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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

Cytotoxicity of EEGE in EAT cells (3 × 105 cells/ml; solid line) and human lymphocytes (1 × 106 cells/ml; dashed line) after 72 hours of incubation. Effects of EEGE on MTT reduction (▲) and protein phosphatase activity (●) is expressed relative to control cell viability (100%) and each point represents the mean ± S.D.
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Figure 1: Cytotoxicity of EEGE in EAT cells (3 × 105 cells/ml; solid line) and human lymphocytes (1 × 106 cells/ml; dashed line) after 72 hours of incubation. Effects of EEGE on MTT reduction (▲) and protein phosphatase activity (●) is expressed relative to control cell viability (100%) and each point represents the mean ± S.D.

Mentions: Cytotoxicity induced by EEGE in EAT cells was evaluated by using MTT reduction and phosphatase activity with different concentrations of EEGE after 72 hours of treatment (Figure 1). EAT cells were exposed to various concentrations of EEGE and it resulted in a significant negative effect in cell proliferation, with the IC50 of 45 μg/ml observed in MTT reduction and phosphatase activity assays. At low concentrations of EEGE, a non-significant acceleration of cell growth was observed (Figure 1). By using trypan blue dye exclusion method, the effect of EEGE in EAT cells in vitro assay we also confirmed the above observation. Cells exposed to EEGE for 72 hours decreased cell viability in a dose-dependent manner (Figure 2). At 50 μg/ml dose the EAT cells viability was close to 65% and the maximum decrease of 15% was observed at 100 μg/ml. From these results, we were convinced that the EEGE potently inhibits the proliferation and viability of EAT cells and we continued with further investigations. EEGE was able to inhibit proliferation of human lymphocytes also, however the potency was not comparable to EAT cells, presenting IC50 nearly 1.5 fold higher as 70 μg/ml than for EAT cells, as observed in the MTT assay after 72 hours of incubation with EEGE in the same range of concentrations (Figure 1). For further in vitro analysis EEGE was used at 25, 50 and 100 μg/ml for cellular assays.


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

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

Cytotoxicity of EEGE in EAT cells (3 × 105 cells/ml; solid line) and human lymphocytes (1 × 106 cells/ml; dashed line) after 72 hours of incubation. Effects of EEGE on MTT reduction (▲) and protein phosphatase activity (●) is expressed relative to control cell viability (100%) and each point represents the mean ± S.D.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Figure 1: Cytotoxicity of EEGE in EAT cells (3 × 105 cells/ml; solid line) and human lymphocytes (1 × 106 cells/ml; dashed line) after 72 hours of incubation. Effects of EEGE on MTT reduction (▲) and protein phosphatase activity (●) is expressed relative to control cell viability (100%) and each point represents the mean ± S.D.
Mentions: Cytotoxicity induced by EEGE in EAT cells was evaluated by using MTT reduction and phosphatase activity with different concentrations of EEGE after 72 hours of treatment (Figure 1). EAT cells were exposed to various concentrations of EEGE and it resulted in a significant negative effect in cell proliferation, with the IC50 of 45 μg/ml observed in MTT reduction and phosphatase activity assays. At low concentrations of EEGE, a non-significant acceleration of cell growth was observed (Figure 1). By using trypan blue dye exclusion method, the effect of EEGE in EAT cells in vitro assay we also confirmed the above observation. Cells exposed to EEGE for 72 hours decreased cell viability in a dose-dependent manner (Figure 2). At 50 μg/ml dose the EAT cells viability was close to 65% and the maximum decrease of 15% was observed at 100 μg/ml. From these results, we were convinced that the EEGE potently inhibits the proliferation and viability of EAT cells and we continued with further investigations. EEGE was able to inhibit proliferation of human lymphocytes also, however the potency was not comparable to EAT cells, presenting IC50 nearly 1.5 fold higher as 70 μg/ml than for EAT cells, as observed in the MTT assay after 72 hours of incubation with EEGE in the same range of concentrations (Figure 1). For further in vitro analysis EEGE was used at 25, 50 and 100 μg/ml for cellular assays.

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