NADPH-diaphorase histochemistry identifies isolated endothelial cells at sites of traumatic injury in the adult rat brain

• Published in: Neuroscience Vol. 53; no. 3; pp. 613 - 624
• Main Authors: Kitchener, P.D., Bourreau, J.-P., Diamond, J.
• Format: Journal Article
• Language: English
• Published: Oxford Elsevier Ltd 01.04.1993; Elsevier
• Subjects: Animals; Antibodies, Monoclonal - immunology; Biological and medical sciences; Brain - pathology; Brain Injuries - pathology; Cerebral Cortex - pathology; Endothelium - cytology; Female; Histocytochemistry; Lectins; Macrophages - ultrastructure; Medical sciences; NADPH Dehydrogenase - analysis; Nervous system involvement in other diseases. Miscellaneous; Neuroglia - ultrastructure; Neurology; Plant Lectins; Rats; Rats, Wistar; NADPH-d; BSI-B 4; NOS; Immunohistochemistry; Nervous system diseases; Endothelial cell; Rat; Enzyme; Rodentia; Biological marker; NADPH dehydrogenase; Trauma; Pathology; Angiogenesis; Vertebrata; Mammalia; Mononuclear cell; Animal; Neovascularization; Brain (vertebrata)
• ISSN: 0306-4522; 1873-7544
• DOI: 10.1016/0306-4522(93)90610-R

In addition to labelling endothelium, some ependymal cells (including tanycytes), and a subpopulation of neurons, nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase histochemistry of stab lesion sites in the neocortex revealed a large population of cells concentrated within several hundred micrometres of the lesion site. To determine the identity of these cells, NADPH-diaphorase reactivity was compared to binding with either the I-B 4 isolectin from Bandeiraea simplicifolia (which has previously been shown to identify endothelial cells and activated mononuclear phagocytes), or a monoclonal antibody (OX-42) that recognizes activated mononuclear phagocytes. Many I-B 4 lectin-labelled cells were also NADPH-diaphorase reactive, and other I-B 4 lectin-labelled cells were also OX-42 immunoreactive, but co-existence of OX-42 immunoreactivity and NADPH-diaphorase reactivity was not observed. Only a small minority of NADPH-diaphorase-reactive cells did not exhibit I-B 4 lectin binding. In contrast to the simple somatic morphology of the majority of NADPH-diaphorase-reactive cells, the I-B 4 lectin-negative cells had a ramified appearance, and while readily observed at two days postlesion, they were only rarely seen at three days postlesion. Primary cultures of bovine aortic endothelial cells also exhibited NADPH-diaphorase reactivity which occupied most of the cytoplasm in a filamentous web pattern. Endothelial cells possess a constitutive form of nitric oxide synthase which, as demonstrated in NADPH-diaphorase-reactive neurons, may be the basis of their NADPH-diaphorase reactivity. These findings indicate that NADPH-diaphorase-reactive cells observed at lesion sites are probably angiogenic endothelial cells not associated with extant blood vessels. Thus, NADPH-diaphorase histochemistry offers an effective method of visualizing neovascularization in the brain and other tissues.