植物细胞中蓝光诱导ROS生成的识别和亚细胞定位检测

以2’,7’-二氯二氢荧光素二乙酯(dichlorofluorescein diacetate,H2DCF-DA)为荧光探针孵育拟南芥叶表皮条,利用荧光光谱和激光共聚焦扫描显微技术,对高辐照蓝光诱导下叶肉细胞活性氧(reactive oxygen spe-cies,ROS)的生成,进行了分子识别和亚细胞定位检测。结果表明:植物细胞在蓝光诱导下,可以产生大量的ROS。过氧化氢酶清除实验表明:高辐照蓝光诱导产生的ROS,主要成分是H2O2,并且主要定位在叶绿体和细胞膜上。     

H(2)O(2) detection from intact mitochondria as a measure for one-electron reduction of dioxygen requires a non-invasive assay system.

Evaluation of the existence of superoxide radicals (O*-(2)), the site of generation and conditions required for one-e(-) transfer to oxygen from biological redox systems is a prerequisite for the understanding of the deregulation of O(2) homeostasis leading to oxidative stress. Mitochondria are increasingly considered the major O*-(2) source in a great variety of diseases and the aging process. Contradictory reports on mitochondrial O*-(2) release prompted us to critically investigate frequently used O*-(2) detection methods for their suitability. Due to the impermeability of the external mitochondrial membrane for most constituents of O*-(2) detection systems we decided to follow the stable dismutation product H(2)O(2). This metabolite was earlier shown to readily permeate into the cytosol. With the exception of tetramethylbenzidine none of the chemical reactants indicating the presence of H(2)O(2) by horseradish peroxidase-catalyzed absorbance change were suited due to solubility problems or low extinction coefficients. Tetramethylbenzidine-dependent H(2)O(2) detection was counteracted by rereduction of the dye through e(-) carriers of the respiratory chain. Although the fluorescent dyes scopoletin and homovanillic acid were found to be suited for the detection of mitochondrial H(2)O(2) release, fluorescence change was strongly affected by mitochondrial protein constituents. The present study has resolved this problem by separating the detection system from H(2)O(2)-producing mitochondria.     

Hydrogen peroxide production in Streptococcus pyogenes: involvement of lactate oxidase and coupling with aerobic utilization of lactate

Streptococcus pyogenes strains can be divided into two classes, one capable and the other incapable of producing H2O2 (M. Saito, S. Ohga, M. Endoh, H. Nakayama, Y. Mizunoe, T. Hara, and S. Yoshida, Microbiology 147:2469-2477, 2001). In the present study, this dichotomy was shown to parallel the presence or absence of H2O2-producing lactate oxidase activity in permeabilized cells. Both lactate oxidase activity and H2O2 production under aerobic conditions were detectable only after glucose in the medium was exhausted. Thus, the glucose-repressible lactate oxidase is likely responsible for H2O2 production in S. pyogenes. Of the other two potential H2O2-producing enzymes of this bacterium, NADH and alpha-glycerophosphate oxidase, only the former exhibited low but significant activity in either class of strains. This activity was independent of the growth phase, suggesting that the protein may serve in vivo as a subunit of the H2O2-scavenging enzyme NAD(P)H-linked alkylhydroperoxide reductase. The activity of lactate oxidase was associated with the membrane while that of NADH oxidase was in the soluble fraction, findings consistent with their respective physiological roles, i.e., the production and scavenging of H2O2. Analyses of fermentation end products revealed that the concentration of lactate initially increased with time and decreased on glucose exhaustion, while that of acetate increased during the culture. These results suggest that the lactate oxidase activity of H2O2-producing cells oxidizes lactate to pyruvate, which is in turn converted to acetate. This latter process proceeds presumably via acetyl coenzyme A and acetyl phosphate with formation of extra ATP.     

Lactococcus lactis SpOx spontaneous mutants: a family of oxidative-stress-resistant dairy strains

Numerous industrial bacteria generate hydrogen peroxide (H(2)O(2)), which may inhibit the growth of other bacteria in mixed ecosystems. We isolated spontaneous oxidative-stress-resistant (SpOx) Lactococcus lactis mutants by using a natural selection method with milk-adapted strains on dairy culture medium containing H(2)O(2). Three SpOx mutants displayed greater H(2)O(2) resistance. One of them, SpOx3, demonstrated better behavior in different oxidative-stress situations: (i) higher long-term survival upon aeration in LM17 and milk and (ii) the ability to grow with H(2)O(2)-producing Lactobacillus delbrueckii subsp. delbrueckii strains. Furthermore, the transit kinetics of the SpOx3 mutant in the digestive tract of a human flora-associated mouse model was not affected.     

产H_2O_2的乳酸杆菌分离培养基的改良

对MRS培养基的使用方法进行了改良。将MRS(含0.25 mg/ml TMB)融化后倾倒入平皿,待其凝固后,在琼脂表明加入200μl(0.01 mg/ml)HRP,涂匀后进行乳酸杆菌划线分离。37℃、厌氧培养48~72 h。结果表明,产生H2O2的乳酸杆菌呈蓝色菌落,和原来的培养方法相比,改良的方法简便经济、HPR的使用量为原来的1%,分离产H2O2的乳酸杆菌的效果良好。     

Hydrogen photoproduction by nutrient-deprived Chlamydomonas reinhardtii cells immobilized within thin alginate films under aerobic and anaerobic conditions

A new technique for immobilizing H2-photoproducing green algae within a thin (<400 microm) alginate film has been developed. Alginate films with entrapped sulfur/phosphorus-deprived Chlamydomonas reinhardtii, strain cc124, cells demonstrate (a) higher cell density (up to 2,000 microg Chl mL(-1) of matrix), (b) kinetics of H2 photoproduction similar to sulfur-deprived suspension cultures, (c) higher specific rates (up to 12.5 micromol mg(-1) Chl h(-1)) of H2 evolution, (d) light conversion efficiencies to H2 of over 1% and (e) unexpectedly high resistance of the H2-photoproducing system to inactivation by atmospheric O2. The algal cells, entrapped in alginate and then placed in vials containing 21% O2 in the headspace, evolved up to 67% of the H2 gas produced under anaerobic conditions. The results indicate that the lower susceptibility of the immobilized algal H2-producing system to inactivation by O2 depends on two factors: (a) the presence of acetate in the medium, which supports higher rates of respiration and (b) the capability of the alginate polymer itself to effectively separate the entrapped cells from O2 in the liquid and headspace and restrict O2 diffusion into the matrix. The strategy presented for immobilizing algal cells within thin polymeric matrices shows the potential for scale-up and possible future applications.     

槲皮素对氧化应激大鼠肝细胞氨基酸代谢的影响

通过检测原代培养的大鼠肝细胞培养液中18种氨基酸含量的变化情况,探讨槲皮素对氧化应激大鼠肝细胞氨基酸代谢的影响。结果表明,与对照组相比,槲皮素组氨基酸质量浓度无明显变化;H2O2组、槲皮素+H2O2组培养液中丙氨酸、蛋氨酸和组氨酸的质量浓度显著减少(P<0.05),而牛磺酸的质量浓度显著增加(P<0.05),上述变化槲皮素+H2O2组较H2O2组更明显,槲皮素+H2O2组M et和H is的质量浓度显著低于H2O2组(P<0.05)。说明槲皮素可影响氧化应激肝细胞某些氨基酸,尤其是含硫氨基酸的代谢。这种影响可能是槲皮素抗氧化应激作用的机制之一。     

Electrospun polystyrene-poly(styrene-co-maleic anhydride) nanofiber as a new aptasensor platform

Here we report on a new approach for the electrochemical detection of hydrogen peroxide (H(2)O(2)) based on the co-immobilization of horseradish peroxidase and methylene blue on the functionalized carbon buckypaper supported by a titanium substrate. Cyclic voltammetry was used to study and optimize the performance of the resulting electrochemical biosensor. The proposed biosensor exhibited high analytical performance towards the quantification of H(2)O(2) at the physiological pH 7.4. Under optimized conditions, the biosensor shows a wide linear response range from 0.1 × 10(-6) to 5 × 10(-4)M concentrations of H(2)O(2). The detection limit was determined to be 7.5 × 10(-8)M (based on S/N=3). Reproducibility and stability of the fabricated biosensor were examined with satisfactory results. The biological relevance of the developed electrochemical biosensor has been further studied by the determination of H(2)O(2) in human urine samples of normal volunteers prior to and following the ingestion of coffee. Increased levels of urinary H(2)O(2) concentration suggest that oxidative stress is induced by coffee drinking in humans. There is considerable interest in oxidative stress as relates to human physiology. The sensitive determination of H(2)O(2) in human urine may serve as a valuable biomarker to effectively elucidate specific levels of oxidative stress in vivo.     

Polyoxometalates as peroxidase mimetics and their applications in H2O2 and glucose detection

Polyoxometalates (H(3)PW(12)O(40), H(4)SiW(12)O(40) and H(3)PMo(12)O(40)) have been proven to possess intrinsic peroxidase-like activity for the first time, which can catalyze oxidation of the peroxidase substrate 3,3',5,5'-tetramethylbenzidine (TMB) by H(2)O(2) to form a blue color in aqueous solution. Among them, H(3)PW(12)O(40) (PW(12)) exhibits higher catalytic activity to TMB than natural enzyme HRP and other two POMs. In addition, H(3)PW(12)O(40)/graphene exhibited higher activity than H(3)PW(12)O(40) in this catalytic oxidation reaction due to the effect of graphene in promoting the electron transfer between the substrate and catalyst. POMs/H(2)O(2)/TMB system provides a simple, accurate approach to colorimetric detection for H(2)O(2) or glucose. The colorimetric method based on POMs showed good response toward H(2)O(2) and glucose detection with a linear range from 1.34×10(-7) to 6.7×10(-5) mol/L and 1×10(-7) to 1×10(-4) mol/L, respectively. The results showed that it is a simple, cheap, more convenient, highly selective, sensitive, and easy handling colorimetric assay.     

Altered O2-/H2O2 production ratio by in vitro and in vivo primed human neutrophils

Human neutrophils were primed by exudation or pretreatment with a synthetic diacylglycerol (diC10), the Ca2+ ionophore ionomycin or lipopolysaccharide (LPS). Compared to control cells, these primed cells showed a significantly decreased O2-/H2O2 ratio when stimulated with formylmethionyl-leucyl-phenylalanine (FMLP). This shift indicates a comparative (and net) increased H2O2 detection in the extracellular medium and can not be explained by a dose-dependent impairment in either O2- or H2O2 detecting capacity. An altered H2O2 degenerating capacity was not observed in the primed cells. We propose that priming enhances the capacity to divalently reduce oxygen and thereby directly produce H2O2.     

Sensitive immunoassay of human chorionic gonadotrophin based on multi-walled carbon nanotube–chitosan matrix

A novel amperometric immunosensor for human chorionic gonadotropin (HCG) assay has been fabricated through incorporating toluidine blue (TB) and hemoglobin (Hb) on the multiwall carbon nanotube (MWNT)-chitosan (CS) modified glassy carbon electrode, followed by electrostatic adsorption of a conducting gold nanoparticles (nanogold) film as sensing interface. The MWNT-CS matrix provided a congenial microenvironment for the immobilization of biomolecules and promoted the electron transfer to enhance the sensitivity of the immunosensor. Due to the strong electrocatalytic properties of Hb and MWNT toward H(2)O(2), the Hb and MWNT significantly amplified the current signal of the antigen-antibody reaction. The immobilized toluidine blue as an electron transfer mediator exhibited excellent electrochemical redox property. After the immunosensor was incubated with HCG solution, the access of activity center of the Hb to toluidine blue was partly inhibited, which leaded to a linear decrease in the catalytic efficiency of the Hb to the oxidation of immobilized toluidine blue by H(2)O(2) over HCG concentration ranges from 0.8 to 500 mIU/mL. Under optimal condition, the detection limit for the HCG immunoassay was 0.3 mIU/mL estimated at a signal-to-noise ratio of 3. Moreover, the proposed immunosensor displayed a satisfactory stability and reproducibility.     

Bovine aortic endothelial cells release hydrogen peroxide.

Endothelial cells grown on microcarriers are able to release H2O2 to the extracellular environment without any added stimulus. The extracellularly released H2O2 can be detected by luminol-amplified chemiluminescence (CL) if horseradish peroxidase is added. The CL response can be reduced by catalase and blocked by superoxide dismutase, indicating that O2- could be a precursor for H2O2. The CL kinetics, i.e., a long lag time followed by a rapid shift to a new level, indicate activation of an O2(-)-producing enzyme. The cells are also able to protect themselves from H2O2 stimulation by both catalase and the glutatione system. Bradykinin stimulates the H2O2 release, but if the effect is directly stimulatory or if it acts by reduction of the protective system is at present unclear. The extracellularly released H2O2 could be a cause of injury to the endothelial cells or to the subendothelial matrix.     

Nitric oxide inhibits blue light-specific stomatal opening via abscisic acid signaling pathways in Vicia guard cells

Recent evidence suggests that nitric oxide (NO) acts as an intermediate of ABA signal transduction for stomatal closure. However, NO's effect on stomatal opening is poorly understood even though both opening and closing activities determine stomatal aperture. Here we show that NO inhibits stomatal opening specific to blue light, thereby stimulating stomatal closure. NO inhibited blue light-specific stomatal opening but not red light-induced opening. NO inhibited both blue light-induced H(+) pumping and H(+)-ATPase phosphorylation. The NO scavenger 2-carboxyphenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (c-PTIO) restored all these inhibitory effects. ABA and hydrogen peroxide (H(2)O(2)) inhibited all of these blue light-specific responses in a manner similar to NO. c-PTIO partially restored the ABA-induced inhibition of all of these opening responses but did not restore inhibition of the responses by H(2)O(2). ABA, H(2)O(2) and NO had slight inhibitory effects on the phosphorylation of phototropins, which are blue light receptors in guard cells. NO inhibited neither fusicoccin-induced H(+) pumping in guard cells nor H(+) transport by H(+)-ATPase in the isolated membranes. From these results, we conclude that both NO and H(2)O(2) inhibit blue light-induced activation of H(+)-ATPase by inhibiting the component(s) between phototropins and H(+)-ATPase in guard cells and stimulate stomatal closure by ABA.     

健康青少年口腔优势菌产生过氧化氢的初步研究

目的研究健康青少年口腔优势菌产生过氧化氢的能力。方法采用过氧化物酶法,在体外测得健康青少年口腔优势菌产生过氧化氢的量。结果在需氧条件下,能产生过氧化氢,产生的量与细菌的生长速度成正比;在厌氧条件下,未测得过氧化氢的产生。结论健康青少年口腔优势菌产生过氧化氢的能力强于S.sATCC10556(P0.05)     

The E-loop is involved in hydrogen peroxide formation by the NADPH oxidase Nox4

In contrast to the NADPH oxidases Nox1 and Nox2, which generate superoxide (O(2)(·-)), Nox4 produces hydrogen peroxide (H(2)O(2)). We constructed chimeric proteins and mutants to address the protein region that specifies which reactive oxygen species is produced. Reactive oxygen species were measured with luminol/horseradish peroxidase and Amplex Red for H(2)O(2) versus L-012 and cytochrome c for O(2)(·-). The third extracytosolic loop (E-loop) of Nox4 is 28 amino acids longer than that of Nox1 or Nox2. Deletion of E-loop amino acids only present in Nox4 or exchange of the two cysteines in these stretches switched Nox4 from H(2)O(2) to O(2)(·-) generation while preserving expression and intracellular localization. In the presence of an NO donor, the O(2)()-producing Nox4 mutants, but not wild-type Nox4, generated peroxynitrite, excluding artifacts of the detection system as the apparent origin of O(2)(·-). In Cos7 cells, in which Nox4 partially localizes to the plasma membrane, an antibody directed against the E-loop decreased H(2)O(2) but increased O(2)(·-) formation by Nox4 without affecting Nox1-dependent O(2)(·-) formation. The E-loop of Nox4 but not Nox1 and Nox2 contains a highly conserved histidine that could serve as a source for protons to accelerate spontaneous dismutation of superoxide to form H(2)O(2). Mutation of this but not of four other conserved histidines also switched Nox4 from H(2)O(2) to O(2)(·-) formation. Thus, H(2)O(2) formation is an intrinsic property of Nox4 that involves its E-loop. The structure of the E-loop may hinder O(2)(·-) egress and/or provide a source for protons, allowing dismutation to form H(2)O(2).     

The mechanism of action of lymphokines. IX. The enzymatic basis of hydrogen peroxide production by lymphokine-activated macrophages

The purpose of this study was to elucidate the biochemical basis of the enhanced hydrogen peroxide (H2O2) production by guinea pig peritoneal macrophages (MP) cultured in lymphokine (LK)-containing medium. The markedly augmented H2O2 generation by these cells, demonstrable by the horseradish peroxidase (HRP)-catalyzed oxidation of phenol red, is distinguished by its lack of dependence on a second stimulus. We demonstrate that H2O2 production is truly spontaneous and is not caused by a stimulant present among the H2O2 assay reagents. The principal candidate for such a role was HRP type II (a mixture of five isoenzymes) that was reported to be capable of eliciting an oxidative burst in MP. Four distinct HRP isoenzymes that were found incapable of provoking an oxidative response were nevertheless adequate for demonstrating H2O2 production by LK-activated MP. Blocking the MP receptor for mannose by the addition of mannan to the assay system resulted in enhanced detection of H2O2 by low concentrations of HRP type II and by three out of four HRP isoenzymes. Treatment of MP with LK-containing medium for 72 hr did not result in a significant change in the activity of cellular superoxide dismutase (SOD) compared with MP cultured for the same length of time in control medium. By using the specific inhibitor of copper, zinc-containing SOD, sodium diethyldithiocarbamate (DDC), and the universal SOD inhibitor, sodium nitroprusside, we found that the predominant enzyme in guinea pig peritoneal MP is probably manganese-containing SOD. Incubation of LK-activated MP with nitroprusside resulted in almost total inhibition of H2O2 production and a simultaneous switch to superoxide (O2-) liberation. Similar exposure to DDC had no effect. These data indicate that H2O2 produced by LK-activated MP is derived exclusively by enzymatic dismutation of O2- mediated by a manganese-containing SOD. The increase in spontaneous H2O2 production induced by LK is therefore secondary to augmented O2- production that occurs at a cellular location where O2- is accessible to SOD. The enzymatic basis of the enhanced oxygen radical production was investigated by determining the kinetic parameters of the O2- -forming NADPH oxidase of resting LK-treated MP in a cellfree system in which O-2 production was induced by sodium dodecyl sulfate. The Km for NADPH and the Vmax of the enzyme of LK-treated MP were not different from those of the enzyme of MP incubated in control medium. We conclude that LK treatment of MP does not modulate the NADPH oxidase itself but, most likely, a process related to activation of the enzyme.     

Superoxide generated by glutathione reductase initiates a vanadate-dependent free radical chain oxidation of NADH.

Vanadate V(V) markedly stimulated the oxidation of NADPH by GSSG reductase and this oxidation was accompanied by the consumption of O2 and the accumulation of H2O2. Superoxide dismutases completely eliminated this effect of V(V), whereas catalase was without effect, as was exogenous H2O2 added to 0.1 mM. These effects could be seen equally well in phosphate- or in 4-(2-hydroxyethyl)1-piperazineethanesulfonic acid-buffered solutions. Under anaerobic conditions there was no V(V)-stimulated oxidation of NADPH. Approximately 4% of the electrons flowing from NADPH to O2, through GSSG reductase, resulted in release of O2-. The average length of the free radical chains causing the oxidation of NADPH, initiated by O2- plus V(V), was calculated to be in the range 140-200 NADPH oxidized per O2- introduced. We conclude that GSSG reductase, and by extension other O2(-)-producing flavoprotein dehydrogenases such as lipoyl dehydrogenase and ferredoxin reductase, catalyze V(V)-stimulated oxidation of NAD(P)H because they release O2- and because O2- plus V(V) initiate a free radical chain oxidation of NAD(P)H. There is no reason to suppose that these enzymes can act as NAD(P)H:V(V) oxidoreductases.     

Proton efflux coupled to dark H2 oxidation in whole cells of a marine sulfur photosynthetic bacterium (Chromatium sp. strain Miami PBS1071).

Whole cells of photoanaerobically grown Chromatium sp. strain Miami PBS1071, a marine sulfur purple bacterium, oxidized H2 in the dark through the oxyhydrogen reaction at rates of up to 59 nmol of H2 per mg (dry weight) per min. H2 oxidation was routinely measured in H2 pulse experiments with air-equilibrated cells. The reaction was accompanied by a reversible H+ efflux from the cells, suggesting an outward H+ translocation reaction coupled to H2 oxidation. The H+/e- ratio, calculated from simultaneous measurements of H2, O2, and H+ changes in the medium, varied with the cultures from 0.7 to 1.2. The ratio increased considerably when the backflow of H+ was taken into account. Anaerobic H2 uptake with 2,5-dimethyl-p-benzoguinone as an oxidant also showed a weak H+-translocating activity. No H+-translocating activity was detected with methylene blue as an oxidant. Carbonylcyanide 3-chlorophenylhydrazone (1 microM) stimulated H2 oxidation and abolished the associated H+ changes when H2 oxidation was observed in O2 pulse experiments with H2-Ar-equilibrated cells. However, the uncoupler inhibited both H2 oxidation and H+ changes when measurements were made in H2 pulse experiments with air-equilibrated cells. It is suggested that in this bacterium the susceptibility of hydrogenase to reversible O2 inactivation in situ is enhanced by the presence of uncoupling agents.     

1,25-Dihydroxyvitamin D3 increases the toxicity of hydrogen peroxide in the human monocytic line U937: the role of calcium and heat shock

1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) increases synthesis of heat shock proteins in monocytes and U937 cells and protects these cells from thermal injury. We examined whether 1,25-(OH)2D3 would also modulate the susceptibility of U937 cells to H2O2-induced oxidative stress. Cell viability was assessed by trypan blue exclusion and [3H]thymidine incorporation into DNA. Prior incubation for 24 h with 1,25-(OH)2D3 (25 pM or higher) unexpectedly increased H2O2 toxicity. Since cellular Ca2+ may be a mediator of cell injury we investigated effects of altering extracellular Ca2+ ([Ca2+]e) on 1,25-(OH)2D3-enhanced H2O2 toxicity as well as effects of 1,25-(OH)2D3 and H2O2 on cytosolic free Ca2+ concentration ([Ca2+]f). Basal [Ca2+]f in medium containing 1.5 mM Ca as determined by fura-2 fluorescence was higher in 1,25-(OH)2D3-pretreated cells than control cells (137 versus 112 nM, P less than 0.005). H2O2 induced a rapid increase in [Ca2+]f (to greater than 300 nM) in both 1,25-(OH)2D3-treated and control cells, which was prevented by a reduction in [Ca2+]e to less than basal [Ca2+]f. The 1,25(OH)2D3-induced increase in H2O2 toxicity was also prevented by preincubation with 1,25-(OH)2D3 in Ca2+-free medium or by exposing the cells to H2O2 in the presence of EGTA. Preexposure of cells to 45 degrees C for 20 min, 4 h earlier, partially prevented the toxic effects of H2O2 particularly in 1,25-(OH)2D3-treated cells, even in the presence of physiological levels of [Ca2+]e. Thus 1,25-(OH)2D3 potentiates H2O2-induced injury probably by increasing cellular Ca2+ stores. The 1,25-(OH)2D3-induced amplification of the heat shock response likely represents a mechanism for counteracting the Ca2+-associated enhanced susceptibility to oxidative injury due to 1,25-(OH)2D3.     

ERK1／2信号通路对黄芪苷Ⅳ抗H2O2诱导H9c2细胞氧化损伤的作用

目的：研究黄芪苷Ⅳ（AST）是否通过细胞外信号调节激酶1／2（ERK1／2）通路发挥对H2O2诱导的H9c2细胞氧化损伤的保护作用。方法：用200μmoL／L的H2O2处理细胞6h,采用MTT法检测细胞存活率,建立H2O2诱导的H9c2细胞氧化损伤模型;比色法测定细胞培养液中乳酸脱氢酶（LDH）活性、总超氧化物歧化酶（T—SOD）和锰超氧化物歧化酶（Mn—SOD）活力以及丙二醛（MDA）含量;Western blot检测H9c2细胞ERK1／2蛋白的磷酸化水平。结果：在H2O2浓度为200μmol／L作用6h条件下,细胞存活率降低程度适中,实验结果重复性好,确定后续实验采用200μmol／L H2O2作用6h建立模型。与H2O2组比较,10mg／L及20mg／L AST均显著提高细胞存活率（P〈0．01）,使细胞培养液中LDH活性显著降低（P〈0．01）,T—SOD及Mn—SOD活力显著提高（P〈0．01）,MDA含量显著降低（P〈0．01）。10mg/L及20mg/L AST均显著增加H2O2损伤的H9c2细胞p—ERK1／2蛋白的表达（P〈0．01）,当用PD98059（ERK1／2的抑制剂）预处理后,AST的作用则被取消。结论：黄芪苷Ⅳ可以通过ERK1／2通路发挥对H2O2诱导的H9c2细胞氧化损伤的保护作用。