The Effect Of MiR-29a In Neointimal Hyperplasia After Balloon Injury And The Correlation Between Ciculating MiR-29a And Coronary Artery Disease

Objective: Under stimulation, such as vascular injury, vascular smoothmuscle cells (VSMC) can transform differentiaed phenotype intodedifferentiated phenotype and obtain proliferation ability. The proliferationand directed migration of vascular smooth muscle cells (VSMCs) from themedia into the intima are key events in pathogenic vascular injuries, such ashypertension, atherosclerosis and restenosis. Coronary artery disease (CAD) isdue to abnormal lipid metabolism. The lipid in blood gradually formed theartery atheromatous plaque, which can lead to arterial lumen, blocked bloodflow and cause disease.MicroRNA is a newly discovered class of endogenous non-coding smallmolecule RNA. Most miRNA by binding to sequences in the3′-untranslatedregion (UTR) of target mRNAs inhibit protein translational level. Recentstudies demonstrated that miRNAs can be detected in circulating blood andmay be useful as biomarkers for disease. MiR-29a is a mature member ofmiR-29family. Recent studies have shown that miR-29a is widely involved inthe occurrence and development of cardiovascular disease. In this study, weaimed to investigate the effect of miR-29a in neointimal hyperplasia afterballoon injury and its molecular mechanism, and the corralation betweencirculating miR-29a and cornary artery disease.Methods: Morphology analysis showed that neointimal formation afterballoon injury, and western bolt analysis showed the expression ofproliferative-related gene. Morphology analysis showed that neointimalformation after overexpression of miR-29a, and western blot analysis showedthe expression of proliferative-related gene. Real-time pcr analysis theexpression of miR-29a in the blood of cornary artery disease. Results:1Neointimal hyperplasia after balloon injury in different timeMorphology analysis showed that neointimal formation after ballooninjury. The I/M (intima/media) ratio in3days injured group was0.03±0.01.The I/M ratio in7days injured group was0.1±0.075. The I/M ratio in14daysinjured group was1±0.35, compared with cntrol group (P <0.05), respectively.These results indicate that the extent of intimal hyperplasia increases afterbolloon injury.2Ballon injury induces the expression of proliferative-related geneWestern blot analysis showed that, the expression of CyclinE, CDK2,PCNA, UBC9and K5gradually increased after balloon injury, whichconsisted with the neointimal hyperplasia.3The expression of miR-29a increases in the intimal hyperplasiaReal-time PCR results showed that the expression of miR-29asignificantly increased in vascular tissue after balloon injury, compared withcontrol group. This result suggests that miR-29a may participate in intimalhyperplasia.4Overexpression of miR-29a promotes neointimal hyperplasia after ballooninjuryMorphology analysis showed that the I/M (intima/media) ratio inpAd-miR-29a group was3.2±0.15, compared with pAd group (P<0.05).Accordingly, the I/M ratio in anti-miR-29a group (0.15±0.03) decreased, theI/M (intima/media) ratio was compared with pAd group (P<0.05). Theseresults indicate that overexpression of miR-29a promote neointimalhyperplasia after balloon injury.5Overexpression of miR-29a promotes the expression of PCNA、CyclinD1Western blot showed that overexpression of miR-29a promoted theexpression of proliferative-ralated gene PCNA and CyclinD1after ballooninjury. Accordingly, inhibiting miR-29a decreased the expression of PCNAand CyclinD1, suggesting miR-29a may inhibit intimal hyperplasia bydeacreasing CyclinD1expression. 6Overexpression of miR-29a increases the expression of Klf5Western blot and RT-PCR results showed that overexpression promotesthe expression of Klf5after balloon injury. Silencing miR-29a expressioninhibits the expression of Klf5, which suggests that Klf5may be an importantfactor participated in the intimal hyperplasia.7The expression of miR-29a in patients with coronary artery disease (CAD)Real-time PCR results showed that circulating miR-29a levels in patientswith coronary artery disease (2.22±0.09mmol/L) decreased significantly,compared with the normal group (6.92±0.12mmol/L).8Validation studyTo verify the reduction of circulating miR-29a in patients with CAD andconfirm the factors influencing the level of circulating miR-29a, weprospectively measured circulating miR-29a in a validation cohort. The resultsshowed that high cholesterol and total cholesterol were associated with thereduction of circulating miR-29a level.Conclusions:1MiR-29a overexpression promotes the expression of PCNA andCyclinD1after balloon injury.2MiR-29a overexpression increases the expression of Klf5after ballooninjury.3The level of circulating miR-29a in patients with coronary arterydisease (CAD) significantly decreased.4High cholesterol and total cholesterol were associated with thereduction of circulating miR-29a level.