Microglia Express Insulin-Like Growth Factor-1 in the Hippocampus of Aged APPswe/PS1ΔE9 Transgenic Mice.

Insulin-like growth factor-1 (IGF-1) is a pleiotropic molecule with neurotrophic and immunomodulatory functions. Knowing the capacity of chronically activated microglia to produce IGF-1 may therefore show essential to promote beneficial microglial functions in Alzheimer's disease (AD). Here, we investigated the expression of IGF-1 mRNA and IGF-1 along with the expression of tumor necrosis factor (TNF) mRNA, and the amyloid-ß (Aß) plaque load in the hippocampus of 3- to 24-month-old APPswe/PS1ΔE9 transgenic (Tg) and wild-type (WT) mice. As IGF-1, in particular, is implicated in neurogenesis we also monitored the proliferation of cells in the subgranular zone (sgz) of the dentate gyrus. We found that the Aß plaque load reached its maximum in aged 21- and 24-month-old APPswe/PS1ΔE9 Tg mice, and that microglial reactivity and hippocampal IGF-1 and TNF mRNA levels were significantly elevated in aged APPswe/PS1ΔE9 Tg mice. The sgz cell proliferation decreased with age, regardless of genotype and increased IGF-1/TNF mRNA levels. Interestingly, IGF-1 mRNA was expressed in subsets of sgz cells, likely neuroblasts, and neurons in both genotypes, regardless of age, as well as in glial-like cells. By double in situ hybridization these were shown to be IGF1 mRNA+ CD11b mRNA+ cells, i.e., IGF-1 mRNA-expressing microglia. Quantification showed a 2-fold increase in the number of microglia and IGF-1 mRNA-expressing microglia in the molecular layer of the dentate gyrus in aged APPswe/PS1ΔE9 Tg mice. Double-immunofluorescence showed that IGF-1 was expressed in a subset of Aß plaque-associated CD11b+ microglia and in several subsets of neurons. Exposure of primary murine microglia and BV2 cells to Aß42 did not affect IGF-1 mRNA expression. IGF-1 mRNA levels remained constant in WT mice with aging, unlike TNF mRNA levels which increased with aging. In conclusion, our results suggest that the increased IGF-1 mRNA levels can be ascribed to a larger number of IGF-1 mRNA-expressing microglia in the aged APPswe/PS1ΔE9 Tg mice. The finding that subsets of microglia retain the capacity to express IGF-1 mRNA and IGF-1 in the aged APPswe/PS1ΔE9 Tg mice is encouraging, considering the beneficial therapeutic potential of modulating microglial production of IGF-1 in AD.

Diverse Protein Profiles in CNS Myeloid Cells and CNS Tissue From Lipopolysaccharide- and Vehicle-Injected APPSWE/PS1ΔE9 Transgenic Mice Implicate Cathepsin Z in Alzheimer's Disease.

Neuroinflammation, characterized by chronic activation of the myeloid-derived microglia, is a hallmark of Alzheimer's disease (AD). Systemic inflammation, typically resulting from infection, has been linked to the progression of AD due to exacerbation of the chronic microglial reaction. However, the mechanism and the consequences of this exacerbation are largely unknown. Here, we mimicked systemic inflammation in AD with weekly intraperitoneal (i.p.) injections of APPSWE/PS1ΔE9 transgenic mice with E. coli lipopolysaccharide (LPS) from 9 to 12 months of age, corresponding to the period with the steepest increase in amyloid pathology. We found that the repeated LPS injections ameliorated amyloid pathology in the neocortex while increasing the neuroinflammatory reaction. To elucidate mechanisms, we analyzed the proteome of the hippocampus from the same mice as well as in unique samples of CNS myeloid cells. The repeated LPS injections stimulated protein pathways of the complement system, retinoid receptor activation and oxidative stress. CNS myeloid cells from transgenic mice showed enrichment in pathways of amyloid-beta clearance and elevated levels of the lysosomal protease cathepsin Z, as well as amyloid precursor protein, apolipoprotein E and clusterin. These proteins were found elevated in the proteome of both LPS and vehicle injected transgenics, and co-localized to CD11b+ microglia in transgenic mice and in primary murine microglia. Additionally, cathepsin Z, amyloid precursor protein, and apolipoprotein E appeared associated with amyloid plaques in neocortex of AD cases. Interestingly, cathepsin Z was expressed in microglial-like cells and co-localized to CD68+ microglial lysosomes in AD cases, and it was expressed in perivascular cells in AD and control cases. Taken together, our results implicate systemic LPS administration in ameliorating amyloid pathology in early-to-mid stage disease in the APPSWE/PS1ΔE9 mouse and attract attention to the potential disease involvement of cathepsin Z expressed in CNS myeloid cells in AD.

Reduced Serotonin Transporter Levels and Inflammation in the Midbrain Raphe of 12 Month Old APPswe/PSEN1dE9 Mice.

BACKGROUND: Although mood and sleep disturbances are nearly universal among patients with Alzheimer's disease (AD), brain structures involved in non-cognitive processing remain under characterized in terms of AD pathology. OBJECTIVES: This study was designed to evaluate hallmarks of AD pathology in the brainstem of the APPswe/PS1dE9 mouse model of familial AD. METHODS: Fresh-frozen sections from female, 12 month old, transgenic and control B6C3 mice (n=6/genotype) were examined for amyloid burden and neurofibrillary alterations, by using 6E10 immunohistochemistry and the Gallyas silver stain, respectively. Serotonin transporter (SERT) densities in the dorsal and the median raphe were quantified by [3H]DASB autoradiography. SERT mRNA expression was measured by RT-PCR and visualized by in situ hybridization. Neuroinflammation was evaluated by immunohistochemical staining for microglia and astrocytes, and by measuring mRNA levels of the proinflammatory cytokines TNF-α, IL-1ß and IL-6. RESULTS: No amyloid- and tau-associated lesions were observed in the midbrain raphe of 12 month old APPswe/PS1dE9 mice. SERT binding levels were reduced in transgenic animals compared to age-matched controls, and SERT mRNA levels were decreased by at least 50% from control values. Intense microglial, but not astrocytic immunoreactivity was observed in APPswe/PS1dE9 vs. wild-type mice. Levels of TNF-α mRNA were two-fold higher than control and correlated positively with SERT mRNA expression levels in transgenic animals. CONCLUSIONS: There was no amyloid accumulation and tau-associated pathology in the midbrain raphe of 12 month old APPswe/PS1dE9 mice. However, there was a local neuroinflammatory response with loss of serotonergic markers, which may partially account for some of the behavioral symptoms of AD.

Cytokine-producing microglia have an altered beta-amyloid load in aged APP/PS1 Tg mice.

Beta-amyloid (Aß) plaques and chronic neuroinflammation are significant neuropathological features of Alzheimer's disease. Microglial cells in aged brains have potential to produce cytokines such as TNF and IL-1 family members (IL-1α, IL-1ß, and IL-1Ra) and to phagocytose Aß in Alzheimer's disease, however the inter-relationship between these processes is poorly understood. Here we show that % Aß plaque load followed a sigmoidal trajectory with age in the neocortex of APPswe/PS1ΔE9 Tg mice, and correlated positively with soluble Aß40 and Aß42. Aß measures were moderately correlated with mRNA levels of CD11b, TNF, and IL-1Ra. Cytokine production and Aß load were assessed in neocortical CD11b(+)(CD45(+)) microglia by flow cytometry. Whereas most microglia in aged mice produced IL-1Ra, relatively low proportions of microglia produced TNF, IL-1α, and IL-1ß. However, microglial production of these latter cytokines was generally increased in APP/PS1 Tg mice. Microglia that phagocytosed endogenously-produced Aß were only observed in APP/PS1 Tg mice. Differences in phagocytic index and total Aß load were observed in microglia with specific cytokine profiles. Both phagocytic index and total Aß load were higher in IL-1α(+) and IL-1Ra(+) microglia, than microglia that did not produce these cytokines. In contrast, total Aß load was lower in IL-1ß(+) and TNF(+) microglia, compared to IL-1ß(-) and TNF(-) microglia, and TNF(+) microglia also had a lower phagocytic index. Using GFP bone marrow chimeric mice, we confirmed that the majority of neocortical CD11b(+)(CD45(+)) microglia were resident cells (GFP(-)) in APP/PS1 Tg mice, even after selectively analysing CD11b(+)CD45(high) cells, which are typically considered to be infiltrating cells. Together, our data demonstrate that cytokine expression is selectively correlated with age and Aß pathology, and is associated with an altered Aß load in phagocytic microglia from APP/PS1 Tg mice. These findings have implications for understanding the regulation of microglial cytokine production and phagocytosis of Aß in Alzheimer's disease.