Role of Complement Factor H Polymorphism and Diet in Neuroinflammation
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BACKGROUND: Multiple lines of evidence point towards an important role for complement factor H (Cfh) in neuroinflammation. Evidence of activated microglia and activated astrocytes has been found in the brains of both Parkinson's disease and Alzheimer's disease patients.1 In addition, Cfh has been shown to be present in amyloid beta plaques in Alzheimer's disease.2 Our laboratory has developed a mouse model of early AMD based on expressing variant Cfh molecules in mice.3 The Cfh transgenic mice develop deposits under the retinal pigment epithelium, which resemble early changes seen on AMD. We believe these findings indicate that the variant Cfh molecules are less able to control chronic low grade inflammation at the tissue level. METHODS: Young (6m old) and aging (18m old) Cfh transgenic and control B6 mice were divided into groups and fed either a control diet or a high-fat diet for 5 months. Brains were collected after perfusion with 2% PFA/PBS and were post-fixed overnight in 4% PFA/PBS. Next, the brains were transferred to a 30% sucrose solution, weighed, and sectioned. Immunohistochemistry was then performed on 30 μm brain sections with antibodies specific for inflammatory, oxidative stress, and microglial markers. Stained images were visualized using a Leica fluorescence microscope with an objective lens of 10x. The microglia over the photographed field were counted and averaged in order to obtain a cells/field value for each mouse. Statistical analysis was then performed on the data. RESULTS: CfhTg brains weighed less than the corresponding B6 brains (0.447 vs. 0.482g; p=0.00024). Many of the tested antibodies, particularly those associated with oxidative stress, did not stain the sections, perhaps due to the fixation method. However, the anti-TREM2 and anti-Iba 1 antibodies stained well. There was no difference in the number of Iba-1+ microglia in the dentate gyrus of CfhTg vs. B6 mice (p=0.607). Younger mice seemed to have higher numbers of these cells compared to older mice (73.3 vs. 52.6 microglia/field; p=0.0352). In addition, the mice fed a high fat diet appeared to have less microglia per field compared to the mice fed a normal control diet (53.1 vs. 72.8 microglia/field; p=0.0469). TREM2 is considered to be a marker for microglial activation. Neither age (p=0.65), nor a high fat diet (p=0.435) appeared to affect the level of TREM2 expression. However, there was a trend towards higher numbers of TREM2+ cells in CfhTg mice compared to B6 mice (p=0.17), particularly in the old group of mice (p=0.12). CONCLUSIONS: Reduced brain mass in CfhTg mice suggests increased CNS oxidative stress and tissue injury. There was no difference in the number of Iba-1+ microglial cells in CfhTg vs B6 mice. However, there was a trend towards increased microglial activation in Tg mice. More brains will be analyzed with alternative methods of tissue collection and additional antibodies to corroborate these findings.