Profiles of Circulating Exosomal microRNAs in Female College Students with Qi Stagnation and Balanced Constitutions by High-Throughput Sequencing.

Introduction: Individual constitutions are classified into nine types in traditional Chinese medical theory, and qi stagnation constitution (QSC) manifests as disrupted Qi circulation and increased susceptibility to emotional disorders and cancers. However, as a pre-disease state mainly affecting women, the biological basis of QSC and its susceptible mechanism to related diseases are still unclear. Exosomal microRNAs (miRNAs) are the stable regulators of gene expression and intercellular communication, and analysis of miRNAs enables us to understand the QSC better. This study profiles plasma exosomal miRNAs in QSC and balanced constitution (BC) females via high-throughput sequencing, aiming to identify the potential biomarkers of QSC and reveal its biological basis and the mechanism of its susceptible disease.

Methods: In this cross-sectional observation, female college students were recruited according to the criterion of QSC and BC in Classification and Determination of Constitution in TCM. Exosomal miRNAs were isolated from blood plasma and then profiled using high-throughput sequencing. Differentially expressed miRNAs (DEMs) were identified with fold change > 2 and P < 0.05, and screened as biomarkers to construct the receiver operating characteristic (ROC) curve. The diagnostic values of these biomarkers in different types of cancers were also validated based on the published data. KEGG and GO functional analysis were explored based on the predicted target genes.

Results: Subjects with QSC showed significantly higher concentrations of albumin (ALB) and alkaline phosphatase (ALP) compared to those with BC, while there was no significant difference in baseline information and other clinical indicators between groups. A total of 54 DEMs were identified, including 30 30 up-regulated and 24 down-regulated miRNAs in the QSC group. The area under the ROC curve (AUC) for 7 specific up-regulated DEMs was 1.0, as well as the AUCs for therein 6 DEMs in various cancers were all above 0.9. The enriched KEGG pathways included "signal transduction," "infectious disease," and "cancers", and the most associating systems included immune, endocrine, and nervous systems, while the GO function was mainly enriched in "protein binding," "nucleus" and "transcription, DNA-templated".

Discussion: These 7 potential biomarkers of QSC have been confirmed to regulate oncogenic processes through epithelial-mesenchymal transition modulation and metabolic reprogramming, as well as therein 1 can also improve depression by lowering the expression of 5-hydroxytryptamine 1A receptor. The results of this study deepen the understanding of the traditional Chinese medicine constitutions. However, the small single-sex sample limits the application of the conclusion, and a large-scale clinical cohort including both sexes is still needed in future.

Conclusion: The expression of exosomal miRNAs in QSC showed unique features that have the potential to serve as biomarkers, and the related functional changes might be the biological basis for the susceptible diseases of QSC.