[Survival analysis of malignant tumors in cancer registration areas of Hubei province in China, 2013 to 2015].

Objective: To analyze the survival of newly diagnosed malignant tumors in cancer registration areas of Hubei Province from 2013 to 2015. Methods: From January 1, 2013 to December 31, 2015, all newly diagnosed malignant tumors were collected from cancer registration areas in Hubei Province, and patients were followed up using a combination of active and passive methods. Cancer survival was analyzed using the strs package in Stata software. Observed and expected survival were calculated using the life table and Ederer Ⅱ methods, and the difference in survival rate of patients with different sex, age, urban and rural areas and different cancer species was compared. Results: From 2013 to 2015, 83 987 new malignant tumors were diagnosed in cancer registration areas in Hubei Province, including 45 742 males (54.46%) and 38245 females (45.54%). The overall 5-year relative survival rate was 41.46%, 34.43% for men and 49.63% for women. With the increase of age, the observed survival rate and relative survival rate of patients of different genders showed a decreasing trend. The 5-year relative survival rate of patients with malignant tumors was 47.58% in urban areas and 26.58% in rural areas. The observed survival rate and relative survival rate in rural areas were significantly lower than those in urban areas. The overall 5-year relative survival rates for common malignancies were 20.61% for lung cancer, 15.36% for liver cancer, 22.89% for esophageal cancer, 34.92% for gastric cancer, and 54.87% for colorectal cancer. In addition, the 5-year relative survival rates of common malignant tumors in women were 78.65% for breast cancer and 52.55% for cervical cancer. Conclusions: In Hubei Province, the survival rate of malignant tumors is different among different genders, regions, age groups and cancer species. Prevention and treatment and health education should be strengthened for malignant tumor patients in rural areas and those with high incidence and low survival rate such as liver cancer and lung cancer, and relevant strategies should be formulated according to the gender and age distribution characteristics of different cancer species.

[Progress on development and application of the diet-related inflammatory index in cancer epidemiological studies].

Cancer is a major public health problem worldwide, causing an more serious burden of disease. Inflammation is considered a predisposing factor for cancer with close relationship with its incidence. In recent years, the public and epidemiologists has paid more attention to the association between nutrition and cancer and other chronic diseases in the perspective of inflammation. This paper summarizes the development and application of the diet-related inflammatory index in cancer epidemiological studies based on the literature retrieval of common diet-related inflammatory index. Firstly, we highlight the common diet-related inflammatory indices and their construction methods, such as the Dietary Inflammatory Index, a literature-derived diet-related inflammatory index, and the Empirical Dietary Inflammatory Index, an empirically derived diet-related inflammatory index, and so on. Secondly, the epidemiological research progress on the commonly used diet-related inflammatory indices is briefly introduced. Finally, the advantages and disadvantages of the two types of this inflammatory indices are also briefly described for the purpose of providing reference for nutrition epidemiological studies of cancer and other chronic diseases in China.

[Choice and application of time scale selection for Cox proportional hazards regression model in cohort studies].

Cox proportional hazards regression model (Cox model) is the most commonly used multivariate approach in time-to-event data analysis. A vital issue in fitting Cox model is choosing the appropriate time scale related to the occurrence of the outcome events. However, few domestic studies have focused on selecting and applying time scales for Cox model in the analysis of cohort study data. This study briefly introduced and compared several time scales in the reports from literature; and used data from the Shanghai Women's Health Study to illustrate the impact of different time scales on data analysis results, using the association between central obesity and the risk of liver cancer as an example. On this basis, several suggestions on selecting time scales in Cox model are proposed to provide a reference for the analysis of cohort study data.

[Statistical methods for relative risk estimation and applications in case-cohort study].

Objective: To systematically introduce the design of case-cohort study and the statistical methods of relative risk estimation and their application in the design. Methods: First, we introduced the basic principles of case-cohort study design. Secondly, Prentice's method, Self-Prentice method and Barlow method were described in the weighted Cox proportional hazard regression models in detail, finally, the data from the Shanghai Women's Health Study were used as an example to analyze the association between obesity and liver cancer incidence in the full cohort and case-cohort sample, and the results of parameters from each method were compared. Results: Significant association was observed between obesity and risk for liver cancer incidence in women in both the full cohort and the case-cohort sample. In the Cox proportional hazard regression model, the partial regression coefficients of the full cohort and the case-cohort sample fluctuated with the adjustment of confounding factors, but the hazard ratio estimates of them were close. There was a difference in the standard error of the partial regression coefficient between the full cohort and the case-cohort sample. The standard error of the partial regression coefficient of the case-cohort sample was larger than that of the full cohort, resulting in a wider 95% confidence interval of the relative risk. In the weighted Cox proportional hazard regression model, the standard error of the partial regression coefficient of Prentice's method was closer to the parameter estimates from full cohort than Self-Prentice method and Barlow method, and the 95% confidence interval of hazard ratio was closer to that of the full cohort. Conclusions: Case-cohort design could yield parameter results closer to the full cohort by collecting and analyzing data from sub-cohort members and patients with the disease, and reduce sample size and improve research efficiency. The results suggested that Prentice's method would be preferred in case-cohort design.

[Report of incidence and mortality of gallbladder cancer in China, 2014].

Objective: The incidence and mortality of gallbladder cancer from Chinese cancer registries in 2014 were analyzed to describe the prevalence of gallbladder cancer in China. Methods: Incidence and mortality data of gallbladder cancer in 2014 derived from registration data in 2017, collected by the National Central Cancer Registry (NCCR). Qualified data from 339 cancer registries were calculated after evaluating. According to the national population data of 2014, the gallbladder cancer incidence and mortality of China in 2014 were stratified by the area, gender and age.The age composition of standard population of Chinese census in 2000 and Segi's population were used for age-standardizes incidence and mortality in China and worldwide. Results: 339 cancer registries cover a total of 288 243 347 population including 146 203 891 males and 142 039 456 females (144 061 915 in urban and 144 181 432 in rural areas). The mortality to incidence ratio of gallbladder cancer was 0.74. The morphologically verified cases (MV%) and death certificate-only cases (DCO%) were 48.38% and 2.66%, respectively. Unclear diagnosis cases (UB%) was 0.48%. The crude incidence of gallbladder cancer in China in 2014 was 3.82/100 000, which accounted for 1.37% of new cancer cases (4.48/100 000 in urban areas and 3.01/100 000 in rural areas, 3.59/100 000 for male and 4.05/100 000 for female). Age-standardized incidence rates by Chinese standard population (ASR China) and world standard population (ASR world) were 2.38/100 000 and 2.37/100 000, respectively, and the cumulative incidence rate (0-74 age years old) was 0.27%.Besides, the crude mortality of gallbladder cancer was 2.86/100 000 (3.47/100 000 in urban areas and 2.12/100 000 in rural areas, 2.59/100 000 for male and 3.14/100 000 for female). Age-standardized mortality rates by ASR China and ASR world were 1.72/100 000 and 1.71/100 000, with a cumulative mortality rate (0-74 age years old) of 0.19%. Conclusion: The incidence and mortality of gallbladder cancer were significantly different between the city and country, while not obviously different between the female and male.

FPR1 interacts with CFH, HTRA1 and smoking in exudative age-related macular degeneration and polypoidal choroidal vasculopathy.

PURPOSE: To determine the genetic association of an inflammation-related gene, formyl peptide receptor 1 (FPR1), in exudative age-related macular degeneration (AMD) and polypoidal choroidal vasculopathy (PCV). METHODS: The coding region of FPR1 gene was sequenced in 554 unrelated Chinese individuals: 155 exudative AMD patients, 179 PCV patients, and 220 controls. Interactions and combined effects of FPR1 with complement factor H (CFH), high temperature requirement factor A1 (HTRA1), and smoking were also investigated. RESULTS: A total of 28 polymorphisms in FPR1 were identified. Single nucleotide polymorphisms (SNP) rs78488639 increased the risk to exudative AMD (P=0.043) and PCV (P=0.016), whereas SNP rs867229 decreased the risk to exudative AMD (P=0.0026), but not PCV. Homozygous G allele of rs1042229 was associated with exudative AMD (P=0.0394, odds ratio (OR)=2.27, 95% confident interval: 1.08-4.74), but not with PCV. Exudative AMD, but not PCV, was associated with the heterozygous genotypes of rs2070746 (P=0.019, OR=0.57) and rs867229 (P=0.0082, OR=0.54). Significantly, interactions were identified among FPR1 rs78488639, CFH rs800292, and HTRA1 rs11200638 in both exudative AMD and PCV. Combined heterozygous risk alleles of CFH rs800292 GA and FPR1 rs78488639 CA were posed to PCV (P=2.22 × 10(-4), OR=10.47), but not exudative AMD. Furthermore, FPR1 rs78488639 CA combining with HTRA1 rs11200638 and smoking was also predisposed risks to exudative AMD and PCV. CONCLUSION: FPR1 is associated with exudative AMD and PCV in a Hong Kong Chinese cohort. FPR1 rs78488639 interacted with CFH rs800292, HTRA1 rs11200638, and smoking, enhancing risk to exudative AMD and PCV.

Chlamydia pneumoniae infection, complement factor H variants and age-related macular degeneration.

BACKGROUND/AIMS: Impaired inhibition of the alternative complement pathway by complement factor H (CFH) is linked to age-related macular degeneration (AMD) based on the strong association between CFH variant and AMD. Chlamydia pneumoniae (C pneumoniae) infection can trigger the alternative pathway, but the evidence for an association between C pneumoniae and AMD is contradictory. This study investigated whether C pneumoniae infection is associated with AMD and whether the presence of C pneumonia modulates AMD risk conferred by CFH variants. METHODS: Genomic DNA extracted from peripheral blood of 148 advanced AMD patients and 162 controls was subjected to Taqman and PCR-RFLP for the CFH polymorphism and PCR for the C pneumoniae gene. Genomic DNA was also examined from microdissected macular cells from 59 AMD and 16 age-matched non-AMD archived slides. chi(2) testing was performed for case-control analysis. RESULTS: C pneumoniae infection was associated with increased risk of AMD (OR = 2.17, p<0.017). A CFH variant was also linked to increased risk of AMD (OR = 1.98, p<0.0001). However, no relationship was found between risk-conferring CFH variant and C pneumoniae (OR = 1.81, p = 0.08). CONCLUSION: There is a possible association between AMD and C pneumoniae infection, although CFH may not be directly involved in the pathogenesis of C pneumoniae infection-mediated AMD.

Detection of CX3CR1 single nucleotide polymorphism and expression on archived eyes with age-related macular degeneration.

There is a significant genetic component in age-related macular degeneration (AMD). CX3CR1, which encodes the fractalkine (chemokine, CX3CL1) receptor, has two single nucleotide polymorphisms (SNPs): V249I and T280M. These SNPs are correlated with other aged-related diseases such as atherosclerosis. We have reported an association of CX3CR1 SNP and AMD. In this study we examined CX3CR1 SNP frequencies and protein expression on archived sections of AMD and normal eyes. We microdissected non-retinal, peripheral retinal and macular cells from archived slides of eyes of AMD patients and normal subjects. CX3CR1 SNP typing was conducted by PCR and restriction fragment length polymorphism analysis. CX3CR1 transcripts from retinal cells were also measured using RT-PCR. CX3CR1 protein expression was evaluated using avidin-biotin complex immunohistochemistry. We successfully extracted DNA from 32/40 AMD cases and 2/2 normal eyes. Among the 32 AMD cases, 18 had neovascular AMD and 14 had non-neovascular AMD. The M280 allele was detected in 19/64 (32 cases x2) with a frequency of 29.7%, which was significantly higher as compared to the frequency in the normal population (11.2%). We detected CX3CR1 expression in the various retinal cells. CX3CR1 transcript and protein levels were diminished in the macular lesions. This study successfully analyzed CX3CR1 SNP and transcript expression in microdissected cells from archived paraffin fixed slides. Our data suggest that the M280 allele, a SNP resulting in aberrant CX3CR1 and CX3CL1 interaction, as well as lowered expression of macular CX3CR1, may contribute to the development of AMD.

The Cockayne Syndrome group B gene product is involved in general genome base excision repair of 8-hydroxyguanine in DNA.

Cockayne Syndrome (CS) is a human genetic disorder with two complementation groups, CS-A and CS-B. The CSB gene product is involved in transcription-coupled repair of DNA damage but may participate in other pathways of DNA metabolism. The present study investigated the role of different conserved helicase motifs of CSB in base excision repair. Stably transformed human cell lines with site-directed CSB mutations in different motifs within its putative helicase domain were established. We find that CSB null and helicase motif V and VI mutants had greater sensitivity than wild type cells to gamma-radiation. Whole cell extracts from CSB null and motif V/VI mutants had lower activity of 8-hydroxyguanine incision in DNA than wild type cells. Also, 8-hydroxyguanine accumulated more in CSB null and motif VI mutant cells than in wild type cells after exposure to gamma-radiation. We conclude that a deficiency in general genome base excision repair of selective modified DNA base(s) might contribute to CS pathogenesis. Furthermore, whereas the disruption of helicase motifs V or VI results in a CSB phenotype, mutations in other helicase motifs do not cause this effect. The biological functions of CSB in different DNA repair pathways may be mediated by distinct functional motifs of the protein.

Importance of guanine nitration and hydroxylation in DNA in vitro and in vivo.

Guanine (Gua) modification by nitrating and hydroxylating systems was investigated in DNA. In isolated calf thymus DNA, 8-NO(2)-Gua and 8-oxo-Gua were dose-dependently formed with peroxynitrite, and 8-NO(2)-Gua was released in substantial amounts. Myeloperoxidase (MPO) with H(2)O(2) and NO(2)(-) reacted with calf thymus DNA to form 8-NO(2)-Gua dose dependently without release of 8-NO(2)-Gua. The frequency of strand breaks was higher than the sum of 8-NO(2)-Gua and 8-oxo-Gua, particularly in the MPO-treated DNA, indicating the importance of other types of damage. The activation of human neutrophils and lymphocytes with phorbol ester did not induce 8-NO(2)-Gua and 8-oxo-Gua in their nuclear DNA. However, 8-NO(2)-Gua was found in calf thymus DNA co-incubated with activated neutrophils in the presence of NO(2)(-). No significant formation of 8-NO(2)-Gua was found in liver DNA from mice treated with Escherichia coli lipopolysaccharide. The incubation of peroxynitrite or MPO-H(2)O(2)-NO(2)(-)-treated DNA with formamidopyrimidine glycosylase (Fpg) released 8-oxo-Gua, but not 8-NO(2)-Gua, indicating that 8-NO(2)-Gua is not a substrate for Fpg. Although 8-NO(2)-Gua was generated in isolated DNA by different nitrating systems, other types of damage were formed in abundance, and the lesion could not be found reliably in nuclear DNA, suggesting that the biological importance is limited.

Enhanced benzene-induced DNA damage in PMA-stimulated cells in vitro and in LPS-treated animals.

The present study investigated the interaction between inflammatory reactions and benzene in vitro and in vivo with respect to oxidative DNA damage. In the in vitro models the oxidative burst of cells was induced by the pretreatment with phorbol myristate acetate (PMA) and in the in vivo models of inflammation mice were pretreated with lipopolysaccharide (LPS). The oxidative DNA damage was indicated by 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) and strand breaks as assessed by alkaline single cell gel electrophoresis (SCGE, Comet assay). The results showed that combination of PMA and benzene enhanced the level of 8-oxodG in DNA from mouse bone marrow cells by 197%, from human lymphocytes by 188% and from human neutrophils by 205% (p < .05). Pretreatment of mice with LPS and benzene resulted in an enhanced Comet score formation in bone marrow cells by 98% and in lymphocytes by 39% in Comet score (p < .05) and in an enhanced 8-oxodG level in bone marrow cells by 290%. The effects of the combined treatment with PMA/LPS and benzene exceeded the sum of the effects induced by PMA/LPS or benzene alone. The production of nitrate/nitrite showed a two fold increase in the supernatant from incubation of benzene and PMA-pretreated neutrophils. The increase in the 8-oxodG level in the human neutrophil incubation system demonstrated a correlation with nitrate/nitrite production, indicating a possible relationship with the generation of reactive nitrogen species.

Dexamethasone ameliorates oxidative DNA damage induced by benzene and LPS in mouse bone marrow.

Mice were grouped to receive vehicle, dexamethasone (DEX), lipopolysaccharide (LPS), benzene (BZ, 200 mg/kg) and combinations: LPS + DEX, BZ + DEX, LPS + BZ, LPS + DEX + BZ. The DNA damage in bone marrow cells from BZ group was enhanced 2.8-fold measured by nuclear 8-hydroxy-2 '-deoxyguanosine (8-oxodG) and 1.4-fold measured by Comet score (index of DNA breaks) (p < 0.05). In the BZ + DEX group, 8-oxodG level and the Comet score were lowered to 65% and 76% respectively of that in the BZ group (p < 0.05). The BZ + LPS caused a 3.9-fold increase in 8-oxodG and a 1.6-fold increase in the Comet score (p < 0.05). The LPS + DEX + BZ lowered 8-oxodG level and the Comet score to 50% and 78% of the values in the LPS + BZ group, respectively (p < 0.05). Nitrate/nitrite levels in serum were higher after BZ + LPS treatment than after all other treatments. Both 8-oxodG level and the Comet scores were correlated to the serum nitrate/nitrite level across all the treatments (r = 0.55, p < 0.01 and r = 0.69, p < 0.01, respectively). In bone marrow cells the 8-oxodG correlated with the Comet scores (r = 0.80, p < 0.01). We conclude that DEX administration can reduce the DNA damage from BZ treatment and from the combination of BZ and LPS. The correlation of DNA damage with nitrate/nitrite indicates the possible involvement of reactive nitrogen species (RNS) in the interaction between BZ and the inflammatory reaction stimulated by LPS. The 8-oxodG determination is more sensitive than strand break analysis by the Comet assay in bone marrow in vivo in mice for measuring the BZ-induced DNA damage.

Prevention of oxidative DNA damage in rats by brussels sprouts.

The alleged cancer preventive effects of cruciferous vegetables could be related to protection from mutagenic oxidative DNA damage. We have studied the effects of Brussels sprouts, some non-cruciferous vegetables and isolated glucosinolates on spontaneous and induced oxidative DNA damage in terms of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) in groups of 6-8 male Wistar rats. Excess oxidative DNA damage was induced by 2-nitropropane (2-NP 100 mg/kg). Four days oral administration of 3 g of cooked Brussels sprouts homogenate reduced the spontaneous urinary 8-oxodG excretion by 31% (p<0.05) whereas raw sprouts, beans and endive (1:1), isolated indolyl glucosinolates and breakdown products had no significant effect. An aqueous extract of cooked Brussels sprouts (corresponding to 6.7 g vegetable per day for 4 days) decreased the spontaneous 8-oxodG excretion from 92 +/- 12 to 52 +/- 15 pmol/24 h (p<0.05). After 2-NP administration the 8-oxodG excretion was increased to 132 +/- 26 pmol/24 h (p<0.05) whereas pretreatment with the sprouts extract reduced this to 102 +/- 30 pmol/24 h (p<0.05). The spontaneous level of 8-oxodG in nuclear DNA from liver and bone marrow was not significantly affected by the sprouts extract whereas the level decreased by 27% in the kidney (p<0.05). In the liver 2-NP increased the 8-oxodG levels in nuclear DNA 8.7 and 3.8 times (p<0.05) 6 and 24 h after dose, respectively. The sprouts extract reduced this increase by 57% (p<0.05) at 6 h whereas there was no significant effect at 24 h. In the kidneys 2-NP increased the 8-oxodG levels 2.2 and 1.2 times (p<0.05) 6 and 24 h after dose, respectively. Pretreatment with the sprouts extract abolished these increases (p<0.05). Similarly, in the bone marrow the extract protected completely (p<0.05) against a 4.9-fold 2-NP induced increase (p<0.05) in the 8-oxodG level. These findings demonstrate that cooked Brussels sprouts contain bioactive substance(s) with a potential for reducing the physiological as well as oxidative stress induced oxidative DNA damage in rats. This could explain the suggested cancer preventive effect of cruciferous vegetables. The correspondence between the urinary excretion and 8-oxodG levels in 2-NP target organs supports its being the main repair product that reflects the rate of guanine oxidation in DNA.

Formation of nitrated and hydroxylated aromatic compounds from benzene and peroxynitrite, a possible mechanism of benzene genotoxicity.

Peroxynitrite, the reaction product of nitric oxide (NO.) and superoxide anion (O2.-) produced during immune activation by a variety of inflammatory cells, may contribute to genotoxicity of benzene through its ability to carry out hydroxylation and nitration. After exposure of benzene to synthesised peroxynitrite, phenol, nitrophenols (p-nitrophenol, o-nitrophenol and m-nitrophenol) and nitrobenzene were identified in the reaction mixture by HPLC separation and single UV wavelength and diode array detection. The formation of phenol, nitrophenols and nitrobenzene showed a linear relationship with both benzene and peroxynitrite concentrations. The molar ratio for phenol/(nitrobenzene and nitrophenols) was approximately 9/5 with a total product yield of 14% hydroxylated and nitrated products as based on peroxynitrite. The physiological relevance of the chemical reaction between benzene and peroxynitrite was tested by detecting the reaction products in human neutrophils (2.5 x 10(7)cells/ml) incubated with 10 mM benzene for 25 min. The concentration of phenol and p-nitrophenol were found to be 1.29+/-0.22 and 1.56+/-0.61 microM (mean+/-SD) in the incubation medium of the neutrophils pretreated with phorbol myristate acetate (500 nM) for 5 min, respectively, whereas no metabolites were detected if the neutrophils were not pretreated. Nitrated aromatic compounds are known to be more carcinogenic than the parent compounds. It is reported that acute and chronic infection increases the risk of cancer at various sites; and that anti-inflammatory agents decrease benzene myelotoxicity. We suggest that the increased production of peroxynitrite during chronic inflammation combined with benzene exposure may increase the carcinogenicity of benzene by a mechanism that includes the formation of metabolites from the chemical reaction between benzene and peroxynitrite. Thus, peroxynitrite mediated hydroxylation and nitration of benzene during immune activation represent a novel in vivo mechanism for generation of proximal carcinogens of benzene.

Experimental study of oxidative DNA damage.

Animal experiments allow the study of oxidative DNA damage in target organs and the elucidation of dose-response relationships of carcinogenic and other harmful chemicals and conditions as well as the study of interactions of several factors. So far the effects of more than 50 different chemical compounds have been studied in animal experiments mainly in rats and mice, and generally with measurement of 8-oxodG with HPLC-EC. A large number of well-known carcinogens induce 8-oxodG formation in liver and/or kidneys. Moreover several animal studies have shown a close relationship between induction of dative DNA damage and tumour formation. In principle the level of oxidative DNA damage in an organ or cell may be studied by measurement of modified bases in extracted DNA by immunohistochemical visualisation, and from assays of strand breakage before and after treatment with repair enzymes. However, this level is a balance between the rates of damage and repair. Until the repair rates and capacity can be adequately assessed the rate of damage can only be estimated from the urinary excretion of repair products albeit only as an average of the entire body. A number of model compounds have been used to induce oxidative DNA damage in experimental animals. The hepatocarcinogen 2-nitropropane induces up to 10-fold increases in 8-oxodG levels in rat liver DNA. The level of 8-oxodG is also increased in kidneys and bone marrow but not in the testis. By means of 2-nitropropane we have shown correspondence between the increases in 8-oxodG in target organs and the urinary excretion of 8-oxodG and between 8-oxodG formation and the comet assay in bone marrow as well potent preventive effects of extracts of Brussels sprouts. Others have shown similar effects of green tea extracts and its components. Drawbacks of the use of 2-nitropropane as a model for oxidative DNA damage relate particularly to formation of 8-aminoguanine derivatives that may interfere with HPLC-EC assays and have unknown consequences. Other model compounds for induction of oxidative DNA damage, such as ferric nitriloacetate, iron dextran, potassium bromate and paraquat, are less potent and/or more organ specific. Inflammation and activation of an inflammatory response by phorbol esters or E. coli lipopolysaccharide (LPS) induce oxidative DNA damage in many target cells and enhance benzene-induced DNA damage in mouse bone marrow. Experimental studies provide powerful tools to investigate agents inducing and preventing oxidative damage to DNA and its role in carcinogenesis. So far, most animal experiments have concerned 8-oxodG and determination of additional damaged bases should be employed. An ideal animal model for prevention of oxidative DNA damage has yet to he developed.

2-Nitropropane-induced DNA damage in rat bone marrow.

DNA damage detected by the comet assay (single cell gel electrophoresis) and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) formation in DNA in the bone marrow has been studied in groups of 6 male Wistar rats treated with a single i.p. injection of the carcinogen 2-nitropropane (2-NP, 100 mg/kg body weight) or vehicle. Twenty-four hours after 2-NP the average tail length in the comet assay in bone marrow cells was increased from 1.46 +/- 0.27 to 9.61 +/- 1.56 microm (mean /- SD, p < 0.01), and 8-oxodG levels in the DNA were increased from 1.04 +/- 0.50 to 5.14 +/- 2.42 per 10(5) dG (p < 0.01). There was a close correlation between the comet tail length and the 8-oxodG level (r = 0.89, p < 0.05). The results indicate that 2-NP inflicts DNA damage in the bone marrow cells and thus could be leukemogenic.

Benzene-induced genotoxicity in mice in vivo detected by the alkaline comet assay: reduction by CYP2E1 inhibition.

The myelotoxic and genotoxic effects of benzene have been related to oxidative DNA damage after metabolism by CYP2E1. Single cell gel electrophoresis (alkaline comet assay) detects DNA damage and may thus be a convenient method for the study of benzene genotoxicity. Benzene exposure to NMRI mice as a single oral gavage at 40, 200 or 450 mg/kg resulted in dose-related DNA damage indicated by an increased comet tail length of peripheral blood lymphocytes and bone marrow nucleated cells sampled 6 h after exposure. After a dose of 40 mg/kg, there was a 1.6-fold increase of 'tail length' in bone marrow nucleated cells in comparison with the control (p < 0.01). There was no significant increase in DNA damage in peripheral blood lymphocytes in the same animals. At 200 mg/kg, the tail length was 4.8-fold and 4.0-fold increased in the two cell types, respectively (p < 0.01). At 450 mg/kg, the tail length was further increased to 5.4-fold and 6.6-fold of the control values, respectively (p < 0.01). Pretreatment with propylene glycol (5 microliters/g b.wt., twice with a 60-min interval), a selective CYP2E1 inhibitor, reduced the increase in the tail length by about half at all doses in both cell types (p < 0.01). By comparing our data with those from genotoxicity studies on benzene using other methods, we conclude that the 'alkaline comet assay' is a sensitive method to detect DNA damage induced by benzene. We also infer that CYP2E1 contributes, at least partly, to the formation of the 'comet'-inducing metabolites in the chosen cell types.