Nobiletin, a hexamethoxyflavonoid from citrus pomace, attenuates G1 cell cycle arrest and apoptosis in hypoxia-induced human trophoblast cells of JEG-3 and BeWo via regulating the p53 signaling pathway
Background: Hypoxia is associated with abnormal cell apoptosis in trophoblast cells, which causes fetal growth restriction and related placental pathologies. Few effective methods for the prevention and treatment of placenta-related diseases exist. Natural products and functional foods have always b...
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Swedish Nutrition Foundation
2020-09-01
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Online Access: | https://foodandnutritionresearch.net/index.php/fnr/article/view/5649/13909 |
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Article |
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DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Mengling Zhang Jian Liu Rui Zhang Zengenni Liang Shenghua Ding Huanling Yu Yang Shan |
spellingShingle |
Mengling Zhang Jian Liu Rui Zhang Zengenni Liang Shenghua Ding Huanling Yu Yang Shan Nobiletin, a hexamethoxyflavonoid from citrus pomace, attenuates G1 cell cycle arrest and apoptosis in hypoxia-induced human trophoblast cells of JEG-3 and BeWo via regulating the p53 signaling pathway Food & Nutrition Research nobiletin p53 molecular docking molecular dynamics spectrum apoptosis trophoblast cells |
author_facet |
Mengling Zhang Jian Liu Rui Zhang Zengenni Liang Shenghua Ding Huanling Yu Yang Shan |
author_sort |
Mengling Zhang |
title |
Nobiletin, a hexamethoxyflavonoid from citrus pomace, attenuates G1 cell cycle arrest and apoptosis in hypoxia-induced human trophoblast cells of JEG-3 and BeWo via regulating the p53 signaling pathway |
title_short |
Nobiletin, a hexamethoxyflavonoid from citrus pomace, attenuates G1 cell cycle arrest and apoptosis in hypoxia-induced human trophoblast cells of JEG-3 and BeWo via regulating the p53 signaling pathway |
title_full |
Nobiletin, a hexamethoxyflavonoid from citrus pomace, attenuates G1 cell cycle arrest and apoptosis in hypoxia-induced human trophoblast cells of JEG-3 and BeWo via regulating the p53 signaling pathway |
title_fullStr |
Nobiletin, a hexamethoxyflavonoid from citrus pomace, attenuates G1 cell cycle arrest and apoptosis in hypoxia-induced human trophoblast cells of JEG-3 and BeWo via regulating the p53 signaling pathway |
title_full_unstemmed |
Nobiletin, a hexamethoxyflavonoid from citrus pomace, attenuates G1 cell cycle arrest and apoptosis in hypoxia-induced human trophoblast cells of JEG-3 and BeWo via regulating the p53 signaling pathway |
title_sort |
nobiletin, a hexamethoxyflavonoid from citrus pomace, attenuates g1 cell cycle arrest and apoptosis in hypoxia-induced human trophoblast cells of jeg-3 and bewo via regulating the p53 signaling pathway |
publisher |
Swedish Nutrition Foundation |
series |
Food & Nutrition Research |
issn |
1654-661X |
publishDate |
2020-09-01 |
description |
Background: Hypoxia is associated with abnormal cell apoptosis in trophoblast cells, which causes fetal growth restriction and related placental pathologies. Few effective methods for the prevention and treatment of placenta-related diseases exist. Natural products and functional foods have always been a rich source of potential anti-apoptotic drugs. Nobiletin (NOB), a hexamethoxyflavonoid derived from the citrus pomace, shows an anti-apoptotic activity, which is a non-toxic constituent of dietary phytochemicals approved by the Food and Drug Administration. However, their effects on hypoxia-induced human trophoblast cells have not been fully studied. Objective: The aim of this study was to investigate the protective effects of NOB on hypoxia-induced apoptosis of human trophoblast JEG-3 and BeWo cells, and their underlying mechanisms. Design: First, the protective effect of NOB on hypoxia-induced apoptosis of JEG-3 and BeWo cells was studied. Cell viability and membrane integrity were determined by CCK-8 assay and lactate dehydrogenase activity, respectively. Real Time Quantitative PCR (RT-qPCR) and Western blot analysis were used to detect the mRNA and protein levels of HIF1α. Propidium iodide (PI)-labeled flow cytometry was used to detect cell cycle distribution. Cell apoptosis was detected by flow cytometry with Annexin V-FITC and PI double staining, and the expression of apoptosis marker protein cl-PARP was detected by Western blot analysis. Then, the molecular mechanism of NOB against apoptosis was investigated. Computer molecular docking and dynamics were used to simulate the interaction between NOB and p53 protein, and this interaction was verified in vitro by Ultraviolet and visible spectrum (UV-visible spectroscopy), fluorescence spectroscopy and circular dichroism. Furthermore, the changes in the expression of p53 signaling pathway genes and proteins were detected by RT-qPCR and Western blot analysis, respectively. Results: Hypoxia treatment resulted in a decreased cell viability and cell membrane integrity in JEG-3 and BeWo cell lines, and an increased expression of HIF1α, cell cycle arrest in the G1 phase, and massive cell apoptosis, which were alleviated after NOB treatment. Molecular docking and dynamics simulations found that NOB spontaneously bonded to human p53 protein, leading to the change of protein conformation. The intermolecular interaction between NOB and human p53 protein was further confirmed by UV-visible spectroscopy, fluorescence spectroscopy and circular dichroism. After the treatment of 100 μM NOB, a down-regulation of mRNA and protein levels of p53 and p21 and an up-regulation of BCL2/BAX mRNA and protein ratio were observed in JEG-3 cells; however, there was also a down-regulation of mRNA and protein levels observed for p53 and p21 in BeWo cells after the treatment of NOB. The BCL2/BAX ratio of BeWo cells did not change after the treatment of 100 μM NOB. Conclusion: NOB attenuated hypoxia-induced apoptosis in JEG-3 and BeWo cell lines and might be a potential functional ingredient to prevent pregnancy-related diseases caused by hypoxia-induced apoptosis. These findings would also suggest the exploration and utilization of citrus resources, and the development of citrus industry. |
topic |
nobiletin p53 molecular docking molecular dynamics spectrum apoptosis trophoblast cells |
url |
https://foodandnutritionresearch.net/index.php/fnr/article/view/5649/13909 |
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AT menglingzhang nobiletinahexamethoxyflavonoidfromcitruspomaceattenuatesg1cellcyclearrestandapoptosisinhypoxiainducedhumantrophoblastcellsofjeg3andbewoviaregulatingthep53signalingpathway AT jianliu nobiletinahexamethoxyflavonoidfromcitruspomaceattenuatesg1cellcyclearrestandapoptosisinhypoxiainducedhumantrophoblastcellsofjeg3andbewoviaregulatingthep53signalingpathway AT ruizhang nobiletinahexamethoxyflavonoidfromcitruspomaceattenuatesg1cellcyclearrestandapoptosisinhypoxiainducedhumantrophoblastcellsofjeg3andbewoviaregulatingthep53signalingpathway AT zengenniliang nobiletinahexamethoxyflavonoidfromcitruspomaceattenuatesg1cellcyclearrestandapoptosisinhypoxiainducedhumantrophoblastcellsofjeg3andbewoviaregulatingthep53signalingpathway AT shenghuading nobiletinahexamethoxyflavonoidfromcitruspomaceattenuatesg1cellcyclearrestandapoptosisinhypoxiainducedhumantrophoblastcellsofjeg3andbewoviaregulatingthep53signalingpathway AT huanlingyu nobiletinahexamethoxyflavonoidfromcitruspomaceattenuatesg1cellcyclearrestandapoptosisinhypoxiainducedhumantrophoblastcellsofjeg3andbewoviaregulatingthep53signalingpathway AT yangshan nobiletinahexamethoxyflavonoidfromcitruspomaceattenuatesg1cellcyclearrestandapoptosisinhypoxiainducedhumantrophoblastcellsofjeg3andbewoviaregulatingthep53signalingpathway |
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doaj-bc32bc6d96fc4b2ab5fe01418a7aebbf2021-10-06T16:47:42ZengSwedish Nutrition FoundationFood & Nutrition Research1654-661X2020-09-01651710.29219/fnr.v65.56495649Nobiletin, a hexamethoxyflavonoid from citrus pomace, attenuates G1 cell cycle arrest and apoptosis in hypoxia-induced human trophoblast cells of JEG-3 and BeWo via regulating the p53 signaling pathwayMengling Zhang0Jian Liu1Rui Zhang2Zengenni Liang3Shenghua Ding4Huanling Yu5Yang Shan6Longping Branch Graduate School, Hunan University, Changsha, Hunan Province, ChinaLongping Branch Graduate School, Hunan University, Changsha, Hunan Province, ChinaSchool of Medical Humanity, Peking University, Beijing, ChinaLongping Branch Graduate School, Hunan University, Changsha, Hunan Province, ChinaLongping Branch Graduate School, Hunan University, Changsha, Hunan Province, ChinaSchool of Public Health, Beijing Key Laboratory of Environmental Toxicology, Capital Medical University, Beijing, ChinaLongping Branch Graduate School, Hunan University, Changsha, Hunan Province, ChinaBackground: Hypoxia is associated with abnormal cell apoptosis in trophoblast cells, which causes fetal growth restriction and related placental pathologies. Few effective methods for the prevention and treatment of placenta-related diseases exist. Natural products and functional foods have always been a rich source of potential anti-apoptotic drugs. Nobiletin (NOB), a hexamethoxyflavonoid derived from the citrus pomace, shows an anti-apoptotic activity, which is a non-toxic constituent of dietary phytochemicals approved by the Food and Drug Administration. However, their effects on hypoxia-induced human trophoblast cells have not been fully studied. Objective: The aim of this study was to investigate the protective effects of NOB on hypoxia-induced apoptosis of human trophoblast JEG-3 and BeWo cells, and their underlying mechanisms. Design: First, the protective effect of NOB on hypoxia-induced apoptosis of JEG-3 and BeWo cells was studied. Cell viability and membrane integrity were determined by CCK-8 assay and lactate dehydrogenase activity, respectively. Real Time Quantitative PCR (RT-qPCR) and Western blot analysis were used to detect the mRNA and protein levels of HIF1α. Propidium iodide (PI)-labeled flow cytometry was used to detect cell cycle distribution. Cell apoptosis was detected by flow cytometry with Annexin V-FITC and PI double staining, and the expression of apoptosis marker protein cl-PARP was detected by Western blot analysis. Then, the molecular mechanism of NOB against apoptosis was investigated. Computer molecular docking and dynamics were used to simulate the interaction between NOB and p53 protein, and this interaction was verified in vitro by Ultraviolet and visible spectrum (UV-visible spectroscopy), fluorescence spectroscopy and circular dichroism. Furthermore, the changes in the expression of p53 signaling pathway genes and proteins were detected by RT-qPCR and Western blot analysis, respectively. Results: Hypoxia treatment resulted in a decreased cell viability and cell membrane integrity in JEG-3 and BeWo cell lines, and an increased expression of HIF1α, cell cycle arrest in the G1 phase, and massive cell apoptosis, which were alleviated after NOB treatment. Molecular docking and dynamics simulations found that NOB spontaneously bonded to human p53 protein, leading to the change of protein conformation. The intermolecular interaction between NOB and human p53 protein was further confirmed by UV-visible spectroscopy, fluorescence spectroscopy and circular dichroism. After the treatment of 100 μM NOB, a down-regulation of mRNA and protein levels of p53 and p21 and an up-regulation of BCL2/BAX mRNA and protein ratio were observed in JEG-3 cells; however, there was also a down-regulation of mRNA and protein levels observed for p53 and p21 in BeWo cells after the treatment of NOB. The BCL2/BAX ratio of BeWo cells did not change after the treatment of 100 μM NOB. Conclusion: NOB attenuated hypoxia-induced apoptosis in JEG-3 and BeWo cell lines and might be a potential functional ingredient to prevent pregnancy-related diseases caused by hypoxia-induced apoptosis. These findings would also suggest the exploration and utilization of citrus resources, and the development of citrus industry.https://foodandnutritionresearch.net/index.php/fnr/article/view/5649/13909nobiletinp53molecular dockingmolecular dynamicsspectrumapoptosistrophoblast cells |