Epigallocatechin-3-gallate induces mesothelioma cell death via H2 O2 -dependent T-type Ca2+ channel opening

Our plain-English summary of Epigallocatechin-3-gallate induces mesothelioma cell death via H2 O2 -dependent T-type Ca2+ channel opening, by E. Ranzato, S. Martinotti, V. Magnelli, et al (Journal of Cellular and Molecular Medicine, 2012).

The study

• Authors: E. Ranzato, S. Martinotti, V. Magnelli, B. Murer, S. Biffo, L. Mutti, B. Burlando
• Journal: Journal of Cellular and Molecular Medicine
• Published: 2012;16(11):2667-78
• Cited by: 40 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Malignant mesothelioma (MMe) is a highly aggressive, lethal tumour requiring the development of more effective therapies. The green tea polyphenol epigallocathechin-3-gallate (EGCG) inhibits the growth of many types of cancer cells. We found that EGCG is selectively cytotoxic to MMe cells with respect to normal mesothelial cells. MMe cell viability was inhibited by predominant induction of apoptosis at lower doses and necrosis at higher doses. EGCG elicited H(2) O(2) release in cell cultures, and exogenous catalase (CAT) abrogated EGCG-induced cytotoxicity, apoptosis and necrosis. Confocal imaging of fluo 3-loaded, EGCG-exposed MMe cells showed significant [Ca(2+) ](i) rise, prevented by CAT, dithiothreitol or the T-type Ca(2+) channel blockers mibefradil and NiCl(2) . Cell loading with dihydrorhodamine 123 revealed EGCG-induced ROS production, prevented by CAT, mibefradil or the Ca(2+) chelator BAPTA-AM. Direct exposure of cells to H(2) O(2) produced similar effects on Ca(2+) and ROS, and these effects were prevented by the same inhibitors. Sensitivity of REN cells to EGCG was correlated with higher expression of Ca(v) 3.2 T-type Ca(2+) channels in these cells, compared to normal mesothelium. Also, Ca(v) 3.2 siRNA on MMe cells reduced in vitro EGCG cytotoxicity and abated apoptosis and necrosis. Intriguingly, Ca(v) 3.2 expression was observed in malignant pleural mesothelioma biopsies from patients, but not in normal pleura. In conclusion, data showed the expression of T-type Ca(2+) channels in MMe tissue and their role in EGCG selective cytotoxicity to MMe cells, suggesting the possible use of these channels as a novel MMe pharmacological target.

© 2012 The Authors Journal of Cellular and Molecular Medicine © 2012 Foundation for Cellular and Molecular Medicine/Blackwell Publishing Ltd.

In plain English

This laboratory study tested epigallocatechin-3-gallate (EGCG), a green tea compound, on human mesothelioma cell lines and non-cancerous mesothelial cells. It did not study asbestos exposure. After 24 hours, the EGCG concentration halving viable cells ranged from 18 micromolar (95% confidence interval 16 to 20) to 85 (76 to 95) across five mesothelioma lines, versus 160 (144 to 178) for primary mesothelial cells and 309 (267 to 359) for immortalized non-cancerous HMC-hTERT mesothelial cells. EGCG produced hydrogen peroxide in the culture medium of REN mesothelioma cells, and a smaller but significant amount even without cells. Adding catalase, an enzyme that removes hydrogen peroxide, blocked the toxicity. EGCG raised calcium and reactive oxygen levels inside REN cells, and drugs that block T-type calcium channels prevented the calcium rise. REN cells expressed more of the Cav3.2 T-type channel than normal mesothelial cells, and silencing that channel reduced EGCG's toxicity. In tissue from eight mesothelioma patients, Cav3.2 staining was more intense than in two normal controls. The authors proposed T-type calcium channels as a possible new drug target.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (in vitro) cell culture experiments, plus immunohistochemical staining of archived patient tissue
• Who was studied: Five human malignant mesothelioma cell lines (REN, MM98, BR95, E198, MPP89) and two non-neoplastic mesothelial cell types: primary mesothelial cells from surgically removed hernial sacs, and HMC-hTERT cells, which are mesothelial cells from patients with congestive heart failure immortalized with the hTERT telomerase subunit. Also archived tissue from eight mesothelioma cases (two sarcomatous, four epithelioid, two biphasic) and two controls, from the Regional Hospital of Mestre-Venice, Italy.
• Exposure: Epigallocatechin-3-gallate (EGCG), a green tea polyphenol, applied to cells at a range of concentrations. Hydrogen peroxide was also applied directly to cells. Asbestos exposure was not studied.
• Main results:
  • EGCG concentrations reducing viability by half at 24 hours (with 95% confidence intervals) were lower for mesothelioma cell lines than for non-neoplastic mesothelial cells:
• MM98: 18 micromolar (16 to 20)
• BR95: 40 (38 to 43)
• REN: 52 (45 to 60)
• E198: 58 (48 to 69)
• MPP89: 85 (76 to 95)
• Primary mesothelium: 160 (144 to 178)
• HMC h-TERT (immortalized): 309 (267 to 359)
  • In REN cells, apoptosis (measured by caspase 3) predominated at lower EGCG concentrations. Necrosis (measured by LDH release) predominated at higher concentrations.
  • EGCG induced hydrogen peroxide release in REN cell cultures. A lower but significant amount was also produced in the culture medium without cells.
  • Adding catalase, which removes hydrogen peroxide, blocked EGCG's toxicity to REN cells. The authors attributed the toxicity largely to extracellular hydrogen peroxide.
  • The T-type calcium channel blocker mibefradil shifted the EGCG concentration reducing REN cell viability by half. It rose from 95 micromolar (95% CI 84 to 106) to 586 micromolar (95% CI 516 to 664).
  • After 45 minutes of 100 micromolar EGCG, intracellular calcium and reactive oxygen species rose significantly in REN cells. For example, REN calcium went from 80 to 771 nM, versus 57 to 169 nM in mesothelium. In mesothelial cells the calcium change was barely significant, and the reactive oxygen increase was significantly lower than in REN cells.
  • The Cav3.2 T-type calcium channel gene and protein were expressed in REN cells but barely detectable in mesothelial cells. Silencing Cav3.2 progressively reduced EGCG toxicity to REN cells compared with control (scramble) silencing.
  • In tissue from the eight mesothelioma patients, Cav3.2 staining was significantly more intense than in the two normal controls.
• The authors' conclusion: The authors concluded that EGCG selectively inhibited mesothelioma cells through a mechanism involving T-type calcium channels. They proposed these channels as a novel potential therapeutic target for mesothelioma.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Mesothelioma.

Medical terms in this study, explained in our Lexicon: Mesothelioma, Mesothelial Cells, Apoptosis, Necrosis, Mesothelium, Pleural Mesothelioma, Pleura.

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