The brain has a limited repertoire of response to injury and disease. Microglial activation is the cornerstone of most of the brains protective responses which are usually associated with inflammatory processes. Aberrant microglial activation in Multiple Sclerosis (MS), stroke, severe head trauma and Alzheimer's Disease (AD) can directly contribute to neurotoxicity and tissue destruction through release of reactive oxygen species, proteases, cytokines and excitatory neurotransmitters. As one of the reactive oxygen species released from microglia and macrophages, NITRIC OXIDE has been reported to be toxic in some, and protective in other experimental paradigms of CNS disease. In peripheral tissues, the induction of nitric oxide release appears to contribute to rheumatoid arthritis and inflammatory bowel disease. Of the 3 different Nitric Oxide Synthases, the inducible form of NOS, known as iNOS or NOS2, is mainly found in microglia and macrophages. The induction of iNOS activity to release nitric oxide is highly regulated and is also regulated in a species specific manner. We have confirmed and extended this by finding that the amyloid-beta peptide (Abeta) associated with AD can stimulate iNOS activity in mouse cells, but fails to stimulate human cells to release nitric oxide. We also find that apolipoprotein- E, another gene associated with Alzheimer s, does stimulate human cells to release nitric oxide. To develop a better animal model of oxyradical in a fashion similar to that found in humans, we specifically propose to make a """"""""Humanized-NOS2"""""""" transgenic mouse expressing only human iNOS2 enzyme. We have recently isolated a PAC clone containing the human NOS2 gene and we propose to insert this piece of genomic DNA into fertilized mouse oocytes by standard injection techniques and mate the resulting human-NOS2 transgenic mice lacking the mouse-NOS2 gene (aka. NOS2-knockout mice, Jackson labs Bar Harbor, ME). The net result will be a transgenic mouse which express human-NOS2 enzyme and whose regulation of nitric release, will be tested for its similitary to the human-specific pattern. Such a mouse model will have broad utility to more accurately predict the role played by oxyradicals in the development of human disease.
| Colton, C A; Chernyshev, O N; Gilbert, D L et al. (2000) Microglial contribution to oxidative stress in Alzheimer's disease. Ann N Y Acad Sci 899:292-307 |
