Transcriptomic Study Of Cardiac Repair During Mouse Embryonic Development

BackgroundCardiovascular disease is the leading cause of death from diseases.The prevailing treatment strategies for cardiac diseases are drug therapy,coronary intervention,etc.These strategies do not achieve satisfied results.A case report shows full cardiac structural and functional recovery cardiac structure and function after MI in neonate.In recent years,myriad researchers have focused on the mechanism of myocardial regeneration and found that the signaling factors involved in the regulation of myocardial regeneration in model animals（zebrafish,Newt,etc.）.partially coincide with those involved in the regulation of heart development.Elucidation of the mechanisms of myocardial regeneration after cardiac injury during the embryonic period in mammals will provide new strategies to solve the regeneration conundrum of mammalian and human adult cardiomyocytes.ObjectiveIn this study,transgenic mice were used to establish a model of middle and late embryonic heart injury in order to study the changes of Transcriptomic profiling of the regenerating heart during mouse embryonic development.Methods1.Inducible Myh6^{Cre ER+/-}male mice were mated with Rosa26-m Tm G^+/+female mice.Tamoxifen was injected into pregnant mice during pregnancy to induce the expression of GFP in Myh6 positive cells.Examine the dynamic expression patterns of Myh6 in middle and late embryos using GFP as a readout.2.Myh6^{Cre ER+/-}male mice were mated with Rosa26-DTA^+/+female mice.Tamoxifen was injected into pregnant mice during pregnancy to induce myocardial cell damage,and the damage of myocardial cells was verified by TUNEL apoptosis assay.3.Myh6^{Cre ER+/-}male mice were mated with Rosa26-DTA^+/+female mice.Tamoxifen was injected into pregnant mice during pregnancy,cell proliferation was analyzed during cardiac repair.4.The embryonic hearts were harvested at different time points,RNA was extracted and RNA-seq was performed in order to study the transcriptimic profile during cardiac regeneration.Results1.Immunofluorescence results showed that GFP was mainly expressed in cardiomyocytes of atria and non-compacted layer of ventricles.GFP was mainly expressed in the noncompacted and compacted layers of ventricular wall after birth.2.TUNEL apoptosis results showed that intraperitoneal injection of tamoxifen induced myocardial cell injury in pregnant mice at embryonic E12.5 and E13.5 days.Sporadic apoptotic cells could be seen in atrium and ventricle on E15.5 day,and the number of apoptotic cells reached the peak on E16.6 days,and the apoptotic cells displayed a cluster distribution,then gradually decreased,and basically disappeared on E19.5 day.Control group(Myh6^{Cre ER-/-};Rosa26-DTA^+/-)showed no obvious apoptosis during the examined development.3.Ed U proliferation results showed that under the same experimental conditions as TUNEL apoptosis experiments,the atrial proliferation rate of the experimental group was lower than that of the control group on E16.5 days,and the atrial and ventricular proliferation rates of the experimental group was were greater than those of the control groups on E19.5 days,respectively,indicating that the fetal heart proliferation rate was enhanced after heart injury.4.After inducing cardiomyocyte apoptosis,cardiac RNA was extracted from embryonic E19.5 days,and the transcriptome was sequenced.The results showed that Wnt9b,Fgf22,Bmp8b,Tnn,Scg5,Myl2 were up-regulated in the central atrium during myocardial regeneration.The expression of Gdf3,Ptgs2 and Ly6c2 decreased.The expression of Wnt2b,Wnt7b,Fzd3 and cdh10 in ventricles were up-regulated.The expression of Hhipl2,BMP2 and Fgf2 decreased.GO enrichment analysis and KEGG signal pathway analysis show that these differential genes are mainly involved in the regulation of cardiac contractility,heart rhythm,metabolism,cell migration,proliferation and differentiation,and some of them have been confirmed to be related to cardiac development and regeneration.ConclusionsCell proliferation is enhanced after cardiac injury in mouse embryos.Examination of Transcriptomic expression revealed upregulation of genes related to cell proliferation,metabolism and electrical signals during Mid-to late-embryonic myocardial regeneration.