Increased extracellular amyloid deposition and neurodegeneration in human amyloid precursor protein transgenic mice deficient in receptor-associated protein - PubMed (original) (raw)

Increased extracellular amyloid deposition and neurodegeneration in human amyloid precursor protein transgenic mice deficient in receptor-associated protein

Emily Van Uden et al. J Neurosci. 2002.

Abstract

The low-density lipoprotein receptor-related protein (LRP) is an abundant neuronal cell surface receptor that regulates amyloid beta-protein (Abeta) trafficking into the cell. Specifically, LRP binds secreted Abeta complexes and mediates its degradation. Previously, we have shown in vitro that the uptake of Abeta mediated by LRP is protective and that blocking this receptor significantly enhances neurotoxicity. To further characterize the effects of LRP and other lipoprotein receptors on Abeta deposition, an in vivo model of decreased LRP expression, receptor-associated protein (RAP)-deficient (RAP-/-) mice was crossed with human amyloid protein precursor transgenic (hAPP tg) mice, and plaque formation and neurodegeneration were analyzed. We found that, although the age of onset for plaque formation was the same in hAPP tg and hAPP tg/RAP-/- mice, the amount of amyloid deposited doubled in the hAPP tg/RAP-/- background. Moreover, these mice displayed increased neuronal damage and astrogliosis. Together, these results further support the contention that LRP and other lipoprotein receptors might be neuroprotective against Abeta toxicity and that this receptor might play an integral role in Abeta clearance.

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Figures

Fig. 1.

Fig. 1.

Characterization of APP and LRP expression in hAPP tg/RAP−/− mice. A, Ribonuclease protection assay. A total of four mice per group was analyzed. Representative autoradiogram for mAPP and hAPP is shown. The_leftmost_ lane represents the undigested (U) radiolabeled probes; the other_lanes_ contain the same riboprobes plus brain RNA samples digested with RNases. Protected mRNAs are indicated on the_right_. The hAPP, mAPP, and actin mRNA bands were detected as fragments of 230, 180, and 85 nt, respectively.B, Analysis of levels of hAPP and mAPP mRNA expression in tg mice; results are expressed as a ratio of APP to actin. Error bars are mean ± SEM. C, Western blot analysis with antibodies against hAPP and LRP. The hAPP-specific antibody identified a broad band at an approximate molecular weight (MW) of 110 kDa. The specific antibody against the C-terminal region of LRP detected a band at an approximate MW of 85 kDa.D, Analysis of hAPP and LRP immunoreactivity; results are expressed as integrated pixel intensity. Error bars are mean ± SEM.

Fig. 2.

Fig. 2.

Immunocytochemical analysis of hAPP, RAP, and LRP expression in hAPP tg/RAP−/− mice. A–D, hAPP immunoreactivity with the 8E5 monoclonal antibody; E–H, RAP immunoreactivity in the frontal cortex of 18-month-old mice;I–L, LRP immunoreactivity in the frontal cortex of 18-month-old mice. A, No hAPP immunoreactivity was observed in non-tg mice. B, hAPP tg mice displayed immunostaining of a subset of pyramidal neurons in the neocortex.C, No hAPP immunoreactivity was observed in RAP−/− mice. D, hAPP tg/RAP−/− tg mice hAPP-immunoreactive pyramidal neurons in the neocortex. E, Non-tg;F, hAPP tg mice showed extensive immunostaining of pyramidal neurons in the neocortex. In the RAP−/− (G) and hAPP tg/RAP−/− (H), no RAP immunoreactivity was observed. Non-tg (I) and hAPP tg (J) mice showed extensive LRP immunostaining of pyramidal neurons in the neocortex. In the RAP−/− (G) and hAPP tg/RAP−/− (H), no LRP levels of LRP immunoreactivity were decreased. Scale bar, 25 μm.

Fig. 3.

Fig. 3.

Patterns of Aβ immunoreactivity in hAPP tg/RAP−/−. A–D, Aβ immunoreactivity in the neocortex; E–H, low-power view (60×) of the hippocampal dentate gyrus; I–L, higher-power view (200×) of the dentate gyrus; M, P, GFAP immunoreactivity in the hippocampus. All images are from 18-month-old mice. A, No Aβ deposits were observed in non-tg controls. B, Mature and diffuse amyloid plaques in hAPP tg/RAP tg mice. C, No Aβ deposits were observed in RAP−/− mice. D, hAPP tg/RAP−/− mice displayed increased Aβ immunoreactive plaques. Scale bar, 20 μm.E, I, No Aβ deposits were observed in the molecular layer of the dentate in non-tg controls.F, J, Abundant diffuse amyloid plaques in hAPP tg mice. G, K, No Aβ deposits were observed in RAP−/− mice. H, L, hAPP tg/RAP−/− mice displayed increased Aβ deposits in the molecular layer of the dentate. Scale bar, 60 μm. M, Mild astrogliosis in non-tg mice. N, Increased astroglial reaction in hAPP mice. O, Mild astroglial immunostaining in RAP−/− mice. P, Enhanced astroglial reaction in hAPP tg/RAP−/− mice. Scale bar, 30 μm.

Fig. 4.

Fig. 4.

Quantitative analysis of amyloid production and neurodegeneration. A, Determination of percentage area of the neuropil occupied by Aβ immunoreactive deposits in the frontal cortex and hippocampus. B, Determination of Aβ levels in the hippocampus by ELISA in 10- and 18-month-old mice. *p < 0.05, different by two-tailed unpaired_t_ test when compared with hAPP tg mice.C, Densitometrical analysis of GFAP immunoreactivity in the hippocampus using the Quantimet 570C in 10- and 18-month-old mice. *p < 0.05 different from non-tg control by one-way ANOVA post hoc Dunnet's test. D, Levels of MAP2-IR in the outer molecular layer of the dentate gyrus in 10- and 18-month-old mice. *p < 0.05, different by one-way ANOVA post hoc Dunnet's test.

Fig. 5.

Fig. 5.

Patterns of dendritic degeneration in hAPP tg/RAP−/−. Sections from 18-month-old mice were immunolabeled with an antibody against MAP2 and imaged with the laser scanning confocal microscope. A–D, Neocortex; E–H, molecular layer of the hippocampal dentate gyrus. Scale bar, 20 μm.

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