Inducible effects of icariin, icaritin, and desmethylicaritin on directional differentiation of embryonic stem cells into cardiomyocytes in vitro
Abstract
Aim: To investigate the possible inducible effects of icariin, icaritin, and
desmethylicaritin on the directional differentiation of embryonic stem (ES) cells
into cardiomyocytes in vitro. Methods: ES cells were cultivated as embryoid
bodies (EBs) in hanging drops with icariin, icaritin, or desmethylicaritin. ES cells
treated with retinoic acid and with solvent were used as positive and negative
controls, respectively. The cardiomyocytes derived from the ES cells were verified
using immunocytochemistry. The expression of cardiac developmentaldependent
genes was detected using the reverse transcription-polymerase chain
reaction (RT-PCR) method. Cell cycle distribution and apoptosis were analyzed
using flow cytometry to determine the partly inducible effect mechanisms involved.
Results: The total percentage of beating EBs treated with 1×10-7 mol/L icariin,
icaritin, or desmethylicaritin was 87% (P<0.01), 59% (P<0.01), and 49%, respectively.
All the beating cardiomyocytes derived from the ES cells expressed cardiac-specific
proteins for α-actinin and troponin T. Among them, 1×10-7 mol/L icariin
treatment resulted in a significantly advanced and increased mRNA level of α-
cardiac major histocompatibility complex (MHC) and myosin light chain 2v (MLC-
2v) in EBs in the early cardiac developmental stage. Before shifting to the
cardiomyocyte phenotype, icariin could evoke the accumulation of ES cells in
G0/G1 and accelerate apoptosis of the cell population (P<0.05). Conclusion:
Icariin facilitated the directional differentiation of ES cells into cardiomyocytes at
a concentration of 1×10-7 mol/L. The promoting effect of icariin on cardiac differentiation
was related to increasing and accelerating gene expression of α-cardiac
MHC and MLC-2v, as well as regulating the cell cycles and inducing apoptosis.
Keywords:
desmethylicaritin on the directional differentiation of embryonic stem (ES) cells
into cardiomyocytes in vitro. Methods: ES cells were cultivated as embryoid
bodies (EBs) in hanging drops with icariin, icaritin, or desmethylicaritin. ES cells
treated with retinoic acid and with solvent were used as positive and negative
controls, respectively. The cardiomyocytes derived from the ES cells were verified
using immunocytochemistry. The expression of cardiac developmentaldependent
genes was detected using the reverse transcription-polymerase chain
reaction (RT-PCR) method. Cell cycle distribution and apoptosis were analyzed
using flow cytometry to determine the partly inducible effect mechanisms involved.
Results: The total percentage of beating EBs treated with 1×10-7 mol/L icariin,
icaritin, or desmethylicaritin was 87% (P<0.01), 59% (P<0.01), and 49%, respectively.
All the beating cardiomyocytes derived from the ES cells expressed cardiac-specific
proteins for α-actinin and troponin T. Among them, 1×10-7 mol/L icariin
treatment resulted in a significantly advanced and increased mRNA level of α-
cardiac major histocompatibility complex (MHC) and myosin light chain 2v (MLC-
2v) in EBs in the early cardiac developmental stage. Before shifting to the
cardiomyocyte phenotype, icariin could evoke the accumulation of ES cells in
G0/G1 and accelerate apoptosis of the cell population (P<0.05). Conclusion:
Icariin facilitated the directional differentiation of ES cells into cardiomyocytes at
a concentration of 1×10-7 mol/L. The promoting effect of icariin on cardiac differentiation
was related to increasing and accelerating gene expression of α-cardiac
MHC and MLC-2v, as well as regulating the cell cycles and inducing apoptosis.