Anti-cancer effects of 3, 3'-diindolylmethane on human hepatocellular carcinoma cells is enhanced by calcium ionophore: The role of cytosolic Ca2+ and P38 mapk

Yuanyue Jiang, Yanfei Fang, Yang Ye, Xinming Xu, Bingfang Wang, Jie Gu, Michael Aschner, Jian Chen, Rongzhu Lu

Research output: Contribution to journalArticle

Abstract

Purpose: 3,3'-Diindolylmethane (DIM), derived from indole-3-carbinol (I3C) in the Brassica species of cruciferous vegetables, has anticancer effects, but its exact underlying mechanism of action is unknown. We explored the roles of cytosolic free calcium ([Ca2+]i) and p38 MAPK in the anti-cancer effects of DIM in human hepatocellular carcinoma cells. Methods: Cell proliferation was measured with a Cell Counting Kit-8 (CCK-8) and the clonogenic formation assay. Cell apoptosis was examined by?ow cytometric analysis and Hoechst dye staining. Cleaved-caspase3, cleaved-PARP, Bax, total, and phosphorylated p38 MAPK were assayed by western blotting. [Ca2+]i was measured with Fluo-3/AM by?uorescence microscopy. A23187, a calcium ionophore, was used to increase [Ca2+]i levels. Results: DIM inhibited cell proliferation in both SMMC-7721 and HepG2 cells in a concentration-and time-dependent manner. DIM also enhanced phosphorylation of p38 MAPK (p-p38), which was attenuated by SB203580. The proliferation inhibition and apoptosis induction by DIM were also blunted. In addition, DIM increased [Ca2+]i in HCC cells, and this effect was inhibited by the calcium chelator, BAPTA-AM, resulting in reduced p-p38 MAPK activation and apoptosis in DIM-treated cells, though the proliferation inhibition by DIM was unchanged. However, the DIM-induced cell proliferation inhibition and apoptosis were signifcantly enhanced by A23187, a selective calcium ionophore, which was attributed to exaggerated p-p38 MAPK. Conclusions: The calcium ionophore enhanced DIM-induced anti-cancer effects in hepatocellular carcinoma cells, secondary to [Ca2+]i-dependent activation of p38 MAPK. Treatment with a combination of DIM and calcium ionophore may offer a new approach to enhance the chemotherapeutic effcacy in liver cancer.

Original languageEnglish (US)
Article number1167
JournalFrontiers in Pharmacology
Volume10
DOIs
StatePublished - Jan 1 2019

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Calcium Ionophores
Hepatocellular Carcinoma
p38 Mitogen-Activated Protein Kinases
Neoplasms
Cell Proliferation
Apoptosis
Phosphorylation
Calcimycin
diindolylmethane
3,3'-diindolylmethane
Brassica
Hep G2 Cells
Liver Neoplasms
Vegetables
Microscopy
Coloring Agents
Western Blotting
Staining and Labeling
Calcium

Keywords

  • 3,3'-diindolylmethane
  • Apoptosis
  • Cytosolic Ca
  • Hepatocellular carcinoma cells
  • P38 MAPK
  • Proliferation

ASJC Scopus subject areas

  • Pharmacology
  • Pharmacology (medical)

Cite this

Anti-cancer effects of 3, 3'-diindolylmethane on human hepatocellular carcinoma cells is enhanced by calcium ionophore : The role of cytosolic Ca2+ and P38 mapk. / Jiang, Yuanyue; Fang, Yanfei; Ye, Yang; Xu, Xinming; Wang, Bingfang; Gu, Jie; Aschner, Michael; Chen, Jian; Lu, Rongzhu.

In: Frontiers in Pharmacology, Vol. 10, 1167, 01.01.2019.

Research output: Contribution to journalArticle

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abstract = "Purpose: 3,3'-Diindolylmethane (DIM), derived from indole-3-carbinol (I3C) in the Brassica species of cruciferous vegetables, has anticancer effects, but its exact underlying mechanism of action is unknown. We explored the roles of cytosolic free calcium ([Ca2+]i) and p38 MAPK in the anti-cancer effects of DIM in human hepatocellular carcinoma cells. Methods: Cell proliferation was measured with a Cell Counting Kit-8 (CCK-8) and the clonogenic formation assay. Cell apoptosis was examined by?ow cytometric analysis and Hoechst dye staining. Cleaved-caspase3, cleaved-PARP, Bax, total, and phosphorylated p38 MAPK were assayed by western blotting. [Ca2+]i was measured with Fluo-3/AM by?uorescence microscopy. A23187, a calcium ionophore, was used to increase [Ca2+]i levels. Results: DIM inhibited cell proliferation in both SMMC-7721 and HepG2 cells in a concentration-and time-dependent manner. DIM also enhanced phosphorylation of p38 MAPK (p-p38), which was attenuated by SB203580. The proliferation inhibition and apoptosis induction by DIM were also blunted. In addition, DIM increased [Ca2+]i in HCC cells, and this effect was inhibited by the calcium chelator, BAPTA-AM, resulting in reduced p-p38 MAPK activation and apoptosis in DIM-treated cells, though the proliferation inhibition by DIM was unchanged. However, the DIM-induced cell proliferation inhibition and apoptosis were signifcantly enhanced by A23187, a selective calcium ionophore, which was attributed to exaggerated p-p38 MAPK. Conclusions: The calcium ionophore enhanced DIM-induced anti-cancer effects in hepatocellular carcinoma cells, secondary to [Ca2+]i-dependent activation of p38 MAPK. Treatment with a combination of DIM and calcium ionophore may offer a new approach to enhance the chemotherapeutic effcacy in liver cancer.",
keywords = "3,3'-diindolylmethane, Apoptosis, Cytosolic Ca, Hepatocellular carcinoma cells, P38 MAPK, Proliferation",
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T1 - Anti-cancer effects of 3, 3'-diindolylmethane on human hepatocellular carcinoma cells is enhanced by calcium ionophore

T2 - The role of cytosolic Ca2+ and P38 mapk

AU - Jiang, Yuanyue

AU - Fang, Yanfei

AU - Ye, Yang

AU - Xu, Xinming

AU - Wang, Bingfang

AU - Gu, Jie

AU - Aschner, Michael

AU - Chen, Jian

AU - Lu, Rongzhu

PY - 2019/1/1

Y1 - 2019/1/1

N2 - Purpose: 3,3'-Diindolylmethane (DIM), derived from indole-3-carbinol (I3C) in the Brassica species of cruciferous vegetables, has anticancer effects, but its exact underlying mechanism of action is unknown. We explored the roles of cytosolic free calcium ([Ca2+]i) and p38 MAPK in the anti-cancer effects of DIM in human hepatocellular carcinoma cells. Methods: Cell proliferation was measured with a Cell Counting Kit-8 (CCK-8) and the clonogenic formation assay. Cell apoptosis was examined by?ow cytometric analysis and Hoechst dye staining. Cleaved-caspase3, cleaved-PARP, Bax, total, and phosphorylated p38 MAPK were assayed by western blotting. [Ca2+]i was measured with Fluo-3/AM by?uorescence microscopy. A23187, a calcium ionophore, was used to increase [Ca2+]i levels. Results: DIM inhibited cell proliferation in both SMMC-7721 and HepG2 cells in a concentration-and time-dependent manner. DIM also enhanced phosphorylation of p38 MAPK (p-p38), which was attenuated by SB203580. The proliferation inhibition and apoptosis induction by DIM were also blunted. In addition, DIM increased [Ca2+]i in HCC cells, and this effect was inhibited by the calcium chelator, BAPTA-AM, resulting in reduced p-p38 MAPK activation and apoptosis in DIM-treated cells, though the proliferation inhibition by DIM was unchanged. However, the DIM-induced cell proliferation inhibition and apoptosis were signifcantly enhanced by A23187, a selective calcium ionophore, which was attributed to exaggerated p-p38 MAPK. Conclusions: The calcium ionophore enhanced DIM-induced anti-cancer effects in hepatocellular carcinoma cells, secondary to [Ca2+]i-dependent activation of p38 MAPK. Treatment with a combination of DIM and calcium ionophore may offer a new approach to enhance the chemotherapeutic effcacy in liver cancer.

AB - Purpose: 3,3'-Diindolylmethane (DIM), derived from indole-3-carbinol (I3C) in the Brassica species of cruciferous vegetables, has anticancer effects, but its exact underlying mechanism of action is unknown. We explored the roles of cytosolic free calcium ([Ca2+]i) and p38 MAPK in the anti-cancer effects of DIM in human hepatocellular carcinoma cells. Methods: Cell proliferation was measured with a Cell Counting Kit-8 (CCK-8) and the clonogenic formation assay. Cell apoptosis was examined by?ow cytometric analysis and Hoechst dye staining. Cleaved-caspase3, cleaved-PARP, Bax, total, and phosphorylated p38 MAPK were assayed by western blotting. [Ca2+]i was measured with Fluo-3/AM by?uorescence microscopy. A23187, a calcium ionophore, was used to increase [Ca2+]i levels. Results: DIM inhibited cell proliferation in both SMMC-7721 and HepG2 cells in a concentration-and time-dependent manner. DIM also enhanced phosphorylation of p38 MAPK (p-p38), which was attenuated by SB203580. The proliferation inhibition and apoptosis induction by DIM were also blunted. In addition, DIM increased [Ca2+]i in HCC cells, and this effect was inhibited by the calcium chelator, BAPTA-AM, resulting in reduced p-p38 MAPK activation and apoptosis in DIM-treated cells, though the proliferation inhibition by DIM was unchanged. However, the DIM-induced cell proliferation inhibition and apoptosis were signifcantly enhanced by A23187, a selective calcium ionophore, which was attributed to exaggerated p-p38 MAPK. Conclusions: The calcium ionophore enhanced DIM-induced anti-cancer effects in hepatocellular carcinoma cells, secondary to [Ca2+]i-dependent activation of p38 MAPK. Treatment with a combination of DIM and calcium ionophore may offer a new approach to enhance the chemotherapeutic effcacy in liver cancer.

KW - 3,3'-diindolylmethane

KW - Apoptosis

KW - Cytosolic Ca

KW - Hepatocellular carcinoma cells

KW - P38 MAPK

KW - Proliferation

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