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Downregulation of long non-coding RNA H19 promotes P19CL6 cells proliferation and inhibits apoptosis during late-stage cardiac differentiation via miR-19b-modulated Sox6

View Article: PubMed Central - PubMed

ABSTRACT

Background: Regulating cardiac differentiation to maintain normal heart development and function is very important. At present, biological functions of H19 in cardiac differentiation is not completely clear.

Methods: To explore the functional effect of H19 during cardiac differentiation. Expression levels of early cardiac-specific markers Nkx-2.5 and GATA4, cardiac contractile protein genes α-MHC and MLC-2v were determined by qRT-PCR and western lot. The levels of lncRNA H19 and miR-19b were detected by qRT-PCR. We further predicted the binding sequence of H19 and miR-19b by online softwares starBase v2.0 and TargetScan. The biological functions of H19 and Sox6 were evaluated by CCK-8 kit, cell cycle and apoptosis assay and caspase-3 activity.

Results: The expression levels of α-MHC, MLC-2v and H19 were upregulated, and miR-19b was downregulated significantly in mouse P19CL6 cells at the late stage of cardiac differentiation. Biological function analysis showed that knockdown of H19 promoted cell proliferation and inhibits cell apoptosis. H19 suppressed miR-19b expression and miR-19b targeted Sox6, which inhibited cell proliferation and promoted apoptosis in P19CL6 cells during late-stage cardiac differentiation. Importantly, Sox6 overexpression could reverse the positive effects of H19 knockdown on P19CL6 cells.

Conclusion: Downregulation of H19 promoted cell proliferation and inhibited cell apoptosis during late-stage cardiac differentiation by regulating the negative role of miR-19b in Sox6 expression, which suggested that the manipulation of H19 expression could serve as a potential strategy for heart disease.

No MeSH data available.


Related in: MedlinePlus

The effect of H19 knockdown and H19 overexpression on P19CL6 cell proliferation and apoptosis. a Transfecting P19CL6 cells with shH19 and pcDNA-H19 efficiently down- and up-regulated the H19 level at day 10, respectively. b CCK-8 assay showed that shH19 resulted in an increase in cell viability at day 8 and 10, whereas pcDNA-H19 had an opposite effect. c Flow cytometry suggested that shH19 significantly increased the cell number in S phase at day 8 and 10, and pcDNA-H19 significantly decreased the cell number. d shH19 significantly reduced the apoptotic rate of P19CL6 cells at day 8 and day 10, whereas pcDNA-H19 had an opposite effect. e Caspase-3 assay indicated that the activity of caspase-3 was significantly reduced in P19CL6 cells transfected with shH19 and significantly increased in cells transfected with pcDNA-H19. *P < 0.05, **P < 0.01, ***P < 0.001
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Fig2: The effect of H19 knockdown and H19 overexpression on P19CL6 cell proliferation and apoptosis. a Transfecting P19CL6 cells with shH19 and pcDNA-H19 efficiently down- and up-regulated the H19 level at day 10, respectively. b CCK-8 assay showed that shH19 resulted in an increase in cell viability at day 8 and 10, whereas pcDNA-H19 had an opposite effect. c Flow cytometry suggested that shH19 significantly increased the cell number in S phase at day 8 and 10, and pcDNA-H19 significantly decreased the cell number. d shH19 significantly reduced the apoptotic rate of P19CL6 cells at day 8 and day 10, whereas pcDNA-H19 had an opposite effect. e Caspase-3 assay indicated that the activity of caspase-3 was significantly reduced in P19CL6 cells transfected with shH19 and significantly increased in cells transfected with pcDNA-H19. *P < 0.05, **P < 0.01, ***P < 0.001

Mentions: Given that H19 might participate in the cardiac differentiation of P19CL6 cells, P19CL6 cells were transfected with the shH19 or pcDNA-H19. qRT-PCR results showed that H19 expression was significantly decreased in P19CL6 cells transfected with shH19 and H19 expression was significantly increased in P19CL6 cells transfected with pcDNA-H19 (Fig. 2a).Fig. 2


Downregulation of long non-coding RNA H19 promotes P19CL6 cells proliferation and inhibits apoptosis during late-stage cardiac differentiation via miR-19b-modulated Sox6
The effect of H19 knockdown and H19 overexpression on P19CL6 cell proliferation and apoptosis. a Transfecting P19CL6 cells with shH19 and pcDNA-H19 efficiently down- and up-regulated the H19 level at day 10, respectively. b CCK-8 assay showed that shH19 resulted in an increase in cell viability at day 8 and 10, whereas pcDNA-H19 had an opposite effect. c Flow cytometry suggested that shH19 significantly increased the cell number in S phase at day 8 and 10, and pcDNA-H19 significantly decreased the cell number. d shH19 significantly reduced the apoptotic rate of P19CL6 cells at day 8 and day 10, whereas pcDNA-H19 had an opposite effect. e Caspase-3 assay indicated that the activity of caspase-3 was significantly reduced in P19CL6 cells transfected with shH19 and significantly increased in cells transfected with pcDNA-H19. *P < 0.05, **P < 0.01, ***P < 0.001
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5120414&req=5

Fig2: The effect of H19 knockdown and H19 overexpression on P19CL6 cell proliferation and apoptosis. a Transfecting P19CL6 cells with shH19 and pcDNA-H19 efficiently down- and up-regulated the H19 level at day 10, respectively. b CCK-8 assay showed that shH19 resulted in an increase in cell viability at day 8 and 10, whereas pcDNA-H19 had an opposite effect. c Flow cytometry suggested that shH19 significantly increased the cell number in S phase at day 8 and 10, and pcDNA-H19 significantly decreased the cell number. d shH19 significantly reduced the apoptotic rate of P19CL6 cells at day 8 and day 10, whereas pcDNA-H19 had an opposite effect. e Caspase-3 assay indicated that the activity of caspase-3 was significantly reduced in P19CL6 cells transfected with shH19 and significantly increased in cells transfected with pcDNA-H19. *P < 0.05, **P < 0.01, ***P < 0.001
Mentions: Given that H19 might participate in the cardiac differentiation of P19CL6 cells, P19CL6 cells were transfected with the shH19 or pcDNA-H19. qRT-PCR results showed that H19 expression was significantly decreased in P19CL6 cells transfected with shH19 and H19 expression was significantly increased in P19CL6 cells transfected with pcDNA-H19 (Fig. 2a).Fig. 2

View Article: PubMed Central - PubMed

ABSTRACT

Background: Regulating cardiac differentiation to maintain normal heart development and function is very important. At present, biological functions of H19 in cardiac differentiation is not completely clear.

Methods: To explore the functional effect of H19 during cardiac differentiation. Expression levels of early cardiac-specific markers Nkx-2.5 and GATA4, cardiac contractile protein genes &alpha;-MHC and MLC-2v were determined by qRT-PCR and western lot. The levels of lncRNA H19 and miR-19b were detected by qRT-PCR. We further predicted the binding sequence of H19 and miR-19b by online softwares starBase v2.0 and TargetScan. The biological functions of H19 and Sox6 were evaluated by CCK-8 kit, cell cycle and apoptosis assay and caspase-3 activity.

Results: The expression levels of &alpha;-MHC, MLC-2v and H19 were upregulated, and miR-19b was downregulated significantly in mouse P19CL6 cells at the late stage of cardiac differentiation. Biological function analysis showed that knockdown of H19 promoted cell proliferation and inhibits cell apoptosis. H19 suppressed miR-19b expression and miR-19b targeted Sox6, which inhibited cell proliferation and promoted apoptosis in P19CL6 cells during late-stage cardiac differentiation. Importantly, Sox6 overexpression could reverse the positive effects of H19 knockdown on P19CL6 cells.

Conclusion: Downregulation of H19 promoted cell proliferation and inhibited cell apoptosis during late-stage cardiac differentiation by regulating the negative role of miR-19b in Sox6 expression, which suggested that the manipulation of H19 expression could serve as a potential strategy for heart disease.

No MeSH data available.


Related in: MedlinePlus