Release of hydrogen peroxide by rat type II pneumocytes in the prolonged culture.

Type II pneumocytes (T II pneumocytes) produce hydrogen peroxide (H(2)O(2)), which may be potentially dangerous for the lung. These cells in culture differentiate to type I-like pneumocytes and it may reflect the differentiation which follows the injury of alveolar epithelium. This work was undertaken to estimate the H(2)O(2) release by T II pneumocytes, freshly isolated and cultured up to 8 days. The light and electron microscopy evaluation confirmed the differentiation of T II pneumocytes to type I-like cells. The release of H(2)O(2), estimated spectrofluorimetrically as homovanillic acid oxidation product obtained in the presence of horseradish peroxidase, was significantly higher at day 4 (0.63+/-0. 68nmol/mg protein/min, P相似文献   

Dose-dependent cysteine-mediated protection of insulin-producing cells from damage by hydrogen peroxide

AIMS/HYPOTHESIS: Oxidative damage is believed to play a key role in the process of pancreatic beta cell destruction leading to type 1 diabetes. The beta cells are sensitive to oxidative stress because their intracellular anti-oxidative defence mechanisms are weak. The defence mechanisms depend heavily on glutathione, the synthesis of which is dependent on the availability of cysteine. We investigated whether an increased amount of cysteine available could protect beta cells from oxidative damage. METHODS: Rat insulinoma cells (RINm5F) were exposed to 50 or 100 microM hydrogen peroxide in the presence of three different cysteine concentrations (0.1, 1 and 5mM). Cell viability was analyzed by vital staining and the cellular metabolic status by C,N-diphenyl-N'-4,5-dimethyl thiazol-2-yl tetrazolium bromide (MTT) analysis. Intracellular insulin, DNA and glutathione contents were measured. The mechanism of death was further clarified by gel electrophoretic DNA fragmentation analysis. RESULTS: Hydrogen peroxide decreased cell viability and induced functional impairment. Vital staining indicated that 1mM cysteine effectively protected the cells. The protective effect was confirmed by the MTT assay showing preserved metabolic integrity, and by measurements of intact intracellular insulin and DNA content. Cysteine increased intracellular glutathione. Gel electrophoretic analysis of DNA revealed hydrogen peroxide-induced apoptotic fragmentation. This was also abolished by 1mM cysteine. The therapeutic window of cysteine was narrow: 0.1mM cysteine provided inadequate protection, and 5mM cysteine was already toxic in this setting. CONCLUSION: A proper dose of cysteine could provide a safe and effective means to protect beta cells from oxidative damage.     

Manganese promotes increased formation of hydrogen peroxide by activated human macrophages and neutrophils in vitro

Although pro-inflammatory mechanisms have been implicated in the pathogenesis of manganese (Mn(2+))-related neurological and respiratory disorders, relatively little is known about the potential of this metal to interact pro-oxidatively with human phagocytes. The primary objective of the current study was to investigate the effects of Mn(2+) as MnCl(2) (0.5-100 μM) on the generation of the reactive oxygen species (ROS), superoxide, hydrogen peroxide (H(2)O(2)), and hypohalous acids by isolated human blood neutrophils and monocyte-derived macrophages following activation of these cells with the chemotactic tripeptide, FMLP (1 μM), or the phorbol ester, PMA (25?ng/mL). Generation of ROS was measured using the combination of oxygen consumption, lucigenin/luminol-enhanced chemiluminescence, spectrofluorimetric detection of oxidation of 2,7-dichlorodihydrofluorescein, radiometric assessment of myeloperoxidase (MPO)-mediated protein iodination, release of MPO by ELISA, and spectrophotometric measurement of nitrite formation. Treatment of activated neutrophils with either FMLP or PMA resulted in significantly decreased reactivity of superoxide in the setting of increased formation of H(2)O(2) and MPO-mediated iodination, with no detectable effects on either oxygen consumption or MPO release. Similar effects of the metal with respect to superoxide reactivity and H(2)O(2) formation were observed with activated macrophages, while generation of NO was unaffected. Taken together with the findings of experiments using cell-free ROS-generating systems, these observations are compatible with a mechanism whereby Mn(2+), by acting as a superoxide dismutase mimetic, increases the formation of H(2)O(2) by activated phagocytes. If operative in vivo, this mechanism may contribute to the toxicity of Mn(2+).     

Manganese promotes increased formation of hydrogen peroxide by activated human macrophages and neutrophils in vitro

Although pro-inflammatory mechanisms have been implicated in the pathogenesis of manganese (Mn²⁺)-related neurological and respiratory disorders, relatively little is known about the potential of this metal to interact pro-oxidatively with human phagocytes. The primary objective of the current study was to investigate the effects of Mn²⁺ as MnCl₂ (0.5–100 µM) on the generation of the reactive oxygen species (ROS), superoxide, hydrogen peroxide (H₂O₂), and hypohalous acids by isolated human blood neutrophils and monocyte-derived macrophages following activation of these cells with the chemotactic tripeptide, FMLP (1 µM), or the phorbol ester, PMA (25?ng/mL). Generation of ROS was measured using the combination of oxygen consumption, lucigenin/luminol-enhanced chemiluminescence, spectrofluorimetric detection of oxidation of 2,7-dichlorodihydrofluorescein, radiometric assessment of myeloperoxidase (MPO)-mediated protein iodination, release of MPO by ELISA, and spectrophotometric measurement of nitrite formation. Treatment of activated neutrophils with either FMLP or PMA resulted in significantly decreased reactivity of superoxide in the setting of increased formation of H₂O₂ and MPO-mediated iodination, with no detectable effects on either oxygen consumption or MPO release. Similar effects of the metal with respect to superoxide reactivity and H₂O₂ formation were observed with activated macrophages, while generation of NO was unaffected. Taken together with the findings of experiments using cell-free ROS-generating systems, these observations are compatible with a mechanism whereby Mn²⁺, by acting as a superoxide dismutase mimetic, increases the formation of H₂O₂ by activated phagocytes. If operative in vivo, this mechanism may contribute to the toxicity of Mn²⁺.     

Augmentation of human neutrophil and alveolar macrophage LTB4 production by N-acetylcysteine: role of hydrogen peroxide

1. The actions of N-acetylcysteine (NAC) on hydrogen peroxide (H₂O₂) and leukotriene B₄ (LTB₄) production by human resting and stimulated peripheral blood neutrophils and alveolar macrophages were investigated.
2. At a concentration of 100 μM, NAC significantly (P<0.01) suppressed the accumulation of H₂O₂ in the incubation medium of resting and opsonized zymosan (OZ; 0.5 mg ml⁻¹)- or N-formylmethionyl-leucyl-phenylalanine (fMLP; 1 μM)-stimulated neutrophils and of resting and OZ-stimulated macrophages. At concentrations of 10 μM and above, NAC augmented significantly the level of LTB₄ in the supernatants of OZ- and fMLP-stimulated neutrophils (P<0.01 and P<0.05, respectively) and OZ-stimulated macrophages (P<0.05 at 10 μM, P<0.01 at 100 μM NAC).
3. NAC (100 μM) caused a significant (P<0.01) reduction in the quantity of measurable H₂O₂ when incubated with exogenous H₂O₂ concentrations equivalent to those released from OZ-stimulated neutrophils and macrophages. At no concentration did NAC affect quantitites of measurable LTB₄ when incubated with exogenous LTB₄.
4. Superoxide dismutase (SOD), which catalyzes the conversion of superoxide anion to H₂O₂ had no significant effect on LTB₄ production by human neutrophils. In contrast, catalase, which catalyzes the conversion of H₂O₂ to H₂O and O₂, caused a pronounced, statistically significant (P<0.01) increase in the levels of LTB₄ measured in the supernatants of OZ- and fMLP-stimulated neutrophils.
5. H₂O₂ (12.5 μM and 25 μM, concentrations equivalent to those measured in the supernatants of activated neutrophils and alveolar macrophages, respectively) caused a small (13%) decrease in the quantity of measurable LTB₄ (P=0.051 and P<0.05 at 12.5 μM and 25 μM, respectively) that was inhibited by NAC (100 μM) but not by catalase (400 u ml⁻¹).
6. In conclusion, the anti-oxidant drug, NAC, increases LTB₄ production by human neutrophils and alveolar macrophages, probably through the elimination of cell-derived H₂O₂. LTB₄ undergoes a H₂O₂-dependent oxidation that is inhibited by NAC but this is unlikely to account fully for the increased levels of LTB₄, suggesting that NAC may increase LTB₄ production by blocking the H₂O₂-dependent inhibition of a synthetic enzyme, such as 5-lipoxygenase.

     

Effects of some natural 5-hydroxy-isoflavones on cultured human endothelial cells in presence and absence of hydrogen peroxide

Due to their biological activity, it has been suggested that consumption of isoflavone-rich diets may prevent prostate and breast cancers, osteoporosis and cardiovascular diseases. Preventive effects of isoflavones on cancer and cardiovascular diseases have been associated with their oestrogenic and antioxidant properties. However, concerns still exist about the potential dangers of consuming high levels of these compounds, since it is known that some of them have cytostatic or cytotoxic properties, depending on the concentration. To evaluate the potential cytotoxic risk and antioxidant benefit of natural 5-hydroxy-isoflavones (5-OH-isoflavones) for human vascular endothelium, the effect of some natural 5-OH-isoflavones was evaluated on cultured human endothelial cells, in the presence and absence of H(2)O(2) (3 mM for 4 h). None of the isoflavones tested were able to prevent oxidative damage to endothelial cells at maximal extracellular concentrations of 1 mM. The low antioxidant capacity of these compounds was also shown by the DPPH (1,1-diphenyl-2-picrylhydrazyl radical) method. On the other hand, genistein and biochanin A, having a free 7-OH group, were toxic to the human endothelial cells in a dose-dependent manner, at concentrations >or=300 microM and >or= 100 microM, respectively. These results indicate that the non-specific cytotoxic effect of 5-OH-isoflavones is associated with the free 7-OH group. In conclusion, we were not able to show that 5-OH-isoflavones are beneficial to human endothelial cells when the cells were exposed to oxidative stress caused by 3 mM of H(2)O(2), but it can be concluded that consumption of 5-OH-isoflavones is of no direct cytotoxic risk to the human vascular endothelium since toxic concentrations are believed to be unreachable in-vivo.     

总丹酚酸对过氧化氢诱导内皮细胞损伤的抑制作用(英文)

目的　研究总丹酚酸对过氧化氢 (H2 O2 )诱导血管内皮细胞损伤的影响。方法　培养的人脐静脉内皮细胞 ,用MTT法测定内皮细胞的存活率。用碘化丙啶 (PI)染色和TUNEL法测定H2 O2 诱导内皮细胞的凋亡作用。用荧光法测定半胱天冬酶 (cas pase) 3的活性。用Fura 2 /AM测定内皮细胞内游离钙离子浓度 ([Ca2 + ] i)。结果　 1 0 0 μmol·L-1 H2 O2可使内皮细胞的存活率下降 40 .2 % ,提前 1h给予总丹酚酸 1 0和 1 0 0mg·L-1 使存活率分别增加8.4%和2 8.6 %。 1 0 0 μmol·L-1 H2 O2 还可引起内皮细胞的凋亡 ,PI染色的结果显示 ,H2 O2 处理内皮细胞 1 8h后 ,凋亡率从 (7.1 %± 0 .7) %升至 (38.1±4.0 ) %。总丹酚酸 1 0 0mg·L-1 提前处理内皮细胞1h,可使内皮细胞凋亡率降至 (1 3 .6± 0 .8) %。TUNEL方法检测结果显示 ,H2 O2 处理内皮细胞后 ,TUNEL染色阳性的细胞从 (3 .0± 0 .8) %升至 (30 .5±2 .9) % ,总丹酚酸 1 0 0mg·L-1 可使TUNEL阳性细胞率降至 (1 9.0± 3 .7) %。此外 ,H2 O2 处理细胞后 ,内皮细胞半胱天冬酶 3的活性增加 1 76% ,[Ca2 + ] i 升高 1 0 1 % ,而总丹酚酸可以抑制H2 O2 引起的半胱天冬酶 3活性的增加和 [Ca2 + ] i 升高。结论　总丹酚酸对H2 O2 引起的内皮细胞损伤具有明显的抑制作     

Lipid peroxidation products inhibit dopamine catabolism yielding aberrant levels of a reactive intermediate

Recent work indicates that oxidative stress is a factor in Parkinson's disease (PD); however, it is unknown how this condition causes selective dopaminergic cell death. The neurotransmitter dopamine (DA) has been implicated as an endogenous neurotoxin to explain the selective neurodegeneration. DA undergoes catabolism by monoamine oxidase (MAO) to the reactive intermediate 3,4-dihydroxyphenylacetaldehyde (DOPAL), which is further oxidized to 3,4-dihydroxyphenylacetic (DOPAC) acid via mitochondrial aldehyde dehydrogenase (ALDH). Previous studies found DOPAL to be more toxic than DA, and the major lipid peroxidation products, that is, 4-hydroxynonenal (4HNE) and malondialdehyde (MDA), potently inhibit ALDH. The hypothesis of this work is that lipid peroxidation products inhibit DOPAL oxidation, yielding aberrant levels of the reactive aldehyde intermediate. Treatment of striatal synaptosomes with 2-100 microM 4HNE or 2-50 microM MDA impaired DOPAL oxidation, resulting in elevated [DOPAL]. The aberrant concentration of DOPAL yielded an increase in protein modification by the DA-derived aldehyde, evident via staining of proteins with nitroblue tetrazolium (NBT). Pretreatment of synaptosomes with an MAO inhibitor significantly decreased NBT staining. On the basis of NBT staining, the order of protein reactivity for DA and metabolites was found to be DOPAL>DOPAC>DA. Mass spectrometric analysis of a model peptide reacted with DOPAL revealed the adduct to be a Schiff base product. In summary, these data demonstrate the sensitivity of DA catabolism to the lipid peroxidation products 4HNE and MDA even at low, physiologic levels and suggest a mechanistic link between oxidative stress and generation of aberrant levels of an endogenous and protein reactive dopaminergic toxin relevant to PD.     

Poly(ADP-ribose) polymerase inhibition improves endothelial dysfunction induced by reactive oxidant hydrogen peroxide in vitro

Reactive oxygen species, such as hydrogen peroxide (H(2)O(2)) induce oxidative stress and DNA-injury. The subsequent activation of poly(ADP-ribose) polymerase (PARP) has been implicated in the pathogenesis of various cardiovascular diseases including ischaemia-reperfusion injury, circulatory shock, diabetic complications and atherosclerosis. We investigated the effect of PARP-inhibition on endothelial dysfunction induced by H(2)O(2). In vascular reactivity measurements on isolated rat aortic rings we investigated the phenylephrine-induced contraction, and endothelium-dependent and -independent vasorelaxation by using cumulative concentrations of acetylcholine and sodium nitroprusside. Endothelial dysfunction was induced by exposing the rings to H(2)O(2) (200 and 400 muM) for 30 min. In the treatment group, rings were preincubated with the potent PARP-inhibitor INO-1001. DNA strand breaks were assessed by the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) method. Immunohistochemical analysis was performed for poly(ADP-ribose) (the enzymatic product of PARP) and for apoptosis inducing factor (a pro-apoptotic factor regulated by PARP). Exposure to H(2)O(2) resulted in reduced contraction forces and a dose-dependent impairment of endothelium-dependent vasorelaxation of aortic rings (maximal relaxation to acetylcholine: 86.21+/-1.574% control vs. 72.55+/-1.984% H(2)O(2) 200 muM vs. 66.86+/-1.961% H(2)O(2) 400 muM; P<0.05). PARP-inhibition significantly improved the acetylcholine-induced vasorelaxation (77.75+/-3.019% vs. 66.86+/-1.961%; P<0.05), while the contractility remained unaffected. The dose-response curves of endothelium-independent vasorelaxation to sodium nitroprusside did not differ in any groups studied. In the H(2)O(2) groups immunohistochemical analysis showed enhanced PARP-activation and nuclear translocation of apoptosis inducing factor, which were prevented by INO-1001. Our results demonstrate that PARP activation contributes to the pathogenesis of H(2)O(2)-induced endothelial dysfunction, which can be prevented by PARP inhibitors.     

Histamine release from serosal mast cells by intermediate products of arachidonic acid metabolism

In the present paper we report the results of experiments carried out to measure the release of histamine from isolated rat mast cells during the metabolic activation of arachidonic acid. Arachidonic acid (10(-8)-10(-4) M) and the terminal products (10(-6) M) of the arachidonic acid pathways were devoid of any significant histamine releasing properties. A substantial amount of histamine was released from rat mast cells by low concentrations of arachidonic acid during incubation with prostanoid generating systems, such as guinea-pig lung microsomes, rat serosal macrophages and polymorphonuclear cells and prostaglandin-H-synthase from calf seminal vesicles. The release of histamine was not accompanied by a leakage of lactate dehydrogenase and was blocked by D-mannitol and by lipoxygenase and cyclooxygenase pathway inhibitors. The data are consistent with the hypothesis that free radical derivatives of arachidonic acid, originating from hydroperoxy fatty acids, are generated during catalysis, causing mast cell histamine release.     

Inhibition of P-glycoprotein by natural products in human breast cancer cells

Multidrug resistance (MDR) is one of the most significant obstacles in cancer chemotherapy. One of the mechanisms involved in the development of MDR is the over-expression of P-glycoprotein (P-gp). It is widely known that natural compounds found in vegetables, fruits, plant-derived beverages and herbal dietary supplements not only have anticancer properties, but may also modulate P-gp activity. Therefore, the purpose of this investigation was to examine the effects of naturally occurring products on P-gp function in human breast cancer cell lines, MCF-7 (sensitive) and MCF-7/ADR (resistant). The accumulation of daunomycin (DNM), a P-gp substrate, was greater in the sensitive cells compared to the resistant cells, while the efflux of DNM was higher in the resistant cells compared to the sensitive cells over a period of 2 h. The IC50 value of DNM in the resistant cells was about 22 times higher than that in the sensitive cells, indicating an over-expression of P-gp in the resistant cells, MCF-7/ADR. All of the compounds tested, with the exception of fisetin, significantly decreased the IC50 value of DNM. Biochanin A showed the greatest increase in [3H]-DNM accumulation, increasing by 454.3 +/- 19.5% in the resistant cells, whereas verapamil, the positive control, increased the accumulation by 229.4 +/- 17.6%. Also, the accumulation of [3H]-DNM was increased substantially by quercetin and silymarin while it was reduced by fisetin. Moreover, biochanin A, silymarin, and naringenin significantly decreased DNM efflux from MCF-7/ADR cells compared with the control. These results suggest that some flavonoids such as biochanin A and silymarin may reverse MDR by inhibiting the P-gp function.     

促红细胞生成素对人视网膜色素上皮细胞氧化损伤的保护作用

目的探讨促红细胞生成素对过氧化氢氧化损伤人视网膜色素上皮细胞的保护作用及其机制。方法采用600μmol.L-1的过氧化氢造成体外培养的人视网膜色素上皮细胞氧化损伤模型。实验设计分为5组:对照组,过氧化氢模型组,过氧化氢+10 kIU.L-1EPO组,过氧化氢+20 kIU.L-1EPO组和过氧化氢+40 kIU.L-1EPO组。MTT法检测细胞活性,测定细胞内超氧化物歧化酶(SOD)和丙二醛(MDA)含量,Annexin V-FITC/PI双染色流式细胞法测定细胞的凋亡率。结果过氧化氢模型组RPE细胞的活性明显降低,不同浓度的EPO药物干预组的细胞活性较模型组明显升高,并随着EPO浓度的升高而升高。过氧化氢模型组RPE细胞SOD活性降低,MDA的含量增加;而EPO药物干预组SOD活性较模型组明显升高,MDA含量减少,差异有显著性。过氧化氢模型组RPE细胞的凋亡率升高,而不同浓度的EPO药物干预组降低RPE的凋亡率,凋亡率随着EPO浓度的升高而降低。结论促红细胞生成素对过氧化氢诱导的人RPE细胞的氧化损伤有保护作用,其机制可能与抑制氧化反应、提高抗氧化酶活性、减少细胞凋亡有关。     

过氧化氢氧化降解法制备低分子玻璃酸

目的制备低分子玻璃酸 (HA)。方法过氧化氢氧化降解法。结果随着过氧化氢浓度增加 ,反应温度的升高 ,降解速率加快。中性条件下HA的氧化降解速率较快 ,而酸性或碱性时却较慢。为了方便降解过程的可控性 ,过氧化氢浓度选 0 .0 5 % ,反应温度定为 5 0℃ ,反应pH为中性。随着HA相对分子质量的降低 ,运动黏度迅速下降 ,而糖醛酸含量基本不变。不同过氧化氢浓度降解时 ,低分子HA收率基本相同。结论过氧化氢氧化降解法可用于制备低分子HA。     

Aminoferrocene-based prodrugs activated by reactive oxygen species

Cancer cells generally generate higher amounts of reactive oxygen species than normal cells. On the basis of this difference, prodrugs have been developed (e.g., hydroxyferrocifen), which remain inactive in normal cells, but become activated in cancer cells. In this work we describe novel aminoferrocene-based prodrugs, which, in contrast to hydroxyferrocifen, after activation form not only quinone methides (QMs), but also catalysts (iron or ferrocenium ions). The released products act in a concerted fashion. In particular, QMs alkylate glutathione, thereby inhibiting the antioxidative system of the cell, whereas the iron species induce catalytic generation of hydroxyl radicals. Since the catalysts are formed as products of the activation reaction, it proceeds autocatalytically. The most potent prodrug described here is toxic toward cancer cells (human promyelocytic leukemia (HL-60), IC(50) = 9 μM, and human glioblastoma-astrocytoma (U373), IC(50) = 25 μM), but not toxic (up to 100 μM) toward representative nonmalignant cells (fibroblasts).     

Bacteriolysis is inhibited by hydrogen peroxide and by proteases

Treatment of Staphylococcus aureus in vitro with cationic agents results in the activation of their autolytic wall enzymes and in the degradation of their cell walls. Exposure of staphylococci either to hydrogen peroxide or the proteinases abolished the autolytic process. This effect was totally reversed by catalase and by proteinase inhibitors, respectively. It is suggested that the failure of neutrophils and macrophages to effectively degrade microbial cell wall components in inflammatory sites might be due to the inactivation of the autolytic wall enzymes of bacteria by hydrogen peroxide and by proteinases generated by the activated leukocytes. This might explain the prolonged chronic inflammatory sequelae seen following infections.     

Targeting cancer stem cells with natural products

The cancer stem cell (CSC) hypothesis presents a fundamentally different paradigm for cancer treatment. CSCs reflect a small fraction of tumor initiating cells capable of sustained self-renewal and differentiation to form the heterogeneous tumor bulk. In order to cure cancer, it is necessary to eliminate cancer stem cells in addition to differentiated cancer cells to decrease metastasis, reduce recurrence, and improve patient survival. In this article, we review cancer stem cell signaling pathways, including Wnt, Hedgehog, and Notch, as well as interactions of CSCs with the tumor microenvironment. We also review methods to isolate CSCs and demonstrate therapeutic efficacy of natural products to modulate these signaling pathways for eliminating CSCs.     

过氧化氢诱导人肝癌细胞BEL7402产生氧化应激细胞模型的建立

目的 研究过氧化氢(H₂O₂)构建体外人肝癌细胞BEL7402氧化应激模型的最佳作用浓度及时间。方法 采用噻唑蓝(MTT)染色法检测以10、100和1 000 μmol/L的H₂O₂处理0、2、4、6、8、10、12和24 h后BEL7402细胞的存活率;采用双氢罗丹明123(DHR123)探针法、单细胞凝胶电泳技术和分光光度法检测以100 μmol/L H₂O₂处理BEL7402细胞4 h后线粒体活性氧(ROS)含量、DNA损伤、丙二醛(MDA)含量、超氧化物歧化酶(SOD)及谷胱甘肽过氧化物酶(GSH-PX)活性。结果 100 μmol/L H₂O₂处理BEL7402细胞4 h后,其存活率为(43.44±5.74)%,与10 μmol/L H₂O₂处理组(92.77±5.51)%相比,细胞存活率降低,差异有统计学意义(P<0.01);ROS含量结果显示100 μmol/L H₂O₂处理BEL7402细胞40 min时其体内的ROS含量达到峰值,之后呈下降趋势;与健康对照组比较,100 μmol/L H₂O₂组OTM值升高(P<0.05),MDA含量升高,差异有统计学意义(P<0.01),SOD和GSH-Px活性均下降(P<0.05或P<0.01)。结论 100 μmol/L H₂O₂、预处理4 h,可成功构建BEL7402细胞体外氧化应激模型,可为通过氧化应激机制抗肝病新药的研发奠定基础。     

M受体对H_2O_2诱导PC12细胞凋亡的保护作用及机制

目的研究毒蕈碱型胆碱受体在抗氧化应激所诱导的凋亡中不同亚型之间是否存在差异。方法用含有M1M4毒蕈碱型胆碱受体亚型的质粒pCDNA3.0转染PC12细胞,加入过氧化氢诱导凋亡,观察氨甲酰胆碱激动毒蕈碱型胆碱受体后各亚型之间抗凋亡能力是否存在差异,同时应用毒蕈碱型胆碱受体抑制剂阿托品,ERK和PI3K信号通路抑制剂PD98059与LY294002观察参与保护作用的相关的信号通路。结果氨甲酰胆碱激动毒蕈碱型胆碱受体后产生抗凋亡效应,各亚型之间没有差异,PI3K信号通路参与了保护作用。结论M1M4亚型毒蕈碱型胆碱受体在对抗过氧化氢诱导的凋亡没有亚型之间的差异,PI3K信号通路参与了保护作用。     

辛伐他汀对过氧化氢诱导的骨髓间充质干细胞凋亡的影响

目的 研究辛伐他汀对过氧化氢(H2O2)诱导的骨髓间充质干细胞(BMSCs)凋亡的影响,并探讨其可能的机制.方法 采用全骨髓贴壁培养法培养SD大鼠BMSCs,随机分成7组:正常对照组(A组)、H2O2处理组(B组)和不同浓度辛伐他汀处理组(C1、C2、C3、C4组,分别为辛伐他汀0.001、0.01、0.1、1 μmol/L)及LY294002干预组(D组).流式细胞术检测各组细胞凋亡率,并用Western blot检测BMSCs中p-Akt、Akt表达.结果 与B组比较,辛伐他汀处理组均抑制H2O2诱导的BMSCs凋亡,以C2组抗凋亡作用最为显著(P<0.05);而LY294002(D组)能明显阻断辛伐他汀的抗凋亡效应(P<0.01).辛伐他汀处理组p-Akt活性显著高于A、B组(P<0.01),而LY294002(D组)阻断p-Akt水平(P<0.01).结论 一定浓度范围内的辛伐他汀能抑制H2O2诱导的BMSCs凋亡,其机制可能与PI3K/Akt信号通路有关.