1 Department of Preventive Medicine and 2 Department of Internal Medicine, College of Medicine, Chungbuk National University, 3 Department of Obstetrics and Gynecology and 4 Department of Preventive Medicine and Institute of Environmental Medicine, College of Medicine, Seoul National University, Korea
5 To whom correspondence should be addressed at: Department of Preventive Medicine, College of Medicine, Chungbuk National University, 48 Gaeshin-dong, Heungduk-gu, Cheongju 361-763, Republic of Korea. Email: jwkang{at}pm.cbu.ac.kr
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Abstract |
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Key words: 1-hydroxypyrene/2-naphthol/PC Game Room/polycyclic aromatic hydrocarbon/testosterone
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Introduction |
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The PC Game Room users are exposed, in a closed space, to various hazardous agents including noise of electronic sound, poor lighting and ventilation, direct smoking and environmental tobacco smoke (ETS), etc. It is most probable that its excessive use will be associated with an elevated exposure to polycyclic aromatic hydrocarbon (PAH), even in non-smokers. Pyrene, one of the major components of PAH, can be absorbed via the respiratory tract as well as through the skin, and the urinary concentration of its major metabolite, 1-hydroxypyrene (1-OHP), has been utilized as a total PAH exposure marker (Jongeneelen et al., 1987). Naphthalene, another major component of PAH, is mainly absorbed by inhalation and metabolized to 2-naphthol; its urinary concentration has been applied as another PAH exposure marker (Kim et al., 1999
).
It is also probable that PC Game Room over-use disturbs sleep and circadian rhythms, which can affect the hormone levels with diurnal variation such as testosterone and LH. Most PC Game Room users are in the reproductive age; however, no association has yet been established between excessive PC Game Room use and potential adverse effects on sex hormonal status. The current cross-sectional study was based on the surveys conducted in PC Game Rooms and was designed to evaluate exposure to PAH and its possible adverse effects on testosterone levels. Urinary concentrations of 1-OHP and 2-naphthol were used as PAH exposure markers, and the correlations of PC Game Room use with PAH exposure and plasma testosterone concentrations were investigated in young male Koreans.
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Materials and methods |
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Questionnaire
Information was collected using a self-administered, structured questionnaire on demographic factors, PC Game Room use, smoking, alcohol intake and PAH exposure-related dietary factors. The duration of PC Game Room use was questioned as either less than 1 h per day, 12 h per day, or more than 2 h per day. Age was divided into 1519 years and 2024 years. Alcohol intake frequency was questioned as either Once a week or less or Twice a week or more. Smoking status was defined as current smoker or non-smoker. Body mass index (BMI) was calculated from height and weight.
Measurement of plasma testosterone and LH concentrations
Commercial kits were used to measure plasma testosterone (TESTO-CT2; CIS bio international, France) and LH (SPAC-S LH kit; Daiichi pharmaceutical, Japan) concentrations. The intra- and inter-assay coefficients of variation were 3.8 and 4.8% for testosterone and 2.8 and 3.7% for LH respectively.
Measurement of urinary 1-hydroxypyrene concentration (1-OHP) and 2-naphthol
Urinary concentrations of 1-OHP and 2-naphthol were measured as markers of exposure to PAH. The analysis of urinary 1-OHP was performed as described by Jongeneelen et al. (1987), and 2-naphthol as we previously described (Kim et al., 2001
).
Briefly, in a dark room, 1 ml urine samples were buffered with 100 ml 0.2 mol/l sodium acetate buffer (pH 5.0), and hydrolysed enzymatically using 10 ml of -glucuronidase with sulphatase activity (Sigma G-8076, USA), for 16 h at 37°C in a shaking water bath. After hydrolysis, 1.5 ml of acetonitrile was added and the samples were centrifuged at 10 000g for 10 min. An high-performance liquid chromatography system, consisting of a pump (Waters 600E; Millipore, USA), a variable fluorescence detector (RF-10AxL; Shimadzu, Japan), an automatic injector (L-2700; Hitachi, Japan), and an integrator (Chromatopac C-R3A; Shimadzu, Japan) was used throughout. A reverse phase column, 150 mm in length (TSK gel ODS-80TM; Tosoh, Japan), was used for the 1-OHP analysis, and a 250 mm-long reverse phase column (J'sphere ODS-H80; YMC, USA) for 2-naphthol analysis. The mobile phase comprised 60% acetonitrile for 1-OHP and 38% acetonitrile for 2-naphthol. The flow rate was 1 ml/min. Excitation/emission wavelengths used in the detection of 1-OHP and 2-naphthol were 242/388 and 227/355 nm respectively. Urinary 1-OHP and 2-naphthol concentrations were standardized versus urinary creatinine concentration.
Statistical analysis
The Statistical Analysis System (SAS) for Windows version 6.12 was used for data analysis. Urinary concentrations of 1-OHP and 2-naphthol, and plasma concentrations of testosterone and LH were log-transformed before statistical analysis. Mean plasma testosterone and LH levels were compared using analysis of variance (ANOVA) and t-test. 2-Test was applied to test the relationships between categorical variables. For analysis between continuous variables, correlation and multiple regression analysis were applied. Either 1-OHP or 2-naphthol was used as the indicator of PAH exposure in the multiple regression analysis.
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Results |
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In correlation analysis, age showed a strong positive correlation with plasma testosterone concentration (r=0.31, P=0.0001) (Table II). Plasma testosterone concentration showed a significant negative correlation with urinary 1-OHP (r=0.22, P=0.0012) and 2-naphthol (r=0.15, P=0.0308) concentrations. Of the two, urinary 1-OHP concentration showed a more prominent negative correlation with plasma testosterone concentration than urinary 2-naphthol concentration. No significant overall correlation was found between plasma testosterone and LH concentrations. However, when stratified with respect to the sample timing, plasma testosterone and LH levels were found to be correlated both in the noon sample group (n=126) (r=0.20, P=0.0280) and in the midnight sample group (n=82) (r=0.29, P=0.0075).
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The younger age group showed a more prominent reduction in plasma testosterone concentration with increasing duration of PC Game Room use than the older age group (r2=0.355, P=0.0301 versus r2=0.213, P=0.0001) (Figure 1). However, midnight testosterone level showed no significant correlations with increasing duration of PC Game Room use, in either age group.
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Discussion |
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Plasma testosterone concentrations in the younger age group (1519 years) were significantly lower than in the older age group (2024 years) (Table I). Testosterone concentrations start to increase with the initiation of puberty and reach adult levels by 17 years of age (Griffin and Wilson et al., 1998
). In the present study, the younger age subgroup was in the last stage of testosterone rise, which may explain the low plasma testosterone concentrations caused by immature testosterone release process. However, the clinical significance of this may be limited since all testosterone concentrations were in the physiological range.
Smoking status was not found to be significantly correlated with plasma testosterone concentrations by univariate analysis in our study (Table I). However, in the older age group (2024 years), the testosterone levels of smokers were significantly lower than those of non-smokers. In the younger age group (1519 years), the testosterone levels were not different in smokers and non-smokers; moreover, the lack of a difference in this group might have masked its significance when all ages were compared. One possible explanation for this observation is that more of the older age group subjects were smokers than in the younger age group (60.7 versus 54.0%), and that smokers show a more remarkable decrease in testosterone levels.
However, there is considerable debate over the correlation between smoking and blood testosterone levels in the literature. Some previous reports have claimed an association between smoking and reduced testosterone levels (Shaarawy and Mahmoud, 1982; Stefanick et al., 1987
; Sofikitis et al., 1995
; Hsieh et al., 1998
), whereas others have produced different results (Klaiber and Broverman, 1988
; Dai et al., 1981
; Field et al., 1994
; Handa et al., 1997; Tamimi et al., 2001
).
Correlation coefficients between plasma testosterone levels and urinary 1-OHP or 2-naphthol concentrations were 0.22 and 0.15 respectively (P<0.05) (Table II). Urinary 1-OHP concentration (representing PAH exposure) showed a significant negative correlation with plasma testosterone concentration after adjusting for other variables (Table III). This is consistent with previous reports on disturbed gonadal testosterone production by PAH exposure in animals (Monteiro et al., 2000; Evanson and Van Der Kraak, 2001
). However, the duration of PC Game Room use, itself, was not found to be an independent significant determinant of plasma testosterone level after adjustment (Table III). Thus, it is probable that PC Game Room use was associated with reduced plasma testosterone levels because of increased PAH exposure.
In the present study, plasma was sampled in the morning (noon samples) and at midnight, considering the diurnal variation of testosterone release. This timing of sampling was found to be the most significant determinant of plasma testosterone levels by multivariate analysis (Table III). The level of plasma testosterone was significantly higher in the noon sample group (Table I), which corresponds with the fact that testosterone release has a circadian rhythm showing a morning peak level.
Individuals vary in terms of their PAH absorption, metabolism and metabolite excretion rates. For example, as we previously reported, genetic polymorphisms of CYP1A1, GSTM1 and GSTT1 are significant determinants of urinary 1-OHP concentration, as is CYP2E1 for urinary 2-naphthol concentration (Nan et al., 2001). In the present study, we did not examine the genetic polymorphisms of these metabolic enzymes.
Alcohol intake showed no significant correlations with plasma testosterone concentration (Table I). There is also controversy concerning the effect of alcohol on blood testosterone levels. Some authors have reported that alcohol intake is related to plasma testosterone concentrations (King et al., 1995; Heikkonen et al., 1996
; Frias et al., 2000
), whereas others did not (Dai et al., 1981
; Handa et al., 1997; Hsieh et al., 1998
). BMI showed no correlations with plasma testosterone concentrations by univariate analysis (Table II); however, a negative correlation was found after adjustment (Table III). Our positive finding is supported by other previous reports (Dai et al., 1981
; Stefanick et al., 1987
; Handa et al., 1997), and this is considered to be probably due to the peripheral conversion of testosterone and a decrease in testosterone-binding globulin (Griffin and Wilson et al., 1998
).
It is noteworthy that, when stratified by age, the younger age group (1519 years) showed a more prominent decrease in plasma testosterone concentration than the older age group (2024 years), with increasing duration of PC Game Room use (Figure 1). The younger the subjects are, the more profoundly affected are the morning (noon) blood testosterone levels, by possible environmental hazards at PC Game Room. This vulnerability of the younger age group requires further longitudinal studies. However, this difference was not observed in the night samples (Figure 1).
In conclusion, our data suggest that the longer the PC Game Room use, the lower the testosterone levels in young male Koreans, which might be mediated by PAH exposure. This effect was found to be more prominent in the younger age group. Further follow-up studies will be necessary to elucidate the temporal relationships between the relevant variables. Comparative data between conventional PC Game Room use and smoke-free PC Game Room use would provide evidence on the role of smoke. In addition, the role of genetic polymorphisms in xenobiotic-metabolizing enzymes in men of susceptible age should be investigated.
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Acknowledgements |
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References |
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Submitted on May 20, 2004; resubmitted on September 28, 2004; accepted on November 11, 2004.
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