Nkx2–5 Regulates the Proliferation and Migration of H9c2 Cells

Figures

• 

  Figure 1. Verification of the knockdown effect of shRNA lentivirus on Nkx2–5 in H9c2 cells by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and western blot analysis, respectively. (A) qRT-PCR detection of the expression of Nkx2–5 in the control group and the Nkx2–5 knockdown group. (B) Western blot detection of the expression of Nkx2–5 in the control group and the Nkx2–5 knockdown group.

• 

  Figure 2. Knockdown of Nkx2–5 inhibited the proliferative capacity of H9c2 cells. The CCK8 method was used to detect the effect of Nkx2–5 on the proliferative capacity of H9c2 cells. (A) Cell growth was detected with the CCK8 method for 4 consecutive days. (B) Statistics of the CCK8 curve are shown.

• 

  Figure 3. Knockdown of Nkx2–5 increase the migration ability of H9c2 cells. The cell scratch test was used to detect the effect of Nkx2–5 on the migration ability of H9c2 cells (0–72 h). Cells were seeded in the culture-insert, which was removed after 24 h, and cells were then continuously observed for 72 h.

• 

  Figure 4. Gene Ontology (GO) analysis of the differential expression of genes caused by knockdown of Nkx2–5 indicated that the genes are enriched in many biological processes, including cardiac epithelial to mesenchymal transition, development of the cardiac bundle of His, and cardiac muscle cell proliferation.

• 

  Figure 5. Pathway analysis of the differential expression of genes caused by knockdown of Nkx2–5 indicated that the genes are enriched in the transforming growth factor (TGF)-β signaling pathway, and pathways related to hypertrophic cardiomyopathy, dilated cardiomyopathy, and arrhythmogenic right ventricular cardiomyopathy.

• 

  Figure 6. Heat map of the differentially expressed miRNAs following knockdown of Nkx2–5.

• 

  Figure 7. Gene Ontology (GO) enrichment analysis of the target genes of the differentially expressed miRNAs caused by knockdown of Nkx2–5 indicated that those genes function in many biological processes.

• 

  Figure 8. Pathway enrichment analysis of the target genes of miRNAs that were differentially expressed owing to knockdown of Nkx2–5 indicated that the target genes are enriched in the MAPK signaling pathway and other signaling pathways.

• 

  Figure 9. (A–D) Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis of the relative expression levels of Tgfb-2, Id2, Ptk2b, Cacna1g, Wt1, Heyl, Olfml, Wnt4, Xdh, Lrp2, Cited1, Syt1, Emp2, and Pou5f in the control group and in the Nkx2–5 knockdown group. The expression level of the genes in the control group was calculated as 1. The bar represents the fold change of the genes in the Nkx2–5 knockdown group compared with the control group. * P<.05; ** P<.01; *** P<.001; **** P<.0001.

• 

  Figure 10. (A–D) Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis of the relative expression levels of Syt13, Itga7, Fos, Slrp1, Nectin3, Tenm4, Bmp10, rno-miR-1-5p, rno-miR-1-3p, rno-let-7a-5p, rno-miR-148b-3p, rno-miR-149-5p, rno-miR-361-3p, rno-miR-455-3p, and rno-miR-25-3p in the control group and in the Nkx2–5 knockdown group. The expression level of the genes in the control group was calculated as 1. The bar represents the fold change of the genes in the Nkx2–5 knockdown group compared with the control group. * P<.05; ** P<.01; *** P<.001; **** P<.0001.