An AIE-Active NIR Fluorescent Probe with Good Water Solubility for the Detection of Aβ1-42 Aggregates in Alzheimer's Disease

Abstract

Alzheimer's disease (AD), an amyloid-related disease, seriously endangers the health of elderly individuals. According to current research, its main pathogenic factor is the amyloid protein, which is a kind of fibrillar aggregate formed by noncovalent self-assembly of proteins. Based on the characteristics of aggregation-induced emission (AIE), a bislactosyl-decorated tetraphenylethylene (TPE) molecule TMNL (TPE + malononitrile + lactose), bearing two malononitrile substituents, was designed and synthesized in this work. The amphiphilic TMNL could self-assemble into fluorescent organic nanoparticles (FONs) with near-infrared (NIR) fluorescence emission in physiological PBS (phosphate buffered saline), achieving excellent fluorescent enhancement (47-fold) upon its combination with Aβ_1-42 fibrils. TMNL was successfully applied to image Aβ_1-42 plaques in the brain tissue of AD transgenic mice, and due to the AIE properties of TMNL, no additional rinsing process was necessary. It is believed that the probe reported in this work should be useful for the sensitive detection and accurate localization mapping of Aβ_1-42 aggregates related to Alzheimer's disease.

Keywords: AIE; Alzheimer’s disease; Aβ; amyloid; fluorescence; lactose; near-infrared imaging; tetraphenylethylene.

Scheme 1

The synthesis route of TMNL.

Figure 1

The UV absorption spectra and fluorescence emission spectra of TMNL.

Figure 2

(a) The fluorescence spectra of different concentrations (μM) of TMNL in PBS buffer solution (pH 7.4, 10 mM). (b) Scatter plot of fluorescence intensity at 645 nm of different concentrations of TMNL in PBS buffer solution (pH 7.4, 10 mM). λ_ex = 360 nm.

Figure 3

Fluorescence spectra of the excitation (a) and emission (b) of ThT (1 μM)-detecting Aβ_1–42 (80 μg·mL⁻¹) fibrils in PBS buffer solution (pH 7.4, 10 mM). λ_ex = 410 nm, λ_em = 470 nm.

Figure 4

(a) Fluorescence spectra of the interaction of TMNL with Aβ_1–42 fibrils after different coincubation times in PBS buffer solution (pH 7.4, 10 mM). Insert: photograph of TMNL after complete interaction with Aβ_1–42 fibrils (left) and in the absence of Aβ_1–42 fibrils (right) in quartz cuvettes under 365 nm UV light. (b) Scatter plot of the relative fluorescence intensity (I/I₀) at 496 nm of the different coincubation times of TMNL and Aβ_1–42 fibrils in PBS buffer solution (pH 7.4, 10 mM). I and I₀ represent the fluorescence intensity in the presence and absence of Aβ_1–42 fibrils, respectively. [TMNL] = 1 μM, [Aβ_1–42] = 80 μg·mL⁻¹, λ_ex = 360 nm.

Figure 5

(a) Fluorescence spectra of the interaction of TMNL with different concentrations of Aβ_1–42 fibrils (μg·mL⁻¹) in PBS buffer solution (pH 7.4, 10 mM). (b) Scatter plot of the relative fluorescence intensity (I/I₀) at 496 nm of the interaction of TMNL and different concentrations of Aβ_1–42 fibrils (μg·mL⁻¹) in PBS buffer solution (pH 7.4, 10 mM). I and I₀ represent the fluorescence intensity in the presence and absence of Aβ_1–42 fibrils, respectively. [TMNL] = 1 μM, λ_ex = 360 nm.

Figure 6

(a) The UV absorption spectra of different concentrations of TMNL in PBS buffer solution (pH 7.4, 10 mM). (b) UV absorption spectra of the interaction of TMNL (1 μM) with different concentrations of Aβ_1–42 fibrils (μg·mL⁻¹) in PBS buffer solution (pH 7.4, 10 mM).

Figure 7

(a) Fluorescence spectra of the interaction of TMNL with different interfering substances and Aβ_1–42 fibrils in PBS buffer solution (pH 7.4, 10 mM). (b) The histogram of relative fluorescence intensity (I/I₀) at 496 nm of the interaction of TMNL with different interfering substances and Aβ_1–42 fibrils in PBS buffer solution (pH 7.4, 10 mM). I represents the fluorescence intensity in the presence of the interfering substances or Aβ_1–42 fibrils, and I₀ represents the fluorescence intensity in the presence of TMNL alone. [Cys] = [GSH] = [HSO₃⁻] = [SO₃²⁻] = [CN⁻] = 100 μM, [BSA] = [Aβ_1–42] = 80 μg·mL⁻¹; [TMNL] = 1 μM; λ_ex = 360 nm.

Figure 8

Saturation binding curves of different concentrations of TMNL interacting with Aβ_1–42 fibrils. [Aβ_1–42] = 80 μg·mL⁻¹; λ_ex = 360 nm.

Figure 9

(a,b) TEM images of TMNL in PBS buffer solution (pH 7.4, 10 mM). (c,d) TEM images of TMNL after binding to Aβ_1–42 fibrils in PBS buffer solution (pH 7.4, 10 mM). [TMNL] = 1 μM; [Aβ_1–42] = 80 μg·mL⁻¹.

Figure 10

(a,b) Fluorescence staining images of TMNL in brain tissue sections of AD transgenic mice.