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Localization of stromelysin gene expression in atherosclerotic plaques by in situ hybridization.

ABSTRACT: The stromelysins are members of a family of extracellular matrix metalloproteinases. These enzymes may erode the connective tissue in atherosclerotic plaques, leading to fissuring and acute thrombotic events. Cell-specific stromelysin expression in human atherosclerotic plaques was studied by in situ hybridization and immunocytochemistry. Sections were taken from nine coronary arteries: eight with well-established plaques and one normal. Unambiguous signals were seen in five plaques, two were inconclusive, and the remaining sample was negative, as was the normal coronary artery. Stromelysin mRNA transcripts were localized to isolated individual cells, some of which were smooth muscle, in the plaque cap, intima, and adventitia, but not the media. Expression was also seen in large clusters of macrophages that contained intracellular lipid deposits. The isolated expression of stromelysin by smooth muscle cells may reflect local connective tissue remodeling associated with growth and the formation of the plaque, whereas the more extensive expression associated with macrophages may be of greater pathological significance, contributing to the destabilization of the extracellular matrix and eventual plaque rupture.

SUBMITTER: Henney AM

PROVIDER: S-EPMC52465 | biostudies-other | 1991 Sep

REPOSITORIES: biostudies-other

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Localization of stromelysin gene expression in atherosclerotic plaques by in situ hybridization.

Henney A M AM Wakeley P R PR Davies M J MJ Foster K K Hembry R R Murphy G G Humphries S S

Proceedings of the National Academy of Sciences of the United States of America 19910901 18

The stromelysins are members of a family of extracellular matrix metalloproteinases. These enzymes may erode the connective tissue in atherosclerotic plaques, leading to fissuring and acute thrombotic events. Cell-specific stromelysin expression in human atherosclerotic plaques was studied by in situ hybridization and immunocytochemistry. Sections were taken from nine coronary arteries: eight with well-established plaques and one normal. Unambiguous signals were seen in five plaques, two were in ...[more]

PMID: 1896464

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Project description:With the advances in genomics research of the past decade, plant biology has seen numerous studies presenting large-scale quantitative analyses of gene expression. Microarray and next generation sequencing approaches are being used to investigate developmental, physiological and stress response processes, dissect epigenetic and small RNA pathways, and build large gene regulatory networks(1-3). While these techniques facilitate the simultaneous analysis of large gene sets, they typically provide a very limited spatiotemporal resolution of gene expression changes. This limitation can be partially overcome by using either profiling method in conjunction with lasermicrodissection or fluorescence-activated cell sorting(4-7). However, to fully understand the biological role of a gene, knowledge of its spatiotemporal pattern of expression at a cellular resolution is essential. Particularly, when studying development or the effects of environmental stimuli and mutants can the detailed analysis of a gene's expression pattern become essential. For instance, subtle quantitative differences in the expression levels of key regulatory genes can lead to dramatic phenotypes when associated with the loss or gain of expression in specific cell types. Several methods are routinely used for the detailed examination of gene expression patterns. One is through analysis of transgenic reporter lines. Such analysis can, however, become time-consuming when analyzing multiple genes or working in plants recalcitrant to transformation. Moreover, an independent validation to ensure that the transgene expression pattern mimics that of the endogenous gene is typically required. Immunohistochemical protein localization or mRNA in situ hybridization present relatively fast alternatives for the direct visualization of gene expression within cells and tissues. The latter has the distinct advantage that it can be readily used on any gene of interest. In situ hybridization allows detection of target mRNAs in cells by hybridization with a labeled anti-sense RNA probe obtained by in vitro transcription of the gene of interest. Here we outline a protocol for the in situ localization of gene expression in plants that is highly sensitivity and specific. It is optimized for use with paraformaldehyde fixed, paraffin-embedded sections, which give excellent preservation of histology, and DIG-labeled probes that are visualized by immuno-detection and alkaline-phosphatase colorimetric reaction. This protocol has been successfully applied to a number of tissues from a wide range of plant species, and can be used to analyze expression of mRNAs as well as small RNAs(8-14).

Dataset Information

Localization of stromelysin gene expression in atherosclerotic plaques by in situ hybridization.

Publications

Localization of stromelysin gene expression in atherosclerotic plaques by in situ hybridization.

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