This APP has been developed to quantify the occurrence of immunohistochemically detectable 5-lipoxygenase (5LO) in atherosclerotic plaques, irrespective of its cellular or extracellular location.
Lipoxygenases are a family of iron-containing enzymes that catalyze the deoxygenation of polyunsaturated fatty acids. 5LO is expressed in inflammatory cells of the myeloid line, such as histiocytes (including foam cells), dendritic reticular cells, mast cells and granulocytes. In resting myeloid cells, 5LO resides in the cytoplasm or the nucleoplasm, but when activated relocates to the nuclear membrane. 5LO is involved in the metabolism of the cell membrane derived arachidonic acid leading to formation of 4-series leukotrienes (LTs) which are major contributors to inflammatory reactions.
In arteriosclerotic disease LTs may enhance the formation of atherosclerotic plaques as well as plaque instability by effects on leukocyte chemotaxis, vascular inflammation and subsequent matrix degeneration. Important clinical endpoints have been studied in genetic association studies, where polymorphisms in the 5LO encoding gene (ALOX5) have been linked to increased risk of myocardial infarction, stroke and increased carotid intima and media thickness.
The expression of 5LO in advanced atherosclerotic plaque seems to be co-localized with CD68 positive macrophages/foam cells/dendritic reticular cells. Apart from mast cells in adventitia, other myeloid cell types contribute very little to the 5LO positive population in the arterial wall. 5LO-positive histiocytes localize to the adventitia in areas of neoangiogenesis and constitute a main component of arteriosclerotic aneurysms.
The rabbit monoclonal antibodies G590.2 (Thermo Scientific) and C49G1 (Cell Signaling) are both useful. Heat induced epitope retrieval (HIER) is mandatory; an alkaline buffer should be preferred. For the Ventana platform, HIER should be followed by a short (3-4 min) proteolytic pre-treatment.
As control tissues, tonsil and liver are useful: In the tonsil, mantle zone B-lymphocytes must show a moderate nuclear reaction, and in the liver, the Kupffer cells must show a moderate cytoplasmic reaction.