Abnormal endothelial tight junctions in active lesions and normal-appearing white matter in multiple sclerosis

Blood-brain barrier (BBB) breakdown, demonstrable in vivo by enhanced MRI is characteristic of new and expanding inflammatory lesions in relapsing remitting and chronic progressive multiple sclerosis (MS). Subtle leakage may also occur in primary progressive MS. However, the anatomical route(s) of BBB leakage have not been demonstrated. We investigated the possible involvement of interendothelial tight junctions (TJ) by examining the expression of TJ proteins (occludin and ZO-1 ) in blood vessels in active MS lesions from 8 cases of MS and in normal-appearing white (NAWM) matter from 6 cases. Blood vessels (10-50 per frozen section) were scanned using confocal laser scanning microscopy to acquire datasets for analysis. TJ abnormalities manifested as beading, interruption, absence or diffuse cytoplasmic localization of fluorescence, or separation of junctions (putative opening) were frequent (affecting 40% of vessels) in oil red-O-positive active plaques but less frequent in NAWM (15%), and in normal (

Absence of aquaporin-4 expression in lesions of neuromyelitis optica but increased expression in multiple sclerosis lesions and normal-appearing white matter.

Aquaporin-4 (AQP4) has recently been implicated in the pathogenesis of neuromyelitis optica(NMO) where it has been identifed as the first defined autoantigen pertinent to an infammatory demyelinating disorder of the human CNS. Furthermore, a recent case report has shown a lack of AQP4 expression in the spinal cord lesions of NMO. However, the pattern of AQP4 expression in multiple sclerosis (MS) tissues has not been well-defned. In the present investigation we have confirmed a lack of expression of AQP4 in optic and spinal cord lesions in NMO which contrasted sharply with the increased levels of AQP4 expression seen in MS lesions. Furthermore a detailed immunohistochemical and semi-quantitative analysis is used to describe the expression pattern of AQP4 on well-characterized tissue microarray samples of MS and control white matter. Anatomically AQP4 was more highly expressed in all categories of MS tissue compared to normal control tissues with the most abundant expression in active lesions. Within active lesions AQP4 expression was significantly correlated with expression of the pro-infammatory cytokine osteopontin. At the cellular level dual-labelling immunofluoresence demonstrated that increased expression of AQP4 was most pronounced at the astrocytic endfeet but was also associated with the cell bodies of astrocytes in the tissue parenchyma. The finding of increased AQP4 expression in MS lesions in contrast to the lack of expression in NMO lesions may suggest different mechanisms of initiation and progression between the two disease states.

An automated machine vision system for the histological grading of cervical intraepithelial neoplasia (CIN)

The histological grading of cervical intraepithelial neoplasia (CIN) remains subjective, resulting in inter- and intra-observer variation and poor reproducibility in the grading of cervical lesions. This study has attempted to develop an objective grading system using automated machine vision. The architectural features of cervical squamous epithelium are quantitatively analysed using a combination of computerized digital image processing and Delaunay triangulation analysis; 230 images digitally captured from cases previously classified by a gynaecological pathologist included normal cervical squamous epithelium (n = 30), koilocytosis (n = 46), CIN 1 (n = 52), CIN 2 (n = 56), and CIN 3 (n=46). Intra- and inter-observer variation had kappa values of 0.502 and 0.415, respectively. A machine vision system was developed in KS400 macro programming language to segment and mark the centres of all nuclei within the epithelium. By object-oriented analysis of image components, the positional information of nuclei was used to construct a Delaunay triangulation mesh. Each mesh was analysed to compute triangle dimensions including the mean triangle area, the mean triangle edge length, and the number of triangles per unit area, giving an individual quantitative profile of measurements for each case. Discriminant analysis of the geometric data revealed the significant discriminatory variables from which a classification score was derived. The scoring system distinguished between normal and CIN 3 in 98.7% of cases and between koilocytosis and CIN 1 in 76.5% of cases, but only 62.3% of the CIN cases were classified into the correct group, with the CIN 2 group showing the highest rate of misclassification. Graphical plots of triangulation data demonstrated the continuum of morphological change from normal squamous epithelium to the highest grade of CIN, with overlapping of the groups originally defined by the pathologists. This study shows that automated location of nuclei in cervical biopsies using computerized image analysis is possible. Analysis of positional information enables quantitative evaluation of architectural features in CIN using Delaunay triangulation meshes, which is effective in the objective classification of CIN. This demonstrates the future potential of automated machine vision systems in diagnostic histopathology. Copyright (C) 2000 John Wiley and Sons, Ltd.

Bcl-2 expression in sequential biopsies of potentially malignant oral mucosal lesions assessed by immunocytochemistry

OBJECTIVE: To examine, for the first time Bcl-2 expression in sequential (autogenous) oral mucosal biopsies taken from the same sites in a gender, risk-factor matched, Caucasoid sample, over a 21-year period,