1. Academic Validation
  2. RA signaling pathway combined with Wnt signaling pathway regulates human-induced pluripotent stem cells (hiPSCs) differentiation to sinus node-like cells

RA signaling pathway combined with Wnt signaling pathway regulates human-induced pluripotent stem cells (hiPSCs) differentiation to sinus node-like cells

  • Stem Cell Res Ther. 2022 Jul 18;13(1):324. doi: 10.1186/s13287-022-03006-8.
Lin Yin 1 2 3 Feng-Yuan Wang 1 2 3 Wei Zhang 1 2 3 Xi Wang 1 2 3 Yan-Hong Tang 1 2 3 Teng Wang 1 2 3 Yu-Ting Chen 1 2 3 Cong-Xin Huang 4 5 6
Affiliations

Affiliations

  • 1 Department of Cardiology, Renmin Hospital of Wuhan University, 238 Jiefang Road, Wuchang, Wuhan, 430060, Hubei, People's Republic of China.
  • 2 Cardiovascular Research Institute, Wuhan University, Wuhan, 430060, People's Republic of China.
  • 3 Hubei Key Laboratory of Cardiology, Wuhan, 430060, People's Republic of China.
  • 4 Department of Cardiology, Renmin Hospital of Wuhan University, 238 Jiefang Road, Wuchang, Wuhan, 430060, Hubei, People's Republic of China. huangcongxin@vip.163.com.
  • 5 Cardiovascular Research Institute, Wuhan University, Wuhan, 430060, People's Republic of China. huangcongxin@vip.163.com.
  • 6 Hubei Key Laboratory of Cardiology, Wuhan, 430060, People's Republic of China. huangcongxin@vip.163.com.
Abstract

Background: The source of SAN is debated among researchers. Many studies have shown that RA and Wnt signaling are involved in heart development. In this study, we investigated the role of retinoic acid (RA) and Wnt signaling in the induction of sinus node-like cells.

Methods: The experimental samples were divided into four groups: control group (CHIR = 0), CHIR = 3, RA + CHIR = 0 andRA + CHIR = 3. After 20 days of differentiation, Western blot, RT-qPCR, immunofluorescence and flow cytometry were performed to identify sinus node-like cells. Finally, whole-cell patch clamp technique was used to record pacing funny current and action potential (AP) in four groups.

Results: The best intervention method used in our experiment was RA = 0.25 µmol/L D5-D9 + CHIR = 3 µmol/L D5-D7. Results showed that CHIR can increase the expression of ISL-1 and TBX3, while RA mainly elevated Shox2. Immunofluorescence assay and flow cytometry further illustrated that combining RA with CHIR can induce sinus node-like cells (CTNT+Shox2+Nkx2.5-). Moreover, CHIR might reduce the frequency of cell beats, but in conjunction with RA could partly compensate for this side effect. Whole cell patch clamps were able to record funny current and the typical sinus node AP in the experimental group, which did not appear in the control group.

Conclusions: Combining RA with Wnt signaling within a specific period can induce sinus node-like cells.

Keywords

Biological pacemaker; Human-induced pluripotent stem cells; Retinoic acid.

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