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Gracilaria edulis extract induces apoptosis and inhibits tumor in Ehrlich ascites tumor cells in vivo.

Patra S, Muthuraman MS - BMC Complement Altern Med (2013)

Bottom Line: Intraperitoneally administration of EEGE to EAT-bearing mice helped to increase the lifespan of the animals significantly inhibited tumor growth and increased survival of mice.Extensive hematology, biochemistry and histopathological analysis of liver and kidney indicated that daily doses of EEGE up to 300 mg/kg for 35 days are well tolerated and did not cause hematotoxicity nor renal or hepatotoxicity.It was evident that the mechanism explains the apoptotic activity of the algae extract.

View Article: PubMed Central - HTML - PubMed

Affiliation: Medical University of the Americas, Charlestown, Nevis, West Indies. s.patra@mua.edu.

ABSTRACT

Background: Marine environment is inestimable for their chemical and biological diversity and therefore is an extraordinary resource for the discovery of new anticancer drugs. Recent development in elucidation of the mechanism and therapeutic action of natural products helped to evaluate for their potential activity.

Methods: We evaluated Gracilaria edulis J. Ag (Brown algae), for its antitumor potential against the Ehrlich ascites tumor (EAT) in vivo and in vitro. Cytotoxicity evaluation of Ethanol Extract of Gracilaria edulis (EEGE) using EAT cells showed significant activity. In vitro studies indicated that EEGE cytotoxicity to EAT cells is mediated through its ability to produce reactive oxygen species (ROS) and therefore decreasing intracellular glutathione (GSH) levels may be attributed to oxidative stress.

Results: Apoptotic parameters including Annexin-V positive cells, increased levels of DNA fragmentation and increased caspase-2, caspase-3 and caspase-9 activities indicated the mechanism might be by inducing apoptosis. Intraperitoneally administration of EEGE to EAT-bearing mice helped to increase the lifespan of the animals significantly inhibited tumor growth and increased survival of mice. Extensive hematology, biochemistry and histopathological analysis of liver and kidney indicated that daily doses of EEGE up to 300 mg/kg for 35 days are well tolerated and did not cause hematotoxicity nor renal or hepatotoxicity.

Conclusion: Comprehensive antitumor analysis in animal model and in Ehrlich Ascites Tumor cells was done including biochemical, and pathological evaluations indicate antitumor activity of the extract and non toxic in vivo. It was evident that the mechanism explains the apoptotic activity of the algae extract.

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Related in: MedlinePlus

Time-course effect of EEGE on ROS generation in EAT cells. The fluorescence intensity of DCF was monitored at 538 nm, with excitation wavelength set at 490 nm, and used to indicate the level of intracellular peroxides formation. Changes in DCF fluorescence in tumor cells were measured at 8, 12, 16, 20 and 24 hours after treatment with 50 μg/ml of EEGE, *p < 0.01 compared to control and 24 h-time points (ANOVA, Tukey test). The results express the mean ± S.D. of three independent experiments run in duplicate.
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Figure 3: Time-course effect of EEGE on ROS generation in EAT cells. The fluorescence intensity of DCF was monitored at 538 nm, with excitation wavelength set at 490 nm, and used to indicate the level of intracellular peroxides formation. Changes in DCF fluorescence in tumor cells were measured at 8, 12, 16, 20 and 24 hours after treatment with 50 μg/ml of EEGE, *p < 0.01 compared to control and 24 h-time points (ANOVA, Tukey test). The results express the mean ± S.D. of three independent experiments run in duplicate.

Mentions: ROS is known to be a key player in highly organized cellular functions such as pathways of signal transduction and apoptosis [25] and a role for oxidative signaling in the cytotoxicity of marine product in cancer cells has been previously reported [26]. In this context we investigated a potential role of oxidative stress in the alteration of cellular sensitivity to EEGE. EAT cells treated with EEGE for 30 min were used for estimation of ROS level after the addition of DCFH-DA. The time-course effect of EEGE on the EAT cell intracellular peroxide levels is presented in Figure 3. Intracellular ROS production was observed at 8–24 hours after incubation of tumor cells with 50 μg/ml of EEGE as compared to control cells, and found to be significantly increased (p < 0.01). Increase in peroxides amounts generated by EAT cells was also noted to be time-dependent, with significantly higher (p < 0.01) at the beginning of treatment such as 8 and 12 hours in comparison with the 24 hours time point and the peroxides levels reached to normal after 24 hours exposure in EAT cells.


Gracilaria edulis extract induces apoptosis and inhibits tumor in Ehrlich ascites tumor cells in vivo.

Patra S, Muthuraman MS - BMC Complement Altern Med (2013)

Time-course effect of EEGE on ROS generation in EAT cells. The fluorescence intensity of DCF was monitored at 538 nm, with excitation wavelength set at 490 nm, and used to indicate the level of intracellular peroxides formation. Changes in DCF fluorescence in tumor cells were measured at 8, 12, 16, 20 and 24 hours after treatment with 50 μg/ml of EEGE, *p < 0.01 compared to control and 24 h-time points (ANOVA, Tukey test). The results express the mean ± S.D. of three independent experiments run in duplicate.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4222716&req=5

Figure 3: Time-course effect of EEGE on ROS generation in EAT cells. The fluorescence intensity of DCF was monitored at 538 nm, with excitation wavelength set at 490 nm, and used to indicate the level of intracellular peroxides formation. Changes in DCF fluorescence in tumor cells were measured at 8, 12, 16, 20 and 24 hours after treatment with 50 μg/ml of EEGE, *p < 0.01 compared to control and 24 h-time points (ANOVA, Tukey test). The results express the mean ± S.D. of three independent experiments run in duplicate.
Mentions: ROS is known to be a key player in highly organized cellular functions such as pathways of signal transduction and apoptosis [25] and a role for oxidative signaling in the cytotoxicity of marine product in cancer cells has been previously reported [26]. In this context we investigated a potential role of oxidative stress in the alteration of cellular sensitivity to EEGE. EAT cells treated with EEGE for 30 min were used for estimation of ROS level after the addition of DCFH-DA. The time-course effect of EEGE on the EAT cell intracellular peroxide levels is presented in Figure 3. Intracellular ROS production was observed at 8–24 hours after incubation of tumor cells with 50 μg/ml of EEGE as compared to control cells, and found to be significantly increased (p < 0.01). Increase in peroxides amounts generated by EAT cells was also noted to be time-dependent, with significantly higher (p < 0.01) at the beginning of treatment such as 8 and 12 hours in comparison with the 24 hours time point and the peroxides levels reached to normal after 24 hours exposure in EAT cells.

Bottom Line: Intraperitoneally administration of EEGE to EAT-bearing mice helped to increase the lifespan of the animals significantly inhibited tumor growth and increased survival of mice.Extensive hematology, biochemistry and histopathological analysis of liver and kidney indicated that daily doses of EEGE up to 300 mg/kg for 35 days are well tolerated and did not cause hematotoxicity nor renal or hepatotoxicity.It was evident that the mechanism explains the apoptotic activity of the algae extract.

View Article: PubMed Central - HTML - PubMed

Affiliation: Medical University of the Americas, Charlestown, Nevis, West Indies. s.patra@mua.edu.

ABSTRACT

Background: Marine environment is inestimable for their chemical and biological diversity and therefore is an extraordinary resource for the discovery of new anticancer drugs. Recent development in elucidation of the mechanism and therapeutic action of natural products helped to evaluate for their potential activity.

Methods: We evaluated Gracilaria edulis J. Ag (Brown algae), for its antitumor potential against the Ehrlich ascites tumor (EAT) in vivo and in vitro. Cytotoxicity evaluation of Ethanol Extract of Gracilaria edulis (EEGE) using EAT cells showed significant activity. In vitro studies indicated that EEGE cytotoxicity to EAT cells is mediated through its ability to produce reactive oxygen species (ROS) and therefore decreasing intracellular glutathione (GSH) levels may be attributed to oxidative stress.

Results: Apoptotic parameters including Annexin-V positive cells, increased levels of DNA fragmentation and increased caspase-2, caspase-3 and caspase-9 activities indicated the mechanism might be by inducing apoptosis. Intraperitoneally administration of EEGE to EAT-bearing mice helped to increase the lifespan of the animals significantly inhibited tumor growth and increased survival of mice. Extensive hematology, biochemistry and histopathological analysis of liver and kidney indicated that daily doses of EEGE up to 300 mg/kg for 35 days are well tolerated and did not cause hematotoxicity nor renal or hepatotoxicity.

Conclusion: Comprehensive antitumor analysis in animal model and in Ehrlich Ascites Tumor cells was done including biochemical, and pathological evaluations indicate antitumor activity of the extract and non toxic in vivo. It was evident that the mechanism explains the apoptotic activity of the algae extract.

Show MeSH
Related in: MedlinePlus