Interactions between amyloid-β and hemoglobin: implications for amyloid plaque formation in Alzheimer's disease - PubMed (original) (raw)

Interactions between amyloid-β and hemoglobin: implications for amyloid plaque formation in Alzheimer's disease

Jia-Ying Chuang et al. PLoS One. 2012.

Abstract

Accumulation of amyloid-β (Aβ) peptides in the brain is one of the central pathogenic events in Alzheimer's disease (AD). However, why and how Aβ aggregates within the brain of AD patients remains elusive. Previously, we demonstrated hemoglobin (Hb) binds to Aβ and co-localizes with the plaque and vascular amyloid deposits in post-mortem AD brains. In this study, we further characterize the interactions between Hb and Aβ in vitro and in vivo and report the following observations: 1) the binding of Hb to Aβ required iron-containing heme; 2) other heme-containing proteins, such as myoglobin and cytochrome C, also bound to Aβ; 3) hemin-induced cytotoxicity was reduced in neuroblastoma cells by low levels of Aβ; 4) Hb was detected in neurons and glial cells of post-mortem AD brains and was up-regulated in aging and APP/PS1 transgenic mice; 5) microinjection of human Hb into the dorsal hippocampi of the APP/PS1 transgenic mice induced the formation of an envelope-like structure composed of Aβ surrounding the Hb droplets. Our results reveal an enhanced endogenous expression of Hb in aging brain cells, probably serving as a compensatory mechanism against hypoxia. In addition, Aβ binds to Hb and other hemoproteins via the iron-containing heme moiety, thereby reducing Hb/heme/iron-induced cytotoxicity. As some of the brain Hb could be derived from the peripheral circulation due to a compromised blood-brain barrier frequently observed in aged and AD brains, our work also suggests the genesis of some plaques may be a consequence of sustained amyloid accretion at sites of vascular injury.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1

Figure 1. Interaction between hemoglobin (Hb) and Aβ.

Freshly dissolved Aβ peptides (50 µM) were incubated with an equal mass of Hb (3.5 µM) at 37°C for 0, 1, 3 and 7 d. A) Aβ1–40; B) Aβ1–42. Left panels, anti-Aβ; right panels, anti-Hb-β chain.

Figure 2

Figure 2. Heme is the Aβ binding moiety of hemoglobin (Hb).

A) Interactions between Aβ and various part of Hb. B) Hemin induced the Aβ oligomerization in a dose-dependent fashion. C) Hemin induced the Aβ oligomerization in a time-dependent fashion. Incubation times: 0, 6, 24 hours. D) Aβ binds to Hb, myoglobin (MyoG) and cytochrome C (CytoC).

Figure 3

Figure 3. Effects of Aβ on hemin induced cytotoxicity.

A) Differentiated neuroblastoma cells, SH-SY5Y, were treated with different doses of pre-incubated Aβ1–42 without (open bars) or with 0.25 µM of hemin (closed bars). B) Differentiated SH-SY5Y cells were treated with different doses of hemin without (open bars) or with 0.25 µM of pre-incubated Aβ1–42 (closed bars). The cell viability was determined by MTT reduction assay. Bonferroni's post-hoc test: * p<0.05 vs. respective hemin group.

Figure 4

Figure 4. The expression of hemoglobin (Hb) in APP/PS1 transgenic (Tg) mice and wild-type (Wt) littermates.

A) Representative immuno-micrographs reveal the intensities of Hb-β+ stains in the coronal brain sections of a 3-month-old and a 14-month-old Tg and Wt mice, respectively. Insert: enlarged micrograph shows the presence of Hb-β+ stains in a plaque-like structure and some brain cells. B) Quantitative results of the optical densities of Hb-β+ stains in primary motor, hippocampus and entorhinal cortex of <5-month-old and >8-month-old Tg and Wt mice. Bonferroni's post-hoc test: * p<0.05 vs. respective Wt group; # p<0.05, ## p<0.01 vs. respective <5-month-old group.

Figure 5

Figure 5. The levels of hemoglobin (Hb)-α and Hb-β in <5-month-old and >8-month-old APP/PS1 transgenic (Tg) mice and wild-type (Wt) littermates.

Representative immunoblots are shown on the top of each panel whereas quantitative results are shown in the respective lower panel. The levels of Hb monomer (∼16 kDa) are quantified by immunoblotting analyses. Bonferroni's post-hoc test: * p<0.05, ** p<0.01, vs. respective Wt group; ## p<0.01 vs. respective <5-month-old group.

Figure 6

Figure 6. Localization of hemoglobin (Hb) immunoreactivity in brains of APP/PS1 transgenic (Tg) mice.

Double immunofluorescent micrographs reveal the presence of Hb-β+ stains in Aβ+ amyloid plaques and some brain cells. Scale bars: 50 µm. Examples of enlarged micrographs are shown in the right. Arrowheads point to Hb-β+, but APP/Aβ− cells; while, arrows show APP/Aβ+, but Hb-β− cells.

Figure 7

Figure 7. The distribution of hemoglobin (Hb) immunoreactivity in inferior temporal gyrus of post-mortem human brain.

Hb-β+ stains are present in most NeuN+ neurons and OSP+ oligodendrocytes; while only a small fractions of GFAP+ astrocytes and Iba-1+ microglia express Hb. Scale bars: 10 µm.

Figure 8

Figure 8. Hemoglobin (Hb) induces Aβ accumulation in the immediate surroundings of Hb.

Human Hb (1 µl, 50 µg/µl) was injected into the dorsal hippocampus of 4-month-old APP/PS1 transgenic (Tg) mice. One week after the injection of Hb or saline, the brain sections were double stained for Hb-β and Aβ. Tg-negative: The sections were stained only by Hb antibodies, but not Aβ antibodies and serve as negative controls for Aβ staining. Scale bars: 30 µm.

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References

    1. Evin G, Weidemann A. Biogenesis and metabolism of Alzheimer's disease Abeta amyloid peptides. Peptides. 2002;23:1285–1297. - PubMed
    1. Selkoe DJ. Alzheimer's disease: genes, proteins, and therapy. Physiol Rev. 2001;81:741–766. - PubMed
    1. Hardy JA, Higgins GA. Alzheimer's disease: the amyloid cascade hypothesis. Science. 1992;256:184–185. - PubMed
    1. Hardy J. Alzheimer's disease: the amyloid cascade hypothesis: an update and reappraisal. J Alzheimers Dis. 2006;9:151–153. - PubMed
    1. Kayed R, Head E, Thompson JL, Mclntire TM, Milton SC, et al. Common structure of soluble amyloid oligomers implies common mechanism of pathogenesis. Science. 2003;300:486–489. - PubMed

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