Rapamycin Promotes Cardiomyocyte Differentiation of Human Induced Pluripotent Stem Cells in a Stage-Dependent Manner

IntroductionCNV is a vital pathogenic factor of congenital heart disease (CHD). However, few CNVs have been reported for total anomalous pulmonary venous connection (TAPVC), which is a rare form of CHD. Using case-control study, we identified 15q11.2 deletion associated with TAPVC. We then used a TAPVC trio as model to reveal possible molecular basis of 15q11.2 microdeletion.MethodsCNVplex and Chromosomal Microarray were used to identify and validate CNVs in samples from 231 TAPVC cases and 200 healthy controls from Shanghai Children’s Medical Center. In vitro cardiomyocyte differentiation of induced pluripotent stem cells from peripheral blood mononuclear cells for a TAPVC trio with paternal inherited 15q11.2 deletion was performed to characterise the effect of the deletion on cardiomyocyte differentiation and gene expression.ResultsThe 15q11.2 microdeletion was significantly enriched in patients with TAPVC compared with healthy control (13/231 in patients vs 0/200 in controls, p=5.872×10−2, Bonferroni adjusted) using Fisher’s exact test. Induced pluripotent stem cells from the proband could not differentiate into normal cardiomyocyte. Transcriptomic analysis identified a number of differentially expressed genes in the 15q11.2 deletion carriers of the family. TAPVC disease-causing genes such as PITX2, NKX2-5 and ANKRD1 showed significantly higher expression in the proband compared with her healthy mother. Knockdown of TUBGCP5 could lead to abnormal cardiomyocyte differentiation.ConclusionWe discovered that the 15q11.2 deletion is significantly associated with TAPVC. Gene expression profile that might arise from 15q11.2 deletion for a TAPVC family was characterised using cell experiments.

Download Full-text

Electrical Stimulation Promotes Cardiac Differentiation of Human Induced Pluripotent Stem Cells

Stem Cells International ◽

10.1155/2016/1718041 ◽

2016 ◽

Vol 2016 ◽

pp. 1-12 ◽

Cited By ~ 35

Author(s):

Damián Hernández ◽

Rodney Millard ◽

Priyadharshini Sivakumaran ◽

Raymond C. B. Wong ◽

Duncan E. Crombie ◽

...

Keyword(s):

Stem Cells ◽

Electrical Stimulation ◽

Induced Pluripotent Stem Cells ◽

Cell Line ◽

Pluripotent Stem Cells ◽

Embryoid Bodies ◽

Cardiac Differentiation ◽

Dependent Manner ◽

Human Ipscs ◽

Induced Pluripotent

Background.Human induced pluripotent stem cells (iPSCs) are an attractive source of cardiomyocytes for cardiac repair and regeneration. In this study, we aim to determine whether acute electrical stimulation of human iPSCs can promote their differentiation to cardiomyocytes.Methods. Human iPSCs were differentiated to cardiac cells by forming embryoid bodies (EBs) for 5 days. EBs were then subjected to brief electrical stimulation and plated down for 14 days.Results. In iPS(Foreskin)-2 cell line, brief electrical stimulation at 65 mV/mm or 200 mV/mm for 5 min significantly increased the percentage of beating EBs present by day 14 after plating. Acute electrical stimulation also significantly increased the cardiac gene expression ofACTC1,TNNT2,MYH7, andMYL7. However, the cardiogenic effect of electrical stimulation was not reproducible in another iPS cell line, CERA007c6. Beating EBs from control and electrically stimulated groups expressed various cardiac-specific transcription factors and contractile muscle markers. Beating EBs were also shown to cycle calcium and were responsive to the chronotropic agents, isoproterenol and carbamylcholine, in a concentration-dependent manner.Conclusions. Our results demonstrate that brief electrical stimulation can promote cardiac differentiation of human iPS cells. The cardiogenic effect of brief electrical stimulation is dependent on the cell line used.

Download Full-text

Differentiation of Mature Eosinophils From Human Embryonic and Induced Pluripotent Stem Cells.

Blood ◽

10.1182/blood.v116.21.1594.1594 ◽

2010 ◽

Vol 116 (21) ◽

pp. 1594-1594

Author(s):

Wenyu Yang ◽

Feng Ma ◽

Kenji Matsumoto ◽

Natsumi Nishihama ◽

Hiroshi Sagara ◽

...

Keyword(s):

Stem Cells ◽

Induced Pluripotent Stem Cells ◽

Pluripotent Stem Cells ◽

Conflicts Of Interest ◽

Allergic Diseases ◽

Time Dependent ◽

Dependent Manner ◽

Hematopoietic Progenitors ◽

Culture Cells ◽

Induced Pluripotent

Abstract Abstract 1594 Eosinophils are multifunctional leukocytes implicated in the pathogenesis of numerous inflammatory processes. As the major effectors, eosinophils function in a variety of biological responses, allergic diseases and helminth infections. It is generally accepted human eosinophils develop through a pathway initially sharing common feature with basophils. However, there lacks a clear chart for early development of human eosinophils, such as during embryonic or fetal stages. We recently established an efficient method for producing eosinophils from human embryonic and induced pluripotent stem cells (hESC/iPSCs). By a two-step induction, we first generated multipotential hematopoietic progenitors by co-culturing hESC/iPSCs with mouse AGM-derived stromal cells for 2 weeks. Then, total co-culture cells were transferred into suspension culture favoring eosinophil development with addition of IL-3 and other factors (SCF, IL-6, TPO, Flt-3 ligand) . The maturation of hESC/iPSC -derived eosinophils was shown in a time-dependent manner, first co-expressing eosinophil-and basophil-specific markers [eosinophil peroxidase (EPO), and 2D7, respectively], then the portion of eosinophil markers gradually increased while that of basophil markers decreased (EPO+ cells from 56.4% at day 7 to 94.4% at day 21, while 2D7+ cells from 62.8% to 25.7%, respectively), typically mimicking the development of eosinophils from human adult hematopoietic progenitors. By flowcytometric analysis, an eosinophil-specific surface marker, Siglec-8, was also expressed on these hESC/iPSC-derived eosinophils in a time-dependent manner (from 10.8% at day 7 to 91.3% at day 21), paralleling to those with EPO. The expression of eosinophil-specific granule cationic proteins (EPO, MBP, ECP, EDN) and IL-5 receptor mRNA was also detected by RT-PCR. Furthermore, transmission electron microscopy (TEM) observation confirmed the eosinophil property. Eosinophils derived from hiPSCs hold similar characteristics as those from hESCs. The function of hES/hiPSC-derived eosinophils is being under investigation. Our study provides an experimental model for exploring early genesis of eosinophils, especially in uncovering the mechanisms controlling the development of the initial innate immune system of human being in normal and diseased individuals. Disclosures: No relevant conflicts of interest to declare.

Download Full-text