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Microglia Endocytose Amyloid β Through the Binding of Transglutaminase 2 and Milk Fat Globule EGF Factor 8 Protein

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Abstract

Activation of glial cells has been observed in neurodegenerative diseases including Alzheimer’s disease (AD). Aggregation of amyloid β (Aβ) is profusely observed as characteristic pathology in AD brain. In our previous study using microglial cell line BV-2, tissue-type transglutaminase (TG2) was found to be involved in phagocytosis (Kawabe et al., in Neuroimmunomodulation 22(4):243–249, 2015; Kawabe et al., Neurochem Res 2017). In the present study, we examined whether TG2 and milk fat globule EGF factor 8 protein (MFG-E8), an adaptor protein promotes macrophage to engulf apoptotic cells, were involved in Aβ endocytosis. When the neuronal/glial mixed culture was stimulated freshly prepared Aβ1−42 for 3 days, the incorporation of Aβ was observed by immunofluorescence staining technique in Iba-1-positive microglia. Cystamine, a broad competitive inhibitor of TGs, suppressed it. When aggregated Aβ was added to the mixed culture, the immunoreactivity of MFG-E8 surrounding Aβ was observed, and then followed by microglial endocytosis. Using western blotting technique, MFG-E8 was detected in cell lysate of astrocyte culture, and was also detected in the medium. When microglia culture was incubated with astrocyte conditioned medium, MFG-E8 levels in microglia tended to increase. It is likely that microglia might utilize MFG-E8 released from astrocytes as well as that expressed in themselves in order to endocytose Aβ aggregation. Furthermore, we confirmed that MFG-E8 could bind with TG2 in microglia culture by immunoprecipitate technique. These results suggest that microglia might uptake Aβ as a complex of aggregated Aβ/MFG-E8/TG2.

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Abbreviations

Aβ:

Amyloid beta

ACM:

Astrocytes conditioned medium;

AD:

Alzheimer’s disease

BSA:

Bovine serum albumin

CBB:

Coomassie brilliant blue

CNS:

Central nervous system

DMEM:

Dulbecco’s modified Eagle medium

DMSO:

Dimethyl sulfoxide

EGF:

Epidermal growth factor

FBS:

Fetal bovine serum

FITC:

Fluorescein isothiocyanate

GFAP:

Glial fibrillary acidic protein

HRP:

Horseradish peroxidase

MFG-E8:

Milk fat globule EGF factor 8 protein

LPS:

Lipopolysaccharide

NO:

Nitric oxide

PBS:

Phosphate-buffered saline

PS:

Phosphatidylserine

TG:

Transglutaminase

VR:

Vitronectin receptor.

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Acknowledgements

This work was supported in part by JSPS KAKENHI Grant No. JP15J12259 to K.K., JP26850209 to K.T., JP26450447 to M.M., and JP15K07768 to Y.N.

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Authors and Affiliations

  1. Laboratory of Integrative Physiology in Veterinary Sciences, Osaka Prefecture University, 1-58, Rinku-Ourai Kita, Izumisano, Osaka, 598-8531, Japan

    Kenji Kawabe, Katsura Takano, Mitsuaki Moriyama & Yoichi Nakamura

Authors
  1. Kenji Kawabe
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  2. Katsura Takano
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  3. Mitsuaki Moriyama
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  4. Yoichi Nakamura
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Correspondence to Katsura Takano.

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Kawabe, K., Takano, K., Moriyama, M. et al. Microglia Endocytose Amyloid β Through the Binding of Transglutaminase 2 and Milk Fat Globule EGF Factor 8 Protein. Neurochem Res 43, 41–49 (2018). https://doi.org/10.1007/s11064-017-2284-y

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  • DOI: https://doi.org/10.1007/s11064-017-2284-y

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