Tags

Type your tag names separated by a space and hit enter

Barakol-induced apoptosis in P19 cells through generation of reactive oxygen species and activation of caspase-9.
J Ethnopharmacol. 2011 Sep 02; 137(2):971-8.JE

Abstract

ETHNOPHARMACOLOGICAL RELEVANCE

Barakol, an anxiolytic agent isolated from Senna siamea leaves which has been traditionally used for producing natural sleep, has been described as toxic to patients.

AIM OF THE STUDY

The aim of current study was to investigate the molecular mechanism of barakol-induced toxicity in mouse embryonal carcinoma P19 cell model.

MATERIALS AND METHODS

XTT assay was used to determine cell viability in P19 cells treated with barakol. Apoptotic cells were detected by Hoechst 33342 staining. Intracellular reactive oxygen species (ROS) generation was analyzed by flow cytometry using a fluorescent dye, DCFH-DA. Detection of apoptotic protein expression in P19 cells was performed by Western blot analysis. Caspase-9 activity was measured using a fluorescent immunosorbent enzyme assay kit.

RESULTS

Treatment with barakol decreased cell viability in a concentration- and time-dependent manner with an IC(50) value of 1.5mM in 24-h treated cells. A Hoechst 33342 assay revealed that barakol cytotoxicity was due to a significant increase in the number of apoptotic cells. Different scavengers to characterize ROS were utilized and revealed that hydroxyl radicals played a major role in ROS-induced apoptosis in barakol-treated cells. Western blot analysis demonstrated that barakol-induced apoptosis was mediated by the increase in expression ratio of Bax/Bcl-2. Furthermore, increase in caspase-9 activity after exposure to barakol for 24h was also observed. Pretreatment of cells with N-acetyl-l-cysteine (NAC) attenuated intracellular ROS generation, the Bax/Bcl-2 protein expression, and apoptosis.

CONCLUSIONS

The mechanism of barakol-mediated toxicity in P19 cells is mainly associated with the ROS generation, followed by the imbalance of the Bax/Bcl-2 ratio, and caspase-9 activation leading to apoptotic cell death. Pretreatment of cells with NAC could antagonize the toxicity produced by barakol.

Authors+Show Affiliations

Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok 10330, Thailand.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

21777666

Citation

Wongtongtair, Supim, et al. "Barakol-induced Apoptosis in P19 Cells Through Generation of Reactive Oxygen Species and Activation of Caspase-9." Journal of Ethnopharmacology, vol. 137, no. 2, 2011, pp. 971-8.
Wongtongtair S, Chanvorachote P, Hutamekalin P, et al. Barakol-induced apoptosis in P19 cells through generation of reactive oxygen species and activation of caspase-9. J Ethnopharmacol. 2011;137(2):971-8.
Wongtongtair, S., Chanvorachote, P., Hutamekalin, P., Chaichantipyuth, C., Lipipun, V., Tiensiwakul, P., & Meksuriyen, D. (2011). Barakol-induced apoptosis in P19 cells through generation of reactive oxygen species and activation of caspase-9. Journal of Ethnopharmacology, 137(2), 971-8. https://doi.org/10.1016/j.jep.2011.07.013
Wongtongtair S, et al. Barakol-induced Apoptosis in P19 Cells Through Generation of Reactive Oxygen Species and Activation of Caspase-9. J Ethnopharmacol. 2011 Sep 2;137(2):971-8. PubMed PMID: 21777666.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Barakol-induced apoptosis in P19 cells through generation of reactive oxygen species and activation of caspase-9. AU - Wongtongtair,Supim, AU - Chanvorachote,Pithi, AU - Hutamekalin,Pilaiwanwadee, AU - Chaichantipyuth,Chaiyo, AU - Lipipun,Vimolmas, AU - Tiensiwakul,Pornthep, AU - Meksuriyen,Duangdeun, Y1 - 2011/07/12/ PY - 2011/02/02/received PY - 2011/06/30/revised PY - 2011/07/03/accepted PY - 2011/7/23/entrez PY - 2011/7/23/pubmed PY - 2012/1/6/medline SP - 971 EP - 8 JF - Journal of ethnopharmacology JO - J Ethnopharmacol VL - 137 IS - 2 N2 - ETHNOPHARMACOLOGICAL RELEVANCE: Barakol, an anxiolytic agent isolated from Senna siamea leaves which has been traditionally used for producing natural sleep, has been described as toxic to patients. AIM OF THE STUDY: The aim of current study was to investigate the molecular mechanism of barakol-induced toxicity in mouse embryonal carcinoma P19 cell model. MATERIALS AND METHODS: XTT assay was used to determine cell viability in P19 cells treated with barakol. Apoptotic cells were detected by Hoechst 33342 staining. Intracellular reactive oxygen species (ROS) generation was analyzed by flow cytometry using a fluorescent dye, DCFH-DA. Detection of apoptotic protein expression in P19 cells was performed by Western blot analysis. Caspase-9 activity was measured using a fluorescent immunosorbent enzyme assay kit. RESULTS: Treatment with barakol decreased cell viability in a concentration- and time-dependent manner with an IC(50) value of 1.5mM in 24-h treated cells. A Hoechst 33342 assay revealed that barakol cytotoxicity was due to a significant increase in the number of apoptotic cells. Different scavengers to characterize ROS were utilized and revealed that hydroxyl radicals played a major role in ROS-induced apoptosis in barakol-treated cells. Western blot analysis demonstrated that barakol-induced apoptosis was mediated by the increase in expression ratio of Bax/Bcl-2. Furthermore, increase in caspase-9 activity after exposure to barakol for 24h was also observed. Pretreatment of cells with N-acetyl-l-cysteine (NAC) attenuated intracellular ROS generation, the Bax/Bcl-2 protein expression, and apoptosis. CONCLUSIONS: The mechanism of barakol-mediated toxicity in P19 cells is mainly associated with the ROS generation, followed by the imbalance of the Bax/Bcl-2 ratio, and caspase-9 activation leading to apoptotic cell death. Pretreatment of cells with NAC could antagonize the toxicity produced by barakol. SN - 1872-7573 UR - https://www.unboundmedicine.com/medline/citation/21777666/Barakol_induced_apoptosis_in_P19_cells_through_generation_of_reactive_oxygen_species_and_activation_of_caspase_9_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0378-8741(11)00489-2 DB - PRIME DP - Unbound Medicine ER -