Nightshift work can induce oxidative DNA damage: a pilot study.

BACKGROUND: Regular sleep is very important for human health; however, the short-term and long-term effects of nightshift with sleep deprivation and disturbance on human metabolism, such as oxidative stress, have not been effectively evaluated based on a realistic cohort. We conducted the first long-term follow-up cohort study to evaluate the effect of nightshift work on DNA damage. METHODS: We recruited 16 healthy volunteers (aged 33 ± 5 years) working night shifts at the Department of Laboratory Medicine at a local hospital. Their matched serum and urine samples were collected at four time points: before, during (twice), and after the nightshift period. The levels of 8-oxo-7,8-dihydroguanosine (8-oxoG) and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), two important nucleic-acid damage markers, were accurately determined based on a robust self-established LC‒MS/MS method. The Mann-Whitney U or Kruskal-Wallis test was used for comparisons, and Pearson's or Spearman's correlation analysis was used to calculate the correlation coefficients. RESULTS: The levels of serum 8-oxodG, estimated glomerular filtration rate-corrected serum 8-oxodG, and the serum-to-urine 8-oxodG ratio significantly increased during the nightshift period. These levels were significantly higher than pre-nightshift work level even after 1 month of discontinuation, but no such significant change was found for 8-oxoG. Moreover, 8-oxoG and 8-oxodG levels were significantly positively associated with many routine biomarkers, such as total bilirubin and urea levels, and significantly negatively associated with serum lipids, such as total cholesterol levels. CONCLUSION: The results of our cohort study suggested that working night shifts may increase oxidative DNA damage even after a month of discontinuing nightshift work. Further studies with large-scale cohorts, different nightshift modes, and longer follow-up times are needed to clarify the short- and long-term effects of night shifts on DNA damage and find effective solutions to combat the negative effects.

8-Hydroxy-2'-deoxyguanosine (8-OHdG) as a biomarker of oxidative DNA damage induced by occupational exposure to nanomaterials: a systematic review.

In nuclear and mitochondrial DNA, 8-hydroxy-2'-deoxyguanosine (8-OHdG) is one of the predominant forms of reactive oxygen species (ROSs) lesions, which commonly used as a biomarker for oxidative stress. Studies showed that the different nanomaterials can induce toxicity by ROSs in human body. So, this study is going to review the studies about oxidative DNA damage caused by occupational exposure to nanomaterials, using 8-OHdG biomarker.Systematic review was managed based on Cochrane systematic review guideline. Literature search was conducted in scientific databases with the main terms of "biomarkers," "biological markers," combined with "occupational exposure" and "nanomaterials." All papers in the field of occupational exposure to nanomaterials until 2020 December were included. To evaluate the quality and bias of studies, GRADE method (Grading of Recommendations, Assessment, Development, and Evaluation) was used.Two hundred twenty-six studies were primarily achieved. By considering the inclusion criteria, overall 8 articles were selected. The majority of the studies were classified as the moderate quality studies (six studies). Also, the study-level bias was critical. This review shows that there is a significant relationship between job title and amount of produced nanomaterials and the existence of 8-OHdG. Also, the levels of 8-OHdG can be measured in urine, blood, and inhalation samples by instrumental procedures.Oxidative damages are an important threat for workers exposed to nanomaterial. Blood and EBC 8-OHdG level can be introduced as a biomarker for metal nanomaterials, but urinary 8-OHdG needs to be taken with caution. So, it is recommended that evaluation not be solely based on one biomarker.

Detection of an alcohol-associated cancer marker by single-molecule quantum sequencing.

Alcoholic beverages are a well-known risk factor for cancer. N2-Ethyl-2'-deoxyguanosine (N2-Et-dG) is a promising biomarker for alcohol-associated cancers. However, the lack of a convenient detection method for N2-Et-dG hinders the development of practical DNA damage markers. Herein, we develop a detection method for N2-Et-dG using a single-molecule quantum sequencing (SMQS) method and machine learning analysis. Our method succeeded in discriminating between N2-Et-dG and dG with an accuracy of 99%, using 20 signals. Our developed method quantified the mixing ratio of N2-Et-dG from a mixed solution of N2-Et-dG and dG. It is shown that our method has the potential to facilitate the development of DNA damage markers, and thus the early detection and prevention of cancers.

Independent Generation and Time-Resolved Detection of 2'-Deoxyguanosin-N2-yl Radicals.

Guanine radicals are important reactive intermediates in DNA damage. Hydroxyl radical (HO. ) has long been believed to react with 2'-deoxyguanosine (dG) generating 2'-deoxyguanosin-N1-yl radical (dG(N1-H). ) via addition to the nucleobase π-system and subsequent dehydration. This basic tenet was challenged by an alternative mechanism, in which the major reaction of HO. with dG was proposed to involve hydrogen atom abstraction from the N2-amine. The 2'-deoxyguanosin-N2-yl radical (dG(N2-H). ) formed was proposed to rapidly tautomerize to dG(N1-H). . We report the first independent generation of dG(N2-H). in high yield via photolysis of 1. dG(N2-H). is directly observed upon nanosecond laser flash photolysis (LFP) of 1. The absorption spectrum of dG(N2-H). is corroborated by DFT studies, and anti- and syn-dG(N2-H). are resolved for the first time. The LFP experiments showed no evidence for tautomerization of dG(N2-H). to dG(N1-H). within hundreds of microseconds. This observation suggests that the generation of dG(N1-H). via dG(N2-H). following hydrogen atom abstraction from dG is unlikely to be a major pathway when HO. reacts with dG.

The roles of polymerases ν and θ in replicative bypass of O6- and N2-alkyl-2'-deoxyguanosine lesions in human cells.

Exogenous and endogenous chemicals can react with DNA to produce DNA lesions that may block DNA replication. Not much is known about the roles of polymerase (Pol) ν and Pol θ in translesion synthesis (TLS) in cells. Here we examined the functions of these two polymerases in bypassing major-groove O6-alkyl-2'-deoxyguanosine (O6-alkyl-dG) and minor-groove N2-alkyl-dG lesions in human cells, where the alkyl groups are ethyl, n-butyl (nBu), and, for O6-alkyl-dG, pyridyloxobutyl. We found that Pol ν and Pol θ promote TLS across major-groove O6-alkyl-dG lesions. O6-alkyl-dG lesions mainly induced G→A mutations that were modulated by the two TLS polymerases and the structures of the alkyl groups. Simultaneous ablation of Pol ν and Pol θ resulted in diminished mutation frequencies for all three O6-alkyl-dG lesions. Depletion of Pol ν alone reduced mutations only for O6-nBu-dG, and sole loss of Pol θ attenuated the mutation rates for O6-nBu-dG and O6-pyridyloxobutyl-dG. Replication across the two N2-alkyl-dG lesions was error-free, and Pol ν and Pol θ were dispensable for their replicative bypass. Together, our results provide critical knowledge about the involvement of Pol ν and Pol θ in bypassing alkylated guanine lesions in human cells.

Effects of Urea on the Reactions of Nucleosides with Hypobromous Acid.

Hypobromous acid (HOBr) is generated not only by eosinophils but also by neutrophils in the presence of Br- at the plasma concentration. Reactivity of HOBr is greatly modulated by coexistent compounds such as amines and amides. In this study, we investigated effects of urea in the reaction of nucleosides with HOBr. When nucleosides were incubated with HOBr without urea in potassium phosphate buffer at pH 7.4 and 37°C, the reactions almost completed within 10 min, with consumptions in the order of 2'-deoxyguanosine > 2'-deoxycytidine > 2'-deoxythymidine > 2'-deoxyadenosine, generating 8-bromo-2'-deoxyguanosine and 5-bromo-2'-deoxycytidine. In the presence of urea, the reaction of nucleosides with HOBr was relatively slow, continuing over several hours. When HOBr was preincubated without urea in potassium phosphate buffer at pH 7.4 and 37°C for 48 h, the preincubated HOBr solution did not react with nucleosides. However, a similar preincubated solution of HOBr with urea reacted with nucleosides to generate 8-bromo-2'-deoxyguanosine and 5-bromo-2'-deoxycytidine. These results imply that a reactive bromine compound with a long life, probably bromourea, is generated by HOBr in neutral urea solution and reacts with nucleosides, resulting in brominated nucleosides.

The Role of DNA Repair Glycosylase OGG1 in Intrahepatic Cholangiocarcinoma.

BACKGROUND/AIM: The effects of oxidative stress on various carcinomas were reported in previous studies, but those in intrahepatic cholangiocarcinoma (ICC) have not been fully elucidated. The purpose of this study was, thus, to reveal the effects of oxidative DNA damage and repair enzymes on ICC. MATERIALS AND METHODS: The levels of 8-hydroxydeoxyguanosine (8-OHdG) and 8-OHdG DNA glycosylase (OGG1) were immunohistochemically evaluated in specimens resected from 63 patients with ICC. RESULTS: Low OGG1 expression was related to tumour depth T4 (p=0.04), venous invasion (p=0.0005), lymphatic vessel invasion (p=0.03), and perineural invasion (p=0.03). Compared to the high-OGG1-expression group, patients with low OGG1 expression had a significantly poorer prognosis (overall survival: p=0.04, recurrence-free survival: p=0.02). Unlike for OGG1, the expression levels of 8-OHdG showed no association with prognosis. CONCLUSION: Oxidative DNA damage and DNA repair enzymes may be closely related to ICC progression.

Electrochemical analysis of 8-hydroxy-2'-deoxyguanosine with enhanced sensitivity based on exonuclease-mediated functional nucleic acid.

In this work, an electrochemical method for sensitive analysis of 8-hydroxy-2'-deoxyguanosine, a key biomarker that is widely used to study oxidative injury-related diseases, is proposed based on exonuclease-mediated functional nucleic acid. In the design, exonuclease can not only distinguish the existence of target, but also suppress the background noise, thus the sensitivity can be enhanced. Moreover, DNAzyme designed in the functional nucleic acid can further improve the sensitivity of the analysis during signal generation process. Therefore, exonuclease-mediated functional nucleic acid may ensure high sensitivity of the assay. Further studies reveal that the detection of 8-hydroxy-2'-deoxyguanosine can be achieved with a linearity from 0.01 nM to 7.0 µM and a detection limit of 6.82 pM. The new method has also been successfully applied to the determination of 8-OHdG in urine with good results, indicating its great potential for practical use.

Predictive value of the ratio of 8­hydroxydeoxyguanosine levels between cancerous and normal tissues in patients with stage II/III colorectal cancer.

Reactive oxygen species (ROS) accumulation is known to induce carcinogenesis and accelerate cancer progression. 8­Hydroxydeoxyguanosine (8­OHdG) is a specific marker of ROS­mediated DNA damage. Therefore, we analysed 8­OHdG levels in cancerous and normal tissue DNA via enzyme­linked immunosorbent assay (ELISA) using 97 tissue specimens obtained from surgically­treated patients with stage II/III colorectal cancer (CRC). Additionally, 8­OHdG levels in these tissues were also assessed via quantitative immunohistochemistry (qIHC). To eliminate individual background variables, the ratio of 8­OHdG levels between cancerous and normal tissues was calculated using both techniques. A comparative analysis demonstrated that the 8­OHdG ratio in DNA was significantly correlated with both lymph node metastasis and lymphatic invasion. Multivariate analysis revealed that a high 8­OHdG ratio in DNA was independently correlated with poor prognosis. These results suggest that the 8­OHdG ratio in DNA reflects ROS­induced cancer progression. Conversely, a low 8­OHdG ratio as estimated via qIHC was an independent factor for poor prognosis. In Kaplan­Meier analysis, the combination of a high 8­OHdG ratio in DNA (ELISA) and a low 8­OHdG ratio in cytoplasm (qIHC) was associated with markedly worse patient prognosis than other combinations. Combined evaluation of the 8­OHdG ratio using ELISA and qIHC may be pivotal for predicting surgical outcomes for patients with stage II/III CRC.

Assessment of Genotoxicity of Levosimendan in Human Cultured Lymphocytes.

BACKGROUND AND OBJECTIVE: Levosimendan is a positive inotropic and a vasodilator agent with pleotropic characteristics that include antioxidation, anti-inflammation and smooth muscle vasodilation. METHODS: In this study, the effects of levosimendan (0, 0.1, 1, 10, and 20 µg/ml) on oxidative DNA damage and sister-chromatid exchanges (SCEs) were evaluated in human cultured lymphocytes. RESULTS: The results showed that levosimendan increased the frequency of SCEs in all examined concentrations (P<0.01) except for 0.1 µg/ml. On the other hand, levosimendan did not induce oxidative DNA damage as measured by the 8-OHdG biomarker (P > 0.05). In addition, neither mitotic arrest nor proliferation index was affected by levosimendan at all examined doses (P > 0.05). CONCLUSION: In conclusion, levosimendan might be associated with increases in sister-chromatid exchanges in cultured human lymphocytes. In vivo studies are required to confirm the present findings.

An ultra-high-performance liquid chromatography tandem mass spectrometry method for oxidative stress biomarker analysis in wastewater.

Reported herein is the development of an analytical method for the detection of four oxidative stress biomarkers in wastewater using ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS) and solid phase extraction (SPE). The following four biomarkers of oxidative stress and lipid peroxidation have been investigated: hydroxynonenal-mercapturic acid (HNE-MA), 8-iso-prostglandin F2beta (8-iso-PGF2ß), 8-nitroguanine (8-NO2Gua) and 8-hydroxy-2-deoxyguanosine (8-OHdG). The method showed very good performance: accuracy (> 87%), precision (> 90%), method quantification limits (1.3-3.0 ng L-1) and biomarker stability in wastewater in the case of HNE-MA, 8-OHdG and 8-iso-PGF2ß. In contrast, 8-NO2Gua was found to be less stable in wastewater, which affected its method performance: accuracy (> 63%), precision (> 91%) and method quantification limits (85.3 ng L-1). Application of the developed method resulted in, for the first time, HNE-MA being successfully observed and quantified within wastewater over a study period of a week (displayed average daily loads per capita of 48.9 ± 4.1 mg/1000/people/day). 8-iso-PGF2ß was detected with good intensity but could not be quantified due to co-elution with other isomers. 8-OHdG was detected, albeit at < MQL. This study demonstrates the potential for expanding on the possible endogenous biomarkers of health used in urban water fingerprinting to aid in measuring health in near-real time on a community-wide scale.

Evaluating clinical and laboratory effects of ozone in non-surgical periodontal treatment: a randomized controlled trial.

OBJECTIVE: This study aims to evaluate the clinical and biochemical (oxidative stress and pro-inflammatory mediators) effects of the gaseous ozone use accompanied by scaling and root planning (SRP) in periodontal treatment. MATERIAL AND METHODS: The study population consisted of 40 patients with chronic periodontitis (CP) randomly sorted into two groups of 20. The experimental group received SRP plus 3 watts gaseous ozone in two separate applications five days apart, whereas the control group received SRP plus placebo. Clinical periodontal parameters were assayed and saliva samples were taken before the initial and one month after the second treatment. Periodontal examination assessed plaque index (PI), gingival index (GI), probing depth, and clinical attachment level (CAL). Total antioxidant status (TAS), total oxidant status (TOS), nitric oxide (NO), 8-hydroxy-2'-deoxyguanosine (8-OHdG), myeloperoxidase (MPO), glutathione (GSH), malondialdehyde (MDA), and transforming growth factor-beta (TGF-ß) levels were evaluated from saliva samples. RESULTS: Changes following treatment in PI, GI, probing depth, and CAL scores were similar for both groups (p>0.05). Of note, TGF-ß levels were observed to be higher in the treatment group than in controls (p<0.05). Changes in 8-OHdG, TAS, TOS, NO, MPO, GSH and MDA levels, however, were not significantly different between groups (p>0.05). CONCLUSION: The findings of this study indicate that SRP plus gaseous ozone versus SRP alone does not correlate to a significant improvement in periodontal recovery.

Neurophysiological responses in the brain tissues of rainbow trout (Oncorhynchus mykiss) treated with bio-pesticide.

The aim of this study was to investigate neurophysiological responses in rainbow trout brain tissue exposed to natural/botanical pesticides. Fish were exposed to botanical and synthetic pesticides over a 21-day period. At the end of the treatment period, oxidative DNA damage (indicated by 8-OHdG (8-hydroxy-2'-deoxyguanosine), AChE activity (acetylcholinesterase) and transcriptional parameters (gpx (glutathione peroxidase), sod (superoxide dismutase), cat (catalase), HSP70 (heat shock protein 70) and CYP1A (cytochromes P450)) was investigated in control and application groups. Our results indicated that brain AChE activities decreased very significantly in fish treated with both insecticide types when compared with control (p < 0.05). 8-OHdG activity increased in a dose/time-dependent situation in the brain tissues of Oncorhynchus mykiss (p < 0.05). In addition, with regards to gene expression, gpx sod and, cat expressions were down-regulated, whereas CYP1A and HSP70 gene expression were up-regulated in fish treated with both insecticides when compared to the control group (p < 0.05). The data for this study suggests that bio-pesticides can cause neurophysiological changes in fish brain tissue.

Evaluating clinical and laboratory effects of ozone in non-surgical periodontal treatment: a randomized controlled trial

Abstract Objective: This study aims to evaluate the clinical and biochemical (oxidative stress and pro-inflammatory mediators) effects of the gaseous ozone use accompanied by scaling and root planning (SRP) in periodontal treatment. Material and Methods: The study population consisted of 40 patients with chronic periodontitis (CP) randomly sorted into two groups of 20. The experimental group received SRP plus 3 watts gaseous ozone in two separate applications five days apart, whereas the control group received SRP plus placebo. Clinical periodontal parameters were assayed and saliva samples were taken before the initial and one month after the second treatment. Periodontal examination assessed plaque index (PI), gingival index (GI), probing depth, and clinical attachment level (CAL). Total antioxidant status (TAS), total oxidant status (TOS), nitric oxide (NO), 8-hydroxy-2'-deoxyguanosine (8-OHdG), myeloperoxidase (MPO), glutathione (GSH), malondialdehyde (MDA), and transforming growth factor-beta (TGF-β) levels were evaluated from saliva samples. Results: Changes following treatment in PI, GI, probing depth, and CAL scores were similar for both groups (p>0.05). Of note, TGF-β levels were observed to be higher in the treatment group than in controls (p<0.05). Changes in 8-OHdG, TAS, TOS, NO, MPO, GSH and MDA levels, however, were not significantly different between groups (p>0.05). Conclusion: The findings of this study indicate that SRP plus gaseous ozone versus SRP alone does not correlate to a significant improvement in periodontal recovery.

Polybrominated Diphenyl Ethers Quinone Induced Parthanatos-like Cell Death through a Reactive Oxygen Species-Associated Poly(ADP-ribose) Polymerase 1 Signaling.

Polybrominated diphenyl ethers (PBDEs) are emerging organic environmental pollutants, which were accused of various toxic effects. Here, we studied the role of a potential PBDEs quinone metabolite, PBDEQ, on cytotoxicity, oxidative DNA damage, and the alterations of signal cascade in HeLa cells. PBDEQ exposure leads to reactive oxygen species (ROS) accumulation, mitochondrial membrane potential (MMP) loss, lactate dehydrogenase (LDH) release, increasing terminal transferase-mediated dUTP-biotin nick end labeling (TUNEL) positive foci, and the elevation of apoptosis rate. Furthermore, we showed PBDEQ exposure result in increased DNA migration, micronucleus frequency, and the promotion of 8-OHdG and phosphorylation of histone H2AX (γ-H2AX) levels. Mechanism study indicated that PBDEQ caused poly(ADP-ribose) polymerase 1 (PARP-1) activation and apoptosis-inducing factor (AIF) nuclear translocation. All together, these results confirmed the occurrence of parthanatos-like cell death upon PBDEQ exposure.

Strong impact of sulfotransferases on DNA adduct formation by 4-aminobiphenyl in bladder and liver in mice.

Bladder cancer risk is 3-4 times higher in men than women, but the reason is poorly understood. In mice, male bladder is also more susceptible than female bladder to 4-aminobiphenyl (ABP), a major human bladder carcinogen; however, female liver is more susceptible than male liver to ABP. We investigated the role of sulfotransferase (Sult) in gender-related bladder and liver susceptibility to ABP. Sulfation reactions of aromatic amine bladder carcinogens catalyzed by Sult may generate highly unstable and toxic metabolites. Therefore, liver Sult may decrease bladder exposure to carcinogens by promoting their toxic reactions in the liver. Notably, the expression of several liver Sults is suppressed by androgen in male mice. Here, we show that two Sults are critical for gender-related bladder susceptibility to ABP in mice. We measured tissue level of N-(deoxyguanosin-8-yl)-4-aminobiphenyl (dG-C8-ABP), a principal ABP-DNA adduct, as readout of tissue susceptibility to ABP. We identified Sutl1a1 and to a lesser extent Sult1d1 as Sults that promote dG-C8-ABP formation in hepatic cells. In mice, gender gap in bladder susceptibility to ABP was narrowed by knocking out Sult1a1 and was almost totally eliminated by knocking out both Sutl1a1 and Sult1d1. This was accompanied by dramatic decrease in ABP genotoxicity in the liver (>97%). These results show the strong impact of the Sults on bladder and liver susceptibility to a human carcinogen. Because liver expression of both Sult1a1 and Sutl1d1 is suppressed by androgen in male mice, our results suggest that androgen renders bladder more exposed to ABP in male mice by suppressing Sult-mediated ABP metabolism in liver, which increases bladder delivery of carcinogenic metabolites.

Improving the fluorometric determination of the cancer biomarker 8-hydroxy-2'-deoxyguanosine by using a 3D DNA nanomachine.

The authors describe a fluorometric method for improving the determination of the cancer biomarker 8-hydroxy-2'-deoxyguanosine (8-OHdG). A nicking endonuclease (NEase)-powered 3-D DNA nanomachine was constructed by assembling hundreds of carboxyfluorescein-labeled single strand oligonucleotides (acting as signal reporter) and tens of swing arms (acting as single-foot DNA walkers) on a gold nanoparticle (AuNP). The activity of this DNA nanomachine was controlled by introducing the protecting oligonucleotides. In the presence of aptamer against 8-OHdG, the protecting oligonucleotides are removed from the swing arms by toehold-mediated strand displacement reaction. In the next step, detached DNA walker hybridizes to the labelled DNA so that the DNA nanomachine becomes activated. Special sequences of signal reporter in the formed duplex can be recognized and cleaved by NEase. As a result, the DNA walker autonomously and progressively moves along the surface of the AuNP, thereby releasing hundreds of signal reporters and causing a rapid increase in green fluorescence. This 3-D nanomachine is highly efficient because one aptamer can release hundreds of signal reporters. These unique properties allowed for the construction of a DNA nanomachine-based method for sensitively detecting 8-OHdG in concentrations as low as 4 pM. This is three orders of magnitude lower compared to previously reported methods. Graphical abstract Schematic of a fluorometric method for determination of the cancer biomarker 8-hydroxy-2'-deoxyguanosine. A nicking endonuclease powered 3D-DNA nanomachine was used to improve the sensitivity. Limit of detection is three orders of magnitude lower than reported methods.

Effect of copper nanoparticles on the mineral content of tissues and droppings, and growth of chickens.

We hypothesised that copper nanoparticles (NanoCu), because of their high physicochemical reactivity and bioavailability, could be used in much smaller quantities than bulk Cu, consequently reducing excretion of Cu into the environment. The objective of the study was to evaluate the effects of various levels of NanoCu on the development and growth of broiler chickens, in order to establish an optimum level of NanoCu dietary supplementation. Broiler chickens were randomly divided into five groups of 10 birds each. The control group received 7.5 mg Cu/kg feed (standard level) as CuSO4, while groups fed with complexes of NanoCu and starch received 25%, 50%, 75% and 100% of the standard level of Cu used in the control group. Chicken growth and excretion of Cu, Fe and Zn were measured during the growth period from d 7 to 42. At d 42, the slaughter characteristics, the content of Cu, Fe and Zn in the breast muscle and liver, and the oxidative status were analysed. The results indicate that using NanoCu can reduce the standard level of Cu from CuSO4 supplementation by 75% without jeopardising animal growth, and at the same time significantly decreasing Cu excretion into the environment.

[Sperm DNA damage is not related to seminal leukocyte subsets in infertile males with asymptomatic genital tract infection].

OBJECTIVE: To study the correlation of leukocyte subsets with sperm DNA damage in the semen of infertile men with asymptomatic genital tract infection (AGTI). METHODS: This study included 111 infertile males with AGTI. After routine semen analysis, we determined the concentration of CD45+ leukocytes in the semen by immunocytochemistry, measured the concentrations of CD14+ cells of the mononuclear / macrophagic system and activated macrophages and HLA-DR+ cells in the semen by flow cytometry, and examined the sperm DNA fragmentation index (DFI) and the expression of 8-hydroxy-2'-deoxyguanosine (8-OHdG) by TUNEL assay. Then we analyzed the correlation of seminal leukocyte subsets with sperm DNA damage and routine semen parameters. RESULTS: The concentration of CD45+ leukocytes was correlated significantly with those of CD14+ and HLA-DR+ cells in the semen (P <0.01), but not that of leukocyte subsets with routine semen parameters, sperm DFI, or the percentage of 8-OHdG-positive cells. The percentage of 8-OHdG-positive sperm was correlated positively with the sperm DFI (r = 0.48, P<0.01) but negatively with sperm concentration (r = －0.44, P <0.01). After adjusted for age, abstinence time and cigarette smoking, the percentage of 8-OHdG-positive sperm was correlated independently with sperm concentration (ß = －0.25, P = 0.008) and DFI (ß = 0.23, P = 0.05). CONCLUSIONS: Sperm DNA damage is associated with poor semen quality but not with seminal leukocyte subsets in infertile males with asymptomatic genital tract infection.