Hyaluronan fragments contribute to the ozone-primed immune response to lipopolysaccharide

Hyaluronan is a high-molecular mass component of pulmonary extracelluar matrix, and lung injury can generate a low-molecular mass hyaluronan (HA) fragment that functions as endogenous ligand to cell surface receptors CD44 and TLR4. This leads to activation of intracellular NF-κB signaling and proinflammatory cytokine production. Based on previous information that ozone exposure causes increased HA in bronchial alveolar lavage fluid and ozone pre-exposure primes immune response to inhaled LPS, we hypothesized that HA production during ozone exposure augments the inflammatory response to LPS. We demonstrate that acute ozone exposure at 1 part per million for 3 h primes the immune response to low-dose aerosolized LPS in C57BL/6J mice, resulting in increased neutrophil recruitment into the airspaces, increased levels of protein and proinflammatory cytokines in the bronchoalveolar lavage fluid, and increased airway hyperresponsiveness. Intratracheal instillation of endotoxin-free HA (25 μg) enhances the biological response to inhaled LPS in a manner similar to ozone pre-exposure. In vitro studies using bone marrow-derived macrophages indicate that HA enhances LPS responses measured by TNF-α production, while immunofluorescence staining of murine alveolar macrophages demonstrates that HA induces TLR4 peripheralization and lipid raft colocalization. Collectively, our observations support that ozone primes macrophage responsiveness to low-dose LPS, in part, due to HA-induced TLR4 peripheralization in lung macrophages.     

Effects of Triiodothyronine (T3) Supplementation Upon Ozone-Induced Lung Injury

Ozone exposure results in an acute decrease in the serum levels of thyroid hormones; the physiologic sequelae of this are unclear. Whereas thyroid hormone supplementation appears to benefit pulmonary function in septic, oxyradical models of injury, thyroid hormone increases ozone toxicity. We demonstrated an increase in metabolic rate and pulmonary injury in lungs from ozone exposed, T₃ treated animals. This was evidenced by an increase in pulmonary weight gain, vascular perfusion pressure, and decrease in compliance in the supplemented animals. However, an increase in alkane generation, as an index of lipid peroxidation, was not seen in the ozone exposed, hormonally treated animals. This suggests that although thyroid hormone supplementation increases metabolic rate and ozone toxicity, an increased rate of lipid peroxidation plays a minimal role.     

Effects of ozone and peroxyacetyl nitrate on polar lipids and Fatty acids in leaves of morning glory and kidney bean

To compare the effects of ozone and peroxyacetyl nitrate (PAN) on leaf lipids, fatty acids and malondialdehyde (MDA), morning glory (Pharbitis nil Choisy cv Scarlet O'Hara) and kidney bean (Phaseolus vulgaris L. cv Gintebo) plants were exposed to either ozone (0.15 microliter per liter for 8 hours) or PAN (0.10 microliter per liter for up to 8 hours). Ozone increased phospholipids in morning glory and decreased in kidney bean at the initial stage (2-4 hours) of exposure, while it scarcely changed glycolipids, the unsaturated fatty acids, and MDA in both plants. A large reduction of glycolipids occurred 1 day after ozone exposure in both plants. PAN caused marked drops in phospholipids and glycolipids in kidney bean at relatively late stage (6-8 hours) of exposure, while it increased phosphatidic acid and decreased the unsaturated fatty acids, an increase which was accompanied by a large increase in MDA. These results suggest that ozone may not directly oxidize unsaturated fatty acids at the initial stage of exposure, but may alter polar lipid metabolism, particularly phospholipids. On the other hand, PAN may abruptly and considerably degrade phospholipids and glycolipids by peroxidation or hydrolysis at the late stage of exposure. The present study shows that ozone and PAN affect polar lipids in different manners.     

Mechanism of oxidative damage to fish red blood cells by ozone

The present study was conducted to elucidate the adverse effects of ozone exposure on rainbow trout (Oncorhynchus mykiss) red blood cells (RBCs). We evaluated whether hemoglobin (Hb) or Hb-derived free iron could participate in the RBC damage using an in vitro ozone exposure system. Ozone exposure induced hemolysis, formation of methemoglobin, and RBC membrane lipid peroxidation. This RBC damage was not suppressed by the addition of a specific iron chelator (deferoxamine mesilate) to the medium but was suppressed by carbon monoxide (CO) treatment before ozone exposure. Generation of hydrogen peroxide (H2O2) in RBC was observed upon ozone exposure but was significantly suppressed by CO treatment before ozone exposure. Thus the Hb status (i.e., Hb redox condition) and H2O2 generation in RBC should play important roles in mediating RBC damage by ozone exposure. In other words, neither ozone nor its derivative directly attacked from the outside of the cell, but ozone that penetrated through the membrane derived the reactive oxygen species from Hb inside of the cell.     

Species- and age-dependent sensitivity to ozone in young plants of pea,wheat and spinach: Effects on acyl lipid and pigment content and metabolism

Acyl lipids and pigments were analyzed in young plants of garden pea, spring wheat and spinach exposed to < 5 or 65 nl l^?1 ozone 12 h per day for 6 days. In one set of experiments, the plants were exposed to ¹⁴CO₂ for 2 h 3 days prior to ozone exposure. The plants responded differently to the moderately enhanced level of ozone used Spinach was not at all sensitive while in both pea and wheat, leaves of different ages differed in ozone sensitivity. In pea, ozone sensitivity increased with leaf age. In the second and third oldest leaves, the amounts of galactolipids per leaf area and the proportions of 18:3 of the total lipid extract and of phosphatidylglycerol decreased. In the second oldest leaf, ozone also caused a decreased proportion of 18:3 of monogalactosyldiacylglycerol. In the fourth oldest leaf, lipid composition and galactolipid unsaturation was unaffected, but ozone caused decreased leaf expansion resulting in increased acyl lipid content per leaf area. In both the first and second leaves of wheat, ozone fumigation caused a marked decrease in the content of monogalactosyldiacylglycerol and in the first leaf, the contents of phosphatidylcholine and phosphatidylethanolamine increased. The proportion of 18:3 in phosphatidylcholine was larger in ozone-fumigated than in control plants, while the reverse applied for phosphatidylglycerol. In the oldest sampled leaves of pea and wheat, ozone caused an increase in the radioactivity associated with β-carotene, indicating increased turnover. Thus, while spinach was unaffected, in both pea and wheat ozone caused a decrease in the proportion of chloroplast membrane lipids to non-chloroplast membrane lipids in older leaves while younger leaves were less sensitive.     

臭氧浓度升高对毛竹叶片光合色素和抗性生理的影响

为了解毛竹对温室气体浓度升高的生理响应,探索竹子应对气候变化的经营策略,运用开顶式同化箱(OTCs)开展了5个O₃浓度梯度(过滤大气CF、环境大气NF、50nl·L^-1、100nl·L^-1、150nl·L^-1对毛竹(Phyllostachys edulis)叶片光合色素、可溶性蛋白、脂质过氧化和抗氧化系统的影响研究。结果表明:随O₃浓度升高,毛竹叶片Chl和Car含量呈下降趋势,可溶性蛋白、MDA和O₂^-含量呈升高趋势。与CF相比,100nl·L^-1和150nl·L^-1O₃,浓度处理毛竹叶片光合色素含量极显著降低,叶片可溶性蛋白、MDA和O₂^-含量极显著升高;随O₃浓度升高,SOD活性呈降低趋势,100nl·L^-1和150nl·L^-1O₃浓度处理较CF极显著下降。POD活性呈升高趋势,100nl·L^-1和150nl·L^-1O₃浓度处理较CF极显著提高;长时间高O₃浓度(100nl·L^-1及以上)胁迫条件下,会导致毛竹叶片光合色素降解或合成受阻,叶片老化加快,膜脂过氧化程度加剧,膜结构和抗氧化系统功能遭到破坏,影响毛竹的正常生长。     

Effects of moderately enhanced levels of ozone on the acyl lipid composition of leaves of garden pea (Pisum sativum)

Plants of garden pea ( Pisum sativum L.) were exposed to charcoal-filtered air with or without addition of 65 ± 5 l⁻¹ ozone. Plants were harvested daily for 9 days and lipids were extracted from the second-oldest leaf. Visible injury of this leaf was evident from day 5 on, while the differences in lipids between ozone and control treatments were observed earlier. Ozone caused large decreases in the contents of monogalactosyldiacylglycerol (MGDG), digalactosyldiacylglycerol (DGDG) and sulfoquinovosyldiacylglycerol (SQDG), a slower decrease in the content of phosphatidylcholine (PC), but an increase in the content of phosphatidylethanolamine (PE) per leaf area, compared with exposure to charcoal-filtered air. The content of phosphatidylglycerol (PG) was unaffected by ozone. Compared with charcoal-filtered air, fumigation with ozone resulted in a decrease in the proportion of linolenic acid (18:3) of the total lipid extract, with a concomitant increase in the proportion of linoleic acid (18:2). For individual lipids, ozone caused a similar pattern of decreased 18:3 and increased 18:2 in MGDG, SQDG, PC and PE, while the fatty acid composition of DGDG was unaffected. In PG, ozone decreased the proportions of 18:3 and trans -Δ³-decenoic acid (16:1_trans), balanced by increased proportions of palmitic and oleic acids. The contents of chlorophylls and carotenoids were unaffected by ozone. Our results show that moderately elevated levels of ozone cause significant changes in the polar lipid composition of garden pea leaves and in the level of unsaturation of the lipid acyl groups and, furthermore, that ozone has different effects, which could be direct or indirect, on chloroplast lipids (MGDG, DGDG, SQDG, PG acylated with 16:1_trans) and cytosolic membrane lipids.     

Inhibition by ozone of the acylation of glycerol 3-phosphate in mitochondria and microsomes from rat lung

The synthesis of lipids from [U-¹⁴C]glycerol 3-phosphate by mitochondrial or microsomal fractions from rat lung was inhibited by ozone. The susceptible reaction was the first acylation of glycerol 3-phosphate. Enzymes unaffected by the ozone exposure included: acyl-CoA thioesterase, acyl-CoA thiokinase, acyl-CoA:acylglycerol 3-phosphate acyltransferase, acyl-CoA:diacylglycerol acyltransferase, and acyl-CoA:acylglycerophosphocholine acyltransferase. The effect of ozone on lipid synthesis is closely comparable to the inhibition by N-ethylmaleimide suggesting that the effect of ozone is the oxidation of enzyme sulfhydryl groups. There was no indication of lipid oxidation caused by ozone and no indication of the production of a stable toxic compound.     

Effect of ozone exposure on prostacyclin synthesis in lung

The effect of ozone exposure on prostacyclin (PGI2) synthesis in the rat lung was studied. Male Wistar rats were exposed to 0.2, 0.4, 0.8, 1.2 and 1.8 ppm ozone for 24h. The higher concentration (1.8 ppm) significantly depressed-PGI2 synthesizing activity of lung homogenates. Time-courses (1, 3, 5, 7, 14 and 28 days) of the effect of ozone (0.4 and 0.8 ppm) exposure on the PGI2-synthesizing activity of lung homogenates were studied. The PGI2-synthesizing activity of the lung decreased, reaching a maximum at 5 days and then gradually returning to normal by day 14, and remaining normal at day 28, even though the ozone exposure continued. The formation of lipid peroxides due to ozone exposure may cause the depression of PGI2-synthesizing activity of lung. Induction of anti-oxidative enzymes may relate to the recovery of the PGI2-synthesizing activity.     

Effects of ozone on photosynthetic CO2 exchange, chlorophyll a fluorescence and antioxidant systems in lettuce leaves

The effects of ozone exposure on lettuce leaves were investigated by means of gas exchange, modulated chlorophyll a fluorescence and antioxidants systems. High ozone concentration decreased the rate of photosynthesis at light saturation level and changes in stomatal conductance and transpiration rate. However, an increase in intercellular CO₂ concentration indicated a decrease in the carboxylation efficiency. These data agreed with a reduction of non-cyclic electron flow and lower capacity to reduce the quinone pool. Ozone affected the ascorbate pool and decreased ascorbate peroxidase activity, increased lipid peroxidation, altered membrane properties and reduced the development of non-photochemical quenching.     

Ozone inhibits prostacyclin synthesis in pulmonary endothelium

The effects of ozone on lung arachidonate metabolism in-vitro were studied in cultured bovine pulmonary endothelial cells exposed for 2 hours to ozone in concentrations up to 1.0 ppm. A concentration-dependent decrease in prostacyclin synthesis was found (90% decrease at the highest ozone level of 1.0 ppm). The inhibition of prostacyclin synthesis was not due to a decreased release of arachidonic acid from membrane lipids. We also examined the hypoxic pulmonary vasoconstrictive response to 10% oxygen inhalation in anesthetized dogs in-vivo after exposure to 1.0 ppm ozone for 1 hour. Pulmonary vascular resistance was significantly increased after ozone exposure, similar to the findings in dogs given indomethacin (15 mg/kg). The percentage change in the hypoxic pulmonary pressor response was similar between the ozone exposure and indomethacin-treated groups, although due to the variance of the pulmonary vascular resistance values during hypoxia the results did not reach statistical significance. These results suggest that ozone inhalation affects pulmonary endothelial arachidonate metabolism in-vivo as well as in-vitro.     

Response of photosynthesis and cellular antioxidants to ozone in populus leaves

Atmospheric ozone causes formation of various highly reactive intermediates (e.g. peroxyl and superoxide radicals, H₂O₂, etc.) in plant tissues. A plant's productivity in environments with ozone may be related to its ability to scavenge the free radicals formed. The effects of ozone on photosynthesis and some free radical scavengers were measured in the fifth emergent leaf of poplars. Clonal poplars (Populus deltoides × Populus cv caudina) were fumigated with 180 parts per billion ozone for 3 hours. Photosynthesis was measured before, during, and after fumigation. During the first 90 minutes of ozone exposure, photosynthetic rates were unaffected but glutathione levels and superoxide dismutase activity increased. After 90 minutes of ozone exposure, photosynthetic rates began to decline while glutathione and superoxide dismutase continued to increase. Total glutathione (reduced plus oxidized) increased in fumigated leaves throughout the exposure period. The ratio of GSH/GSSG also decreased from 12.8 to 1.2 in ozone exposed trees. Superoxide dismutase levels increased twofold in fumigated plants. After 4 hours of ozone exposure, the photosynthetic rate was approximately half that of controls while glutathione levels and superoxide dismutase activity remained above that of the controls. The elevated antioxidant levels were maintained 21 hours after ozone exposure while photosynthetic rates recovered to about 75% of that of controls. Electron transport and NADPH levels remained unaffected by the treatment. Hence, elevated antioxidant metabolism may protect the photosynthetic apparatus during exposure to ozone.     

Induction by ozone of ethylene production and an ACC oxidase cDNA in rice (Oryza sativa L.) leaves

Exposure to ozone at 1 µl l^–1 for 6 h induced ethylene production in rice (Oryza sativa L. cv. Hitomebore) leaves. The stimulation of ethylene production was detectable 2 h after the start of the exposure to ozone, and lasted for 6 h after the exposure. A 429-bp cDNA fragment encoding ACC oxidase was obtained by RT-PCR from ozone-treated rice leaves. Its nucleotide sequence and deduced amino-acid sequence had 97.2% and 94.4% identity, respectively, to those of OS1A1COX, which was previously obtained from deepwater rice. The abundance of the cDNA increased in accordance with the induction of ethylene production by the exposure to ozone.     

毛竹对大气臭氧浓度倍增的生理响应

为了给气候变化背景下的竹林培育应对策略提供理论依据,以毛竹 （Phyllostachys edulis） 为材料,运用开顶式气室 （OTCs） 模拟当前环境背景大气O3浓度（NF,40~45nL·L-1）的倍增1倍（TR1,92~106nL·L-1）和倍增2倍（TR2, 142~160nL·L-1）情景,分析叶片光合色素、脂质过氧化及抗氧化酶等生理指标的变化规律。结果表明：大气O3浓度倍增处理90d时,毛竹叶片Chl、Car含量和SOD活性显著降低,叶片相对电导率和可溶性蛋白、MDA、O2-含量及POD活性显著升高。叶片O2-含量与SOD活性、光合色素含量呈极显著负相关,而与叶片相对电导率、POD活性和MDA、可溶性蛋白含量呈显著正相关。研究表明大气O3浓度倍增会导致毛竹叶片老化加快,光合色素降解或合成受阻,膜脂过氧化,膜结构和氧化系统破坏,影响毛竹的正常生长。本文从抗性生理方面揭示了毛竹对大气臭氧胁迫的耐受性,为竹子应对气候变化研究提供了参考。     

The Physiological Response of Phyllostachys edulis under the Elevated Atmospheric Ozone

In order to provide theoretical evidences for bamboo cultivation under the background of climate change, the method of open top chambers (OTCs) was employed to approach the effects of elevated ozone on photosynthetic pigments content, lipid peroxidation and anti oxidation enzymes in the leaves of Phyllostachys edulis. The result indicated that with the increasing of ozone concentration, the content of the chlorophyll, carotenoid and SOD activity decreased, while the content of souble protein, relative electrical conductivity, contents of MDA, O2- and the activity of POD increased.Correlation analyses showed that there was significant negative correlation between O2- content and SOD activity as well as photosynthetic pigment,whereas significant positive correlation between O2- and relative electeical conductivity, MDA, POD as well as soluble protein. It was summarried that, on one hand, long time exposure to high ozone concentration exposure, accelerated the aging of leaves, intensified the damage to membrane lipid, and destroyed the structure of cell membrane and the function of anti oxidation system as well. It affected the regular growth of P.edulis. This research was conducted to provide theoretical evidence for bamboo cultivation during this time of possible climate change.     

Decreased levels of ascorbic acid in lung following exposure to ozone

Mice were exposed to concentrations of 20, 40 and 200 ppm ozone in air for 30 min. Ozone exposure decreased lung ascorbic acid levels and increased lung weight by up to 50% in a dose related manner. On incubation in Krebsphosphate solution, lung slices from mice exposed to 200 ppm ozone released a smaller fraction of their content of ascorbic acid into the medium than did lung slices from control mice, suggesting that there was a preferential loss of extracellular ascorbic acid during ozone exposure. These results are consistent with the proposed function of ascorbic acid as an extracellular antioxidant in lungs.     

Pre-activating wounding response in tobacco prior to high-level ozone exposure prevents necrotic injury

In tobacco, both wounding and treatment with jasmonates prior to exposure of the tissue to high concentrations of ozone (250 to 500 p.p.b.) produce a dramatic decrease in ozone injury. A systemic pattern of increased ozone tolerance developed within 3–6 h after wounding and also after local application of jasmonates. Ozone treatment of transgenie (NahG) tobacco plants showed that the inability of these plants to accumulate salicylic acid is also accompanied by increased ozone tolerance. Expression of mRNA encoding the anti-oxidant enzyme ascorbate peroxidase is upregulated by ozone challenge, wounding and by methyl jasmonate exposure within 3–4 h, while levels of carbonic anhydrase mRNA are simultaneously depressed following ozone exposure and methyl jasmonate treatment. The pattern of these results shows that the response to ozone challenge in tobacco involves signalling mechanisms similar to those induced in plants by other environmental stresses that generate reactive oxygen species.     

臭氧急性暴露对大鼠血管的损伤及其机制

目的:探索不同浓度臭氧(O3)急性暴露对雄性Wistar大鼠血管的损伤效应和可能的机制。方法:120只雄性Wistar大鼠随机分为6组,每组20只;实验动物置于气体染毒柜中,对照组暴露于过滤后空气,处理组分别暴露于浓度为0.12ppm,0.5ppm,1.0ppm,2.0ppm和4.0ppm的臭氧,持续暴露4h。利用PC-lab医学生理信号采集系统获得动脉血压数据;血流变指标和血生化指标由天津迪安诊断实验室检测;血清中内皮素(ET-1)、同型半胱氨酸(HCY)、血管性血友病因子(vWF)、8-羟基脱氧鸟苷(8-OhdG)、白介素(IL-6)和肿瘤坏死因子α(TNF-α)采用酶联免疫(ELISA)微孔板法检测;氧化应激指标超氧化物歧化酶(SOD)活力和丙二醛(MDA)分别采用黄嘌呤氧化酶法、硫代巴比妥酸(TBA)法测定,还原型谷胱甘肽(GSH)和一氧化氮(NO)采用微孔板比色法;取胸主动脉组织制备石蜡切片,经HE染色后观察血管结构改变。结果:0.12ppm臭氧急性暴露可导致动脉收缩血压(SBP)显著升高;不同浓度臭氧暴露均可导致血浆粘度显著升高,1.0ppm臭氧暴露组血沉(ESR)方程K值显著升高,全血高切相对指数和还原粘度均在臭氧浓度为0.5ppm和4.0ppm时显著降低,而红细胞变形指数在臭氧浓度为0.12ppm、0.5ppm、1.0ppm和2.0ppm时显著升高;急性臭氧暴露可导致总胆固醇含量降低,高密度脂蛋白胆固醇(HDL-C)在0.12ppm臭氧暴露组显著降低;当臭氧浓度高于1.0ppm时还可导致机体出现炎症反应(TNF-α升高)和氧化应激反应(MDA升高、GSH降低);臭氧急性暴露可导致血液中ET-1含量升高,在4.0ppm浓度组具有显著性差异,而HCY水平呈现先降低后升高的趋势,在1.0ppm浓度组达到最高值,胸主动脉未见明显的病理改变。结论:臭氧急性暴露可影响大鼠的动脉血压、血流变及胆固醇代谢,可能的机制是臭氧暴露导致炎症反应和氧化应激反应,引起血管内皮功能损伤,并且随着臭氧暴露浓度升高血管内皮细胞功能损伤越显著。     

急性臭氧暴露对大鼠肺部细胞遗传毒性的影响*

目的: 探讨不同浓度臭氧急性暴露对大鼠肺部细胞的遗传毒性的影响。方法: 36只wistar大鼠随机分为对照组(过滤空气暴露)、臭氧暴露组(0.12 ppm、0.5 ppm、1.0 ppm、2.0 ppm、4.0 ppm)共6组,每组6只。以不同浓度的臭氧对大鼠进行动态染毒4 h后,取肺组织并分离单细胞,采用酶联免疫吸附法检测8-羟基脱氧鸟苷(8-OHdG),利用彗星实验、微核试验和DNA-蛋白质交联实验进行DNA和染色体损伤分析。结果: 与对照组相比,肺组织中8-OHdG含量从臭氧暴露浓度为0.12 ppm起即显著增加,在0.5 ppm时达到最高值。随着臭氧暴露浓度升高,彗星拖尾率逐渐上升,且存在明显的剂量-效应关系;DNA-蛋白质交联率有先升高后下降的趋势,且在2.0 ppm时达到最大值;而肺部细胞微核率尽管呈现出上升趋势,但与对照组相比无显著性差异。结论: 急性臭氧暴露在较低浓度(0.12 ppm)时即可导致大鼠肺部细胞的DNA损伤;而在较高浓度(4 ppm)时却未见显著的染色体损伤。