Reduced serum miR-202 may promote the progression of Alzheimer's disease patients via targeting amyloid precursor protein

Abstract

The current study investigated whether the expression of miR-202 was abnormal in the serum of patients with Alzheimer's disease (AD) and evaluated the potential clinical significance, thereby shedding light on the diagnosis of AD. Here, our data showed that the level of miR-202 decreased significantly in the serum of AD patients (n = 121) compared with that of healthy controls (n = 86). Further analysis showed that the level of serum miR-202 was gradually decreased in the mild AD group (n = 31), moderate AD group (n = 52) and severe AD group (n = 38) compared with the healthy control group. Receiver operating characteristic (ROC) curve analysis demonstrated that serum miR-202 could differentiate AD patients from healthy controls, with an AUC of 0.892. Spearman correlation analysis showed that serum miR-202 was positively correlated with the Mini-Mental State Examination (MMSE). Based on TargetScan, a conserved binding site was identified in the 3'UTR of amyloid precursor protein (APP). The dual luciferase assay showed that miR-202 suppressed the relative luciferase activity of pmirGLO-APP-3'UTR. Western blot assays indicated that overexpression of miR-202 suppressed the expression of APP, while the expression of APP was enhanced after inhibition of miR-202 in PC12 cells, indicating that APP was a possible target gene of miR-202. Moreover, the cell apoptosis induced by transfection of miR-202 inhibitor was abolished by silencing APP. In summary, we showed novel data that downregulation of serum miR-202 may be used as a potential biomarker for AD and may promote the development of AD by targeting APP.

Keywords: Alzheimer's disease; amyloid precursor protein; diagnosis; miR-202.

FIGURE 1

Serum miR‐202 was reduced in the serum of Alzheimer's disease (AD) patients compared with that of healthy controls. (A) Reverse transcription polymerase chain reaction (RT‐PCR) analysis showed that the level of miR‐202 was decreased in the serum of AD patients. (B) The level of serum miR‐202 was significantly decreased based on the severity of AD. ***p < 0.001 versus control. ##p < 0.01, ###p < 0.001 versus as indicated

FIGURE 2

Spearman correlation analysis showed that serum miR‐202 positively correlated with the Mini‐Mental State Examination (MMSE) in Alzheimer's disease (AD) patients

FIGURE 3

Receiver operating characteristic (ROC) analysis demonstrated that serum miR‐202 could differentiate Alzheimer's disease (AD) patients from healthy controls

FIGURE 4

miR‐202 reduced PC12 cell apoptosis and increased cell viability. (A) In the Aβ‐induced PC12 cellular Alzheimer's disease (AD) model, the level of miR‐202 was decreased compared with the control. (B) The MTT assay showed that overexpression of miR‐202 reduced the toxicity and side effects of Aβ. (C) TUNEL staining showed that overexpression of miR‐202 can reduce the apoptotic rate induced by Aβ. *p < 0.05, **p < 0.01 versus control, #p < 0.05 versus as indicated

FIGURE 5

Amyloid precursor protein (APP) was a target gene of miR‐202. (A) Based on TargetScan, a conserved binding site was identified in the 3′UTR of APP. (B) Dual luciferase assays showed that miR‐202 suppressed the relative luciferase activity of pmirGLO‐APP‐3′UTR. (C) Reverse transcription polymerase chain reaction (RT‐PCR) analysis showed that transfection with miR‐202 mimic significantly increased the relative level of miR‐202. (D) Western blot assays indicated that overexpression of miR‐202 suppressed the expression of APP in PC12 cells. (E) Transfection with miR‐202 inhibitor significantly suppressed the level of miR‐202. (F) The expression of APP was enhanced after inhibition of miR‐202 in PC12 cells. **p < 0.01, ***p < 0.001 versus control

FIGURE 6

Silencing amyloid precursor protein (APP) abolished miR‐202 inhibition‐induced cell apoptosis. (A) Western blot analysis showed that the expression of APP was obviously reduced by si‐APP, even in PC12 cells transfected with miR‐202 inhibitor. (B) TUNEL staining demonstrated that miR‐202 inhibition‐induced PC12 cell apoptosis could be largely reversed by knockdown of APP. **p < 0.01, ***p < 0.001 versus control