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Mitochondrial Apoptotic Pathway Is Activated by H2O2-Mediated Oxidative Stress in BmN-SWU1 Cells from Bombyx mori Ovary.

Chen P, Hu YF, Wang L, Xiao WF, Bao XY, Pan C, Yi HS, Chen XY, Pan MH, Lu C - PLoS ONE (2015)

Bottom Line: H2O2-induced apoptosis also increased intracellular ROS level, changed mitochondrial distribution, reduced mitochondrial membrane potential and increased the release of cytochrome c from mitochondria.These results show that H2O2 treatment induced apoptosis in BmN-SWU1 cells via the mitochondrial apoptotic pathway.Taken together, these findings improve our knowledge of apoptosis in silkworm and the apoptotic pathways in insects.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Silkworm Genome Biology, Southwest University, Chongqing, 400716, China; Key Laboratory for Sericulture Functional Genomics and Biotechnology of Agricultural Ministry, Southwest University, Chongqing, 400716, China.

ABSTRACT
Apoptosis is a known regulator of morphogenetic events. In mammals, the critical role of oxidative stress-induced apoptosis has been well-studied; however, in insects the role of oxidative stress in apoptosis is not clear. In a previous study, we showed that apoptosis-related genes are present in the silkworm Bombyx mori, an important lepidopteran insect model. In this study, we evaluated the effect of H2O2-induced oxidative stress on apoptosis, reactive oxygen species (ROS) levels, mitochondrial response, cytochrome c release and apoptosis-related gene expression in the BmN-SWU1 cell line from B. mori ovaries. Our results showed that BmN-SWU1 cells exposed to H2O2 showed cell protuberances, cytoplasmic condensation, apoptotic bodies, DNA ladder formation and caspase activities indicating apoptosis. H2O2-induced apoptosis also increased intracellular ROS level, changed mitochondrial distribution, reduced mitochondrial membrane potential and increased the release of cytochrome c from mitochondria. Furthermore, western blot analysis revealed a significant increase in p53 and cytochrome c expression, and a decrease in Bcl-2 expression compared to the controls. Moreover, quantitative real-time PCR (qRT-PCR) showed an increase in the transcript levels of BmICE, Bmapaf-1 and BmEndoG by 439.5%, 423.9% and 42.2%, respectively, after treatment with 1 μM H2O2 for 24 h. However, the transcript levels of Bmbuffy declined by 41.4% after 24 h of exposure to 1 μM H2O2. These results show that H2O2 treatment induced apoptosis in BmN-SWU1 cells via the mitochondrial apoptotic pathway. Further, it appears that oxidative stress induced by H2O2 activates both caspase-dependent and caspase-independent mitochondrial apoptotic pathways in silkworm cells. Taken together, these findings improve our knowledge of apoptosis in silkworm and the apoptotic pathways in insects.

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qRT-PCR analysis of apoptosis-related genes in BmN-SWU1 cells after exposure to H2O2.All treated groups were compared with the control (0 h). Values indicate mean ± SD (n = 3). **, P < 0.01, Student’s t-test.
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pone.0134694.g005: qRT-PCR analysis of apoptosis-related genes in BmN-SWU1 cells after exposure to H2O2.All treated groups were compared with the control (0 h). Values indicate mean ± SD (n = 3). **, P < 0.01, Student’s t-test.

Mentions: Moreover, we investigated the mRNA levels of four other apoptosis-related genes to determine whether apoptosis induced by H2O2 in BmN-SWU1 cells is through the caspase-dependent or caspase-independent pathway. We first examined the expression levels of BmICE, Bmapaf-1 and Bmbuffy involved in the caspase-dependent apoptosis. We found that there was a 439.5% and 423.9% increase in the BmICE and Bmapaf-1 mRNA expression, respectively after exposure to 1 μM H2O2 for 24 h (Fig 5A and 5B). On the other hand, Bmbuffy expression decreased by 41.4% after 24 h treatment with 1 μM H2O2 (Fig 5C). Next, we evaluated the expression level of B. mori endonuclease G (BmEndoG), which is implicated in the caspase-independent apoptotic pathway in the mitochondria [31]. The results showed that there was a 42.2% increase after exposure of BmN-SWU1 cells to 1 μM H2O2 from 12–24 h (Fig 5D) indicating that oxidative stress induced by H2O2 may also activate caspase-independent apoptotic pathway in BmN-SWU1 cells.


Mitochondrial Apoptotic Pathway Is Activated by H2O2-Mediated Oxidative Stress in BmN-SWU1 Cells from Bombyx mori Ovary.

Chen P, Hu YF, Wang L, Xiao WF, Bao XY, Pan C, Yi HS, Chen XY, Pan MH, Lu C - PLoS ONE (2015)

qRT-PCR analysis of apoptosis-related genes in BmN-SWU1 cells after exposure to H2O2.All treated groups were compared with the control (0 h). Values indicate mean ± SD (n = 3). **, P < 0.01, Student’s t-test.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4520666&req=5

pone.0134694.g005: qRT-PCR analysis of apoptosis-related genes in BmN-SWU1 cells after exposure to H2O2.All treated groups were compared with the control (0 h). Values indicate mean ± SD (n = 3). **, P < 0.01, Student’s t-test.
Mentions: Moreover, we investigated the mRNA levels of four other apoptosis-related genes to determine whether apoptosis induced by H2O2 in BmN-SWU1 cells is through the caspase-dependent or caspase-independent pathway. We first examined the expression levels of BmICE, Bmapaf-1 and Bmbuffy involved in the caspase-dependent apoptosis. We found that there was a 439.5% and 423.9% increase in the BmICE and Bmapaf-1 mRNA expression, respectively after exposure to 1 μM H2O2 for 24 h (Fig 5A and 5B). On the other hand, Bmbuffy expression decreased by 41.4% after 24 h treatment with 1 μM H2O2 (Fig 5C). Next, we evaluated the expression level of B. mori endonuclease G (BmEndoG), which is implicated in the caspase-independent apoptotic pathway in the mitochondria [31]. The results showed that there was a 42.2% increase after exposure of BmN-SWU1 cells to 1 μM H2O2 from 12–24 h (Fig 5D) indicating that oxidative stress induced by H2O2 may also activate caspase-independent apoptotic pathway in BmN-SWU1 cells.

Bottom Line: H2O2-induced apoptosis also increased intracellular ROS level, changed mitochondrial distribution, reduced mitochondrial membrane potential and increased the release of cytochrome c from mitochondria.These results show that H2O2 treatment induced apoptosis in BmN-SWU1 cells via the mitochondrial apoptotic pathway.Taken together, these findings improve our knowledge of apoptosis in silkworm and the apoptotic pathways in insects.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Silkworm Genome Biology, Southwest University, Chongqing, 400716, China; Key Laboratory for Sericulture Functional Genomics and Biotechnology of Agricultural Ministry, Southwest University, Chongqing, 400716, China.

ABSTRACT
Apoptosis is a known regulator of morphogenetic events. In mammals, the critical role of oxidative stress-induced apoptosis has been well-studied; however, in insects the role of oxidative stress in apoptosis is not clear. In a previous study, we showed that apoptosis-related genes are present in the silkworm Bombyx mori, an important lepidopteran insect model. In this study, we evaluated the effect of H2O2-induced oxidative stress on apoptosis, reactive oxygen species (ROS) levels, mitochondrial response, cytochrome c release and apoptosis-related gene expression in the BmN-SWU1 cell line from B. mori ovaries. Our results showed that BmN-SWU1 cells exposed to H2O2 showed cell protuberances, cytoplasmic condensation, apoptotic bodies, DNA ladder formation and caspase activities indicating apoptosis. H2O2-induced apoptosis also increased intracellular ROS level, changed mitochondrial distribution, reduced mitochondrial membrane potential and increased the release of cytochrome c from mitochondria. Furthermore, western blot analysis revealed a significant increase in p53 and cytochrome c expression, and a decrease in Bcl-2 expression compared to the controls. Moreover, quantitative real-time PCR (qRT-PCR) showed an increase in the transcript levels of BmICE, Bmapaf-1 and BmEndoG by 439.5%, 423.9% and 42.2%, respectively, after treatment with 1 μM H2O2 for 24 h. However, the transcript levels of Bmbuffy declined by 41.4% after 24 h of exposure to 1 μM H2O2. These results show that H2O2 treatment induced apoptosis in BmN-SWU1 cells via the mitochondrial apoptotic pathway. Further, it appears that oxidative stress induced by H2O2 activates both caspase-dependent and caspase-independent mitochondrial apoptotic pathways in silkworm cells. Taken together, these findings improve our knowledge of apoptosis in silkworm and the apoptotic pathways in insects.

No MeSH data available.


Related in: MedlinePlus