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Combination of IGF‑1 gene manipulation and 5‑AZA treatment promotes differentiation of mesenchymal stem cells into cardiomyocyte‑like cells.

Li J, Zhu K, Wang Y, Zheng J, Guo C, Lai H, Wang C - Mol Med Rep (2014)

Bottom Line: Our results demonstrated that 5‑AZA treatment alone induced a limited cardiomyocyte‑like differentiation effect in vitro.Overexpression of the IGF‑1 gene in MSCs improved the induction effect of 5‑AZA, while knockdown of the IGF‑1 gene attenuated the differentiation.These results suggest that IGF‑1 is a significant stimulus affecting the cardiomyocyte‑like differentiation of porcine MSCs.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiac Surgery, Shanghai Institute of Cardiovascular Disease, Zhongshan Hospital, Fudan University, Shanghai 200032, P.R. China.

ABSTRACT
Mesenchymal stem cell (MSC) transplantation has been proposed as a promising therapeutic strategy for ischemic myocardium repair following myocardial infarction. Differentiation of MSCs into cardiomyocyte‑like cells prior to cell transplantation is advantageous in improving their potential clinical benefits for cardiac repair. In the present study, we isolated and cultured porcine MSCs and evaluated the synergistic effect of 5‑azacytidine (5‑AZA) treatment and insulin‑like growth factor‑1 (IGF‑1) gene manipulation on MSC differentiation into cardiomyocyte‑like cells. Our results demonstrated that 5‑AZA treatment alone induced a limited cardiomyocyte‑like differentiation effect in vitro. Overexpression of the IGF‑1 gene in MSCs improved the induction effect of 5‑AZA, while knockdown of the IGF‑1 gene attenuated the differentiation. These results suggest that IGF‑1 is a significant stimulus affecting the cardiomyocyte‑like differentiation of porcine MSCs. In addition, the combination of IGF‑1 gene manipulation and 5‑AZA treatment provides a new strategy to obtain more committed differentiated cardiomyocyte‑like cells from porcine MSCs prior to cell transplantation.

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Morphology and characterization of isolated porcine mesenchymal stem cells (MSCs). (A) Morphology of MSCs at passage 2 (P2) on day 1 (D1), day 4 (D4), day 7 (D7) and day 10 (D10) show an adherent fibroblast-like shape and begin to form cell colonies after 10 days of culture. Passage 1 (P1) exhibited low growth ability. Scale bar, 50 μm. (B) Fluorescence-activated cell sorting analysis of CD90, CD44, CD34 and CD45 expression in MSCs. The histogram shows the average positive percentage of different antibodies from three independent experiments. Columns represent mean values and error bars represent SD. **P<0.01, vs. positive antibody expression. (C) The proliferation rate of MSCs in different passages (P2, P5 and P8) was detected by proliferation assay. (D) The cell viability of MSCs in different passages (P2, P5 and P8) was detected by trypan blue staining. Columns represent mean values and error bars represent SD. *P<0.05 and **P<0.01, vs. P8.
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f1-mmr-11-02-0815: Morphology and characterization of isolated porcine mesenchymal stem cells (MSCs). (A) Morphology of MSCs at passage 2 (P2) on day 1 (D1), day 4 (D4), day 7 (D7) and day 10 (D10) show an adherent fibroblast-like shape and begin to form cell colonies after 10 days of culture. Passage 1 (P1) exhibited low growth ability. Scale bar, 50 μm. (B) Fluorescence-activated cell sorting analysis of CD90, CD44, CD34 and CD45 expression in MSCs. The histogram shows the average positive percentage of different antibodies from three independent experiments. Columns represent mean values and error bars represent SD. **P<0.01, vs. positive antibody expression. (C) The proliferation rate of MSCs in different passages (P2, P5 and P8) was detected by proliferation assay. (D) The cell viability of MSCs in different passages (P2, P5 and P8) was detected by trypan blue staining. Columns represent mean values and error bars represent SD. *P<0.05 and **P<0.01, vs. P8.

Mentions: The morphology of isolated porcine MSCs (passage 2) is shown in Fig. 1A. During the first two days, the culture was heterogeneous, containing round and non-adherent cells with lipid micelles in the supernatant. After 10 days, the cell population became more homogeneous, presenting an adherent fibroblast-like shape, and began to form cell colonies.


Combination of IGF‑1 gene manipulation and 5‑AZA treatment promotes differentiation of mesenchymal stem cells into cardiomyocyte‑like cells.

Li J, Zhu K, Wang Y, Zheng J, Guo C, Lai H, Wang C - Mol Med Rep (2014)

Morphology and characterization of isolated porcine mesenchymal stem cells (MSCs). (A) Morphology of MSCs at passage 2 (P2) on day 1 (D1), day 4 (D4), day 7 (D7) and day 10 (D10) show an adherent fibroblast-like shape and begin to form cell colonies after 10 days of culture. Passage 1 (P1) exhibited low growth ability. Scale bar, 50 μm. (B) Fluorescence-activated cell sorting analysis of CD90, CD44, CD34 and CD45 expression in MSCs. The histogram shows the average positive percentage of different antibodies from three independent experiments. Columns represent mean values and error bars represent SD. **P<0.01, vs. positive antibody expression. (C) The proliferation rate of MSCs in different passages (P2, P5 and P8) was detected by proliferation assay. (D) The cell viability of MSCs in different passages (P2, P5 and P8) was detected by trypan blue staining. Columns represent mean values and error bars represent SD. *P<0.05 and **P<0.01, vs. P8.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4262510&req=5

f1-mmr-11-02-0815: Morphology and characterization of isolated porcine mesenchymal stem cells (MSCs). (A) Morphology of MSCs at passage 2 (P2) on day 1 (D1), day 4 (D4), day 7 (D7) and day 10 (D10) show an adherent fibroblast-like shape and begin to form cell colonies after 10 days of culture. Passage 1 (P1) exhibited low growth ability. Scale bar, 50 μm. (B) Fluorescence-activated cell sorting analysis of CD90, CD44, CD34 and CD45 expression in MSCs. The histogram shows the average positive percentage of different antibodies from three independent experiments. Columns represent mean values and error bars represent SD. **P<0.01, vs. positive antibody expression. (C) The proliferation rate of MSCs in different passages (P2, P5 and P8) was detected by proliferation assay. (D) The cell viability of MSCs in different passages (P2, P5 and P8) was detected by trypan blue staining. Columns represent mean values and error bars represent SD. *P<0.05 and **P<0.01, vs. P8.
Mentions: The morphology of isolated porcine MSCs (passage 2) is shown in Fig. 1A. During the first two days, the culture was heterogeneous, containing round and non-adherent cells with lipid micelles in the supernatant. After 10 days, the cell population became more homogeneous, presenting an adherent fibroblast-like shape, and began to form cell colonies.

Bottom Line: Our results demonstrated that 5‑AZA treatment alone induced a limited cardiomyocyte‑like differentiation effect in vitro.Overexpression of the IGF‑1 gene in MSCs improved the induction effect of 5‑AZA, while knockdown of the IGF‑1 gene attenuated the differentiation.These results suggest that IGF‑1 is a significant stimulus affecting the cardiomyocyte‑like differentiation of porcine MSCs.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiac Surgery, Shanghai Institute of Cardiovascular Disease, Zhongshan Hospital, Fudan University, Shanghai 200032, P.R. China.

ABSTRACT
Mesenchymal stem cell (MSC) transplantation has been proposed as a promising therapeutic strategy for ischemic myocardium repair following myocardial infarction. Differentiation of MSCs into cardiomyocyte‑like cells prior to cell transplantation is advantageous in improving their potential clinical benefits for cardiac repair. In the present study, we isolated and cultured porcine MSCs and evaluated the synergistic effect of 5‑azacytidine (5‑AZA) treatment and insulin‑like growth factor‑1 (IGF‑1) gene manipulation on MSC differentiation into cardiomyocyte‑like cells. Our results demonstrated that 5‑AZA treatment alone induced a limited cardiomyocyte‑like differentiation effect in vitro. Overexpression of the IGF‑1 gene in MSCs improved the induction effect of 5‑AZA, while knockdown of the IGF‑1 gene attenuated the differentiation. These results suggest that IGF‑1 is a significant stimulus affecting the cardiomyocyte‑like differentiation of porcine MSCs. In addition, the combination of IGF‑1 gene manipulation and 5‑AZA treatment provides a new strategy to obtain more committed differentiated cardiomyocyte‑like cells from porcine MSCs prior to cell transplantation.

Show MeSH
Related in: MedlinePlus