Summary

Alzheimer’s disease (AD) is an age-related neurodegenerative disorder marked by memory loss and cognitive impairment; it is the leading cause of dementia in the elderly. Due to the paucity of biological indicators for AD diagnosis in the region, predicting and diagnosing AD, particularly in its early stages, remains challenging. This study assessed the expression of specific microRNAs (miR-146a-5p and miR-125b-5p) and telomerase genes (human telomerase reverse transcriptase (hTERT) and telomerase RNA component (TERC) as an early, non-invasive, new biomarker for the diagnosis and prediction of Alzheimer’s disease.

In the present work, 60 participants were recruited, consisting of 30 dementia included 11 mild cognitive impairment (MCI) and 19 AD and 30 non-dementia collected from private neurological clinics in Erbil city between November 2021 and March 2022. Total RNA was extracted from the plasma samples using a specific kit to include miRNAs and mRNAs isolation. A two-step RT qPCR reactions kit was used for miRNAs expression, and a One-step RT qPCR reaction kit was unitized for telomerase genes, using specific primers to target (miR-125b-5p, miR-146a-5p) for miRNA and (hTERT, TERC) gene for telomerase. The reference genes small nuclear RNA U6-2 (RNU6-2) miRNA and Beta-actin (ACTB) mRNA were used for comparison and normalization of miRNAs and telomerase gene expression. The relative quantification method calculated the fold changes in the gene expression levels between patients and healthy control.

The mini-mental state examination MMSE scores show a significant difference between the demented and non-demented groups.

The miR-146a-5p levels were significantly down-regulated in AD patients compared to healthy subjects. Similarly, the miR-125b-5p was detected at a low expression level in the dementia cohort. The diagnostic value of miRNAs and telomerase differed between people with dementia and those without dementia. It was estimated by Receiver operating characteristic (ROC) curve analysis; to detect the area under the curve AUC= 0.737 and 0.841, respectively. The AUC of miR-146a-5p and miR-125b-5p revealed that miRNAs are considered promising diagnostic biomarkers for early AD prediction. The positive correlation coefficient was detected between the relative expression of miR-146a-5p and miR-125b-5p of AD cases with MMSE scores.

The relative RT-qPCR results show that hTERT and TERC gene expression is significantly down-regulated in the Alzheimer’s cohort compared to the non-dementia subject. The correlation coefficient analysis revealed a positive correlation of the telomerase gene expression of AD cases with MMSE scores. The area under curve AUC was 0.773 for hTERT and 0.703 for TERC, indicating a good diagnostic value for AD prediction.

Finally, our findings revealed that alterations in circulating levels of miR-146a-5p and miR-125b-5p might serve as early non-invasive diagnostic markers for the prediction of AD patients. Also, it shows down-regulation in hTERT and TERC gene expression levels in AD. We propose that telomerase gene expression in circulation can serve as a non-invasive, early, and novel diagnostic marker of AD.