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Anandamide-induced Ca2+ elevation leading to p38 MAPK phosphorylation and subsequent cell death via apoptosis in

Shu-Shong Hsu1, Chun-Jen Huang, He-Hsiung Cheng

  • 1Department of Surgery, Kaohsiung Veterans General Hospital, Kaohsiung 813, Taiwan.

Toxicology
|January 16, 2007
PubMed

Insights

Anandamide induces apoptosis in human osteosarcoma cells by increasing intracellular calcium and activating p38 MAPK. This pathway leads to caspase-3 activation and cell death, impacting osteoblast research.

Area of Science:

  • Cell Biology
  • Biochemistry
  • Pharmacology

Background:

  • The role of anandamide in human osteoblast function remains largely uncharacterized.
  • Osteoblasts are crucial for bone formation and remodeling.
  • Understanding cellular responses to anandamide is vital for potential therapeutic applications.

Purpose of the Study:

  • To investigate the effects of anandamide on human osteosarcoma MG63 cell viability, apoptosis, and intracellular signaling pathways.
  • To elucidate the specific mitogen-activated protein kinase (MAPK) pathways involved in anandamide-induced cellular responses.
  • To determine the role of intracellular calcium (Ca2+) in anandamide-mediated effects on osteoblasts.

Main Methods:

  • MG63 cells were treated with varying concentrations of anandamide.
  • Cell viability was assessed using propidium iodide staining.
  • Apoptosis was evaluated by caspase-3 activation assays.
  • Mitogen-activated protein kinase (MAPK) expression (ERK, JNK, p38) was analyzed via immunoblotting.
  • Intracellular Ca2+ ([Ca2+]i) levels were measured.
  • Pharmacological inhibitors (SB203580, PD98059, SP600125) and BAPTA were used to probe signaling pathways.

Main Results:

  • Anandamide (50-200 microM) decreased cell viability and induced apoptosis, evidenced by caspase-3 activation.
  • Anandamide upregulated ERK, JNK, and p38 MAPK expression.
  • SB203580, a p38 MAPK inhibitor, reversed anandamide-induced apoptosis, while PD98059 (ERK) and SP600125 (JNK) did not.
  • Anandamide (1-100 microM) increased intracellular Ca2+ levels, involving both influx and release.
  • Chelation of intracellular Ca2+ with BAPTA inhibited anandamide-induced cell death and p38 MAPK phosphorylation.

Conclusions:

  • Anandamide triggers apoptosis in MG63 osteosarcoma cells through a mechanism involving increased intracellular Ca2+.
  • The observed calcium increase activates p38 MAPK, which subsequently phosphorylates caspase-3, leading to apoptosis.
  • Anandamide's effects are mediated by the p38 MAPK pathway, not ERK or JNK, highlighting a specific signaling cascade in osteosarcoma cells.

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