Anda Zhao1, Kena Zhao1, Yuanqing Xia1, Jiajun Lyu1, Yiting Chen1, Shenghui Li1,2. 1. School of Public Health, Shanghai Jiao Tong University School of Medicine, Shanghai, China. 2. MOE-Shanghai Key Laboratory of Children's Environmental Health, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
Abstract
BACKGROUND: Our recent epidemiological study revealed that maternal sleep during the periconceptional period should be involved in the risk of congenital heart disease (CHD) in offspring. Melatonin, a sleep related hormone, has been suggested to play a crucial role in embryonic development based on the emerging evidence. In this study, we set out to assess the effect of melatonin on the embryonic cardiac cell growth and to explore the underlying mechanisms. METHODS: We observed the effect of different gradient doses of melatonin as 10, 100, or 1,000 μM on cell proliferation in H9c2 embryonic rat cardiac cells. Furthermore, flow cytometry was applied to evaluate the impact on apoptosis and cell cycle. RNA-seq was conducted to screen the changes in expression of mRNA and signaling pathways. Quantitative Real-Time-PCR (qRT-PCR) was then conducted to validate the results. RESULTS: It was observed that melatonin could inhibit H9c2 cell growth, at the doses of 100 and 1,000 μM, but not at 10 μM. Moreover, melatonin ranged from 100 to 1,000 μM could instigate cell cycle arrest at G1 phase and simulate apoptosis, in a dose-dependent manner. In addition, melatonin was found to down-regulate the expression of a number of genes, which are related to heart development (SPARC, IFITM3, TNNT2, LOX), and PI3K-Akt signaling pathway activation (FN1, HSP90B1, THBS1, MFGE8, and CLU). CONCLUSIONS: Our findings suggested that high level of melatonin could be capable of inhibiting growth through the induction of apoptosis and cell cycle arrest via PI3K-AKT signaling pathway, thereby interfering with embryonic heart development. Considering this study is based on H9c2 embryonic rat cardiac cells, future additional studies using human embryonic cardiac cell are warranted.
BACKGROUND: Our recent epidemiological study revealed that maternal sleep during the periconceptional period should be involved in the risk of congenital heart disease (CHD) in offspring. Melatonin, a sleep related hormone, has been suggested to play a crucial role in embryonic development based on the emerging evidence. In this study, we set out to assess the effect of melatonin on the embryonic cardiac cell growth and to explore the underlying mechanisms. METHODS: We observed the effect of different gradient doses of melatonin as 10, 100, or 1,000 μM on cell proliferation in H9c2 embryonic rat cardiac cells. Furthermore, flow cytometry was applied to evaluate the impact on apoptosis and cell cycle. RNA-seq was conducted to screen the changes in expression of mRNA and signaling pathways. Quantitative Real-Time-PCR (qRT-PCR) was then conducted to validate the results. RESULTS: It was observed that melatonin could inhibit H9c2 cell growth, at the doses of 100 and 1,000 μM, but not at 10 μM. Moreover, melatonin ranged from 100 to 1,000 μM could instigate cell cycle arrest at G1 phase and simulate apoptosis, in a dose-dependent manner. In addition, melatonin was found to down-regulate the expression of a number of genes, which are related to heart development (SPARC, IFITM3, TNNT2, LOX), and PI3K-Akt signaling pathway activation (FN1, HSP90B1, THBS1, MFGE8, and CLU). CONCLUSIONS: Our findings suggested that high level of melatonin could be capable of inhibiting growth through the induction of apoptosis and cell cycle arrest via PI3K-AKT signaling pathway, thereby interfering with embryonic heart development. Considering this study is based on H9c2 embryonic rat cardiac cells, future additional studies using human embryonic cardiac cell are warranted.