The Potential Role Of Plasma MicroRNA As Biomarkers For Amnestic Mild Cognitive Impairment And Their Modulation On BACE1

OBJECT:To explore the potential diagnostic value of plasma micro RNAs（miRNAs）as biomarkers for amnestic mild cognitive impairment（a MCI）,and to explore the modulation of miRNAs onβ-site amyloid precursor protein cleaving enzyme 1（BACE1）.METHOD:Mild cognitive impairment（MCI）subjects were selected from the China Longitudinal Aging Study.Normal controls（NC）were age-,gender-,and educational years-matched elderly.Plasma samples were collected from all participants.The miRNA expression profile was performed in discovery set(five NC and five MCI due to Alzheimer’s diasease subjects that confirmed by[18F]-3’-F-PIB-PET using microarray sequencing.Target genes of differentially expressed miRNAs were predicted by bioinformatics analyses.Combined the fold change value and P value,we selected five miRNAs into advanced validation.In analysis set（20 a MCI and 10 NC）,real-time quantitative PCR（q PCR）was used to verify the miRNA sequencing results.Significantly down-regulated miRNAs in a MCI group and miR-107 were selected as candidate biomarkers.In validation set（40 a MCI and 40 NC）,we analyzed the correlation between candidate miRNAs and clinical phenotype,and explored the diagnostic value of these candidates.Human neuroblastoma cells which stably expressed BACE1 protein(SH-SY5Y^BACE1)was transfected with miRNA inhibitor or mimics lentivirus to knock-down or over-expressed the corresponding miRNA,then BACE1 protein level was measured by Western Blot（WB）.Primary hippocampal neurons from SD rat were transfected with inhibitor or mimics lentivirus,then BACE1 m RNA and protein levels were measured by q PCR and WB separately.Human primary hippocampal neurons were transfected with miRNA inhibitor lentivirus,then BACE1protein level was measured by WB.RESULTS:We identified 13 significantly up-regulated and 31 down-regulated human miRNAs in plasma of MCI due to AD subjects.Bioinformatics analyses revealed that BACE1 was one of the target genes of 15 significantly down-regulated miRNAs.The optimal five miRNAs（miR-5691,miR-1185-2-3p,miR-1909-3p,miR-22-5p,and miR-134-3p）were selected for further verification.Except miR-5691,the other four miRNAs which were significantly decreased in a MCI and miR-107 which was found to decline in AD previously,were selected as candidates.Five miRNAs together had an outstanding diagnostic accuracy with sensitivity and specificity was 80.00%and 82.50%separately.Except for miR-134-3p,other four miRNAs modulate BACE1 expression and C-terminal fragments-β（CTF-β）production effectively in rat primary hippocampal neurons in vitro.Similar modulation of miR-1185-2-3p and miR-1909-3p was testified in human hippocampal neurons in vitro.CONCLUSIONS:Combination of five miRNAs（miR-1185-2-3p,miR-1909-3p,miR-22-5p,miR-134-3p,and miR-107）had outstanding diagnostic value in a MCI.These miRNAs regulated BACE1 expression effectively.Overexpression of four miRNAs suppressed BACE1 and CTF-βexpression,suggesting their potential therapeutic value.