Quercetin-Conjugated Superparamagnetic Iron Oxide Nanoparticles Protect AlCl3-Induced Neurotoxicity in a Rat Model of Alzheimer's Disease via Antioxidant Genes, APP Gene, and miRNA-101

Abstract

Alzheimer's disease (AD) is a neurodegenerative disease with cognitive impairment. Oxidative stress in neurons is considered as a reason for development of AD. Antioxidant agents such as quercetin slow down AD progression, but the usage of this flavonoid has limitations because of its low bioavailability. We hypothesized that quercetin-conjugated superparamagnetic iron oxide nanoparticles (QT-SPIONs) have a better neuroprotective effect on AD than free quercetin and regulates the antioxidant, apoptotic, and APP gene, and miRNA-101. In this study, male Wistar rats were subjected to AlCl₃, AlCl₃ + QT, AlCl₃ + SPION, and AlCl₃ + QT-SPION for 42 consecutive days. Behavioral tests and qPCR were used to evaluate the efficiency of treatments. Results of behavioral tests revealed that the intensity of cognitive impairment was decelerated at both the middle and end of the treatment period. The effect of QT-SPIONs on learning and memory deficits were closely similar to the control group. The increase in expression levels of APP gene and the decrease in mir101 led to the development of AD symptoms in rats treated with AlCl₃ while these results were reversed in the AlCl₃ + QT-SPIONs group. This group showed similar results with the control group. QT-SPION also decreased the expression levels of antioxidant enzymes along with increases in expression levels of anti-apoptotic genes. Accordingly, the antioxidant effect of QT-SPION inhibited progression of cognitive impairment via sustaining the balance of antioxidant enzymes in the hippocampus of AD model rats.

Keywords: AlCl3; Alzheimer’s disease; antioxidant; miR-101; quercetin; superparamagnetic iron oxide nanoparticle.

FIGURE 1

Timeline of experiments. MWM, Morris water maze; qRT-PCR, quantitative real-time polymerase chain reaction.

FIGURE 2

Characterization of QT-SPION. (A) Scanning electron microscopy (SEM) image for QT-SPION. (B) SEM-EDX spectrum for QT-SPION. Arrows show the existence of Fe in nanocomposition. (C) FTIR spectra of dextran-coated SPION (a), QT (b), and QT-SPION (c). (D) XRD pattern obtained for dextran-coated SPION (a) and QT-SPION (b). QT-SPION, quercetin-superparamagnetic conjugate; FTIR, Fourier transform infrared; XRD: X-ray diffraction.

FIGURE 3

Results of one-way ANOVA for (A) body weight of rats in six groups at 1, 14, 28, and 42 days of treatment. (B) Liver weight of rats after sacrifice on the 42nd day. (C) Brain weight of rats after sacrifice on the 42nd day. Data are presented as mean ± SEM (n = 8); ^∗∗∗p < 0.001, ^∗∗p < 0.01, and ^∗p < 0.05 are statistically different in comparison with all groups. ANOVA, analysis of variance.

FIGURE 4

Effect of QT-SPION on rats treated with AlCl₃ simultaneously in the MWM test. Test results are presented as mean ± SEM (n = 8; p < 0.001): (A) The escape latency during the acquisition phase including four training days of treatment (17th, 18th, 19th, and 20th day of treatment): a significant increase in AlCl₃ group in comparison with control and AlCl3 + QT-SPION groups. (B) The time spent in the target zone at both probe trial days (21st and 42nd): shorter time spent by the AlCl₃ group compared to control and AlCl3 + QT-SPION groups. (C) The number of plate crosses in the AlCl₃ group is significantly more than that in the control group. Data are presented as mean ± SEM (n = 8); ^∗∗∗p < 0.001, ^∗∗p < 0.01, and ^∗p < 0.05 are statistically different in comparison with all groups. SEM, standard error of mean; MWM, Morris water maze.

FIGURE 5

Effect of QT-SPION treatment on step-through latency of rats using passive avoidance in AlCl₃-treated groups. Data are presented as mean ± SEM (n = 8); ^∗∗∗p < 0.001 in comparison with all groups. (A) Step-through latency on the training day. (B) Time spent in the white box on days 21 and 42. (C) Retention latency via cross number between boxes on days 21 and 42. AlCl₃, aluminum chloride; QT-SPION, quercetin-superparamagnetic iron oxide nanoparticles; SEM, standard error of mean.

FIGURE 6

(A) Effect of QT-SPION treatment on acetylcholinesterase activity in the hippocampus of AlCl₃-treated rats with induced oxidative stress. (B) Concentration of Al³⁺ ions in the hippocampus of rats in AlCl₃, AlCl₃ + SPION, AlCl₃ + QT, and AlCl₃ + QT-SPION groups. (C) The concentration of iron in the hippocampus of rats in AlCl₃ + SPION and AlCl₃ + QT-SPION groups. Data are presented as mean ± SEM (n = 8); ^∗∗∗p < 0.001, ^∗∗p < 0.01, and ^∗p < 0.05 are statistically different in comparison with all groups. QT-SPION, quercetin-superparamagnetic iron oxide nanoparticle; AlCl₃, aluminum chloride; SEM, standard error of mean; ICP, inductively coupled plasma mass spectroscopy.

FIGURE 7

Effect of QT-SPION treatment on relative gene expression levels in the hippocampus of rats treated with AlCl₃. (A) A significant decrease in expression levels of APP in the hippocampus of AlCl₃ + QT-SPION group rats in comparison with the AlCl₃ group. (B) A significant increase in expression levels of mir101 in the hippocampus of AlCl₃ + QT-SPION group rats in comparison with the AlCl₃ group. (C) A significant decrease in expression levels of iNOS in the hippocampus of AlCl₃ + QT-SPION group rats in comparison with the AlCl₃ group. (D) A significant increase in expression levels of SOD1 in the hippocampus of AlCl₃ + QT-SPION group rats in comparison with the AlCl₃ group. (E) A significant increase in expression levels of GPX1 in the hippocampus of AlCl₃ + QT-SPION group rats in comparison with the AlCl₃ group. (F) A significant increase in expression levels of CAT in the hippocampus of AlCl₃ + QT-SPION group rats in comparison with the AlCl₃ group. (G) A significant increase in expression levels of BCL2 in the hippocampus of AlCl₃ + QT-SPION group rats in comparison with the AlCl₃ group. (H) A significant increase in expression levels of BAX in the hippocampus of AlCl₃ + QT-SPION group rats in comparison with the AlCl₃ group. Data are expressed as mean ± SEM (n = 8); ^∗∗∗p < 0.001, ^∗∗p < 0.01, and ^∗p < 0.05 are statistically different in comparison with all groups. QT-SPION, quercetin-superparamagnetic iron oxide nanoparticle; AlCl₃, aluminum chloride; SEM, standard error of mean; APP, amyloid precursor protein; mir101, microRNA 101; iNOS, inducible nitric oxide synthase; SOD1, superoxide dismutase 1; GPX1, glutathione peroxidase 1; CAT, catalase.