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The generation of germline competent rat embryonic stem cells (rESCs) allows the study of their lineage commitment. Here, we developed a highly efficient system for rESC-derived cardiomyocytes, and even the formation of three-dimensional (3D)-like cell clusters with cTNT and α-Actinin. We have validated that laminin can interact with membrane integrin to promote the phosphorylation of both phosphatidylinositol 3-kinase (PI3K) p85 and the focal adhesion kinase (FAK). In parallel, GATA4 was up-regulated. Upon inhibiting the integrin, laminin loses the effect on cardiomyocyte differentiation, accompanied with a down-regulation of phosphorylation level of PI3K p85 and FAK. Meanwhile, the expression of Gata4 was inhibited as well. Taken together, laminin is a crucial component in the differentiation of rESCs into cardiomyocytes through increasing their proliferation via interacting with integrin pathway. These results provide new insights into the pathways mediated by extracellular laminin involved in the fate of rESC-derived cardiomyocytes., (© 2019 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

Background: Pluripotent stem cell-derived cardiomyocytes (PSC-CMs) are widely used models for regenerative medicine and disease research. However, PSC-CMs are usually immature in morphology and functionality and the maturity of PSC-CMs could not be determined accurately. In order to reasonably interpret the experimental results obtained by PSC-CMs, it is necessary to evaluate the maturity of PSC-CMs and find the key genes related to maturation., Methods: Using the gene expression profiles of normal adult cardiac tissue and embryonic stem cell (ESC) samples, we identified gene pairs with identically relative expression orderings (REOs) within adult cardiac tissue but reversely identical in ESCs. Then, for a PSC-CM model, we calculated the maturity score as the percentage of these gene pairs that exhibit the same REOs in adult cardiac tissue. Lastly, the CellComp method was used to identify the maturation-related genes., Results: The maturity score increased gradually from 0.8401 for 18-week fetal cardiac tissue to 0.9997 for adult cardiac tissue. For four human PSC-CM models, the mature scores increased with prolonged culture time but were all below 0.8. The genes involved in energy metabolism, angiogenesis, immunity, and proliferation were dysregulated in the 1-year PSC-CMs compared with adult cardiac tissue., Conclusion: We proposed a qualitative transcriptional signature to score the maturity degree of PSC-CMs. This score can reasonably track the maturity of PSC-CMs and be used to compare different PSC-CM culture methods.

Background and Objectives: The control of cardiomyocyte differentiation is tightly linked to microRNAs (miRNAs), which have been emerging as important players in heart development. However, the regulation mechanisms mediated by miRNAs in early heart development remains speculative. Here, we evaluated the impact of miR-26b during the progression of cardiomyocyte differentiation from the P19 cell line., Materials and Methods: The overexpression of miR-26b in P19 cells was performed by transduction with lentivirus vector. The levels of cardiac-related genes during P19 cell differentiation were detected using quantitative real-time PCR for mRNA abundance and Western blots for protein expression. ICG-001 was applied to elucidate the role of β-catenin on P19 cells differentiation. The Cell Counting kit-8 (CCK-8) was used to monitor the cell proliferation. The target genes of miR-26b were validated using the dual luciferase reporter system., Results: Overexpression of miR-26b upregulates the expression level of cardiomyocyte-related genes such as Gata4, cTNT, α-MHC and α-Actinin that comprehensively represent cardiomyocyte differentiation by effecting Wnt5a signalling and Gsk3β activity. However, ICG-001 blocks the differentiation along with inhibition of the cell proliferation. In addition, miR-26b also regulates CyclinD1 to promote P19 cell proliferation, thereby, demonstrating the rapid aggregation and differentiation programming of these cells into cardiomyocytic types., Conclusions: Our results indicated that miR-26b exerts a role on promoting cardiomyocyte differentiation of P19 cells by controlling the canonical and non-canonical Wnt signalling., (© 2017 John Wiley & Sons Ltd.)

Heart is the first organ to function during mammalian embryogenesis. The differentiation of embryonic stem cells (ESCs) into cardiomyocyte is complex and dynamic, which involves 4 differentiation stages including ESCs, mesoderm, cardiac precursor, and terminal cardiomyocytes. Abnormal expression of certain genes can lead to congenital heart diseases during cardiomyocyte differentiation. Epigenetic regulation plays a crucial role on the switch of gene activation and deactivation during cardiomyocyte differentiation. Non-coding RNA, particularly microRNA and long non-coding RNA, may significantly influence gene expression. Exploring the regulatory roles of non-coding RNA in cardiomyocyte differentiation may contribute to the understanding of the functions of myocardial cells and mechanism of congenital heart diseases.

The aryl hydrocarbon receptor (AhR) is the receptor for 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), benzo(a)pyrene (BaP) and other exogenous compounds. In human sebocytes, TCDD and BaP were found to activate the expression of multiple genes, including cytochrome P450 1A1 (CYP1A1), and inhibit lipid synthesis via AhR, while little is known about endogenous functions of the AhR. In order to expand this knowledge, we analyzed the impact of AhR knockdown on lipid synthesis as well as on cell differentiation of SZ95 sebocytes in vitro and observed that lipid synthesis was significantly induced in AhR silenced SZ95 sebocytes. In line with this result, expression of lipogenesis-associated genes, such as peroxisome proliferator activated receptor (PPAR) δ and PPARγ, was increased. Morphological changes with smaller cells in size but more abundant cytoplasmic lipids were observed in AhR silenced SZ95 sebocytes compared with the AhR activated cells. Besides, the expression of keratin 7, an early sebaceous differentiation marker, was increased, while the expression of the terminal sebocyte differentiation marker epithelial membrane antigen (EMA) was reduced. Moreover, the terminal keratinocyte differentiation markers keratin 10 and involucrin, and the AhR downstream protein CYP1A1 were reduced after AhR silencing. To the best of our knowledge, we provide evidence that in the absence of exogenous ligands, the AhR inhibits lipid synthesis and involves in cell differentiation of human SZ95 sebocytes, which indicates the physiological function of this receptor in human sebocytes., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Background: Neural crest defects lead to congenital heart disease involving outflow tract malformation. Integrin-linked-kinase (ILK) plays important roles in multiple cellular processes and embryogenesis. ILK is expressed in the neural crest, but its role in neural crest and outflow tract morphogenesis remains unknown., Results: We ablated ILK specifically in the neural crest using the Wnt1-Cre transgene. ILK ablation resulted in abnormal migration and overpopulation of neural crest cells in the pharyngeal arches and outflow tract and a significant reduction in the expression of neural cell adhesion molecule (NCAM) and extracellular matrix components. ILK mutant embryos exhibited an enlarged common arterial trunk and ventricular septal defect. Reduced smooth muscle differentiation, but increased ossification and neurogenesis/innervation were observed in ILK mutant outflow tract that may partly be due to reduced transforming growth factor β2 (TGFβ2) but increased bone morphogenetic protein (BMP) signaling. Consistent with these observations, microarray analysis of fluorescence-activated cell sorting (FACS)-sorted neural crest cells revealed reduced expression of genes associated with muscle differentiation, but increased expression of genes of neurogenesis and osteogenesis., Conclusions: Our results demonstrate that ILK plays essential roles in neural crest and outflow tract development by mediating complex crosstalk between cell matrix and multiple signaling pathways. Changes in these pathways may collectively result in the unique neural crest and outflow tract phenotypes observed in ILK mutants.

In the past few decades, embryonic stem cells (ESCs) were of great interest as a model system for studying early developmental processes and because of their potential therapeutic applications in regenerative medicine. However, the underlying mechanisms of ESC differentiation remain unclear, which limits our exploration of the therapeutic potential of stem cells. Fortunately, the increasing quantity and diversity of biological datasets can provide us with opportunities to explore the biological secrets. However, taking advantage of diverse biological information to facilitate the advancement of ESC research still remains a challenge. Here, we propose a scalable, efficient and flexible function prediction framework that integrates diverse biological information using a simple weighted strategy, for uncovering the genetic determinants of mouse ESC differentiation. The advantage of this approach is that it can make predictions based on dynamic information fusion, owing to the simple weighted strategy. With this approach, we identified 30 genes that had been reported to be associated with differentiation of stem cells, which we regard to be associated with differentiation or pluripotency in embryonic stem cells. We also predicted 70 genes as candidates for contributing to differentiation, which requires further confirmation. As a whole, our results showed that this strategy could be applied as a useful tool for ESC research.

microRNAs (miRNAs) play essential roles in cardiogenesis. The altered expression of miRNAs can result in cardiac malformations by inducing abnormalities in the behavior of cardiac cells. However, the role of miR-10a in the regulation of cardiomyocyte progenitor cells (CMPCs) remains undetermined. In the present study, we found that up- or down-regulation of miR-10a inhibited or promoted the proliferation of human CMPCs, respectively, without affecting their differentiation toward cardiomyocytes. miR-10a bound to GATA6 directly and reduced GATA6 expression. Over-expression of GATA6 greatly attenuated the miR-10a-mediated inhibitory effect on the proliferation of human CMPCs. Thus, our results indicate that miR-10a could effectively modulate the proliferation of human CMPCs by targeting GATA6. The finding provides novel insights into the potency of miR-10a during heart development. [ABSTRACT FROM AUTHOR]