Study On Changes In The Expression Profiles Of MRNAs In Right Atrial Appendage And LncRNAs In Serum Of Patients With Pulmonary Hypertension Secondary To Ventricular Septal Defect

Objective: Based on the preliminary results of changes in the expression profiles of mRNAs and lncRNAs in right atrial appendage of patients with pulmonary hypertension secondary to ventricular septal defect（VSD）,this study was to further analyze these data to obtain the co-expression of lncRNAs and mRNAs.By selecting key signaling pathway nodes,fluorescent real-time quantitative PCR was performed on mRNAs in right atrial appendage tissue and lncRNAs in serum to obtain the biomarker in serum that might predict the rate of progress towards pulmonary artery hypertension（PAH）.Methods: Total RNA was extracted from 4 samples of moderate-severe PAH secondary to VSD in experiment group and 4 samples with simply VSD,these samples were acquired from auricula dextra tissue during surgical operation before cardio-pulmonary bypass,and the serum samples were acquired at the same times.After quality inspection,the qualified RNA was used to hybrid synthesize cDNA.Scanning the hybridization chip to get the preliminary results,which were converted and analyzed to obtain the data of mRNA and lncRNA.Fold Change>2,P<0.05 as the difference of gene expression inclusion criteria to get lncRNAs and mRNAs expression profile.For all differentially expressed lncRNAs and differential known encoded protein mRNAs,the Pearson Correlation Coefficient between differential lncRNAs and differential known encoded protein mRNAs was calculated.The co-expression regulating network between differential lncRNAs and differential known encoded protein mRNAs was built according to the absolute value of the Pearson Correlation Coefficient（0.9）and the significance of correlation coefficient P（0.01）.The highest twenty levels of mRNAs differential gene expression and the highest twenty levels of lncRNAs differential gene expression were selected for network demonstration.The regulating role of a gene in network was evaluated by K-core,and the correlation density between such gene and adjacent genes was measured based on the Clustering Coefficient.According to the results of K-core and Clustering Coefficient,mRNAs and lncRNAs with more important values of regulation in network were selected based on GOs functional analysis and KEGGs signal pathway analysis.The expressions of mRNAs selected were detected by performing PCR on the right atrial appendage tissues of patients with PHA secondary to VSD and patients with no PHA,and PCR was performed to detect lncRNAs expression level in serum obtained during the same period.Results: Gene expression profile comparison between the experimental group and the control group indicated that 813 lncRNA expression levels were significantly up-regulated,and 541 were significantly down-regulated;527 mRNAs expression levels were significantly up-regulated,and 268 were significantly down-regulated.These results suggested that significantly differential expressions of lncRNAs might take part in the regulation of PHA,which might be realized by extensive participation in the regulation of mRNAs.According to the regulatory network degrees of genes and K-core,a total of 75 key regulatory genes（including lncRNAs and mRNAs）were screened out and analyzed by combining with GOs functional analysis and KEGGs signaling pathway analysis.TNNT3、SAA1 、TCONS₀0008552 and ENST00000433673 were selected as candidate genes.According to Pearson correlation coefficient,the correlation coefficient between TNNT3 and ENST00000433673 was 0.998965,the correlation coefficient between SAA1 and TCONS₀0008552 was 0.998903.PCRs on the right atrial appendage tissues of both groups suggested that TNNT3 and SAA1 expressopms in the experimental group were significantly increased than those of the control group,showing statistically significant differences;these two groups showed no obvious difference in ENST00000433673 expression.Conclusion: Right atrial appendage tissue is a good resource for researches on the mechanism of VSD and PAH.Differentially expressed lncRNAs or mRNAs may participate in the development of PAH,and for lncRNAs,it might be realized by extensive participations in the regulation of mRNAs.The biological information of 75 key regulatory genes such as HP、TF、TNNT3、SAA1、FGFBP1、FABP2、FAP、S100A12、NR₀38286 and TCONS₀0008552 are consistent well with the regulatory process of PAH.There are statistically significant differences in TNNT3 and SAA1 expressions in the right atrial appendage tissue between the experimental group and the control group;TCONS₀0008552 expression in the experimental group is much higher than that in the control group,showing a statistically significant difference;there is no obvious difference in NST00000433673 expression between these two groups.Therefore,TCONS₀0008552 may serve as a candidate serum biomarker to predict the rate of progress towards PHA.