Semen quality of male smokers and nonsmokers in infertile couples

Transcription

1 FERTILITY AND STERILITY VOL. 79, NO. 2, FEBRUARY 2003 Copyright 2003 American Society for Reproductive Medicine Published by Elsevier Science Inc. Printed on acid-free paper in U.S.A. Semen quality of male smokers and nonsmokers in infertile couples Robert Künzle, M.D., a Michael D. Mueller, M.D., a Willy Hänggi, M.D., a Martin H. Birkhäuser, M.D., a Heinz Drescher, M.D., b and Nick A. Bersinger, Ph.D. a University of Berne, Berne, and MCL Laboratories, Dudingen, Switzerland Objective: To investigate the effect of cigarette smoking on main sperm variables. Design: Cohort study. Setting: Men attending the andrology laboratory in the context of infertility investigation in the couple. Patient(s): Eight hundred thirty-nine smokers and 1,266 non-smokers were enrolled; 655 smokers and 1,131 nonsmokers fulfilled the inclusion criteria and were compared. Main Outcome Measure(s): Standard clinical analysis of semen. Result(s): Cigarette smoking was associated with a significant decrease in sperm density ( 15.3%), total sperm count ( 17.5%), total number of motile sperm ( 16.6%), and citrate concentration ( 22.4%). The percentage of normal forms was significantly reduced in smokers, and sperm vitality, ejaculate volume, and fructose concentration were slightly but nonsignificantly affected. Conclusion: Cigarette smoking is associated with reduced semen quality. (Fertil Steril 2003;79: by American Society for Reproductive Medicine.) Key Words: Smoking, semen quality, fertility, lifestyle Received February 7, 2002; revised and accepted June 17, Reprint requests: Nick A. Bersinger, Ph.D., Reproductive and Perinatal Medicine Research Laboratory, Inselspital KKL G3-825, Berne CH-3010, Switzerland (FAX: ; nick. bersinger@dkf2.unibe.ch). a Department of Obstetrics and Gynaecology, University of Berne. b MCL Laboratories /03/$30.00 PII S (02) According to the World Health Organization, approximately one third of the world s population older than 15 years of age smokes. Tobacco smoking is recognized as a general health hazard, and evidence indicates that in both men and women, cigarette smoking affects reproductive health more than does consumption of caffeine or alcohol (1). The combustion of tobacco yields about 4,000 chemical compounds. In the gaseous fraction, carbon monoxide, nitrogen oxide, ammonia, and hydrocarbons are found, whereas the main component of the particulate phase is composed of aggregates of nicotine. Polycyclic aromatic hydrocarbons activate a proapoptotic protein in mice (2), which leads to damage of oocytes and to reduced fertility. Given that cigarette smoke contains more than 30 chemical agents known to be mutagens, aneugens, or carcinogens in model systems, direct deleterious effects on human embryos and female and male germ cells are plausible (3). The mechanisms by which tobacco smoke affects spermatozoa are poorly understood. The effect of cigarette smoke on sperm function was demonstrated almost 2 decades ago (4) and has been confirmed by various investigators using classic microscopic analysis (5 8) or computer-assisted sperm analysis, which provides detailed information on sperm kinetics (9, 10). Some of the studies focused on the relation between cigarette smoking and principal semen analysis variables, such as density, progressive motility, and morphology; results were conflicting, particularly with regard to the sperm concentration and morphology (5, 6, 11, 12). This inconsistency in the literature, which led to a wide range of conclusions about the effects of smoking on male fertility, appears to stem from the small samples in some early studies (5, 6) and the differences in the thresholds of daily smoked cigarettes used to distinguish smokers from nonsmokers. We therefore sought to investigate the effect of cigarette smoking on main semen variables in large, well-defined groups of smokers and nonsmokers, in which the criteria defining smokers was strict ( 1 cigarette/d). 287

2 MATERIALS AND METHODS Study Participants and Sample Collection Men attending the andrology laboratory between 1991 and 1997 for reduced fecundity in the couple were included. Only the first sperm evaluation was used in this analysis. Samples from patients who had had vasectomy or vasostomy were excluded. Before semen analysis, a questionnaire was distributed to obtain information on age, smoking habits, alcohol use (regular, irregular, or total abstinence), and use or abuse of other substances and drugs (yes or no). Patients were also asked about history of orchitis, testicular trauma, sexually transmitted disease, varicocele, inguinal hernia operation and cryptorchism. A total of 2,105 sperm samples from 839 smokers and 1,266 nonsmokers fulfilled the inclusion criteria and were included in the study. Approval by the institutional review board was not necessary because all analyses were part of the routine infertility investigation. Semen samples were collected by ejaculation into a special sterile container (Telast SA, Préverenges, Switzerland) after 2 to 7 days of sexual abstinence. Analysis was performed within 2 hours after collection. Semen Analysis Semen analysis consisted of determination of sample volume, sperm density (concentration), progressive motility, vitality (eosin exclusion), morphology, ph, and concentrations of citrate and fructose. Standard clinical semen analysis was performed according to World Health Organization criteria (13). Motility was assessed by computer-assisted sperm analysis using an ATS 20-J.C. analyser (Diffusion Internationale, Fresnele, France) and was based on the principle of negative phase contrast microscopy. Oligospermic samples were manually analyzed in a Makler chamber by using an Olympus BH-2 microscope (Olympus, Volketswil, Switzerland). Morphology was assessed under the microscope after staining with a kit based on fast green, eosin, and thiacin (Diff-Quick; Dade Diagnostica, Dudingen, Switzerland). Citrate and fructose were measured as tests of function of the prostate and seminal vesicles, respectively. Statistical Analyses The associations between smoking status and lifestyle or medical history, which may introduce a bias in case of positive correlation, were tested by 2 2 contingency tables using Fisher s exact test. The effect of smoking on sperm and ejaculate quality parameters was analyzed by using the Student t-test if the SDs were similar or the Welch approximate test if SDs were dissimilar. RESULTS The initial study sample (available semen analysis data with returned questionnaires) consisted of 2,105 participants, of whom 839 (39.9%) were smokers and 1,266 (60.1%) were nonsmokers. The two groups were first analyzed for differences in and associations with lifestyle characteristics and disease history. The response rate for any given question was similar in the two groups, that is, no bias was introduced owing to the proportion of patients who did not respond to a particular question. Statistically significant associations were observed between smoking and alcohol consumption (P.0021) or past sexually transmitted disease (P.0204). Both alcohol consumption and past sexually transmitted disease were found to be more frequent in smokers. For this reason, the few patients with past sexually transmitted disease (41 smokers and 36 nonsmokers) were excluded. After we excluded men who consumed one or more standard drinks per day (130 smokers and 81 nonsmokers) and those who did not respond to the question on alcohol use (31 men: 13 smokers and 18 nonsmokers), the sample consisted of 655 smokers (36.7%) and 1,131 nonsmokers (63.3%). This sample comprised a large majority of social drinkers (88.4%) and nondrinkers. The association between smoking and moderate alcohol intake remained statistically significant; however, moderate drinking has been shown to have no negative effect on fertility in men (1, 11) or in women (1, 14). Varicocele compromises sperm quality (15). In our sample, nonsmokers had a more frequent incidence of past varicocele (which was surgically treated in all cases); however, the numbers of affected participants was low (31 of 379 responders) and nonsignificant. Thus, these individuals were retained in the sample. Smokers were slightly but significantly younger than nonsmokers ( and years; P.0010, Student t-test). Table 1 shows the clinical history and characteristics of the final sample. Table 2 shows semen quality variables. Statistical analysis demonstrated significantly lower semen quality in smokers compared with nonsmokers. Sperm concentration was also strongly affected. Ejaculate volume was not reduced, and progressive motility was slightly but nonsignificantly reduced. The total number of progressively motile sperm differed substantially and significantly between smokers and non-smokers (Table 2). Smoking had deleterious effects on sperm morphology. The ph of ejaculate from smokers was significantly higher than that of ejaculate from nonsmokers, as also reflected by a reduced citrate concentration. Smoking did not affect the fructose concentration. After logarithmic transformation of the absolute numbers (sperm density, total sperm count, total motile sperm, and citrate and fructose concentrations) from raw data and subsequent statistical analysis, the significance of the observed effects became more pronounced. 288 Künzle et al. Cigarette smoking reduces semen quality Vol. 79, No. 2, February 2003

3 TABLE 1 Characteristics of the study sample. Characteristic No. of smokers (n 655) DISCUSSION No. of nonsmokers (n 1,131) P value a Moderate alcohol consumption Yes No Cryptorchidism Responded Yes No Operation for hernia Responded Yes No Use of drugs or medicines Responded Yes No Mumps Responded Yes No Testicular injury Responded Yes No Orchitis Responded Yes No Varicocele Responded Yes No a Fisher exact test. Künzle. Cigarette smoking reduces semen quality. Fertil Steril Many studies have suggested that cigarette smoking is associated with altered semen quality, but conclusions about the extent of the deleterious effects vary widely. Affected variables include motility, sperm concentration, total sperm count, semen volume, and morphology (6, 8, 10, 11, 16 19). Advancing age is associated with decreased semen volume and motility and increase in abnormal morphology but with no consistent effect on sperm concentration (20). Sperm morphology is an indicator of the state of the seminal epithelium; degenerative changes due to ageing may affect spermatogenesis and thus alter sperm morphology. We studied a large sample consisting of men attending an andrology laboratory. Although age differed significantly between smokers and nonsmokers, our findings are highly representative: Smokers were younger than nonsmokers, and we classified men who consumed one or more cigarettes daily as smokers. We found that cigarette smoking was associated with reduced sperm quality (lower total sperm count combined with increased abnormal morphology) and decreased citrate concentration (increased ph). Vine et al. (6) found a negative association between decreased sperm quality and smoking only among men 22 years of age or older (n 60). In younger men (n 28 [32%]), sperm and semen quality were not reduced. Our results largely confirm those of Vine et al. in a much larger sample (1,770 of our participants were 22 years of age or older). However, our sample included only 16 men (0.9% [6 smokers and 10 nonsmokers]) who were younger than 22 years, and this group could not be analyzed separately. Because our participants were recruited in an infertility clinic, the mean age of the sample was relatively high. Our data show that cigarette smoking has significant negative effects on various sperm and fertility assessment variables. This is not the case with moderate alcohol consumption (1, 11). One group of investigators even observed better sperm morphology in men who drank moderately (21). In our sample (smokers as well as nonsmokers), we found no differences in sperm variables between moderate drinkers and nondrinkers. The mechanisms by which cigarette smoking affect semen quality are not fully understood. The fact that nicotine and its water-soluble metabolite cotinine are detectable in the seminal plasma of smokers suggests that other harmful components of tobacco smoke would pass through the blood testis barrier (22). Using the immunoperoxidase method, Zenzes et al. (7) demonstrated the presence of adducts formed between benzo(a)pyrene and sperm DNA in smoking men. Lower motility has also been associated with abnormalities in the ultrastructure of the flagellum and the axonemal structures of the sperm tail (23). Pacifici et al. (5) found that total motility of spermatozoa was significantly and negatively correlated with concentrations of cotinine and hydroxycotinine in the seminal plasma. We did not measure cotinine, and the mechanisms responsible for the correlation between the plasma concentration of this metabolite and reduced sperm function parameters remain unexplained. Zavos et al. (24) investigated the effect of smoking on the ability of seminal plasma to maintain sperm viability and found that seminal plasma from smokers had a strong detrimental effect on motility of spermatozoa from nonsmokers. Washing of sperm from smokers and exposure to seminal plasma from nonsmokers restored motility (24). Cigarette smoke contains several chemical agents, many of which are carcinogenic or mutagenic. These agents affect the production and function of healthy normal sperm via different mechanisms. Smoking has also been found to affect accessory glands (prostate, epididymis, and seminal vesicles) (25). We observed a significantly reduced citrate concentration in smokers, indicating impaired prostate function. FERTILITY & STERILITY 289

4 TABLE 2 Sperm and semen quality variables, by smoking status. Characteristic Smokers Nonsmokers P value No. of participants 655 1,131 Age (y) b Sperm density ( 10 6 cells/ml) Mean ( SD) c Range Sample volume (ml) b Total sperm count ( 10 6 cells) b Progressive motility (%) b Total motile sperm a ( 10 6 cells) b Morphology (% normal forms) c Vitality (%) by eosin exclusion b ph c Citrate concentration ( mol/l) c Fructose concentration ( mol/l) b Note: Values are means ( SD), unless otherwise indicated. a Calculated as total sperm count progressive motility/100. b Student t-test. c Welch t-test (for different SDs). Künzle. Cigarette smoking reduces semen quality. Fertil Steril Cigarette smoke may decrease male fertility through a direct effect on the testis and its ability to produce progressively motile, vital sperm (spermatogenesis). Comparison of smokers and nonsmokers by subgroups of men with past (operated) varicocele and cryptorchidism did not yield statistically different results than those found in the full sample (Table 2). However, the number of patients analyzed, particularly for smokers with varicocele, were low (Table 1). Sperm morphology, total sperm count, and sperm density (concentration) are prognostic for fertility in vivo and in vitro (11, 26, 27). However, the thresholds used to assess semen quality for discrimination of infertile from fertile men are being reexamined and redefined (28). In conclusion, sperm variables in smokers remain in the normal range, but a clear negative trend is observed. Therefore, men with borderline semen quality who wish to have children should especially benefit from quitting smoking. Acknowledgments: The authors thank Ms. Eva Zbinden and her assistants at the andrology laboratories of the MCL Institute for skillful technical assistance with the sperm analysis. References 1. Curtis KM, Savitz DA, Arbuckle TE. Effects of cigarette smoking, caffeine consumption, and alcohol intake on fecundability. Am J Epidemiol 1997;146: Matikainen T, Perez GI, Jirisicova A, Pru JK, Schlezinger JJ, Ryu HY, et al. Aromatic hydrocarbon receptor-driven Bax gene expression is required for premature ovarian failure caused by biohazardous environmental chemicals. Nat Genet 2001;28: Zenzes MT. Smoking and reproduction: gene damage to human gametes and embryos. Hum Reprod Update 2000;6: Osser S, Liedholm P, Ranstam J. Depressed semen quality: a study over two decades. Arch Androl 1984;12: Pacifici R, Altieri I, Gandini L, Lenzi A, Pichini S, Rosa M, et al. Nicotine, cotinine, and trans-3-hydroxycotinine levels in seminal plasma of smokers: effects on sperm parameters. Ther Drug Monitor 1993;15: Vine MF, Tse CJ, Hu PC, Truong KY. Cigarette smoking and semen quality. Fertil Steril 1996;65: Zenzes MT, Bielecki R, Reed TE. Detection of benzo(a)pyrene diol epoxide-dna adducts in sperm of men exposed to cigarette smoke. Fertil Steril 1999;72: Zhang JP, Meng QY, Wang Q, Zhang LJ, Mao YL, Sun ZX. Effect of smoking on semen quality of infertile men in Shandong, China. Asian J Androl 2000;2: Gandini L, Lombardo F, Lenzi A, Culasso F, Pacifici R, Zuccaro P, et al. The in-vitro effects of nicotine and cotinine on sperm motility. Hum Reprod 1997;12: Rubes J, Lowe X, Moore D, Perreault S, Slott V, Evenson D, et al. Smoking cigarettes is associated with increased sperm disomy in teenage men. Fertil Steril 1998;70: Chia SE, Lim ST, Tay SK, Lim ST. Factors associated with male infertility: a case-control study of 218 infertile and 240 fertile men. Brit J Obstet Gynaecol 2000;107: Karagounis CS, Papanikolaou NA, Zavos PM. Semen parameters compared between smoking and non-smoking men: smoking intensity and semen parameters. Infertility 1985;8: WHO laboratory manual for the examination of human semen and semen-cervical mucus interaction, 3rd ed. Cambridge (UK): Cambridge University Press: Juhl M, Anderson AM, Gronbaek M, Olsen J. Moderate alcohol consumption and waiting time to pregnancy. Hum Reprod 2001;16: Lund L, Rasmussen HH, Ernst E. Asymptomatic varicocele testis. Scand J Urol Nephrol 1993;27: Vine MF, Margolin BH, Morrison HI, Hulka BS. Cigarette smoking and sperm density: a metaanalysis. Fertil Steril 1994;61: Oldereid NB, Rui H, Purvis K. Life styles of men in barren couples and their relationship to sperm quality. Int J Fertil 1992;37: Osser S, Beckman-Ramirez A, Liedholm P. Semen quality of smoking and non-smoking men in infertile couples in a Swedish population. Acta Obstet Gynecol 1992;71: Goverde HJ, Dekker HS, Janssen HJ, Bastiaans BA, Rolland R, Zielhuis GA. Semen quality and frequency of smoking and alcohol con- 290 Künzle et al. Cigarette smoking reduces semen quality Vol. 79, No. 2, February 2003

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