Accumulation of amyloid beta in human glioblastomas

A Zayas-Santiago, A Díaz-García, R Nuñez-Rodríguez, M Inyushin, A Zayas-Santiago, A Díaz-García, R Nuñez-Rodríguez, M Inyushin

Abstract

Many cancer types are intrinsically associated with specific types of amyloidosis, in which amyloid is accumulated locally inside tumors or systemically. Usually, this condition relates to the hyperproduction of specific amylogenic proteins. Recently, we found that the accumulation of amyloid beta (Aβ) peptide immunofluorescence is linked to glioma cells in mouse tumors. Here we report that amyloid-specific histochemical dyes reveal amyloid accumulation in all human glioma samples. Application of two different antibodies against Aβ peptide (a polyclonal antibody against human Aβ1-42 and a monoclonal pan-specific mAb-2 antibody against Aβ) showed that the amyloid in glioma samples contains Aβ. Amyloid was linked to glioma cells expressing glial-specific fibrillary acidic protein (GFAP) and to glioma blood vessels. Astrocytes close to the glioma site and to affected vessels also accumulated Aβ. We discuss whether amyloid is produced by glioma cells or is the result of systemic production of Aβ in response to glioma development due to an innate immunity reaction. We conclude that amyloid build-up in glioma tumors is a part of the tumor environment, and may be used as a target for developing a novel class of anti-tumor drugs and as an antigen for glioma visualization.

Keywords: brain; cancer; human; microscopy; tumor immunology.

Conflict of interest statement

The authors declare that they have no conflicts of interest.

© 2020 The Authors. Clinical & Experimental Immunology published by John Wiley & Sons Ltd on behalf of British Society for Immunology.

Figures

Fig. 1
Fig. 1
Top row: glioma samples from three patients stained with thioflavin S (Th‐S, green fluorescence). (a) Glioma patient 17; (b) glioma patient 11; (c) glioma patient 7. These images were acquired at the same amplification. Cells with Th‐S fluorescence have different sizes in different patients. Some extracellular staining is also present, showing possible extracellular deposition. Bottom row: glioma removed from patient 1 stained with different amyloid‐detecting dyes: D = thioflavin S (Th‐S, green); E = Congo red (red); F = Coomassie brilliant blue G (BBG, red). All stains for amyloid beta peptides (Aβ) aggregates yield the same pattern of sparsely or compactly distributed cells as well as extracellular aggregates. White arrow = extracellular aggregates; black arrow = cells with amyloid; scale bar = 40 μm.
Fig. 2
Fig. 2
Top row: double immunostaining in patient 1. (a) Compactly distributed glioma cells containing amyloid beta peptides (Aβ) in the main tumor (green fluorescence in cells labeled with rabbit polyclonal antibody against human Aβ1–42) double‐stained with antibody against neuronal hexaribonucleotide binding protein‐3 (NeuN) (red). This region contains low levels of red staining. (b) Human glioma tumor periphery (normal tissue), which contains neurons labeled with NeuN antibodies (red) but almost no anti‐Aβ green staining. (c) Glioma tumor periphery (normal tissue), which contains reactive astrocytes labeled with glial‐specific fibrillary acidic protein (GFAP) antibodies (red), while some astrocytes sending endfeet to blood vessels have low levels of green + red = yellow staining or co‐localization of glial marker anti‐GFAP with anti‐Aβ. Bottom row: in patient 1, Aβ accumulates in cells of glial origin expressing GFAP. (d1) Green, polyclonal antibody against Aβ; blue, 4′,6‐diamidino‐2‐phenylindole (DAPI) nuclear marker; (d2) red, antibody against glial marker GFAP; (d3) composite. There is perfect co‐localization of the majority of Aβ‐expressing cells with GFAP‐expressing cells (see white arrows). Green = Aβ; red (in b) = NeuN (neurons); red (in c, d2, d3) = GFAP; blue (DAPI) = nuclei; scale bar, 40 μm.
Fig. 3
Fig. 3
Amyloid around blood vessels in glioma. (a) Congo red staining of amyloid (green) at low magnification (×10) showing amyloid‐containing cells concentrated around the large blood vessel (red), on the inner surface of the vessel and in the perivascular space. (b) Anti‐amyloid beta peptides (Aβ) antibody (green) shows that Aβ‐containing cells are co‐localized with glial‐specific fibrillary acidic protein (GFAP) (red)‐containing cells of glial origin around blood vessels. Some Aβ‐containing cells are present at the inner surface of the blood vessel wall and in the perivascular space, and these cells do not always coincide with GFAP. (c) Aβ‐containing cells (green) around small blood vessels in samples containing normal tissue. Some reactive astrocytes in the layer on top of the glioma cell layer had started to accumulate amyloid. Aβ is found in the blood vessel walls and in the astrocytic endfeet on the vessel walls (green + red = yellow). Arrow in B = perivascular space; blue = 4′,6‐diamidino‐2‐phenylindole (DAPI) staining (nuclei). Scale bars = 240 μm; (a) 40 μm (b,c).

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