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Tbx18-dependent differentiation of brown adipose tissue-derived stem cells toward cardiac pacemaker cells

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Abstract

A cell-sourced biological pacemaker is a promising therapeutic approach for sick sinus syndrome (SSS) or severe atrial ventricular block (AVB). Adipose tissue-derived stem cells (ATSCs), which are optimal candidate cells for possible use in regenerative therapy for acute or chronic myocardial injury, have the potential to differentiate into spontaneous beating cardiomyocytes. However, the pacemaker characteristics of the beating cells need to be confirmed, and little is known about the underlying differential mechanism. In this study, we found that brown adipose tissue-derived stem cells (BATSCs) in mice could differentiate into spontaneous beating cells in 15% FBS Dulbecco’s modified Eagle’s medium (DMEM) without additional treatment. Subsequently, we provide additional evidence, including data regarding ultrastructure, protein expression, electrophysiology, and pharmacology, to support the differentiation of BATSCs into a cardiac pacemaker phenotype during the course of early cultivation. Furthermore, we found that silencing Tbx18, a key transcription factor in the development of pacemaker cells, terminated the differentiation of BATSCs into a pacemaker phenotype, suggesting that Tbx18 is required to direct BATSCs toward a cardiac pacemaker fate. The expression of Tbx3 and shox2, the other two important transcription factors in the development of pacemaker cells, was decreased by silencing Tbx18, which suggests that Tbx18 mediates the differentiation of BATSCs into a pacemaker phenotype via these two downstream transcription factors.

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Acknowledgements

This work was supported by grants from the Natural Science Foundation of China (31170934, 31271050, and 81271717).

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    Authors and Affiliations

    1. Research Center of Regenerative Medicine, Second Military Medical University, Shanghai, China

      Lei Chen, Zi-Jun Deng, Rui-Juan Ji, Xi Zhang, Chuan-Sen Zhang, Yu-Quan Li & Xiang-Qun Yang

    2. Biochemistry and Molecular Biology Department, Second Military Medical University, Shanghai, China

      Jian-Sheng Zhou

    3. Department of Anatomy, Second Military Medical University, 800 Xiangyin Road, Shanghai, 200433, China

      Yu-Quan Li & Xiang-Qun Yang

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    1. Lei Chen
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    Correspondence to Yu-Quan Li or Xiang-Qun Yang.

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    Lei Chen and Zi-Jun Deng have contributed equally to this work.

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    11010_2017_3016_MOESM1_ESM.tif

    Supplement Fig. 1 The expression of markers at transcriptional levels in ATSCs, qRT-PCR was used to detected the gene expression of LPL, Leptin, PPAR γ2(a); Collagen II, Aggrecan, Sox9(b); OP, OC, Runx2(c) after 2-week induced. Expression levels were normalized to α-actin. Error bars represent by ± SD. One-tailed Student’s t-tests were used as appropriate to evaluate the statistical significance of differences between two groups.*P <0.05 vs WATSCs, #P <0.05 vs BATSCs. (TIF 4034 KB)

    11010_2017_3016_MOESM2_ESM.tif

    Supplementary Fig. 2 Western blot analysis of Tbx18, Tbx3, sarcomeric α-actin (Sr) and HCN4 expression in shTbx18-treated cells transfected with Tbx18. (a) Western blot analysis of Tbx18, Tbx3, Sr and HCN4 protein expression; (b) quantitative assessment of Tbx18, Tbx3, Sr and HCN4 protein levels using integrated optical density analyses. *P <0.01 compared to the corresponding values for the shTbx18 groups. #P <0.01 compared to the corresponding values for the untreated groups. (TIF 6712 KB)

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    Chen, L., Deng, ZJ., Zhou, JS. et al. Tbx18-dependent differentiation of brown adipose tissue-derived stem cells toward cardiac pacemaker cells. Mol Cell Biochem 433, 61–77 (2017). https://doi.org/10.1007/s11010-017-3016-y

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