甲基紫精对丹参培养细胞抗氧化防护系统的影响

doi:10.3724/SP.J.1258.2014.00047

摘要/Abstract

摘要：

为了探讨甲基紫精(MV)对丹参(Salvia miltiorrhiza)体内抗氧化防护系统的影响及其生理机制。以MV为诱导剂, 以敌草隆(DCMU)为抑制剂, 考察了MV与DCMU处理后丹参悬浮培养细胞中H₂O₂、丙二醛、还原型谷胱甘肽的含量以及抗氧化防护酶(超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT))活性变化和同工酶的表达差异。结果表明, MV处理显著提高了丹参培养细胞内H₂O₂、丙二醛以及还原型谷胱甘肽含量; MV处理使CAT、POD活性增强, 谱带颜色更亮, 条带增加。DCMU处理显著抑制了MV诱导的H₂O₂、丙二醛、还原型谷胱甘肽含量的增加, 抗氧化酶活性的升高和同工酶的表达。以上结果说明, MV可诱导丹参培养细胞叶绿体产生H₂O₂, H₂O₂激活了丹参培养细胞抗氧化防护系统以维持细胞正常的生理活动。

关键词: 抗氧化防护系统, 叶绿体, 敌草隆, 甲基紫精, 聚丙烯酰胺凝胶电泳, 丹参

Abstract:

Aims We studied the effects of the methyl viologen (MV) in the culture medium on the antioxidant enzyme system in Salvia miltiorrhiza cells and investigated changes in the contents of H₂O₂, malonaldehyde (MDA), glutathione (GSH), and antioxidant enzyme. The differential expressions of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) were discussed. Our objective was to explore if the MV could significantly affect the antioxidant system in the cultured S. miltiorrhiza cells.
Methods We treated the S. miltiorrhiza cells in suspension cultures with MV as an elicitor on the accumulation of H₂O₂, and used the diuron (DCMU) as an electron transporting antagonist. The biosynthesis of H₂O₂, MDA, and GSH, and the activities of SOD, CAT, and POD were determined by ultraviolet spectrophotometry. The specific isoenzymes bands of SOD, CAT, and POD were examined by acrylamide gel electrophoresis technique.
Important findings We found that MV significantly increased the accumulation of H₂O₂, MDA, and GSH, along with increased activities of CAT and POD as well as enhanced expressions and increased number of isoenzymes bands of CAT and POD; whereas DCMU inhibited the effects of MV. Results in this study suggest that MV could significantly enhance H₂O₂ accumulation in chloroplasts and regulate the antioxidant system in S. miltiorrhiza cells in suspension cultures, in order to maintain normal physiological conditions.

Key words: antioxidant system, chloroplast, DCMU, methyl viologen, polyacrylamide gel electrophoresis, Salvia miltiorrhiza

行冰玉,朱楠,张洪培,杨喜玲,董娟娥. 甲基紫精对丹参培养细胞抗氧化防护系统的影响. 植物生态学报, 2014, 38(5): 507-514. DOI: 10.3724/SP.J.1258.2014.00047

XING Bing-Yu,ZHU Nan,ZHANG Hong-Pei,YANG Xi-Ling,DONG Juan-E. Effects of methyl viologen on the antioxidant system in cultured Salvia miltiorrhiza cells. Chinese Journal of Plant Ecology, 2014, 38(5): 507-514. DOI: 10.3724/SP.J.1258.2014.00047

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链接本文: https://www.plant-ecology.com/CN/10.3724/SP.J.1258.2014.00047

https://www.plant-ecology.com/CN/Y2014/V38/I5/507

图/表 6

图1 不同浓度甲基紫精对丹参培养细胞中H2O2含量的影响(平均值±标准误差)。试验材料为培养6天的丹参悬浮培养细胞。不同小写字母表示差异显著(p < 0.05)。

Fig. 1 Effects of methyl viologen on the accumulation of H2O2 in cultured Salvia miltiorrhiza cells (mean ± SE). Test materials are S. miltiorrhiza cells in suspension cultures for six days. Different small letters indicate significant differences (p < 0.05).

图2 敌草隆(DCMU)和甲基紫精(MV)对丹参培养细胞中H2O2含量的影响(平均值±标准误差)。试验材料为培养6天的丹参悬浮培养细胞。DCMU在MV处理前30 min添加。不同小写字母表示差异显著(p < 0.05)。

Fig. 2 Effects of diuron (DCMU) and methyl viologen (MV) on the accumulation of H2O2 in cultured Salvia miltiorrhiza cells (mean ± SE). Test materials are S. miltiorrhiza cells in suspension cultures for six days. DCMU is added 30 min prior to the MV treatments. Different small letters indicate significant differences (p < 0.05).

图3 不同处理对丹参培养细胞中丙二醛含量的影响(平均值±标准误差)。DCMU, 敌草隆; MV, 甲基紫精。试验材料为培养6天的丹参悬浮培养细胞。DCMU在MV处理前30 min添加。不同小写字母表示差异显著(p < 0.05)。

Fig. 3 Effects of different treatments on the accumulation of malondialdehyde (MDA) in cultured Salvia miltiorrhiza cells (mean ± SE). DCMU, diuron; MV, methyl viologen. Test materials are S. miltiorrhiza cells in suspension cultures for six days. DCMU is added 30 min prior to the MV treatments. Different small letters indicate significant differences (p < 0.05).

图4 不同处理对丹参培养细胞中还原型谷胱甘肽含量的影响(平均值±标准误差)。DCMU, 敌草隆; MV, 甲基紫精。试验材料为培养6天的丹参悬浮培养细胞。DCMU在MV处理前30 min添加。不同小写字母表示差异显著(p < 0.05)。

Fig. 4 Effects of different treatments on the accumulation of glutathione (GSH) in cultured Salvia miltiorrhiza cells (mean ± SE). DCMU, diuron; MV, methyl viologen. Test materials are S. miltiorrhiza cells in suspension cultures for six days. DCMU is added 30 min prior to the MV treatments. Different small letters indicate significant differences (p < 0.05).

图5 不同处理对丹参培养细胞中超氧化物歧化酶(SOD)(A)、过氧化氢酶(CAT)(B)和过氧化物酶(POD)(C)活性的影响(平均值±标准误差)。DCMU, 敌草隆; MV, 甲基紫精。试验材料为培养6天的丹参悬浮培养细胞。DCMU在MV处理前30 min添加。不同小写字母表示差异显著(p < 0.05)。

Fig. 5 Effects of different treatments on the activities of superoxide dismutase (SOD) (A), catalase (CAT) (B), and peroxidase (POD) (C) in cultured Salvia miltiorrhiza cells (mean ± SE). DCMU, diuron; MV, methyl viologen. Test materials are S. miltiorrhiza cells in suspension cultures for six days. DCMU is added 30 min prior to the MV treatments. Different small letters indicate significant differences (p < 0.05).

图6 不同处理对丹参培养细胞中超氧化物歧化酶(SOD) (A)、过氧化氢酶(CAT)(B)和过氧化物酶(POD)(C)同工酶的影响。1, 蒸馏水处理; 2, 甲基紫精处理; 3, 敌草隆处理; 4, 甲基紫精+敌草隆联合处理。a-e, SOD, CAT, POD的同工酶条带。

Fig. 6 Effects of different treatments on the isoenzymes spectrum of superoxide dismutase (SOD) (A), catalase (CAT) (B), and peroxidase (POD) (C) in culture Salvia miltiorrhiza cells. 1, distilled water treatment; 2, methyl viologen treatment; 3, diuron treatment; 4, methyl viologen + diuron treatment. a-e, soenzyme bands of SOD, CAT and POD.

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