Alterations in sugar residues in squamous metaplasia in hamster tracheal explants induced by benzo(a)pyrene and its reversal by retinoic ACID

Summary We have demonstrated that squamous metaplasia induced by benzo(a)pyrene (BP) in the hamster tracheal explants accompany distinct alterations in carbohydrate moieties in the epithelial mucosa. Most prominent alterations were the preferential binding of peanut agglutinin (PNA) and wheat germ agglutinin (WGA) in the basal cell layer in metaplastic lesions. In this study we examined if reversal of BP-induced lesions by all-trans retinoic acid (RA) results in the acquisition of normal carbohydrate composition by the tissue. Four lectins, PNA, WGA, Dolichos biflorus agglutinin, and Concanavalin A, in their horseradish peroxidase conjugates were used. In control explants the intercellular plasma membrane of basal and mucous cells exhibited no significant reaction with any of the lectins tested. In the metaplastic lesions induced by BP, PNA and WGA intensely stained the plasma membrane and intercellular spaces of basal and intermediate cell layers; the granular layer cells did not bind PNA whereas they were stained moderately with WGA. RA, which reversed the metaplasia, also conferred the tissue with lectin binding patterns similar to that of control explants. These results thus show that the reversal of metaplasia is accompanied by acquisition of the tissue’s original carbohydrate composition. This research was supported by grant RO1-HL32308 from the National Heart, Lung and Blood Institute, Bethesda, MD.     

乳杆菌吸附苯并芘的特性

[目的]探讨植物乳杆菌(Lactobacillus plantarum)121和戊糖乳杆菌(Lactobacillus pentosus)ML32的苯并芘吸附作用与机制.[方法]采用高效液相色谱检测菌体对苯并芘的吸附率.[结果]菌株121和ML32对苯并芘的吸附率分别为65.9%和64.9%,这种吸附特性与菌体活力无关,随培养时间延长、温度提高以及细胞浓度的上升而增加.菌株121和ML32的吸附率在pH 4和5时达到最大,分别为87.6%和89.0%.当培养液中Ca2+或Mg2+浓度大于0.05mol/L时,菌体吸附率与盐离子浓度呈正相关.苯洗脱会导致乳杆菌所吸附的苯并芘减少90%.经碱性蛋白酶、中性蛋白酶、溶菌酶及TCA和SDS等方法处理后,菌体吸附率上升,且不易被苯去除.在胆盐及胃酸环境下,两株菌的吸附率均提高至70%以上,而胰蛋白酶的存在仅对菌株121的吸附率有较大影响.[结论]两株乳杆菌可以通过吸附作用从环境中清除苯并芘,其吸附效果与细菌细胞壁的结构和组成有关.     

Biodegradation of benzo(a)pyrene by a newly isolated Fusarium sp

Benzo(a)pyrene (BaP) is a five-ring polycyclic aromatic hydrocarbon produced by the incomplete combustion of organic materials. It is one of the priority pollutants listed by the US Environmental Protection Agency. This study describes a fungal isolate that is able to biodegrade benzo(a)pyrene. The filamentous fungus, isolated from leaves of Pterocarpus macrocarpus Kurz., was identified as a Fusarium sp. (strain E033). Fusarium sp. E033 was able to survive in the presence of benzo(a)pyrene concentrations up to 1.2 mM (300 mg L(-1)). Biodegradation experiments using 0.4 mM (100 mg L(-1)) benzo(a)pyrene demonstrated that Fusarium sp. E033 was able to degrade 65-70% of the initial benzo(a)pyrene provided, and two transformation products, a dihydroxy dihydro-benzo(a)pyrene and a benzo(a)pyrene-quinone, were detected within 30 days of incubation at 32 degrees C. The factors affecting biodegradation efficiency were also investigated. While increasing aeration promoted better fungal growth and benzo(a)pyrene biodegradation, increasing the glucose concentration from 5 to 50 mM had an adverse effect on biodegradation. Ethanol and methanol, provided at 5 mM to increase benzo(a)pyrene water solubility, increased the fungal biomass yield but did not promote degradation. The Fusarium sp. E033 isolated in this study can tolerate and degrade relatively high concentrations of benzo(a)pyrene, suggesting its potential application in benzo(a)pyrene bioremediation.     

Some properties of vicinal diol-epoxides derived from benzo(a)anthracene and benzo(a)pyrene

The alkylating properties of pairs of syn- and anti-isomers of 2 diol-epoxides derived from benzo(a)pyrene (BP) and of 1 derived from benz(a)anthracene (BA) have been investigated. Of the anti-diol-epoxides, anti-BP 7,8-diol-9,10-oxide was the most reactive compound towards DNA, towards sodium p-nitrothiophenolate in a non-aqueous solvent system, and towards 4-(p-nitrobenzyl)pyridine in aqueous solution; anti-BP 9,10,-diol-7,8-oxide was of intermediate reactivity and anti-BA 8,9-diol-10,11-oxide was least reactive. The syn-diol-epoxides gave unsatisfactory results with DNA and 4-(p-nitrobenzyl)pyridine because of their rapid solvolysis in aqueous solution, but with sodium p-nitrothiophenolate showed the order of reactivity syn-BP 7,8-diol-9,10-oxide greater than syn-BA 8,9-diol-10,11-oxide greater than syn-BP 9,10-diol-7,8-oxide. The products of the reaction between diol-epoxides and nucleic acids were examined by Sephadex LH-20 chromatography followed by high-pressure liquid chromatography (HPLC) and the diol-epoxides were shown to react principally with the guanosine and adenosine moieties of RNA.     

Protective effect of vanillic acid against benzo(a)pyrene induced lung cancer in Swiss albino mice

Vanillic acid (VA) is found in high concentrations in various plants and used as traditional medicine for various diseases. The aim of the existing study is to illustrate the protective effects of VA against benzo(a)pyrene (B(a)P)‐induced lung cancer in Swiss albino mice. B(a)P (50 mg/kg b.wt.) was given orally to induce lung cancer in mice. The body weight, tumor incidence, carcinoembryonic antigen (CEA), neuron‐specific enolase (NSE), and enzymatic/nonenzymatic antioxidants (superoxide dismutase, catalase, glutathione reductase, glutathione peroxidase, and glutathione) were estimated. Further histochemical investigation through hematoxylin and eosin staining was also carried out. B(a)P administered groups showed increased levels of serum pathological markers CEA, NSE along with reduced final body weight as well as decreased tissue enzymatic and nonenzymatic antioxidants activities, whereas VA treatment (200mg/kg/b.wt) along with B(a)P showed significantly reverted the above changes, which proves as prominent anticancer effects in experimentally induced lung cancer. Overall, these results suggest that VA has an efficient preventive action against B(a)P‐induced lung cancer, and this is attributed to its free‐radical scavenging antioxidant activities.     

乳杆菌在模拟肉制品条件下结合苯并芘的效果

【背景】乳杆菌对众多致癌物具有吸附作用,但关于乳杆菌结合吸附苯并芘特性的研究并不多。【目的】探讨戊糖乳杆菌(Lactobacillus pentosus) ML32和植物乳杆菌(Lactobacillus plantarum)121对加工肉制品中苯并芘的吸附能力与吸附机制。【方法】基于HPLC检测菌体对不同模拟加工处理方式肉品中的苯并芘的吸附率。【结果】植物乳杆菌121和戊糖乳杆菌ML32对模拟油炸、烟熏或烧烤方式处理肉中苯并芘的吸附率均在30%以上。菌株121对直接烟熏肉中的苯并芘吸附率为41.21%,直接油炸肉中吸附率为38.71%,直接烧烤肉中吸附率为37.51%;菌株ML32对间接烟熏肉中的苯并芘吸附率为40.02%,间接烧烤肉中吸附率为38.01%。植物乳杆菌121适合于去除高温长时间加工肉中的苯并芘,戊糖乳杆菌ML32则相反。另外,乳杆菌细胞壁中的肽聚糖或许在吸附过程中发挥了主要作用。【结论】两株乳杆菌121和ML32具有吸附某些加工肉制品中苯并芘的效果,或许可以作为一种方法用于消除某些肉制品中因苯并芘过量带来的风险。     

Analysis of fish bile with HPLC — fluorescence to determine environmental exposure to benzo(a)pyrene

Brown bullhead from the Black River, Ohio, have a high incidence of liver neoplasia which is associated with elevated concentrations of polynuclear aromatic hydrocarbons (PAHs) in the sediment. We evaluated the use of biliary concentrations of benzo(a)pyrene [B(a)P] equivalents as a means for determining PAH exposure. Bile was collected from 16 brown bullheads and 8 common carp taken from each of two Lake Erie tributaries in Ohio, the industrialized Black River and the non-industrialized Old Woman Creek. Hatchery bullhead (n = 8) were used to determine base levels of PAHs. A high performance liquid chromatography (HPLC) — fluorescence technique was used to determine the concentration of B(a)P equivalents in the bile samples. The area of all peaks fluorescing at 380/430 nm was summed to give a single value for B(a)P equivalents in each sample. Concentrations of B(a)P equivalents generally reflected concentrations of PAH in sediment where fish were collected. Bile taken from Black River carp contained the highest concentration of B(a)P equivalents and was significantly different from all other groups. The value obtained for Black River bullhead was also high and was found to be significantly different from hatchery bullhead. B(a)P equivalents varied between carp and bullhead from the same habitat possibly because of differing food habits or metabolic pathways. However, our results indicate that relative levels of B(a)P equivalents in the bile of fish correspond well to B(a)P levels in sediment and may offer a means of determining environmental exposure of fish to the parent compound.     

Enhanced phytoremediation of cadmium and/or benzo(a)pyrene contaminated soil by hyperaccumlator Solanum nigrum L.

The role of same amendment on phytoremediating different level contaminated soils is seldom known. Soil pot culture experiment was used to compare the strengthening roles of cysteine (CY), EDTA, salicylic acid (Sa), and Tween 80 (TW) on hyperaccumulator Solanum nigrum L. phytoremediating higher level of single cadmium (Cd) or Benzo(a)pyrene (BAP) and their co-contaminated soils. Results showed that the Cd capacities (ug pot^?1) in shoots of S. nigrum in the combined treatment T_{0.1EDTA+0.9CY} were the highest for the 5 and 15 mg kg^?1 Cd contaminated soils. When S. nigrum remediating co-contaminated soils with higher levels of Cd and BAP, that is, 5 mg kg^?1 Cd + 1 mg kg^?1 BAP and 15 mg kg^?1 Cd + 2 mg kg^?1 BAP, the treatment T_{0.9CY+0.9Sa+0.3TW} showed the best enhancing remediation role. This results were different with co-contaminated soil with 0.771 mg kg^?1 Cd + 0.024 mg kg^?1 BAP. These results may tell us that the combine used of CY, SA, and TW were more useful for the contaminated soils with higher level of Cd and/or BAP. In the combined treatments of Sa+TW, CY was better than EDTA.     

Pyrene and benzo(a)pyrene Metabolism by an Aspergillus Terreus Strain Isolated from a Polycylic Aromatic Hydrocarbons Polluted Soil

A strain of Aspergillus terreus was isolated from a polycyclic aromatic hydrocarbons (PAHs) polluted soil. The metabolism of pyrene and benzo(a)pyrene by this fungus was investigated in liquid submerged culture added of 50 and 25 ppm respectively of each compound. Depletion of pyrene and Benzo(a)pyrene was evident during the first stages of growth and was 60% and 27.5% respectively of the added amount after nine days of culture. Solvent extracts of the fermentation broth and mycelium were analysed for presence of metabolites by HPLC-MS technique. Under the present cultural conditions pyrene was mainly metabolised to pyrenylsulfate similarly to benzo(a)pyrene that led to benzo(a)pyrenylsulfate. The structure of 1-pyrenilsulfate was determined after purification of extracts and H-NMR analysis. The result show that the isolated A. terreus strain metabolises PAHs by reaction similar to those previously reported for non lignolinolytic fungi with a mechanism that suggests the hydroxylation by a cytochrome P-450 monooxygenase followed by conjugation with sulfate ion.     

Differences in the repair of DNA strand breaks induced by 9-hydroxy-benzo(a)pyrene and trans-7,8-dihydro-7,8-dihydroxy-benzo(a)pyrene in cultured human fibroblasts

Two benzo(a)pyrene metabolites were found to induce DNA strand breaks in cultured human fibroblasts. DNA strand breaks induced by the non- or weakly carcinogenic 9-hydroxy-benzo(a)pyrene were repaired within two hours, while those induced by the strongly carcinogenic trans-7,8-dihydro-7,8-dihydroxy-benzo(a)pyrene were repaired at a much slower rate.     

Formation of the 11,12-diol as a metabolite of benzo(a)pyrene by rat skin in vivo

The dihydrodiols present as metabolites in rat skin after topical application of ³H-labelled benzo(a)pyrene included a significant amount of radioactivity that cochromatographed with synthetic $\text{trans}$-11,12-dihydro-11,12-dihydroxybenzo(a)pyrene. Treatment of the radioactive metabolite with hot mineral acid gave a product that had chromatographic properties identical to those of the phenol similarly formed from the synthetic dihydrodiol and acetylation of the metabolite yielded a product that cochromatographed with the diacetate of the synthetic dihydrodiol. These observations show that the 11,12-dihydrodiol is formed as a metabolite of BP in rat skin $\text{in vivo}$. The metabolite was not detected in mouse skin.     

Role of poly(ADP-ribose) glycohydrolase silencing in DNA hypomethylation induced by benzo(a)pyrene

Benzo(a)pyrene (BaP) is a known carcinogen cytotoxic which can trigger extensive cellular responses. Many evidences suggest that inhibitors of poly(ADP-ribose) glycohydrolase (PARG) are potent anticancer drug candidates. However, the role of PARG in BaP carcinogenesis is less understood. Here we used PARG-deficient human bronchial epithelial cell line (shPARG cell) as an in vitro model, and investigated the role of PARG silencing in DNA methylation pattern changed by BaP. Our study shows, BaP treatment decreased global DNA methylation levels in 16HBE cells in a dose-dependent manner, but no dramatic changes were observed in shPARG cells. Further investigation revealed PARG silencing protected DNA methyltransferases (DNMTs) activity from change by BaP exposure. Interestingly, Dnmt1 is PARylated in PARG-null cells after BaP exposure. The results show a role for PARG silencing in DNA hypomethylation induced by BaP that may provide new clue for cancer therapy.     

Anticancer effect of Limonin against benzo(a)pyrene‐induced lung carcinogenesis in Swiss albino mice and the inhibition of A549 cell proliferation through apoptotic pathway

The main purpose of the current study is to reveal the anticancer action of limonin against benzo(a)pyrene [B(a)P]‐treated lung carcinogenesis in Swiss albino mice and A549 lung cancer cells. B(a)P was orally supplemented (50 mg/kg body weight) twice a week for four weeks induction of lung cancer in mice. The lung weight, body weight, incidence of tumor, lipid peroxidation, carcinoembryonic antigen (CEA), enzymatic and nonenzymatic antioxidants (superoxide dismutase, GPx, glutathione, glutathione reductase, catalase, and glutathione S‐transferase), serum marker enzymes (aryl hydroxylase, lactate dehydrogenase, 5′‐nucleotidases, and γ‐glutamyl transpeptidase), and inflammatory mediators (interleukin‐1β, interleukin‐6, and tumor necrosis factor‐α) were estimated. Moreover, a histopathological study of lung tissues was supported by the biochemical analysis. Furthermore, the anticancer activity of limonin on A549 cells was measured by cell viability, production of reactive oxygen species (ROS), apoptotic morphological changes by AO/EtBr staining. Additionally, the status of apoptosis protein (caspase‐9 and ‐3) expressions was analyzed by the colorimetric analysis. B(a)P‐induced mice showed increased lipid peroxidation, CEA, serum marker enzymes and inflammatory cytokines levels with simultaneously decreased in the nonenzymatic and enzymatic antioxidants levels. Limonin supplements significantly reverted back to all these changes in this manner, showing the efficiency of anticancer effect. Furthermore, our in vitro study also supported the anticancer effect of the treatment of limonin‐enhanced apoptosis by loss of cell viability, improved ROS production, apoptotic morphological changes, and apoptosis protein expression were analyzed. Overall, these results suggest the anticancer potential of limonin against B(a)P‐induced lung cancer in Swiss albino mice and A549 lung cancer cells.     

苯并(a)芘对鲫鱼肝脏EROD活性的影响

研究了典型多环芳烃类有机污染物苯并(a)芘(BaP)对鲫鱼(Carassius auratus)肝脏7-乙氧基-3-异吩唑酮-脱乙基酶(EROD)活性的影响。结果表明,注射后96 h,10和100 mg.kg-1处理组肝脏EROD活性被明显诱导,分别为对照组的2.3(P<0.05)和3.1倍(P<0.01)。肝脏EROD活性随着时间延长继续升高,至注射后14 d,1 mg.kg-1处理组鲫鱼肝脏EROD活性为对照的3.0倍(P<0.001),而100 mg.kg-1处理组则高达5.8倍(P<0.001)。鲫鱼肝脏EROD活性可作为反映BaP暴露水平的生物标志物。BaP对鲫鱼的最低效应浓度为1 mg.kg-1(鱼体重)。     

模拟淀粉条件下乳杆菌吸附苯并芘的能力

【目的】研究了模拟淀粉条件下嗜酸乳杆菌(Lactobacillus acidophilus)NCFM、植物乳杆菌(Lactobacillus plantarum)121以及戊糖乳酸菌(Lactobacillus pentosus)ML32吸附苯并芘的能力,为利用乳杆菌去除苯并芘提供一定的理论指导。【方法】基于苯并芘的HPLC检测方法,考察了淀粉含量及类型、培养时间和p H等因素对乳杆菌吸附苯并芘能力的影响,研究了淀粉水解产物及菌体活性影响乳杆菌吸附苯并芘的效果。【结果】淀粉含量在2%–10%的范围内,乳杆菌吸附苯并芘的能力与淀粉含量的增加呈正相关性,且与淀粉种类关系不大,但经糊化处理的淀粉可以促进菌体吸附苯并芘。在模拟淀粉体系中,培养前4 h时乳杆菌吸附苯并芘的效率增长快,此后其吸附率增加缓慢。淀粉经酸性(p H为3–4)和碱性(p H为8–9)处理,乳杆菌吸附苯并芘的能力提升。淀粉的水解产物麦芽糖和葡萄糖都能显著改善乳杆菌吸附苯并芘的能力。与活细胞相比,经灭活处理后乳杆菌细胞吸附苯并芘的能力降低。【结论】在淀粉体系中,乳杆菌依然表现出良好的苯并芘吸附能力,且一定范围内淀粉含量增多、糊化作用以及麦芽糖和葡萄糖的存在可促进其吸附苯并芘的能力。因此,本研究中的乳杆菌或许可以用作生物脱除剂来减少淀粉食物中的苯并芘。     

Modulation of aortic protein phosphorylation by benzo(a)pyrene: Implications in PAH-induced atherogenesis

The toxicity of polycyclic aromatic hydrocarbons such as benzo(a)pyrene, 7,12-dimethylbenz(a)anthracene, and 3-methylcholanthrene has been associated with alterations in the proliferation of vascular smooth muscle cells and the development of lesions of mesenchymal origin. Because phosphorylation of endogenous substrates plays a central role in the regulation of smooth muscle cell growth, the present studies were conducted to evaluate the phosphorylation pattern of medial aortic protein upon repeated in vivo exposure of Japanese quail to benzo(a)pyrene (BaP). Medial aortic homogenates from quail treated for 10 weeks with 10 mg/kg benzo(a)pyrene or vehicle were processed for in vitro measurements of protein phosphorylation. In vitro phosphorylation of endogenous or exogenous proteins stimulated in vitro by phorbol myristate acetate/phosphatidyl-serine or cyclic AMP, known activators of protein kinase C and cyclic AMP-dependent protein kinase, respectively, was examined in the cytosolic and particulate fractions of homogenates from control and treated animals. Benzo(a)pyrene treatment significantly enhanced the basal phosphorylation of Mr 113, 35, and 23 kDa proteins in the cytosolic fraction. Modest increases in the phosphorylation of Mr 71, 52, and 38 kDa were also observed under basal conditions. No changes in the basal phosphorylation of particulate proteins were observed. Phosphorylation of endogenous protein substrates by protein kinase C in the cytosolic fraction was not altered by benzo(a)pyrene treatment. In contrast, inhibition of C-kinase-mediated phosphorylation of endogenous Mr 272, 72, and 45 kDa proteins was observed in the particulate fraction of aortic homogenates from benzo(a)pyrene-treated quail relative to controls. Exogenous histone phosphorylation by PKC in the particulate, but not cytosolic fraction, was decreased by benzo(a)pyrene treatment. The effects of benzo(a)pyrene on the C-kinase system were specific, since cAMP-mediated phosphorylation of endogenous proteins, as well as exogenous histone, was not altered by benzo(a)pyrene. Interestingly, benzo(a)pyrene treatment was associated with a selective increase of Mr 200, 80, and 67 kDa proteins in the cytosolic fraction. Collectively, these data are consistent with the hypothesis that medial protein phosphorylation is a significant molecular target of benzo(a)pyrene within the vascular wall.     

用EROD研究苯并芘和六氯苯的联合毒性作用

用7-乙氧基异叻唑酮-脱乙基酶(EROD)检测的方法,研究了苯并芘和六氯苯对日本青鳉肝脏EROD酶的比活力的影响。结果表明,苯并芘和六氯苯对EROD酶的比活力均有激活作用,在实验浓度范围内,EROD酶的比活力与两者浓度之间存在剂量-效应关系。苯并芘和六氯苯表现为一定的协同作用。实验同时发现日本青鳉在六氯苯和苯并芘中暴露后,EROD酶的比活力开始有一个短暂的降低,然后持续升高。对六氯苯和苯并芘暴露的最佳时间进行了探讨。     

Metabolism of Benzo(a)pyrene by Human Epithelial and Fibroblastic Cells: Metabolite Patterns and DNA Adduct Formation

We demonstrate in cell culture that mammary epithelial cells from normal human breast specimens metabolize benzo(a)pyrene (BaP) and form adducts with the bases of their DNA more readily and at lower concentrations of BaP than do fibroblasts from the same specimens. BaP metabolism and adduct formation was determined in the same incubations with epithelial cells grown out in early passage from each of three specimens and with fibroblasts from one of these specimens. The metabolite pattern of the epithelial cells was indicative of preferential formation of 7, 8-dihydrodiol-9, 10-dihydroepoxybenzo(a)pyrene the ultimate carcinogen. In contrast, fibroblasts formed mainly mono- and dihydroxide derivatives of BaP. The metabolite pattern from epithelial cells was compatible with the ease in which adducts between DNA and the diolepoxide of benzo(a)pyrene were formed. These results provide evidence that chemical carcinogens should be considered as possible factors in the induction of breast cancer in women.     

The carcinogen benzo(e)pyrene is metabolized by DM15 cells without an uncoupling effect on their gap junctions

Benzo(e)pyrene (B(e)P) promotes carcinogenesis in the skin. Unlike some other promoters however, B(e)P does notproduce an uncoupling effect on gap junction permeability in DM15 transformedfibroblasts. This study demonstrates thatDM15 cells exhibit a relatively high level of B(e)P metabolism. Moreover, although pretreatment of DM15 cells with benz(a)anthracene results in an 8-fold increase of arylhydrocarbon hydroxylase activity and a 2-fold increase in the rate ofB(e)P metabolism, it did not enable B(e)P to affectLucifer Yellow transfer between DM15 cells. We conclude that neitherB(e)P nor its metabolites are capable of uncoupling gap junction permeability in DM15 cells.Abbreviations AHH aryl hydrocarbon hyroxylase - BA benz(a)anthracene - B(a)P benzo(a)pyrene - B(e)P benzo(e)pyrene - LY Lucifer Yellow - MFO mixed-function oxidases - PAH polycyclic aromatic hydrocarbons     

熏烤肉制品中苯并芘的危害及控制措施

本文从食品安全的角度出发,对熏烤肉制品中的有害物质苯并(a)芘的产生,对人体的危害作用,以及控制熏烤肉制品中苯并(a)芘残留的方法进行了初步的研究.这对提高熏烤肉制品品质和对苯并(a)芘进行深入的研究具有重要意义.