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Islet-1 promotes the cardiac-specific differentiation of mesenchymal stem cells through the regulation of histone acetylation.

Yin N, Lu R, Lin J, Zhi S, Tian J, Zhu J - Int. J. Mol. Med. (2014)

Bottom Line: The AcH3 relative amount increased following transfection with Lenti-Islet-1, which was associated with the enhanced expression of Gata4, Nkx2.5 and Mef2c in these cells.The expression of Gata4, Nkx2.5 and Mef2c in the C3H10T1/2 cells transfected with Lenti-Islet-1 and treated with EGCG was reduced following treatment with EGCG.The data presented in this study indicate that Islet-1 specifically induces the differentiation of C3H10T1/2 cells into cardiomyocyte-like cells, and one of the mechanisms involved is the regulation of histone acetylation.

View Article: PubMed Central - PubMed

Affiliation: Ministry of Education Key Laboratory of Child Development and Disorders, Pediatric Research Institute, Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China.

ABSTRACT
The aim of the present study was to investigate the effects of Islet-1 on the process of mesenchymal stem cell (MSC) differentiation into cardiomyocyte-like cells and to elucidate the possible mechanisms involved. Lentiviral vectors expressing Islet-1 (Lenti-Islet-1) were constructed and used for C3H10T1/2 cell transfection. Cell morphology was observed. Cardiac-related genes and proteins were detected by qPCR and western blot analysis. Epigallocatechin gallate (EGCG) was used as an inhibitor of acetylated histone H3 (AcH3). AcH3 was detected by chromatin immunoprecipitation. Cells overexpressing Islet-1 tended to change into fibroblast-like cells and were arranged in the same direction. The enhanced expression of GATA binding protein 4 (Gata4), NK2 homeobox 5 (Nkx2.5), myocyte enhancer factor 2C (Mef2c) and cardiac troponin T (cTnT) was observed in the cells overexpressing Islet-1 following transfection with Lenti-Islet-1. However, the expression of hepatocyte-, bone- and neuronal-specific markers was not affected by Islet-1. The AcH3 relative amount increased following transfection with Lenti-Islet-1, which was associated with the enhanced expression of Gata4, Nkx2.5 and Mef2c in these cells. The expression of Gata4, Nkx2.5 and Mef2c in the C3H10T1/2 cells transfected with Lenti-Islet-1 and treated with EGCG was reduced following treatment with EGCG. The data presented in this study indicate that Islet-1 specifically induces the differentiation of C3H10T1/2 cells into cardiomyocyte-like cells, and one of the mechanisms involved is the regulation of histone acetylation.

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Differences in histone acetylation levels in the untransfected C3H10T1/2 cells, the C3H10T1/2 cells transfected with Lenti-N and the C3H10T1/2 cells transfected with Lenti-Islet-1. (A) Acetylated histone H3 (AcH3) was detected by western blot analysis in the untranstected C3H10T1/2 cells, the C3H10T1/2 cells transfected with Lenti-N and the C3H10T1/2 cells transfected with Lenti-Islet-1. The AcH3 relative amount in the C3H10T1/2 cells transfected with Lenti-Islet-1 was higher than that in the untransfected C3H10T1/2 cells and the C3H10T1/2 cells transfected with Lenti-N (*P<0.05). (B) Chromatin immunoprecipitation (ChIP) and qPCR were performed to reveal the acetylation levels of histone H3 on the cardiac-specific genes, GATA binding protein 4 (Gata4), NK2 homeobox 5 (Nkx2.5) and myocyte enhancer factor 2C (Mef2c), at their peak expression times (2 weeks after transfection). The expression of Gata4, Nkx2.5 and Mef2c combined with AcH3 in the C3H10T1/2 cells transfected with Lenti-Islet-1 was higher than that in the untransfected C3H10T1/2 cells and the C3H10T1/2 cells transfected with Lenti-N (*P<0.05). (C) Gata4, Nkx2.5 and Mef2c expression was found to be reduced 3 h following treatment with 120 μmol/l epigallocatechin gallate (EGCG) (*P<0.05). (D) Gata4, Nkx2.5 and Mef2c expression in the C3H10T1/2 cells transfected with Lenti-Islet-1 and treated with EGCG was lower compared with the cells not treated with EGCG (*P<0.05).
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f5-ijmm-33-05-1075: Differences in histone acetylation levels in the untransfected C3H10T1/2 cells, the C3H10T1/2 cells transfected with Lenti-N and the C3H10T1/2 cells transfected with Lenti-Islet-1. (A) Acetylated histone H3 (AcH3) was detected by western blot analysis in the untranstected C3H10T1/2 cells, the C3H10T1/2 cells transfected with Lenti-N and the C3H10T1/2 cells transfected with Lenti-Islet-1. The AcH3 relative amount in the C3H10T1/2 cells transfected with Lenti-Islet-1 was higher than that in the untransfected C3H10T1/2 cells and the C3H10T1/2 cells transfected with Lenti-N (*P<0.05). (B) Chromatin immunoprecipitation (ChIP) and qPCR were performed to reveal the acetylation levels of histone H3 on the cardiac-specific genes, GATA binding protein 4 (Gata4), NK2 homeobox 5 (Nkx2.5) and myocyte enhancer factor 2C (Mef2c), at their peak expression times (2 weeks after transfection). The expression of Gata4, Nkx2.5 and Mef2c combined with AcH3 in the C3H10T1/2 cells transfected with Lenti-Islet-1 was higher than that in the untransfected C3H10T1/2 cells and the C3H10T1/2 cells transfected with Lenti-N (*P<0.05). (C) Gata4, Nkx2.5 and Mef2c expression was found to be reduced 3 h following treatment with 120 μmol/l epigallocatechin gallate (EGCG) (*P<0.05). (D) Gata4, Nkx2.5 and Mef2c expression in the C3H10T1/2 cells transfected with Lenti-Islet-1 and treated with EGCG was lower compared with the cells not treated with EGCG (*P<0.05).

Mentions: Acetylated histone H3 (AcH3) was detected by western blot analysis in the untransfected C3H10T1/2 cells (controls), the C3H10T1/2 cells transfected with Lenti-N and the C3H10T1/2 cells transfected with Lenti-Islet-1. The relative amount of AcH3 in the C3H10T1/2 cells transfected with Lenti-Islet-1 was higher than that of the other cells (Fig. 5A). ChIP and qPCR were performed to determine the acetylation levels of histone H3 at the cardiac-specific genes, Gata4, Nkx2.5 and Mef2c, at their peak expression times (2 weeks after transfection). The expression of Gata4, Nkx2.5 and Mef2c combined with AcH3 in the C3H10T1/2 cells transfected with Lenti-Islet-1 was higher than that in the untransfected C3H10T1/2 cells and in the C3H10T1/2 cells transfected with Lenti-N (Fig. 5B). Gata4, Nkx2.5 and Mef2c expression was found to be reduced 3 h following treatment with 120 μmol/l EGCG (Fig. 5C). Gata4, Nkx2.5 and Mef2c expression in the C3H10T1/2 cells transfected with Lenti-Islet-1 and treated with EGCG was lower compared with the cells not treated with EGCG (Fig. 5D).


Islet-1 promotes the cardiac-specific differentiation of mesenchymal stem cells through the regulation of histone acetylation.

Yin N, Lu R, Lin J, Zhi S, Tian J, Zhu J - Int. J. Mol. Med. (2014)

Differences in histone acetylation levels in the untransfected C3H10T1/2 cells, the C3H10T1/2 cells transfected with Lenti-N and the C3H10T1/2 cells transfected with Lenti-Islet-1. (A) Acetylated histone H3 (AcH3) was detected by western blot analysis in the untranstected C3H10T1/2 cells, the C3H10T1/2 cells transfected with Lenti-N and the C3H10T1/2 cells transfected with Lenti-Islet-1. The AcH3 relative amount in the C3H10T1/2 cells transfected with Lenti-Islet-1 was higher than that in the untransfected C3H10T1/2 cells and the C3H10T1/2 cells transfected with Lenti-N (*P<0.05). (B) Chromatin immunoprecipitation (ChIP) and qPCR were performed to reveal the acetylation levels of histone H3 on the cardiac-specific genes, GATA binding protein 4 (Gata4), NK2 homeobox 5 (Nkx2.5) and myocyte enhancer factor 2C (Mef2c), at their peak expression times (2 weeks after transfection). The expression of Gata4, Nkx2.5 and Mef2c combined with AcH3 in the C3H10T1/2 cells transfected with Lenti-Islet-1 was higher than that in the untransfected C3H10T1/2 cells and the C3H10T1/2 cells transfected with Lenti-N (*P<0.05). (C) Gata4, Nkx2.5 and Mef2c expression was found to be reduced 3 h following treatment with 120 μmol/l epigallocatechin gallate (EGCG) (*P<0.05). (D) Gata4, Nkx2.5 and Mef2c expression in the C3H10T1/2 cells transfected with Lenti-Islet-1 and treated with EGCG was lower compared with the cells not treated with EGCG (*P<0.05).
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f5-ijmm-33-05-1075: Differences in histone acetylation levels in the untransfected C3H10T1/2 cells, the C3H10T1/2 cells transfected with Lenti-N and the C3H10T1/2 cells transfected with Lenti-Islet-1. (A) Acetylated histone H3 (AcH3) was detected by western blot analysis in the untranstected C3H10T1/2 cells, the C3H10T1/2 cells transfected with Lenti-N and the C3H10T1/2 cells transfected with Lenti-Islet-1. The AcH3 relative amount in the C3H10T1/2 cells transfected with Lenti-Islet-1 was higher than that in the untransfected C3H10T1/2 cells and the C3H10T1/2 cells transfected with Lenti-N (*P<0.05). (B) Chromatin immunoprecipitation (ChIP) and qPCR were performed to reveal the acetylation levels of histone H3 on the cardiac-specific genes, GATA binding protein 4 (Gata4), NK2 homeobox 5 (Nkx2.5) and myocyte enhancer factor 2C (Mef2c), at their peak expression times (2 weeks after transfection). The expression of Gata4, Nkx2.5 and Mef2c combined with AcH3 in the C3H10T1/2 cells transfected with Lenti-Islet-1 was higher than that in the untransfected C3H10T1/2 cells and the C3H10T1/2 cells transfected with Lenti-N (*P<0.05). (C) Gata4, Nkx2.5 and Mef2c expression was found to be reduced 3 h following treatment with 120 μmol/l epigallocatechin gallate (EGCG) (*P<0.05). (D) Gata4, Nkx2.5 and Mef2c expression in the C3H10T1/2 cells transfected with Lenti-Islet-1 and treated with EGCG was lower compared with the cells not treated with EGCG (*P<0.05).
Mentions: Acetylated histone H3 (AcH3) was detected by western blot analysis in the untransfected C3H10T1/2 cells (controls), the C3H10T1/2 cells transfected with Lenti-N and the C3H10T1/2 cells transfected with Lenti-Islet-1. The relative amount of AcH3 in the C3H10T1/2 cells transfected with Lenti-Islet-1 was higher than that of the other cells (Fig. 5A). ChIP and qPCR were performed to determine the acetylation levels of histone H3 at the cardiac-specific genes, Gata4, Nkx2.5 and Mef2c, at their peak expression times (2 weeks after transfection). The expression of Gata4, Nkx2.5 and Mef2c combined with AcH3 in the C3H10T1/2 cells transfected with Lenti-Islet-1 was higher than that in the untransfected C3H10T1/2 cells and in the C3H10T1/2 cells transfected with Lenti-N (Fig. 5B). Gata4, Nkx2.5 and Mef2c expression was found to be reduced 3 h following treatment with 120 μmol/l EGCG (Fig. 5C). Gata4, Nkx2.5 and Mef2c expression in the C3H10T1/2 cells transfected with Lenti-Islet-1 and treated with EGCG was lower compared with the cells not treated with EGCG (Fig. 5D).

Bottom Line: The AcH3 relative amount increased following transfection with Lenti-Islet-1, which was associated with the enhanced expression of Gata4, Nkx2.5 and Mef2c in these cells.The expression of Gata4, Nkx2.5 and Mef2c in the C3H10T1/2 cells transfected with Lenti-Islet-1 and treated with EGCG was reduced following treatment with EGCG.The data presented in this study indicate that Islet-1 specifically induces the differentiation of C3H10T1/2 cells into cardiomyocyte-like cells, and one of the mechanisms involved is the regulation of histone acetylation.

View Article: PubMed Central - PubMed

Affiliation: Ministry of Education Key Laboratory of Child Development and Disorders, Pediatric Research Institute, Children's Hospital of Chongqing Medical University, Chongqing 400014, P.R. China.

ABSTRACT
The aim of the present study was to investigate the effects of Islet-1 on the process of mesenchymal stem cell (MSC) differentiation into cardiomyocyte-like cells and to elucidate the possible mechanisms involved. Lentiviral vectors expressing Islet-1 (Lenti-Islet-1) were constructed and used for C3H10T1/2 cell transfection. Cell morphology was observed. Cardiac-related genes and proteins were detected by qPCR and western blot analysis. Epigallocatechin gallate (EGCG) was used as an inhibitor of acetylated histone H3 (AcH3). AcH3 was detected by chromatin immunoprecipitation. Cells overexpressing Islet-1 tended to change into fibroblast-like cells and were arranged in the same direction. The enhanced expression of GATA binding protein 4 (Gata4), NK2 homeobox 5 (Nkx2.5), myocyte enhancer factor 2C (Mef2c) and cardiac troponin T (cTnT) was observed in the cells overexpressing Islet-1 following transfection with Lenti-Islet-1. However, the expression of hepatocyte-, bone- and neuronal-specific markers was not affected by Islet-1. The AcH3 relative amount increased following transfection with Lenti-Islet-1, which was associated with the enhanced expression of Gata4, Nkx2.5 and Mef2c in these cells. The expression of Gata4, Nkx2.5 and Mef2c in the C3H10T1/2 cells transfected with Lenti-Islet-1 and treated with EGCG was reduced following treatment with EGCG. The data presented in this study indicate that Islet-1 specifically induces the differentiation of C3H10T1/2 cells into cardiomyocyte-like cells, and one of the mechanisms involved is the regulation of histone acetylation.

Show MeSH