathological and clinical pattern.

The cytogenetic analysis of 67 meningiomas (58 intracranial and 9 spinal tumors) identified chromosomal abnormalities in 63% of cases. When chromosomes involved in numerical and structural changes with a frequency of more than one standard deviation above the mean were considered, distinct cytogenetic patterns could be identified according to sex, anatomical location and histology. The chromosomes more frequently affected were 1, 2, 3, 4, 8, 14, 15, 19, 22, Y. No conclusion could be drawn regarding the prognostic significance of these karyotypic alterations.

Evaluation of the monoclonal antibody antithyroperoxidase MoAb47 in the diagnostic decision of cold thyroid nodules by fine-needle aspiration.

Fine needle aspiration (FNA) of cold thyroid nodules has become the first line diagnostic decision for electing which patients need surgery. In order to improve FNA accuracy, the monoclonal antithyroperoxidase (TPO) antibody (MoAb47) was tested. A total of 554 patients were included in this study and among them, 208 were referred to surgery. The results of FNA compared to the final histological diagnosis revealed a sensitivity and a specificity of 94 and 55% respectively, while the sensitivity and specificity of TPO immunodetection on the same cases reached 98 and 83% respectively. By combining the two methods, the 3 false-negative of FNA and 60% of suspicious cytology corresponding to histological benign lesions were correctly identified by immunocytochemistry. With better results than FNA alone, TPO immunodetection with MoAb47 represents a useful adjunct to conventional cytology for selecting patients for surgery.

Evaluation of morphological criteria in the cytological diagnosis of thyroid cold nodules. A preliminary study.

Although cytological diagnosis plays a significant role in the management of thyroid cold nodules, the rather high rates of false negative cases diminishes its usefulness. The purpose of this preliminary study is to evaluate the utility of numerous morphological criteria used by the cytologist to exclude benign tumours. Thirty-one cytological criteria were routinely scored as binary (yes/no) or as categories: 6 referred to the general arrangement and frequency of thyroid cells, 9 to the associated cellular and cell product elements, and 16 to the morphological features of the cells. We examined the manner in which these criteria, alone or combined, contributed to the diagnosis. The data base consisted of 171 intraoperative imprint cytological samples (143 histologically benign, 1 atypical adenoma and 27 cancers), as well as 257 thyroid cold nodule aspirates from another set of patients (198 histologically benign, 7 atypical adenomas and 52 cancers). For the imprint cytology, the diagnostic power of each criterion was individually assessed by the likelihood ratio (LR) which eliminated 11 as being undiscriminatory. The remaining independent criteria were subjected to logistic regression analysis to determine the most discriminant. Three were selected: Cellular clustering organisation, nuclear hypertrophy and colloid quantity with the latter being somewhat less powerful. Furthermore, it appears that the diagnostic power of the criteria was significantly lower when there was at least one nucleolus (number of nucleoli greater than 0). The smears gave essentially the same results as the imprints.(ABSTRACT TRUNCATED AT 250 WORDS)

Micro-Raman spectroscopy for optical pathology of oral squamous cell carcinoma.

Micro-Raman spectra of formalin-fixed oral squamous normal and carcinoma tissues, stored at room temperature for 2 months, have been recorded. Spectra were recorded both in the epithelial and subepithelial regions of the tissues. No noticeable spectral contamination due to formalin was observed. Very significant differences between spectra of normal epithelial and malignant epithelial samples were found. No such differences in spectra of subepithelial malignant and subepithelial normal samples could be observed. This study shows that spectra from the epithelial region changes drastically because of malignancy-induced biochemical changes in this region. Major differences between normal and malignant spectra seem to arise from the protein composition, conformational/structural changes, and possible increase in protein content in malignant epithelia. The differences between normal epithelial and subepithelial spectra, as expected, arise mainly from the collagen in subepithelial tissue. Principal component analysis of the combined sets of spectra-epithelial and subepithelial, normal and malignant- showed that very good discrimination can be achieved by Raman microspectroscopy. This study thus validates the suitability of formalin-fixed tissues for optical pathology in oral malignancy.

Comparison of benzo[a]pyrene DNA adduct formation in the skin of two strains of mice selected for resistance (DBA/2) or susceptibility (C3H/HeN) to contact dermatitis.

Two strains of mice were selected for resistance (DBA/2) or susceptibility (C3H/HeN) to contact dermatitis. Benzo[a]pyrene-DNA adduct formations was compared in the two mouse strains by a postlabeling procedure to determine if there was a significant effect. Results showed that adduct profiles in DBA/2 and C3H/HeN dermis were qualitatively similar. The total binding levels were higher in DBA/2 mice on the d 2 and the d 10. DNA adduct formation has been shown to inversely correlate with skin allergy induction. Data suggest that the expression of the genes responsible for the differences in responsiveness to chemical induced contact dermatitis in mouse may play an important role in benzo[a]pyrene-DNA adduct formation.

Molecular characterization of Charcot-Marie-Tooth patients in 15 pedigrees from France.

Molecular characterization of Charcot-Marie-Tooth patients in 15 pedigree from France: We collected 15 Charcot-Marie-Tooth (CMT) pedigrees from France. DNA polymorphisms analysis by Southern blotting with probes at the D17S122 locus demonstrated 17p duplication in three CMT1a families and in one sporadic case. Two families affected by CMT2 showed no evidence of the duplication.