[Identification and validation of plasma biomarkers for mild cognitive impairment based on mRNA N6-methyladenosine methylome analysis].

Objective

To identify and validate differential mRNA N6-methyladenosine (m6A) methylation sites associated with mild cognitive impairment (MCI), providing potential molecular evidence for its early prevention and intervention.

Methods

A case-control study was conducted among older adults recruited from multiple community health centers in Shenzhen, including a pilot cohort (5 pairs) and a validation cohort (25 pairs). Single-based m6A quantitative polymerase chain reaction was used to quantify plasma mRNA m6A methylation levels. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses were performed, followed by the construction of a protein-protein interaction network to identify hub genes. The differential methylation levels of key genes were validated in an expanded cohort. For normally distributed data, t-tests were used; for non-normal data, Mann-Whitney U or χ2 tests were applied. Partial correlation analysis and Firth logistic regression were conducted to verify the associations further.

Results

A total of 126 differential mRNA m6A methylation sites were identified, including 73 upregulated and 53 downregulated sites. Five hub genes (ZFHX3, PARD3, ARHGEF2, GLB1, and LRIG3) were selected, based on protein-protein interaction network topology. Validation experiments showed that the mRNA m6A methylation levels of these five genes were significantly higher in the MCI group than in healthy controls (all P<0.001). After adjustment for potential confounders, partial correlation analysis revealed that the methylation levels of all five genes were negatively correlated with MMSE scores (r=-0.719- -0.426, all P<0.05). Firth logistic regression confirmed that these genes were independently associated with MCI (OR=1.57-2.88, all P<0.001).

Conclusion

This study screened and verified a range of MCI-related differential m6A methylation sites, suggesting that abnormal m6A methylation of specific gene mRNAs may be associated with MCI, providing research clues for further exploration of the molecular mechanism of MCI and potential plasma molecular markers.