Rapid communication3D-Reconstruction of microglia and amyloid in APP23 transgenic mice: no evidence of intracellular amyloid
Introduction
The formation of amyloid plaques in the brain is a principal feature of Alzheimer’s disease [13], [24], [34]. These extracellular amyloid deposits are associated with activated glial cells [6], [8], [9], [14], [16] which produce a large variety of proinflammatory cytokines and cytotoxic mediators [1], [3]. Thus, a chronic inflammatory process occurs in the vicinity of plaques during the course of the disease and many of the detrimental effects of amyloid deposition may be the result of this secondary inflammation [1], [12], [18]. In line with this hypothesis, epidemiologic studies reported a reduced risk for AD in patients using nonsteroidal anti-inflammatory drugs [17], [26].
In addition to their role in inflammation, microglia have been thought to contribute to amyloidosis by participating in the formation [5], [21], [29], [30] and/or phagocytosis of amyloid [2], [19], [31]. These hypotheses arose from electron microscopic studies reporting that amyloid fibrils are often completely engulfed in microglial cytoplasm whose surface membranes form deep infoldings filled with amyloid fibers [22], [30]. This observation indicated that β-amyloid (Aβ) is either produced or taken up by microglial cells surrounding a plaque.
In order to better understand the relationship between microglia cells and amyloid in AD, transgenic mouse models of cerebral amyloidosis have been intensively studied in recent years. Very similar to AD, a robust activation and proliferation of microglia in the vicinity of congophilic amyloid deposits with production of various inflammatory mediators have been reported in APP transgenic mice [6], [11], [25]. Interestingly, a recent study demonstrated that ibuprofen, a nonsteroidal anti-inflammatory drug, not only suppresses inflammation, but also significantly decreases amyloid deposition in APP transgenic mice [15]. Thus, the analysis of transgenic mouse models of AD promises new insights into basic mechanisms underlying the interaction between microglia and amyloid. In the present study, we reinvestigate the role of microglia in cerebral amyloidosis by using serial immunogold electron microscopy and three-dimensional reconstruction analysis of microglia and compact amyloid plaques in APP23 transgenic mice.
Section snippets
Animals
The generation of APP23 transgenic mice has been described elsewhere [27]. Briefly, a murine Thy-1 promoter element was used to drive neuron-specific expression of mutated human APP751 (Swedish double mutation) in B6D2 mice. Six 15–20 months old hemizygous APP23 mice and two age-matched nontransgenic controls were used. The mice were from the F8-F10 generation of backcrossing to B6 mice.
Tissue preparation
Mice were injected intraperitoneally (i.p.) by an overdose of pentobarbital (50 mg/ml Nembutal; Abbott
Dense core plaques are surrounded by hypertrophic microglia
We used an antibody to the microglia cell surface marker CD11b to study microglia in the brains of APP23 transgenic mice. Light microscopic examination revealed that clusters of activated and hypertrophic microglia are associated with dense core amyloid deposits throughout the neocortex (Fig. 1a). Analysis of semithin sections confirmed the tight association between microglia and compact amyloid. Microglia cell bodies were found in close vicinity of the amyloid with microglia processes
Discussion
We have previously described a close association between activated microglia and compact amyloid plaques in the brains of adult and aged APP23 transgenic mice [25]. In the present study, we have extended these observations using serial ultrathin sectioning, postembedding immunogold labeling, and 3D-reconstruction of microglia cells in the vicinity of a dense core amyloid plaque. Our data demonstrate that amyloid fibrils are exclusively extracellular deposits that form a myriad of finger-like
Acknowledgements
We thank A. Schneider (University of Freiburg, Germany) for excellent experimental help. We would also like to acknowledge the support of C. Sturchler-Pierrat, D. Abramowski, K.H. Wiederhold (Novartis, Basel, Switzerland), M. Pfeifer, A. Probst (Institute of Pathology, Basel, Switzerland), M. Dürrenberger (Biocenter, University of Basel, Switzerland), and M. Frotscher (University of Freiburg, Germany). This study was supported by Grants 31-56753.99 from the Swiss National Science Foundation,
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These authors contributed equally to this work.