Oxidative DNA damage and defence gene expression in the mouse lung after short-term exposure to diesel exhaust particles by inhalation

Exposure to diesel exhaust particles (DEP) is suspected to contribute to lung cancer and cardiopulmonary diseases. In recent years generation of reactive oxygen species capable of inducing cellular oxidative stress has been in focus as one of the underlying mechanisms behind the genotoxic effects of particles. However, the role of the antioxidative defence system still needs to be clarified, especially in relation to low-dose DEP exposures. The aim of this study was to characterize the effects of short-term exposure to DEP in terms of DNA damage and expression of key response genes towards oxidative stress in lungs of mice. Mice were exposed by inhalation to 20 or 80 mg/m3 DEP inhaled as either a single dose, or four lower doses (5 and 20 mg/m3) inhaled on four consecutive days. Our results indicate that HO-1 mRNA expression in lung tissue was up-regulated after both types of DEP exposures, whereas OGG1 expression was only up-regulated after repeated exposures. The level of oxidative DNA damage in terms of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) was increased in the lung tissue after a single exposure, whereas increased levels of DNA strand breaks was observed in bronchoalveolar lavage cells after repeated DEP exposures. The levels of 8-oxodG and OGG1 mRNA in lung tissue were mirror images. This suggests that after repeated exposures, up-regulation of DNA repair counteracts an increased rate of 8-oxodG formation leaving the steady state level of 8-oxodG in DNA unchanged. In conclusion, this study indicates that a single high dose of DEP generates 8-oxodG in lung tissue, whereas the same dose inhaled as four low-exposures may up-regulate the antioxidative defence system and protect against generation of 8-oxodG.     

Increased 8-Hydroxyguanine in DNA and Its Repair Activity in Hamster and Rat Lung after Intratracheal Instillation of Crocidolite Asbestos

Asbestos and man-made-mineral fibers are known to increase one type of oxidative DNA damage, 8-hydroxyguanine (8-OH-Gua), in vitro. In this study, we analyzed the 8-OH-Gua level in DNA and its repair activity after a single intratracheal instillation of fibers (crocidolite or glass) or saline to Syrian hamsters or Wistar rats. The 8-OH-Gua level was measured with a high-performance liquid chromatography-electrochemical detector (HPLC-ECD) system. The 8-OH-Gua repair enzyme activity was determined with an endonuclease nicking assay using a ³²P-labeled or fluorescently labeled 22mer DNA that contains 8-OH-Gua at a specific position. A significant increase in the 8-OH-Gua level in the lung DNA was observed 1 day after the exposure to crocidolite, as compared to the saline control. The repair activity was increased significantly at 7 days. On the other hand, after exposure to glass fibers, little or no increase of these carcinogenicity indicators was detected. These assays of 8-OH-Gua and its repair activity in short-term animal experiments will be useful for evaluating the carcinogenicity of fibers. This is the first report of the increase of 8-OH-Gua and its repair activity in the animal lung after the instillation of asbestos fibers.     

Increased 8-hydroxyguanine in DNA and its repair activity in hamster and rat lung after intratracheal instillation of crocidolite asbestos.

Asbestos and man-made-mineral fibers are known to increase one type of oxidative DNA damage, 8-hydroxyguanine (8-OH-(Gua), in vitro. In this study, we analyzed the 8-OH-Gua level in DNA and its repair activity after a single intratracheal instillation of fibers (crocidolite or glass) or saline to Syrian hamsters or Wistar rats. The 8-OH-Gua level was measured with a high-performance liquid chromatography-electrochemical detector (HPLC-ECD) system. The 8-OH-Gua repair enzyme activity was determined with an endonuclease nicking assay using a 32P-labeled or fluorescently labeled 22mer DNA that contains 8-OH-Gua at a specific position. A significant increase in the 8-OH-Gua level in the lung DNA was observed 1 day after the exposure to crocidolite, as compared to the saline control. The repair activity was increased significantly at 7 days. On the other hand, after exposure to glass fibers, little or no increase of these carcinogenicity indicators was detected. These assays of 8-OH-Gua and its repair activity in short-term animal experiments will be useful for evaluating the carcinogenicity of fibers. This is the first report of the increase of 8-OH-Gua and its repair activity in the animal lung after the instillation of asbestos fibers.     

DNA adduct formation and oxidative stress in colon and liver of Big Blue rats after dietary exposure to diesel particles

Exposure to diesel exhaust particles (DEP) via the gastrointestinal route may impose risk of cancer in the colon and liver. We investigated the effects of DEP given in the diet to Big Blue rats by quantifying a panel of markers of DNA damage and repair, mutation, oxidative damage to proteins and lipids, and antioxidative defence mechanisms in colon mucosa cells, liver tissue and the blood compartment. Seven groups of rats were fed a diet with 0, 0.2, 0.8, 2, 8, 20 or 80 mg DEP/kg feed for 21 days. DEP induced a significant increase in DNA strand breaks in colon and liver. There was no effect on oxidative DNA damage (8-oxodG) in colon or liver DNA or in the urine. However, the mRNA expression of OGG1, encoding an enzyme involved in repair of 8-oxodG, was increased by DEP in both liver and colon. DNA adduct levels measured by 32P-post-labelling were elevated in colon and liver, and the expression of ERCC1 gene was affected in liver, but not in colon. In addition to these effects, DEP exposure induced apoptosis in liver. There was no significant change in mutation frequency in colon or liver. The levels of oxidative protein modifications (oxidized arginine and proline residues) were increased in liver accompanied by enhanced vitamin C levels. In plasma, we found no significant effects on oxidative damage to proteins and lipids, antioxidant enzymes or vitamin C levels. Our data indicate that gastrointestinal exposure to DEP induces DNA adducts and oxidative stress resulting in DNA strand breaks, enhanced repair capacity of oxidative base damage, apoptosis and protein oxidation in colon mucosa cells and liver.     

Analyses of oxidative DNA damage and its repair activity in the livers of 3'-methyl-4-dimethylaminoazobenzene-treated rodents.

We measured the levels of 8-hydroxyguanine (8-OH-Gua) and its repair activity in the livers of the Donryu rat, the carcinogen-resistant DRH rat, and the ddy mouse, which were fed a 0.06% 3'-methyl-4-dimethylaminoazobenzene (3'-MeDAB)-containing diet. In a short-term rat experiment (maximum 2 months), 3'-MeDAB did not increase the 8-OH-Gua levels in the livers of the two rat strains, although it significantly increased the repair activity in only the Donryu rat liver at 1 and 2 months. After long-term 3'-MeDAB administration to the ddy mouse (8 months), the levels of 8-OH-Gua and its repair activity were increased in the liver by 3.6-fold and 1.6-fold, respectively. These experiments suggest that 3'-MeDAB increases 8-OH-Gua generation in rodent liver DNA and the 8-OH-Gua repair assay is a reliable marker of cellular oxidative stress induced by carcinogens.     

Detection of a smaller, 32-kDa 8-oxoguanine DNA glycosylase 1 in 3'-methyl-4-dimethylamino-azobenzene-treated mouse liver

We previously reported that 3'-methyl-4-dimethylaminoazobenzene (3'-MeDAB) increased the 8-hydroxyguanine (8-OH-Gua) content in nuclear DNA and the base excision repair activity in mouse liver. However, to understand the mechanism of 3'-MeDAB carcinogenesis, a further investigation of the 8-OH-Gua repair systems was necessary. In this report, we examined the expression of the repair enzyme, 8-oxoguanine DNA glycosylase 1 (OGG1), in 3'-MeDAB-treated mouse liver. We prepared four kinds of anti-peptide polyclonal antibodies raised against mouse OGG1 (mOGG1). The sequences used as epitopes were designed from positions located close to the N-terminus, the nuclear localization signal (NLS), and the regions containing Lys²⁴⁹ and Asp²⁶⁷, which are involved in the catalytic mechanisms of mOGG1 (glycosylase and lyase, respectively). Immunoblotting, using all four antibodies, revealed a 32-kDa protein (mOGG1-32) in addition to the 38-kDa mOGG1 in the 3'-MeDAB-treated mouse liver. Moreover, immunostaining with mOGG1 antibody yielded strong, positive signals in the 3'-MeDAB-treated mouse liver nuclei. However, we could not detect any difference in the Ogg1 mRNA expression pattern. Although the function of mOGG1-32 remains unclear, these findings suggest that 3'-MeDAB may alter the function of the DNA repair protein, and this action may be related to 3'-MeDAB carcinogenesis.     

Analyses of Oxidative DNA Damage and Its Repair Activity in the Livers of 3'-Methyl-4-dimethylaminoazobenzene-treated Rodents

We measured the levels of 8-hydroxyguanine (8-OH-Gua) and its repair activity in the livers of the Donryu rat, the carcinogen-resistant DRH rat, and the ddy mouse, which were fed a 0.06% 3'- methyl-4-dimethylaminoazobenzene (3'-MeDAB)-containing diet. In a short-term rat experiment (maximum 2 months), 3'-MeDAB did not increase the 8-OH-Gua levels in the livers of the two rat strains, although it significantly increased the repair activity in only the Donryu rat liver at 1 and 2 months. After long-term 3'-MeDAB administration to the ddy mouse (8 months), the levels of 8-OH-Gua and its repair activity were increased in the liver by 3.6-fold and 1.6-fold, respectively. These experiments suggest that 3'-MeDAB increases 8-OH-Gua generation in rodent liver DNA and the 8-OH-Gua repair assay is a reliable marker of cellular oxidative stress induced by carcinogens.     

非小细胞肺癌细胞中OGG1调控顺铂诱导的细胞毒性的作用机制探讨

目的:探讨8-氧鸟嘌呤DNA糖基化酶(OGG1)在人肺癌细胞中调控顺铂诱导的细胞毒性的作用机制。方法:采用酶联免疫吸附试验、免疫印迹、流式细胞术和免疫荧光法、线粒体拷贝数和膜电位检测等方法,进行相关实验。结果:顺铂可诱导8-羟基-2'-脱氧鸟苷（8-OHdG）浓度升高,8-OHdG的浓度及细胞凋亡分别与OGG1 mRNA的表达存在显著相关性。外源性表达OGG1促进其DNA修复活性,可降低顺铂诱导的8-OHdG水平,提高非小细胞肺癌（non-small cell lung cancer,NSCLC）细胞活力,并抑制顺铂诱导的NSCLC细胞凋亡,而OGG1沉默则表现相反的结果。此外,过表达OGG1减少顺铂导致的线粒体DNA损伤和功能障碍,沉默OGG1增强顺铂诱导的MAPK/p38通路的激活,导致NSCLC细胞发生凋亡。结论:OGG1的下调有利于提高肺癌细胞对顺铂的敏感性。因此,本研究通过在体外证实OGG1介导的DNA修复在顺铂耐药中的作用,为肺癌的治疗提供了理论支持。     

Increased 8-hydroxyguanine levels in DNA and its repair activity in rat kidney after administration of a renal carcinogen, ferric nitrilotriacetate

The renal carcinogen, ferric nitrilotriacetate (Fe-NTA), isknown to induce oxidative stress and the subsequent formationof a type of oxidative DNA damage, 8-hydroxygu-anine (8-OH-Gua),in the rat kidney (Umemura et al., 1990). Using an improvedDNA isolation method (Nakae et al., 1995), which reduces thebackground level of 8-OH-Gua, we found a five-fold increasein the 8-OH-Gua level in kidney DNA after a single i.p. injectionof Fe-NTA. On the basis of the report that 8-OH-Gua repair activityis enhanced after cells are exposed to oxidative stress dueto ionizing radiation (Bases et al., 1992), the measurementof 8-OH-Gua repair activity will also be useful to assess cellularoxidative stress. The 8-OH-Gua repair enzyme activity was determinedwith an endonuclease assay using a 22 mer DNA that contains8-OH-Gua at a specific position. A five-fold increase in the8-OH-Gua repair activity as compared with the control, was observedin the target organ, the rat kidney, 120 h after Fe-NTA administration.In the non-target organ, the liver, the increase was not aslarge (two-fold). This simple assay of oxidative DNA damagerepair will be useful for evaluating the carcinogenicity ofoxygen radical forming chemicals, in addition to chemical analysesof oxidative DNA damage.     

Induction of cytochrome P450 1B1 in lung, liver and kidney of rats exposed to diesel exhaust.

We have shown previously that diesel exhaust particle (DEP) extracts (DEPE) and 1-nitropyrene were genotoxically activated by human cytochrome P450 1B1 in SOS/umu assay. In this study, the in vivo induction of P450 family 1 enzymes in rats by exposure to diesel exhaust was investigated with regard to mRNA levels, P450 enzyme content, drug oxidation activities in the microsomes and umu gene expression of typical P450 substrates and DEPE itself catalyzed by the microsomes. Male Fischer 344 rats (4 weeks old) were exposed to 0.3 and 3.0 mg/m(3) DEP for 12 h per day for 4 weeks; the former dose corresponded to the typical daily airborne particle concentration. The levels of mRNA of rat P450 1B1 and P450 1A1 in the lung and liver were significantly increased 1.1-1.4-fold by exposure to 0.3 mg/m(3) DEP. Diesel exhaust particle extracts induced umu gene expression in Salmonella typhimurium TA1535/pSK1002 in the absence of a functional P450 system and were further activated by human recombinant P450 1B1. Using an O-acetyltransferase overexpressing Salmonella strain, genotoxic activation of P450 1B1 marker chemicals (1-nitropyrene, 1-aminopyrene and DEPE) by lung, liver and kidney microsomes was increased 1.7-4.2-, 1.4-1.5- and 1.0-1.3-fold, respectively, by exposure to 0.3 mg/m(3) DEP. Activation of 3-amino-1,4-dimethyl-5H-pyrido [4,3-b]indole (Trp-P-1; marker for P450 1A1) by lung microsomes and the P450 1A2 content in liver microsomes were slightly increased by exposure to 3.0 mg/m(3) DEP. This is the first report to suggest that typical daily contaminant levels (0.3 mg particle/m(3)) of diesel exhaust can induce P450 1B1 in rats and that the induced P450 1B1 may catalyze the genotoxic activation of DEP.     

Oxygen radicals in lung carcinogenesis accompanying phagocytosis of diesel exhaust particles

We sought to examine the involvement of oxygen radicals derived from phagocytosis process in lung carcinogenesis induced by diesel exhaust particles (DEP). The carcinogenic response and formation of 8-hydroxydeoxyguanosine (8-OHdG) were examined in the lungs of mice intratracheally injected with washed DEP (WDEP), DEP, or nontoxic control particles of titanium dioxide (TiO2). After 10 weekly treatments with these particles, the formation of 8-OHdG in the lungs of mice treated with WDEP or DEP showed a significant increase, but not in those treated with TiO2. After 12 months, the incidence of lung tumors in mice treated with WDEP or DEP was higher than that of mice treated with vehicle by 2.3- and 3.1-fold, respectively. A significant difference in the incidence of tumors was found between the vehicle group and DEP-treated group. Treatment with TiO2 had no effect on the incidence of lung tumors. The formation of 8-OHdG in mice treated with these particles was significantly correlated with the development of lung tumors. These results suggest that the induction of DNA damage by oxygen radicals may be an important factor in the initiation of WDEP- and DEP-induced lung carcinogenesis, and that oxygen radicals derived from the phagocytic process may play a role in 8-OHdG formation induced by DEP.     

Inter-individual variation,seasonal variation and close correlation of OGG1 and ERCC1 mRNA levels in full blood from healthy volunteers

The mRNA levels of the nucleotide excision DNA repair gene ERCC1 and the base excision DNA repair gene OGG1 were quantified in 43 healthy volunteers in a dietary intervention trial as markers for the DNA repair capacity. Nine samples were collected from each subject over a period of 52 days. Sampling took place from January to May. The mRNA levels of OGG1 and ERCC1 correlated closely (r = 0.86, P < 0.0001) after normalization to either 18S ribosomal RNA or to beta-actin mRNA. The levels of OGG1 and ERCC1 mRNA were relatively constant within an individual with intra-individual correlation (R(2) = 0.45-0.46) in a General Linear Model. The amounts of ERCC1 and OGG1 relative to 18S RNA were doubled in May compared with January. This coincided with an increase in the monthly influx of sunlight from 18 MJ/m(2) in January to 242 MJ/m(2) in May. The mRNA levels of both ERCC1 and OGG1 were positively correlated to the average daily influx of sunlight in the previous 30 and 5 days (r = 0.49; r = 0.37, respectively, P < 0.001). There were no significant effects of the dietary interventions. The inter-individual variation was 5-10-fold, which is more than the observed 2-3-fold seasonal variation. Thus, despite seasonal variation of the individual mRNA levels, the inter-person variation is still far larger than the intra-person variation, supporting the use as biomarkers.     

Analysis of 8-hydroxyguanine in rat kidney genomic DNA after administration of a renal carcinogen, ferric nitrilotriacetate.

The frequency of oxidative base damage, such as 8-hydroxyguanine (8-OH-Gua), was determined at the nucleotide level of resolution using the ligation-mediated PCR technique. Administration of a renal carcinogen, ferric nitrilotriacetate (Fe-NTA), is known to induce oxidative stress and subsequent formation of 8-OH-Gua in the rat kidney. Whole genomic DNA was isolated from the rat kidney after or without Fe-NTA treatment and then cleaved with hot piperidine. In order to assess the frequency of 8-OH-Gua formation, we chose three genes, the tumor suppressor gene p53, the heat shock protein 70 (HSP70-1) gene and the Na,K-ATPase alpha1 subunit gene. No alteration in the cleavage profile was observed in the p53 and HSP70 genes after Fe-NTA treatment. In the case of the p53 gene, a low incidence of point mutations has been observed in this carcinogenesis system. On the other hand, time-dependent alterations, corresponding to the time course of overall 8-OH-Gua formation and repair, were detected in the promoter region of the Na,K-ATPase alpha1 subunit gene. GpG and GpGpG in specific regions seem to be hotspots for the formation of 8-OH-Gua. These results were confirmed by formamidopyrimidine-DNA glycosylase-dependent DNA cleavage patterns. Thus, oxidative base damage, such as 8-OH-Gua, was not distributed uniformly along the whole genome, but seemed to be restricted to particular genes and regions.     

Formation of 8-hydroxydeoxyguanosine and cell-cycle arrest in the rat liver via generation of oxidative stress by phenobarbital: association with expression profiles of p21(WAF1/Cip1), cyclin D1 and Ogg1

To evaluate the risk of exposure to so-called non-genotoxic chemicals and elucidate mechanisms underlying their promoting activity on rat liver carcinogenesis the formation of 8-hydroxy-2'-deoxyguanosine (8-OHdG), cytochrome P-450 (P-450) and hydroxyl radicals induction, DNA repair and alteration to cellular proliferation and apoptosis in the rat liver were investigated during 2 weeks of phenobarbital (PB) administration at a dose of 0.05%. Significant increase of hydroxyl radical levels by day 4 of PB exposure accompanied the accumulation of 8-OHdG in the nucleus and P-450 isoenzymes CYP2B1/2 and CYP3A2 in the cytoplasm of hepatocytes. Conspicuous elevation of 8-OHdG and apoptosis in the liver tissue were associated with reduction of the proliferating cell nuclear antigen (PCNA) index after 8 days of PB application. Thereafter, 8-OHdG levels decreased with an increase in mRNA expression for the 8-OHdG repair enzyme, DNA glycosylase 1 (Ogg1). Analysis with LightCycler quantitative 2-step RT-PCR demonstrated induction of cyclin D1 (CD1) and p21(WAF1/Cip1) mRNA expression on days 4 and 6, respectively, preceding marked elevation of PCNA and apoptotic indices. These results suggest that similar to genotoxic, non-genotoxic chemicals might induce reversible alteration to nuclear 8-OHdG in the rat liver after several days of continuous application; however, by a different mechanism. Increased 8-OHdG formation is caused by developing oxidative stress or apoptotic degradation of DNA and coordinated with enhanced expression of CD1 mRNA and cell proliferation, subsequent increase of p21(WAF1/Cip1) mRNA expression, cell-cycle arrest and apoptosis, while activation of 8-OHdG repair mechanisms contributes to protection of tissue against reactive oxygen species-induced cell death.     

Down-regulation of the DNA-repair endonuclease 8-oxo-guanine DNA glycosylase 1 (hOGG1) by sodium dichromate in cultured human A549 lung carcinoma cells.

Hexavalent chromium is a genotoxic human pulmonary carcinogen that elevates DNA oxidation, apparently through the generation of reactive DNA-damaging intermediates including Cr(V), Cr(IV) and reactive oxygen species. We tested the hypothesis that elevation of DNA oxidation may also be through inhibition of the expression of the repair glycosylase for 8-oxo deoxyguanine (hOGG1) in cultured A549 human lung epithelial cells. Treatment with sodium dichromate (0-100 microM, 16 h) resulted in a concentration-dependent decrease in the levels of OGG1 mRNA as measured by both RT-PCR and RNase protection assay. Sodium dichromate at 25 microM and above gave a marked reduction of OGG1 mRNA expression which was not seen at 1 microM and below. No effect on the expression of the apurinic endonuclease hAPE or the house-keeping gene GAPDH was observed at any of the concentrations of sodium dichromate investigated. Treatment of cells with the pro-oxidant H(2)O(2) (0-200 microM) for 16 h had no detectable effect on the levels of OGG1 mRNA or protein expression suggesting that the effect of sodium dichromate is not mediated by H(2)O(2). Western blotting demonstrated that sodium dichromate (100 microM; 16 h and >25 microM; 28 h) markedly reduced levels of OGG1 protein in nuclear cell extracts. Additionally, treatment of cells with sodium dichromate (>25 microM, 28 h) resulted in a concentration-dependent decrease in the ability of nuclear extracts to nick a synthetic oligonucleotide containing 8-oxo deoxyguanine (8-oxo dG). We conclude that the elevation of 8-oxo dG levels observed in A549 cells treated with sodium dichromate may be, at least in part, due to a reduced capacity to repair endogenous and hexavalent chromium-induced 8-oxo dG.     

THE DUAL EFFECT OF THE PARTICULATE AND ORGANIC COMPONENTS OF DIESEL EXHAUST PARTICLES ON THE ALTERATION OF PULMONARY IMMUNE/INFLAMMATORY RESPONSES AND METABOLIC ENZYMES

ABSTRACT

Exposure to diesel exhaust particles (DEP) is an environmental and occupational health concern. This review examines the cellular actions of the organic and the particulate components of DEP in the development of various lung diseases. Both the organic and the particulate components cause oxidant lung injury. The particulate component is known to induce alveolar epithelial damage, alter thiol levels in alveolar macrophages (AM) and lymphocytes, and activate AM in the production of reactive oxygen species (ROS) and pro-inflammatory cytokines. The organic component, on the other hand, is shown to generate intracellular ROS, leading to a variety of cellular responses including apoptosis. There are a number of differences between the biological actions exerted by these two components. The organic component is responsible for DEP induction of cytochrome P450 family 1 enzymes that are critical to the polycyclic aromatic hydrocarbons (PAH) and nitro-PAH metabolism in the lung as well as in the liver. The particulate component, on the other hand, causes a sustained down-regulation of CYP2B1 in the rat lung. The significance of this effect on pulmonary metabolism of xenobiotics and endobiotics remains to be seen, but may prove to be an important factor governing the interplay of the pulmonary metabolic and inflammatory systems. Long-term exposures to various particles including DEP, carbon black (CB), TiO₂, and washed DEP devoid of the organic content, have been shown to produce similar tumorigenic responses in rodents. There is a lack of correlation between tumor development and DEP chemical-derived DNA adduct formation. But the organic component has been shown to generate ROS that produce 8-hydroxydeoxyguanosine (8-OHdG) in cell culture. The organic, but not the particulate, component of DEP suppresses the production of pro-inflammatory cytokines by AM and the development of Th1 cell-mediated immunity. The mechanism for this effect is not yet clear, but may involve the induction of heme oxygenase-1 (HO-1), a cellular genetic response to oxidative stress. Both the organic and the particulate components of DEP enhance respiratory allergic sensitization. Part of the DEP effects may be due to a depletion of glutathione in lymphocytes. The organic component, which is shown to induce IL-4 and IL-10 productions, may skew the immunity toward Th2 response, whereas the particulate component may stimulate both the Th1 and Th2 responses. In conclusion, the literature shows that the particulate and organic components of DEP exhibit different biological actions but both involve the induction of cellular oxidative stress. Together, these effects inhibit cell-mediated immunity toward infectious agents, exacerbate respiratory allergy, cause DNA damage, and under long-term exposure, induce the development of lung tumors.     

Somatic mutations and single nucleotide polymorphisms of base excision repair genes involved in the repair of 8-hydroxyguanine in damaged DNA

To elucidate the involvement of 8-hydroxyguanine (oh(8)G) repair genes in human lung carcinogenesis, 47 lung cancer cell lines and 55 primary lung cancers were examined for somatic mutations and genetic polymorphisms in all coding exons of the MYH and APEX genes, and exon 8 of the OGG1 gene by polymerase chain reaction-single strand conformation polymorphism analysis. In the MYH gene, one missense mutation was detected in a cell line, NCI-H157, whereas no mutations were detected in primary cancers. There were no mutations in the APEX and OGG1 genes in the cell lines or primary cancers. Ten single nucleotide polymorphisms (SNPs) were identified, and seven of them were accompanied by amino acid substitutions. Differences in the oh(8)G repair activities of MYH, APEX and OGG1 proteins due to somatic mutations and SNPs can be involved in human carcinogenesis.     

非小细胞肺癌患者hOGG1基因启动子区域突变的研究

背景与目的 8-羟基鸟嘌呤DNA糖苷酶(8-hydroxygumine DNA glycosylase1,OGG1)是一种DNA修复酶,可以从DNA切除修复8-羟基鸟嘌呤(8-dihydro-8-oxoguanine,8-OH-G)。人类OGG1基因(hOGG1)的多态性可能会改变酶的活性而影响个体修复损伤DNA的能力,促进癌变。然而,hOGG1基因启动子区域的突变与非小细胞肺癌(non-small cell lung cancer,NSCLC)的关系尚不明晰。我们拟探讨hOGG1基因启动子区域的突变与NSCLC发生发展的潜在关系。方法选取苏州大学附属第一医院2003年1月-2005年12月新鲜手术切除的40例NSCLC组织标本,采用PCR-SSCP和直接测序的方法检测NSCLC及其对应的癌旁组织中hOGG1基因启动子区域的突变。结果在40例NSCLC患者中未发现hOGG1基因启动子区域的异常突变,但发现单核苷酸多态位点rs159153与TNM分期明显相关(P=0.008);同时发现吸烟者中淋巴结转移率明显较低(P=0.034)。结论单核苷酸多态位点rs159153和吸烟史可能对NSCLC的侵袭和转移潜在性提供预测。