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Bee Venom Protects against Rotenone-Induced Cell Death in NSC34 Motor Neuron Cells.

Jung SY, Lee KW, Choi SM, Yang EJ - Toxins (Basel) (2015)

Bottom Line: In this study, we tested the effects of BV on neuronal cell death by examining rotenone-induced mitochondrial dysfunction.Pretreatment with 2.5 μg/mL BV had a neuroprotective effect against 10 μM rotenone-induced cell death in NSC34 motor neuron cells.Pre-treatment with BV significantly enhanced cell viability and ameliorated mitochondrial impairment in rotenone-treated cellular model.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Research, Korea Institute of Oriental Medicine, 483 Expo-ro, Yuseong-gu, Daejeon 305-811, Korea. syzzim84@gmail.com.

ABSTRACT
Rotenone, an inhibitor of mitochondrial complex I of the mitochondrial respiratory chain, is known to elevate mitochondrial reactive oxygen species and induce apoptosis via activation of the caspase-3 pathway. Bee venom (BV) extracted from honey bees has been widely used in oriental medicine and contains melittin, apamin, adolapin, mast cell-degranulating peptide, and phospholipase A₂. In this study, we tested the effects of BV on neuronal cell death by examining rotenone-induced mitochondrial dysfunction. NSC34 motor neuron cells were pretreated with 2.5 μg/mL BV and stimulated with 10 μM rotenone to induce cell toxicity. We assessed cell death by Western blotting using specific antibodies, such as phospho-ERK1/2, phospho-JNK, and cleaved capase-3 and performed an MTT assay for evaluation of cell death and mitochondria staining. Pretreatment with 2.5 μg/mL BV had a neuroprotective effect against 10 μM rotenone-induced cell death in NSC34 motor neuron cells. Pre-treatment with BV significantly enhanced cell viability and ameliorated mitochondrial impairment in rotenone-treated cellular model. Moreover, BV treatment inhibited the activation of JNK signaling and cleaved caspase-3 related to cell death and increased ERK phosphorylation involved in cell survival in rotenone-treated NSC34 motor neuron cells. Taken together, we suggest that BV treatment can be useful for protection of neurons against oxidative stress or neurotoxin-induced cell death.

No MeSH data available.


Related in: MedlinePlus

BV pretreatment regulates the activation of rotenone-mediated signaling in NSC34 neuronal cells. Effect of rotenone on the phosphorylation of the MAPK proteins ERK and JNK in NSC34 cells. (A) NSC34 cells were treated with 10 µM rotenone for the indicated time. Total cell lysates were separated with SDS-PAGE and Western blots were performed using anti-phospho JNK, anti-phospho ERK1/2, JNK, and ERK antibodies; (B) NSC34 cells were pretreated with 2.5 μg/mL BV for 24 h and then stimulated with 10 μM rotenone for 1 h in the presence or absence of BV. Western blots were performed with specific antibodies, including those for the phosphorylated forms of ERK and JNK. Total ERK and JNK were used as loading controls for the cell lysates; (C) Immune blots were quantified with the relative phospho-/nonphospho ratio. The values shown are the means ±S.E.M. of data obtained from three independent experiments. *p < 0.05, **p < 0.01.
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toxins-07-03715-f002: BV pretreatment regulates the activation of rotenone-mediated signaling in NSC34 neuronal cells. Effect of rotenone on the phosphorylation of the MAPK proteins ERK and JNK in NSC34 cells. (A) NSC34 cells were treated with 10 µM rotenone for the indicated time. Total cell lysates were separated with SDS-PAGE and Western blots were performed using anti-phospho JNK, anti-phospho ERK1/2, JNK, and ERK antibodies; (B) NSC34 cells were pretreated with 2.5 μg/mL BV for 24 h and then stimulated with 10 μM rotenone for 1 h in the presence or absence of BV. Western blots were performed with specific antibodies, including those for the phosphorylated forms of ERK and JNK. Total ERK and JNK were used as loading controls for the cell lysates; (C) Immune blots were quantified with the relative phospho-/nonphospho ratio. The values shown are the means ±S.E.M. of data obtained from three independent experiments. *p < 0.05, **p < 0.01.

Mentions: For this study, NSC34 cells were treated with 10 μM rotenone for various time periods (0, 0.25, 6, and 24 h). JNK phosphorylation, indicating JNK activation, increased after 1 h of rotenone treatment in NSC34 cells. However, the expression of ERK1/2 phosphorylation, which promoted cell survival, was significantly reduced at 15 min by rotenone treatment compared to untreated cells (Figure 2A). Both significant increase in JNK phosphorylation and decrease in ERK phosphorylation were observed after 1 h of rotenone treatment in NSC34 cells. To examine the BV effect on cell death signaling, we pretreated with 2.5 μg/mL BV for 24 h and then stimulated with 10 μM rotenone for 1 h in the presence or absence of BV. As shown in Figure 2B,C, pretreatment with BV attenuated by 1.2 fold JNK phosphorylation compared to rotenone-treated cells. In addition, BV pretreatment recovered by 1.4 fold ERK under-phosphorylation compared rotenone-treated NSC34 cells (Figure 2B,C). These results suggest that BV pretreatment could prevent oxidative stress induced-neuronal cell death by inhibiting cell death-related MAPK signaling.


Bee Venom Protects against Rotenone-Induced Cell Death in NSC34 Motor Neuron Cells.

Jung SY, Lee KW, Choi SM, Yang EJ - Toxins (Basel) (2015)

BV pretreatment regulates the activation of rotenone-mediated signaling in NSC34 neuronal cells. Effect of rotenone on the phosphorylation of the MAPK proteins ERK and JNK in NSC34 cells. (A) NSC34 cells were treated with 10 µM rotenone for the indicated time. Total cell lysates were separated with SDS-PAGE and Western blots were performed using anti-phospho JNK, anti-phospho ERK1/2, JNK, and ERK antibodies; (B) NSC34 cells were pretreated with 2.5 μg/mL BV for 24 h and then stimulated with 10 μM rotenone for 1 h in the presence or absence of BV. Western blots were performed with specific antibodies, including those for the phosphorylated forms of ERK and JNK. Total ERK and JNK were used as loading controls for the cell lysates; (C) Immune blots were quantified with the relative phospho-/nonphospho ratio. The values shown are the means ±S.E.M. of data obtained from three independent experiments. *p < 0.05, **p < 0.01.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4591667&req=5

toxins-07-03715-f002: BV pretreatment regulates the activation of rotenone-mediated signaling in NSC34 neuronal cells. Effect of rotenone on the phosphorylation of the MAPK proteins ERK and JNK in NSC34 cells. (A) NSC34 cells were treated with 10 µM rotenone for the indicated time. Total cell lysates were separated with SDS-PAGE and Western blots were performed using anti-phospho JNK, anti-phospho ERK1/2, JNK, and ERK antibodies; (B) NSC34 cells were pretreated with 2.5 μg/mL BV for 24 h and then stimulated with 10 μM rotenone for 1 h in the presence or absence of BV. Western blots were performed with specific antibodies, including those for the phosphorylated forms of ERK and JNK. Total ERK and JNK were used as loading controls for the cell lysates; (C) Immune blots were quantified with the relative phospho-/nonphospho ratio. The values shown are the means ±S.E.M. of data obtained from three independent experiments. *p < 0.05, **p < 0.01.
Mentions: For this study, NSC34 cells were treated with 10 μM rotenone for various time periods (0, 0.25, 6, and 24 h). JNK phosphorylation, indicating JNK activation, increased after 1 h of rotenone treatment in NSC34 cells. However, the expression of ERK1/2 phosphorylation, which promoted cell survival, was significantly reduced at 15 min by rotenone treatment compared to untreated cells (Figure 2A). Both significant increase in JNK phosphorylation and decrease in ERK phosphorylation were observed after 1 h of rotenone treatment in NSC34 cells. To examine the BV effect on cell death signaling, we pretreated with 2.5 μg/mL BV for 24 h and then stimulated with 10 μM rotenone for 1 h in the presence or absence of BV. As shown in Figure 2B,C, pretreatment with BV attenuated by 1.2 fold JNK phosphorylation compared to rotenone-treated cells. In addition, BV pretreatment recovered by 1.4 fold ERK under-phosphorylation compared rotenone-treated NSC34 cells (Figure 2B,C). These results suggest that BV pretreatment could prevent oxidative stress induced-neuronal cell death by inhibiting cell death-related MAPK signaling.

Bottom Line: In this study, we tested the effects of BV on neuronal cell death by examining rotenone-induced mitochondrial dysfunction.Pretreatment with 2.5 μg/mL BV had a neuroprotective effect against 10 μM rotenone-induced cell death in NSC34 motor neuron cells.Pre-treatment with BV significantly enhanced cell viability and ameliorated mitochondrial impairment in rotenone-treated cellular model.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Research, Korea Institute of Oriental Medicine, 483 Expo-ro, Yuseong-gu, Daejeon 305-811, Korea. syzzim84@gmail.com.

ABSTRACT
Rotenone, an inhibitor of mitochondrial complex I of the mitochondrial respiratory chain, is known to elevate mitochondrial reactive oxygen species and induce apoptosis via activation of the caspase-3 pathway. Bee venom (BV) extracted from honey bees has been widely used in oriental medicine and contains melittin, apamin, adolapin, mast cell-degranulating peptide, and phospholipase A₂. In this study, we tested the effects of BV on neuronal cell death by examining rotenone-induced mitochondrial dysfunction. NSC34 motor neuron cells were pretreated with 2.5 μg/mL BV and stimulated with 10 μM rotenone to induce cell toxicity. We assessed cell death by Western blotting using specific antibodies, such as phospho-ERK1/2, phospho-JNK, and cleaved capase-3 and performed an MTT assay for evaluation of cell death and mitochondria staining. Pretreatment with 2.5 μg/mL BV had a neuroprotective effect against 10 μM rotenone-induced cell death in NSC34 motor neuron cells. Pre-treatment with BV significantly enhanced cell viability and ameliorated mitochondrial impairment in rotenone-treated cellular model. Moreover, BV treatment inhibited the activation of JNK signaling and cleaved caspase-3 related to cell death and increased ERK phosphorylation involved in cell survival in rotenone-treated NSC34 motor neuron cells. Taken together, we suggest that BV treatment can be useful for protection of neurons against oxidative stress or neurotoxin-induced cell death.

No MeSH data available.


Related in: MedlinePlus