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Cytotoxic effects of oosporein isolated from endophytic fungus Cochliobolus kusanoi.

Ramesha A, Venkataramana M, Nirmaladevi D, Gupta VK, Chandranayaka S, Srinivas C - Front Microbiol (2015)

Bottom Line: After 24 h of exposure histopathological observations were made to know the effects of oosporein on target organs.Further, oosporein treatment to studied cell lines indicated significant suppression of oxidative stress related gene (Superoxide dismutase1 and Catalase ) expression, and increased levels of mRNA expression in apoptosis or oxidative stress inducing genes HSP70, Caspase3, Caspase6, and Caspase9 as measured by quantitative real time-PCR assay.The high previlance of these fungi in temperate climates further warrants the need of safe food grain storage and processing practices to control the toxic effects of oosporein to humans and live stock.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Biotechnology, Bangalore University Bangalore, India.

ABSTRACT
In the present study, oosporein, a fungal toxic secondary metabolite known to be a toxic agent causing chronic disorders in animals, was isolated from fungus Cochliobolus kusanoi of Nerium oleander L. Toxic effects of oosporein and the possible mechanisms of cytotoxicity as well as the role of oxidative stress in cytotoxicity to Madin-Darby canine kidney kidney cells and RAW 264.7 splene cells were evaluated in vitro. Also to know the possible in vivo toxic effects of oosporein on kidney and spleen, Balb/C mouse were treated with different concentrations of oosporein ranging from 20 to 200 μM). After 24 h of exposure histopathological observations were made to know the effects of oosporein on target organs. Oosporein induced elevated levels of reactive oxygen species (ROS) generation and high levels of malondialdehyde, loss of mitochondrial membrane potential, induced glutathione hydroxylase (GSH) production was observed in a dose depended manner. Effects oosporein on chromosomal DNA damage was assessed by Comet assay, and increase in DNA damage were observed in both the studied cell lines by increasing the oosporein concentration. Further, oosporein treatment to studied cell lines indicated significant suppression of oxidative stress related gene (Superoxide dismutase1 and Catalase ) expression, and increased levels of mRNA expression in apoptosis or oxidative stress inducing genes HSP70, Caspase3, Caspase6, and Caspase9 as measured by quantitative real time-PCR assay. Histopathological examination of oosporein treated mouse kidney and splenocytes further revealed that, oosporein treated target mouse tissues were significantly damaged with that of untreated sam control mice and these effects were in directly proportional to the the toxin dose. Results of the present study reveals that, ROS is the principle event prompting increased oosporein toxicity in studied in vivio and in vitro animal models. The high previlance of these fungi in temperate climates further warrants the need of safe food grain storage and processing practices to control the toxic effects of oosporein to humans and live stock.

No MeSH data available.


Related in: MedlinePlus

(A) Dose dependent effects of oosporein on Oxidative biomarker gene expression of Superoxide dismutase1 (SOD1) and Catalase (CAT) genes quantified by real time PCR. (B) Dose dependent effects of oosporein on apoptosis inducing gene expression of HSP70, Cas3, Cas6, and Cas9 genes quantified by real time PCR in MDCK cellline. (C) Dose dependent effects of oosporein on apoptosis inducing gene expression of HSP70, Cas3, Cas6, and Cas9 genes quantified by real time PCR in RAW 264.7 cellline. Values represent mean ± SD of three parallel experiments. In the figure each series mean values followed by the different letter are significantly different according to DMRT at p < 0.05.
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Figure 5: (A) Dose dependent effects of oosporein on Oxidative biomarker gene expression of Superoxide dismutase1 (SOD1) and Catalase (CAT) genes quantified by real time PCR. (B) Dose dependent effects of oosporein on apoptosis inducing gene expression of HSP70, Cas3, Cas6, and Cas9 genes quantified by real time PCR in MDCK cellline. (C) Dose dependent effects of oosporein on apoptosis inducing gene expression of HSP70, Cas3, Cas6, and Cas9 genes quantified by real time PCR in RAW 264.7 cellline. Values represent mean ± SD of three parallel experiments. In the figure each series mean values followed by the different letter are significantly different according to DMRT at p < 0.05.

Mentions: To explore whether oosporein intervened on the Oxidative biomarker, gene expression of antioxidant enzymes of SOD1 and CAT in cells were measured. The CAT and SOD1 gene response were assessed by Q-RT-PCR. Compared to control cells, the oosporein treated cells strongly down-regulated CAT and SOD1 mRNA levels with increasing concentration of oosporein (Figure 5A). The expression of CAT gene decreased by ∼3 to ∼7.5-fold and ∼2.5 to ∼6-fold in MDCK cells and RAW 264.7 cells, respectively, with increasing concentration of oosporein from 25 to 100 μM. There was ∼3 to ∼8.4 and ∼2.8 to ∼6.5-fold decrease in the expression of SOD1 gene in MDCK cells and RAW 264.7 cells, respectively, on exposure to 25–100 μM of oosporein (Figure 5A).


Cytotoxic effects of oosporein isolated from endophytic fungus Cochliobolus kusanoi.

Ramesha A, Venkataramana M, Nirmaladevi D, Gupta VK, Chandranayaka S, Srinivas C - Front Microbiol (2015)

(A) Dose dependent effects of oosporein on Oxidative biomarker gene expression of Superoxide dismutase1 (SOD1) and Catalase (CAT) genes quantified by real time PCR. (B) Dose dependent effects of oosporein on apoptosis inducing gene expression of HSP70, Cas3, Cas6, and Cas9 genes quantified by real time PCR in MDCK cellline. (C) Dose dependent effects of oosporein on apoptosis inducing gene expression of HSP70, Cas3, Cas6, and Cas9 genes quantified by real time PCR in RAW 264.7 cellline. Values represent mean ± SD of three parallel experiments. In the figure each series mean values followed by the different letter are significantly different according to DMRT at p < 0.05.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4556033&req=5

Figure 5: (A) Dose dependent effects of oosporein on Oxidative biomarker gene expression of Superoxide dismutase1 (SOD1) and Catalase (CAT) genes quantified by real time PCR. (B) Dose dependent effects of oosporein on apoptosis inducing gene expression of HSP70, Cas3, Cas6, and Cas9 genes quantified by real time PCR in MDCK cellline. (C) Dose dependent effects of oosporein on apoptosis inducing gene expression of HSP70, Cas3, Cas6, and Cas9 genes quantified by real time PCR in RAW 264.7 cellline. Values represent mean ± SD of three parallel experiments. In the figure each series mean values followed by the different letter are significantly different according to DMRT at p < 0.05.
Mentions: To explore whether oosporein intervened on the Oxidative biomarker, gene expression of antioxidant enzymes of SOD1 and CAT in cells were measured. The CAT and SOD1 gene response were assessed by Q-RT-PCR. Compared to control cells, the oosporein treated cells strongly down-regulated CAT and SOD1 mRNA levels with increasing concentration of oosporein (Figure 5A). The expression of CAT gene decreased by ∼3 to ∼7.5-fold and ∼2.5 to ∼6-fold in MDCK cells and RAW 264.7 cells, respectively, with increasing concentration of oosporein from 25 to 100 μM. There was ∼3 to ∼8.4 and ∼2.8 to ∼6.5-fold decrease in the expression of SOD1 gene in MDCK cells and RAW 264.7 cells, respectively, on exposure to 25–100 μM of oosporein (Figure 5A).

Bottom Line: After 24 h of exposure histopathological observations were made to know the effects of oosporein on target organs.Further, oosporein treatment to studied cell lines indicated significant suppression of oxidative stress related gene (Superoxide dismutase1 and Catalase ) expression, and increased levels of mRNA expression in apoptosis or oxidative stress inducing genes HSP70, Caspase3, Caspase6, and Caspase9 as measured by quantitative real time-PCR assay.The high previlance of these fungi in temperate climates further warrants the need of safe food grain storage and processing practices to control the toxic effects of oosporein to humans and live stock.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Biotechnology, Bangalore University Bangalore, India.

ABSTRACT
In the present study, oosporein, a fungal toxic secondary metabolite known to be a toxic agent causing chronic disorders in animals, was isolated from fungus Cochliobolus kusanoi of Nerium oleander L. Toxic effects of oosporein and the possible mechanisms of cytotoxicity as well as the role of oxidative stress in cytotoxicity to Madin-Darby canine kidney kidney cells and RAW 264.7 splene cells were evaluated in vitro. Also to know the possible in vivo toxic effects of oosporein on kidney and spleen, Balb/C mouse were treated with different concentrations of oosporein ranging from 20 to 200 μM). After 24 h of exposure histopathological observations were made to know the effects of oosporein on target organs. Oosporein induced elevated levels of reactive oxygen species (ROS) generation and high levels of malondialdehyde, loss of mitochondrial membrane potential, induced glutathione hydroxylase (GSH) production was observed in a dose depended manner. Effects oosporein on chromosomal DNA damage was assessed by Comet assay, and increase in DNA damage were observed in both the studied cell lines by increasing the oosporein concentration. Further, oosporein treatment to studied cell lines indicated significant suppression of oxidative stress related gene (Superoxide dismutase1 and Catalase ) expression, and increased levels of mRNA expression in apoptosis or oxidative stress inducing genes HSP70, Caspase3, Caspase6, and Caspase9 as measured by quantitative real time-PCR assay. Histopathological examination of oosporein treated mouse kidney and splenocytes further revealed that, oosporein treated target mouse tissues were significantly damaged with that of untreated sam control mice and these effects were in directly proportional to the the toxin dose. Results of the present study reveals that, ROS is the principle event prompting increased oosporein toxicity in studied in vivio and in vitro animal models. The high previlance of these fungi in temperate climates further warrants the need of safe food grain storage and processing practices to control the toxic effects of oosporein to humans and live stock.

No MeSH data available.


Related in: MedlinePlus