Inducible effects of icariin, icaritin, and desmethylicaritin on directional differentiation of embryonic stem cells into cardiomyocytes in vitro.
OBJECTIVE
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 developmental-dependent 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 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 a-actinin and troponin T. Among them, 10(-7) mol/L icariin treatment resulted in a significantly advanced and increased mRNA level of a-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).


CONCLUSIONS
Icariin facilitated the directional differentiation of ES cells into cardiomyocytes at a concentration of 10(-7) mol/L. The promoting effect of icariin on cardiac differentiation was related to increasing and accelerating gene expression of a-cardiac MHC and MLC-2v, as well as regulating the cell cycles and inducing apoptosis.
Zhu
Dan-yan
DY
Department of Pharmacology and Toxicology, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou 310031, China.
Lou
Yi-jia
YJ
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002453:Cell Cycle / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects*; D002478:Cells, Cultured; D004622:Embryo, Mammalian / Q000166:cytology; D034181:Epimedium / Q000737:chemistry; D005260:Female; D005419:Flavonoids / Q000302:isolation & purification / Q000494:pharmacology*; D005786:Gene Expression Regulation; D008297:Male; D051379:Mice; D032383:Myocytes, Cardiac / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D018995:Myosin Heavy Chains / Q000096:biosynthesis* / Q000235:genetics; D018994:Myosin Light Chains / Q000096:biosynthesis / Q000235:genetics; D010946:Plants, Medicinal / Q000737:chemistry; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D013234:Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism
D005419:Flavonoids; D018994:Myosin Light Chains; D012333:RNA, Messenger; C499404:desmethylicaritin; D018995:Myosin Heavy Chains; C499403:icaritin; C056599:icariin