| Objective:To investigate the value of exosomal circular RNAs(exo-circ RNAs)in the diagnosis,treatment and prognosis of myocardial injury associated with multiple myeloma(MM)by high-throughput sequencing,bioinformatics analysis,clinical correlation analysis and cell experiment,the possible mechanism of myocardial injury induced by abnormal exo-circ RNAs expression in MM patients was further investigated.Methods:Referring to the TCM diagnostic efficacy criteria for heart-kidney yang deficiency and heart qi deficiency in thoracic palsy self-designed TCM evidence scoring scale for patients with heart-kidney yang deficiency multiple myeloma.5 healthy controls(HC)and 20 MM patients who met the criteria of deficiency of heart-kidney yang deficiency were included.Extract the serum exosomes of 6 MM patients and 5 HC,and obtain the differently expressed exo-circ RNAs spectrum by high-throughput sequencing.Real-time quantitative PCR(q RT-PCR)verified the abnormal expression of circ-G042080 in the serum secretion of 20 MM patients.Correlation of clinical characteristics of serum exosome circ-G042080 with MM-related myocardial injury in MM patients using ROC curves,one-way COX regression models and multi-way COX regression models.Prediction and bioinformatics of differential expression of exo-circ RNAs and downstream target genes using Gene Ontology(GO),Kyoto Encyclopedia of Genes and Genomes(KEGG),mi Randa and Target Scan informatics analysis.The circ RNA-mi RNA-m RNA interaction network was constructed based on the predictions,and the predictions were validated by dual luciferase assays.U266 cell-derived exosomes were co-incubated with H9C2 cells to explore the effect of circG042080 overexpression on cardiomyocytes and to verify the mechanism.Results:1.High-throughput sequencing analysis showed that a total of 1035 significantly up-regulated circ RNAs and 507 significantly down-regulated circ RNAs were identified in the serum exosomes of MM patients with heartkidney yang deficiency compared with the HC group.q RT-PCR verified the abnormally high expression of circ-G042080 in the serum exosomes of 20 MM patients.2.Clinical correlation analysis showed that circ-G042080 expression levels were positively correlated with troponin T(TNT)and pro-B-type natriureticpeptide(pro BNP)levels and negatively correlated with ventricular ejection fraction(EF)and systolic blood pressure(SPB)in MM patients with heart-kidney yang deficiency.3.The GO and KEGG enrichment analysis of the abnormally expressed circ RNAs showed that the up-regulated exo-circ RNAs were closely related to the progression of MM and the pathogenesis of cardiac injury,mainly manifested in the enrichment of " activation of transmembrane receptor protein tyrosine kinase activity","P38 signal pathway","PI3K-Akt signal pathway","glutamate receptor pathway" and "calcium ion pathway".Target gene prediction and functional enrichment analysis revealed that circG042080 overexpressed in exosomes of MM patients with heart-kidney yang deficiency may interfere with downstream TLR4 expression through ce RNA mechanism to induce MM-related myocardial injury.The existence of this ce RNA mechanism was further confirmed by the dual luciferase assay.4.Compared with the control group,H9C2 cells co-incubated with U266 cell exosomes showed significantly higher levels of circ-G042080 and TLR4,significantly lower levels of hsa-mi R-4268,significantly lower cell viability,higher expression levels of autophagy-related proteins LC3 and Beclin1,lower expression levels of P62,and significantly higher numbers of intracellular autophagic vesicles,all of which effects were reversed after blocking exosomes with GW4869.Conclusion:The serum of MM patients with heart-kidney-yang deficiency type contains a large number of abnormally expressed exo-circ RNAs,of which circ-G042080 expression level is significantly elevated,which can be used as a novel biomarker for clinical diagnosis and an independent prognostic indicator for MM-related myocardial injury patients with heart-kidney-yang deficiency type.In addition,exo-circ-G042080 may be involved in the occurrence and progression of MM-related myocardial injury through the circ-G042080/hsa-mi R-4268/TLR4 axis,and is a potential therapeutic target for MM-related myocardial injury. |
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