This Article Abstract Full Text (PDF) Submit a related Letter to the Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Copyright Permission Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Andersson, A.-M. Articles by Skakkebæk, N. E. Search for Related Content PubMed PubMed Citation Articles by Andersson, A.-M. Articles by Skakkebæk, N. E. Related Collections Male Endocrinology

Secular Decline in Male Testosterone and Sex Hormone Binding Globulin Serum Levels in Danish Population Surveys

Department of Growth and Reproduction (A.-M.A., T.K.J., A.J., J.H.P., N.E.S.), Copenhagen University Hospital, Copenhagen DK-2100, Denmark; Department of Environmental Medicine (T.K.J.), University of Southern Denmark, Odense DK-5230, Denmark; Department of Biostatistics (J.H.P.), University of Copenhagen, DK-2400 Copenhagen, Denmark; and Research Centre for Prevention and Health (T.J.), Glostrup University Hospital, Glostrup DK-2600, Denmark

Address all correspondence and requests for reprints to: Anna-Maria Andersson, Department of Growth and Reproduction, Copenhagen University Hospital, Rigshospitalet, Sect. GR 5064, Blegdamsvej 9, DK-2100 Copenhagen, Denmark. E-mail: anna{at}rh.dk.

	Abstract

Top Abstract Introduction Subjects and Methods Results Discussion References

 
Context: Adverse secular trends in male reproductive health have been reported to be reflected in increased testicular cancer risk and decreased semen quality in more recently born men. These secular trends may also be reflected by changes in Leydig cell function.

Objective: The objective of the study was to examine whether an age-independent time trend in male serum testosterone levels exists.

Design and Setting: Testosterone and SHBG were analyzed in 5350 male serum samples from four large Danish population surveys conducted in 1982–1983, 1986–1987, 1991–1992, and 1999–2001. Free testosterone levels were calculated. The effects of age, year of birth, and time period on hormone levels were estimated in a general linear statistical model.

Main Outcome Measures: Testosterone, SHBG, and calculated free testosterone levels in Danish men in relation to age, study period, and year of birth were measured.

Results: Serum testosterone levels decreased and SHBG levels increased with increasing age. In addition to this expected age effect, significant secular trends in testosterone and SHBG serum levels were observed in age-matched men with lower levels in the more recently born/studied men. No significant age-independent effect was observed for free testosterone. Adjustment for a concurrent secular increase in body mass index reduced the observed cohort/period-related changes in testosterone, which no longer were significant. The observed cohort/period-related changes in SHBG levels remained significant after adjustment for body mass index.

Conclusions: The observed age-independent changes in SHBG and testosterone may be explained by an initial change in SHBG levels, which subsequently lead to adjustment of testosterone at a lower level to sustain free testosterone levels.

	Introduction

Top Abstract Introduction Subjects and Methods Results Discussion References

 
THERE IS GENERAL agreement that male serum testosterone levels decline with age, but the data have not been conclusive. Cross-sectional surveys have demonstrated very different rates of decline, ranging from 0.2 to 0.8%/yr (1, 2, 3, 4, 5, 6), and some have even failed to observe a significant decrease in testosterone levels with increasing age (7, 8, 9, 10, 11). Furthermore, longitudinal studies have shown more steep age-related declines in serum testosterone than cross-sectional studies (12, 13). A suggested explanation for this apparent discrepancy between cross-sectional and longitudinal studies on age-related declines has been that poor health might accelerate an age-related decline in individual longitudinal testosterone levels (13). Alternatively, cross-sectional data on the older age ranges may be biased toward containing a higher proportion of healthy men, compared with the younger age ranges, implying that unhealthy men are less likely to become old. However, yet another alternative explanation could be that a secular decline in testosterone levels exists, because men studied 20 yr ago had higher serum testosterone levels than men of the same age studied today. The existence of such an age-independent time trend may blunt the effect of increasing age in cross-sectional studies.

If such a secular decline in male testosterone levels exists, this may not only lead to misinterpretation of the effect of age and compromise the durability of normal reference data, but also more importantly, it may be a sign of general changes in male reproductive health. A recently published study showing an age-independent population-level decline in male serum testosterone levels in American men (14) strengthens the suspicion that population trends in male testosterone levels may exist in certain countries, and further scrutiny of these effects is warranted.

SHBG has more conclusively been shown to increase in men with increasing age in both cross-sectional (3, 5, 15, 16) and longitudinal (13) studies. However, as for testosterone, age-independent time trends may exist for SHBG levels and may blunt or even exaggerate the associations between age and hormone levels, depending on the interaction between age and period on SHBG serum levels.

To study age and secular trends in testosterone levels among Danish men, we analyzed serum samples from more than 5300 men participating in four large population-based surveys conducted at Glostrup University Hospital (Glostrup, Denmark) during 1982–2001 for testosterone and SHBG and calculated free testosterone using the equation by Vermeulen et al. (17).

	Subjects and Methods

Top Abstract Introduction Subjects and Methods Results Discussion References

 
Study population

The present study includes available serum samples from 5350 male participants of four large population-based surveys conducted at the Research Centre for Prevention and Health, Glostrup University Hospital, (Glostrup, Denmark) during 1982–2001, namely the three Danish Monitoring Cardiovascular (MONICA) risk factors cohorts examined in 1982–1983 (MONICA I), 1986–1987 (MONICA II), and 1991–1992 (MONICA III) and the Inter99 cohort examined in 1999–2001 (18, 19). An overview of the different cohorts and the number of serum samples available for this study is given in Table 1. In contrast to the MONICA studies, the Inter99 study was an intervention study aimed to look at effect of changes in lifestyle. However, the Inter99 samples used for this study was from baseline before intervention. Blood samples were taken in the morning after an overnight fast, and serum was stored in aliquots at –20 C. Samples from some of the surveys had been thawed for other analyses before this study, whereas others had been kept in the freezer exclusively. Information on year of birth, age, sex, and body mass index (BMI; weight/squared height) of the participants was available from all surveys.

Testosterone was measured by time-resolved fluoroimmunoassays (DELFIA; Wallac Oy, Turku, Finland) with a detection limit of 0.23 nmol/liter and intra- and interassay coefficients of variation were less than 12%. SHBG was measured by time-resolved immunofluorometric assays (DELFIA; Wallac). The detection limit was 0.23 nmol/liter, and the intra- and interassay coefficients of variation were less than 8%. The detection limit was defined as the concentration corresponding to the signal that is 2 SD above or below the mean of the zero standard measurement in the immunofluorometric assays and competitive immunoassays (fluoroimmunoassays), respectively.

All samples were analyzed during the same period (spring to autumn 2004), and samples from the different surveys were analyzed and mixed in the different assay runs to eliminate any influence of assay variation.

Validation of the integrity of hormone levels in stored samples

At the time of hormone analysis, samples had been stored between 3 and 21 yr at –20 C.

First, because an aliquot of 434 samples (not included in the present study) that had been analyzed in the same laboratory 10 yr previously (in 1994 when the samples were new) was available (11), this allowed a direct comparison of the hormone levels obtained in fresh samples and the same samples after 10 yr of storage at –20 C. This approach allowed us to test whether storage for 10 yr at –20 C resulted in major changes in the levels of the hormones studied, whereas it would not be possible to discriminate whether minor changes were due to storage or assay variation because all immunoassays have a certain day-to-day variation, which may be even higher over a long time span due to the use of different batches of regents over time. For SHBG the same assay format was used 10 yr ago, whereas testosterone in 1994 had been analyzed in a different assay format than used in the present study. For SHBG the median of the ratio between the first and second measure was 1.04 (95% confidence interval 0.88–1.21), and the correlation coefficient between the two measures was 0.98. For testosterone the median of the ratio between the first and second measure was 0.95 (95% confidence interval 0.63–1.47), and the correlation coefficient was 0.83. The difference observed between the two measurements was for both hormones within the expected day-to-day assay variation (see Fig. 1).

View larger version (10K): [in this window] [in a new window]   FIG. 1. Comparison of testosterone (left) and SHBG (right) levels measured in the same samples 10 yr apart (measured the first time in 1994 and measured again in 2004). Different testosterone assays were used at the two times of measurement, whereas the same SHBG assay format was used at both time points. The drawn line represents the regression line (and 95% mean confidence interval) between the two measurements. The dotted line represents the identity line.

In the samples from the four population surveys, we generally measured higher hormone levels in the samples that had been stored the longest. Thus, we were concerned about whether any evaporation had occurred during previous handling of the samples, which could lead to concentration of the samples. We therefore measured serum Na⁺ in 25 randomly selected samples from each of the surveys (total n = 100 samples) as an estimate of whether any concentration of samples had occurred. Serum Na⁺ was measured by indirect potentiometry (ISE system; Roche/Hitachi, Basel, Switzerland) at the clinical chemistry department at our hospital. We found a significant negative correlation between year of sampling and the Na⁺ concentration, indicating that evaporation of samples during storage or previous handling may have occurred (Fig. 2). The mean Na⁺ concentrations in the samples from the MONICA I, MONICA II, MONICA III, and Inter99 surveys were 172, 169, 162, and 154 mmol/liter, respectively, which all were above the normal range for serum Na⁺ concentrations. To normalize serum concentration to a normal mean serum Na⁺ concentration of 140 mmol/liter, hormone levels measured in samples from the MONICA I, MONICA II, MONICA III, and Inter99 surveys were multiplied with a correction factor of 0.81, 0.83, 0.86, and 0.91, respectively (and thereby adjusted for any effect of evaporation). The variation in Na⁺ levels was larger than the normal biological variation for Na⁺, indicating that samples from the same sampling period were not identically affected by storage or handling of the samples. Thus, by using a general correction factor for all samples from the same sampling period, some samples may be undercorrected and some may be overcorrected. Unfortunately, serum was not available for Na⁺ measurements in all samples, and thus, individual adjustment was not possible. However, due to the large number of samples available and the fact that we in this study were interested in general trends rather than individual levels, it can be justified to use a general correction factor for each sampling period.

View larger version (24K): [in this window] [in a new window]   FIG. 2. Correlation between date of sampling and serum Na+ level measured. The shaded area represents the normal range for serum Na+.

Hormone levels were initially corrected for evaporation (see above). Free testosterone was calculated from the testosterone and SHBG concentrations using the method by Vermeulen et al. (17), with the assumption of an average serum albumin concentration of 43 g/liter. Hormone levels of the different age groups were compared using one-way ANOVA with Bonferroni post hoc test.

To disentangle the effects of age, study time (period), and year of birth (cohort), age-period-cohort modeling was used (21). Hormone levels were natural logarithm transformed to obtain approximate homoscedacity and an approximate normal distribution of residuals. The following general linear model was used:

where _i represents the age effect in the i^th age group, β_j the period effect in the j^th period, and _k the cohort (birth year) effect of the k^th cohort. The remainder term _ijk is the residual representing the noise. The model included age (30, 40, 50, 60, and 70 yr), period (1982–1983, 1986–1987, 1991–1992, and 1999–2001), and year of birth (1921–1926, 1931–1939, 1940–1946, 1949–1952, 1956–1961, and 1969–1970) as categorical variables. An inherent problem in age-period-cohort modeling, stemming from the fact that period can be calculated as the sum of a man’s age and birth year (and vice versa) is that a linear trend in mean level of the response cannot be attributed specifically to the period or the cohort (22). This is the fundamental nonidentifiable problem to which there is no solution. The adequacy of the model can be investigated by introducing a general interaction term. In the presence of a general interaction term, the model does not lend itself to simple interpretation in terms of age, period, or cohort effects. A particular age-cohort interaction is obtained by including period in the model with the restriction that the first and last category are identical. Likewise a particular age-period interaction is obtained by including cohort in the model with the same restriction. This constitutes a main-effects model in which the effects of age, period, and cohort, apart from the linear trend, can be identified. In subsequent analyses, BMI was included in the statistical model as an additional explanatory variable to investigate whether the hormonal changes could partly be explained by a change in BMI. The final models were subjected to standard checks of the residuals.

	Results

Top Abstract Introduction Subjects and Methods Results Discussion References

 
Median testosterone, SHBG, and free testosterone levels in each age group for all periods together as well as stratified according to study period and year of birth are shown in Figs. 3 and 4. In Fig. 3 the data for testosterone are plotted both without and with adjustment for evaporation.

View larger version (22K): [in this window] [in a new window]   FIG. 3. Median testosterone levels plotted in relation to age (A and D) and with stratification by study period (B and E) and year of birth (C and F). A–C, Data without adjustment for evaporation during storage; D–F, data after adjustment for evaporation during storage.

View larger version (21K): [in this window] [in a new window]   FIG. 4. Median SHBG (A-C) and free testosterone (D–F) serum levels plotted in relation to age (A and D) and with stratification by study period (B and E) and year of birth (C and F).

When testosterone levels were not stratified, a modest decrease in testosterone levels with increasing age was observed (Fig. 3D). Bonferroni post hoc test showed the levels at 30 yr > 40 yr > 50 yr = 60 yr = 70 yr. However, when data were plotted stratified according to study period or year of birth, a steeper decrease in testosterone levels with increasing age was evident within each category. When stratified according to study period, the age-related changes differed between the different study periods. Thus, whereas the levels and the age-related decline was very similar between the different study periods for the 30- and 40-yr-old men, this was not true for the older men (Fig. 3E). When stratified according to birth cohort, the same age-related decline was observed within all cohorts, revealing a steady decline over all age groups. However, a significant cohort effect was evident with higher levels observed in the oldest cohort (Fig. 3F). This cohort effect was evident between the men born in the 1920s, 1930s, and 1940s, whereas there was no difference between later-born cohorts.

The influence of age, birth year, and study period on serum testosterone levels were further investigated in age-period-cohort general linear models. The parameter estimates of the age-period-cohort models are shown in Table 2. In the model focusing on the age-cohort relation (linear component of period nullified), the estimated age-related decline in serum testosterone on average varied from –0.7 to –1.0% per year of age with the steepest decline from 30 to 40 yr of age. In the age-period model (linear component of birth year nullified), the estimated age-related decline in serum testosterone was slightly smaller, varying from –0.5 to –0.7% per year of age. The age-cohort model also showed a statistically significant decline in serum testosterone with later year of birth. In the age-period model, a declining trend in serum testosterone levels with later study year was found, but this trend was statistically significant only between the oldest and latest study periods.

The graphical displays for SHBG show a significant increase with increasing age when data were not stratified with median SHBG levels being more than 50% higher in the 70-yr age group, compared with the 30-yr age group (Fig. 4A). When the data were stratified according to study period, a similar age-related increase was observed in all four study periods, although with differences in absolute levels indicating a period effect, with higher levels in the earliest study periods (Fig. 4B). When the data were stratified according to birth cohorts, most of the apparent age-related increase in SHBG levels seemed to be due to a cohort effect, with higher levels in older cohorts (Fig. 4C). However, the age-related trend in SHBG differed between the different cohorts.

The parameter estimates from the age-period-cohort models for SHBG are shown in Table 3. Age, study period, and birth year were all statistically significant confounding variable for SHBG serum levels. The estimates of the age-related increase in serum SHBG levels differed quite significantly between the age-cohort and the age-period models, but in both models the increase seemed to set off between 40 and 50 yr of age and onward. In the age-cohort model, the estimated average increase was 0.6–0.7% per year from 40 yr of age and onward; in the age-period model, the estimated average increase was 1.2–1.5% per year during the same age range. In the age-cohort model, a significant decrease in SHBG levels between the cohort born between 1921 and 1926 and the cohort born between 1931 and 1939, with a further but more moderate decline in more recent cohorts, was seen. In addition, a trend of lower SHBG levels in the more resent study period was found in the age-period model, with the levels obtained in the 1999–2001 study period being 12% lower than the levels obtained in the 1982–1984 study period.

Free testosterone levels decreased with increasing age, and the rate was the same irrespective of whether the data were stratified according to period of sampling, birth cohort, or not stratified (Fig. 4, D and E). Thus, in contrast to testosterone and SHBG, no significant difference between the different study periods or between the different birth cohorts was evident for free testosterone. This was confirmed in the age-period-cohort models in which neither birth year nor study period was a significant confounding variable (Table 4). The average decline in free testosterone varied from –1.0 to –1.2% per year of age.

BMI was used as a crude index of the body composition. Serum levels of testosterone and SHBG were associated with BMI, even after adjustment for age, period, and cohort reflected in this study by a significant negative correlation between testosterone and SHBG toward BMI (P < 0.01). Because BMI may change with increasing age as well as changes in year of birth, we also included BMI in the statistical model to evaluate how much of the age- and cohort/period-related changes could be explained by concurrent changes in BMI. The estimated changes in hormone levels after adjustment also for BMI are given in Table 5. When adding BMI as a confounding variable in the age-period-cohort models, birth year and study period no longer remained statistically significant confounding variables for serum testosterone levels and were therefore omitted from the final model. In contrast, although the estimated effect of birth year and period on serum SHBG levels diminished when adjusted for changes in BMI, they remained statistically significant confounding variables. Both testosterone and SHBG serum levels were estimated to decrease –4% per increase in BMI. The estimated age-related decline in testosterone levels diminished when the effect of increasing BMI with increasing age was adjusted for. Thus, after adjustment for BMI, the estimated decline in serum testosterone per increasing year varied between –0.3 and –0.5%. In contrast, the estimated age-related effect on SHBG was even more pronounced when the confounding effect of BMI was adjusted for. After adjustment for BMI, a very steady age-related decline in free testosterone of –1% per year was estimated.

	Discussion

Top Abstract Introduction Subjects and Methods Results Discussion References

The secular decline was most pronounced for SHBG and less so for testosterone. The observed age-independent decline in male serum testosterone levels could be explained by a concurrent secular increase in BMI. In contrast, the concurrent increase in BMI explained only part of the secular trend in male SHBG serum levels and the impact of BMI were more pronounced in the later cohorts, whereas changes in BMI seemed to play only a minor role in the changes seen in SHBG between the older cohorts. The fact that the free testosterone level did not seem to be affected by cohort or period effects indicates that whatever causes the secular trend, it apparently is primarily affecting the SHBG serum levels. We speculate that the secular decline in testosterone serum levels could be secondary to the decline in SHBG levels, simply adjusting the pituitary-gonadostat to a lower level to sustain the same level of free testosterone.

The regulation of SHBG is complex and only partly understood. Sex steroids stimulate SHBG production and secretion in vitro (22), and SHBG levels increase during pharmacological oral estrogen treatment (23, 24, 25) and decrease during oral androgen treatment (26). However, parenteral administration of sex steroids as well as physiological changes in sex steroids has only moderate effects on serum SHBG levels (26, 27, 28, 29), and endogenous sex steroid levels presumably play a minor role in the regulation of SHBG. Correlations between serum sex steroids and SHBG levels are thus more likely due to the indirect regulation of sex steroid levels by serum SHBG levels than vice versa. Serum SHBG levels are negatively associated with obesity and various measures of insulin resistance (30) and has been suggested as a marker of the metabolic syndrome. Furthermore, insulin decreases SHBG production and secretion in vitro (22). It could be speculated that the observed secular decline in serum SHBG levels could be linked to increased incidence of obesity/metabolic syndrome in the later cohorts, although changes in BMI in our study could not explain the observed effects. On the other hand, BMI is a rather crude estimate of obesity. Also IGF-I and thyroid hormones have been indicated as regulators of serum SHBG levels (22, 31), but in the present study, information on these hormones was not available. Thus, the etiology of the observed secular decline in male SHBG serum levels remains unknown.

The secular decline in SHBG was most pronounced for the oldest cohorts, indicating that whatever caused this cohort effect seems to have leveled out in the more recent cohorts. Large changes in lifestyle as well as the environment occurred in Denmark during the 20th century with a general significant increase in standard of life along with an increased industrialization. A declining trend in male reproductive health manifested as an increase in testicular cancer, and a declining sperm quality has been reported for the same period in many Western countries (32, 33, 34, 35, 36). These adverse trends in male reproductive health and the observed changes in male reproductive hormone levels presented here could be interrelated.

The secular decline in SHBG and testosterone serum levels did not lead to a change in the level of free testosterone, which often is believed to determine the androgen activity. Nevertheless, a general decreased testosterone production will lead to lower intratesticular testosterone levels, and thus, the paracrine effects of testosterone within the testes may be blunted. This is in line with our previous findings of decreased serum SHBG and testosterone levels as well as decreased sperm concentration in men with BMI greater than 25 kg/m² (37). Furthermore, recently the existence of specific binding sites for SHBG at the cell membrane of steroid-responsive tissues has been shown (38), challenging the dogma that SHBG merely acts as a carrier protein. Binding of SHBG to its cell membrane receptor and subsequently to steroid hormone has been shown to initiate a steroid receptor-independent downstream signal through increased intracellular cAMP (for review see Ref. 39). The free hormone hypothesis, according to which only free steroids are biologically relevant, has been further challenged by the finding that SHBG may play an active role in the cellular uptake of steroids through binding and subsequently internalization of the protein/steroid complex via the cell membrane receptor megalin (40). Thus, the observed decline in serum SHBG levels may in itself have an impact on steroid action.

Studies on long-term trends like ours have inherent challenges related to the integrity of the samples or the methods of measurement over time. In our study design, the integrity of the samples over time was an important issue, which we acknowledge as a limitation of these kinds of studies. However, although the observed magnitude of the decline in SHBG (and testosterone) in later-born cohorts may be slightly affected by storage issues, the overall conclusion that a decline has occurred remains robust. An alternative design could be to measure samples as they are collected to avoid the issue of stability of samples during long-term storages. However, this approach is, on the other hand, vulnerable to the long-term integrity and stability of the methods of measurement over several decades. Thus, there is no simple solution to the design of studies on long-term effects of hormone levels. Nevertheless, taking all these difficulties into consideration, it is interesting that a recent American study adapting the later approach (e.g. measuring samples at different time points over a long period) also found a significant decline in serum testosterone levels in later-born men when matched by age (14). In contrast to our data, a secular decline in calculated free testosterone of similar magnitude to that in total testosterone was also observed in the American study, implying that no secular decline in SHBG had occurred in the American men (14). Thus, although a decline in total testosterone serum levels was observed in both the Danish and American study groups, the etiology for this decline may differ between the two populations. The age-independent changes in male serum testosterone levels observed in our Danish population and the American population may be too small to be of any clinical relevance for the individual man, but in a population-level perspective, it is alarming that changes of this magnitude can be detected over such a relatively short time, evolutionarily speaking. Similar population studies from other countries are much warranted to explore whether the observed trends in male total testosterone levels is a general trend in Western countries. Clarification of the causes for these changes, whether they are related to changes in lifestyle or environment, should provide information on which preventive action can be taken.

	Acknowledgments

 
We thank the personnel at the Research Center for Prevention and Health for making the samples from their large population studies available for this study and skilled technicians Ole Nielsen and Birgitte Schou at the hormone laboratory, Department of Growth and Reproduction, Rigshospitalet, for their assistance and great efforts of running hormone analyses of all the samples.

	Footnotes

 
This work was supported by the European Commission (contracts QLK4–1999-01422 and QLK4-CT-2002-00603, EDEN), the Danish Research Council (Grant 22-03-0198), and the Sven Andersen Foundation.

Disclosure Information: A.-M.A., T.K.J., A.J., J.H.P., T.J., and N.E.S. have nothing to declare.

First Published Online September 25, 2007

Abbreviations: BMI, Body mass index; MONICA, Monitoring Cardiovascular.

Received November 29, 2006.

Accepted September 13, 2007.

	References

Top Abstract Introduction Subjects and Methods Results Discussion References

 

1. Drafta D, Schindler AE, Stroe E, Neacsu E 1982 Age-related changes of plasma steroids in normal adult males. J Steroid Biochem 17:683–687[CrossRef][Medline]
2. Deslypere JP, Vermeulen A 1984 Leydig cell function in normal men: effect of age, life-style, residence, diet, and activity. J Clin Endocrinol Metab 59:955–962[Abstract/Free Full Text]
3. Gray A, Feldman HA, McKinlay JB, Longcope C 1991 Age, disease, and changing sex hormone levels in middle-aged men: results of the Massachusetts male aging study. J Clin Endocrinol Metab 73:1016–1025[Abstract/Free Full Text]
4. Simon D, Preziosi P, Barrett-Connor E, Roger M, Saint-Paul M, Nahoul K, Papoz L 1992 The influence of aging on plasma sex hormones in men: the Telecom Study. Am J Epidemiol 135:783–791[Abstract/Free Full Text]
5. Svartberg J, Midtby M, Bonaa KH, Sundsfjord J, Joakimsen RM, Jorde R 2003 The associations of age, lifestyle factors and chronic disease with testosterone in men: the Tromso Study. Eur J Endocrinol 149:145–152[Abstract]
6. Boyce MJ, Baisley KJ, Clark EV, Warrington SJ 2004 Are published normal ranges of serum testosterone too high? Results of a cross-sectional survey of serum testosterone and luteinizing hormone in healthy men. BJU Int 94:881–885[CrossRef][Medline]
7. Harman SM, Tsitouras PD 1980 Reproductive hormones in aging men. I. Measurement of sex steroids, basal luteinizing hormone, and Leydig cell response to human chorionic gonadotropin. J Clin Endocrinol Metab 51:35–40[Abstract/Free Full Text]
8. Sparrow D, Bosse R, Rowe JW 1980 The influence of age, alcohol consumption, and body build on gonadal function in men. J Clin Endocrinol Metab 51:508–512[Abstract/Free Full Text]
9. Nieschlag E, Lammers U, Freischem CW, Langer K, Wickings EJ 1982 Reproductive functions in young fathers and grandfathers. J Clin Endocrinol Metab 55:676–681[Abstract/Free Full Text]
10. McKinlay JB, Longcope C, Gray A 1989 The questionable physiologic and epidemiologic basis for a male climacteric syndrome: preliminary results from the Massachusetts Male Aging Study. Maturitas 11:103–115[CrossRef][Medline]
11. Gyllenborg J, Rasmussen SL, Borch-Johnsen K, Heitmann BL, Skakkebæk NE, Juul A 2001 Cardiovascular risk factors in men: the role of gonadal steroids and sex hormone-binding globulin. Metabolism 50:882–888[CrossRef][Medline]
12. Morley JE, Kaiser FE, Perry III HM, Patrick P, Morley PM, Stauber PM, Vellas B, Baumgartner RN, Garry PJ 1997 Longitudinal changes in testosterone, luteinizing hormone, and follicle-stimulating hormone in healthy older men. Metabolism 46:410–413[CrossRef][Medline]
13. Feldman HA, Longcope C, Derby CA, Johannes CB, Araujo AB, Coviella AD, Bremner WJ, McKinlay JB 2002 Age trends in the level of serum testosterone and other hormones in middle-aged man: longitudinal results from the Massachusetts Male Aging Study. J Clin Endocrinol Metab 87:589–598[Abstract/Free Full Text]
14. Travison TG, Araujo AB, O’donnell AB, Kupelian V, McKinlay JB 2007 A population-level decline in serum testosterone levels in American men. J Clin Endocrinol Metab 92:196–202[Abstract/Free Full Text]
15. Field AE, Colditz GA, Willett WC, Longcope C, McKinlay JB 1994 The relation of smoking, age, relative weight, and dietary intake to serum adrenal steroids, sex hormones, and sex hormone-binding globulin in middle-aged men. J Clin Endocrinol Metab 79:1310–1316[Abstract]
16. Muller M, den Tonkelaar I, Thijssen JH, Grobbee DE, van der Schouw YT 2003 Endogenous sex hormones in men aged 40–80 years. Eur J Endocrinol 149:583–589[Abstract]
17. Vermeulen A, Verdonck l, Kaufman JM 1999 A critical evaluation of simple methods for the estimation of free testosterone in serum. J Clin Endocrinol Metab 84:3666–3672[Abstract/Free Full Text]
18. Gerdes LU, Bronnum-Hansen H, Madsen M, Borch-Johnsen K, Jorgensen T, Sjol A, Schroll M 2000 Trends in selected biological risk factors for cardiovascular diseases in the Danish MONICA population, 1982–1992. J Clin Epidemiol 53:427–434[CrossRef][Medline]
19. Jørgensen T, Borch-Johnsen K, Thomsen TF, Ibsen H, Glumer C, Pisinger C 2003 A randomized non-pharmacological intervention study for prevention of ischaemic heart disease: baseline results Inter99. Eur J Cardiovasc Prev Rehabil 10:377–386[CrossRef][Medline]
20. Harman SM, Metter EJ, Tobin JD, Pearson J, Blackman MR 2001 Longitudinal effects of aging on serum total and free testosterone levels in healthy men. Baltimore Longitudinal Study of Aging. J Clin Endocrinol Metab 86:724–731[Abstract/Free Full Text]
21. Clayton D, Schifflers E 1987 Models for temporal variation in cancer rates. I: Age-period and age-cohort models. Stat Med 6:449–467[Medline]
22. Loukovaara M, Carson M, Adlercreutz H 1995 Regulation of production and secretion of sex hormone-binding globulin in HepG2 cell cultures by hormones and growth factors. J Clin Endocrinol Metab 80:160–164[Abstract]
23. Bergink EW, Crona N, Dahlgren E, Samsioe G 1981 Effect of oestriol, oestradiol valerate and ethinyloestradiol on serum proteins in oestrogen-deficient women. Maturitas 3:241–247[CrossRef][Medline]
24. Helgason S, Damber JE, Damber MG, von Schoultz B, Selstam G, Sodergard R 1982 A comparative longitudinal study on sex hormone binding globulin capacity during estrogen replacement therapy. Acta Obstet Gynecol Scand 61:97–100[Medline]
25. El Makhzangy MN, Wynn V, Lawrence DM 1979 Sex hormone binding globulin capacity as an index of oestrogenicity or androgenicity in women on oral contraceptive steroids. Clin Endocrinol (Oxf) 10:39–45[Medline]
26. Conway AJ, Boylan LM, Howe C, Ross G, Handelsman DJ 1988 Randomized clinical trial of testosterone replacement therapy in hypogonadal men. Int J Androl 11:247–264[Medline]
27. Handelsman DJ, Conway AJ, Boylan LM 1990 Pharmacokinetics and pharmacodynamics of testosterone pellets in man. J Clin Endocrinol Metab 71:216–222[Abstract/Free Full Text]
28. Maruyama Y, Aoki N, Suzuki Y, Sinohara H, Yamamoto T 1984 Variation with age in the levels of sex-steroid-binding plasma protein as determined by radioimmunoassay. Acta Endocrinol (Copenh) 106:428–432[Abstract/Free Full Text]
29. Carlstrom K, Collste L, Eriksson A, Henriksson P, Pousette A, Stege R, von Schoultz B 1989 A comparison of androgen status in patients with prostatic cancer treated with oral and/or parenteral estrogens or by orchidectomy. Prostate 14:177–182[CrossRef][Medline]
30. Kalme T, Seppala M, Qiao Q, Koistinen R, Nissinen A, Harrela M, Loukovaara M, Leinonen P, Tuomilehto J 2005 Sex hormone-binding globulin and insulin-like growth factor-binding protein-1 as indicators of metabolic syndrome, cardiovascular risk, and mortality in elderly men. J Clin Endocrinol Metab 90:1550–1556[Abstract/Free Full Text]
31. Mercier-Bodard C, Baville F, Bideux G, Binart N, Chambraud B, Baulieu EE 1989 Regulation of SBP synthesis in human cancer cell lines by steroid and thyroid hormones. J Steroid Biochem 34:199–204[CrossRef][Medline]
32. Adami H-O, Bergström R, Möhner M, Zatonski W, Storm H, Ekbom A, Tretli S, Teppo L, Ziegler H, Rahu M, Gurevicius R, Stengrevics A 1994 Testicular cancer in nine Northern European countries. Int J Cancer 59:33–38[Medline]
33. Carlsen E, Giwercman A, Keiding N, Skakkebæk NE 1995 Declining semen quality and increasing incidence of testicular cancer: is there a common cause? Environ Health Perspect 103:137–139[CrossRef][Medline]
34. Møller H, Jørgensen N, Forman D 1995 Trends in incidence of testicular cancer in boys and adolescent men. Int J Cancer 61:761–764[Medline]
35. Swan SH, Elkin EP, Fenster L 2000 The question of declining sperm density revisited: an analysis of 101 studies published 1934–1996. Environ Health Perspect 108:961–966[Medline]
36. Andersen AG, Jensen TK, Carlsen E, Jørgensen N, Andersson A-M, Krarup T, Keiding N, Skakkebæk NE 2000 High frequency of sub-optimal semen quality in an unselected population of young men. Hum Reprod 15:366–372[Abstract/Free Full Text]
37. Jensen TK, Andersson A-M, Jørgensen N, Andersen A-G, Carlsen E, Petersen JH, Skakkebæk NE 2004 Body mass index in relation to semen quality and reproductive hormones among 1,558 Danish men. Fertil Steril 82:863–870[CrossRef][Medline]
38. Frairia R, Fortunati N, Fissore F, Fazzari A, Zeppegno P, Varvello L, Orsello M, Berta L 1992 The membrane receptor for sex steroid binding protein is not ubiquitous. J Endocrinol Invest 15:617–619[Medline]
39. Rosner W, Hryb DJ, Khan MS, Nakhla AM, Romas NA 1998 Androgens, estrogens, and second messengers. Steroids 63:278–281[CrossRef][Medline]
40. Hammes A, Andreassen TK, Spoelgen R, Raila J, Hubner N, Schulz H, Metzger J, Schweigert FJ, Luppa PB, Nykjaer A, Willnow TE 2005 Role of endocytosis in cellular uptake of sex steroids. Cell 122:751–762[CrossRef][Medline]

This article has been cited by other articles:

				J. D. Meeker, A. M. Calafat, and R. Hauser Urinary Metabolites of Di(2-ethylhexyl) Phthalate Are Associated With Decreased Steroid Hormone Levels in Adult Men J Androl, May 1, 2009; 30(3): 287 - 297. [Abstract] [Full Text] [PDF]

				A. Juul, K. Sorensen, L. Aksglaede, I. Garn, E. Rajpert-De Meyts, I. Hullstein, P. Hemmersbach, and A. M. Ottesen A Common Deletion in the Uridine Diphosphate Glucuronyltransferase (UGT) 2B17 Gene Is a Strong Determinant of Androgen Excretion in Healthy Pubertal Boys J. Clin. Endocrinol. Metab., March 1, 2009; 94(3): 1005 - 1011. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Submit a related Letter to the Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Copyright Permission Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Andersson, A.-M. Articles by Skakkebæk, N. E. Search for Related Content PubMed PubMed Citation Articles by Andersson, A.-M. Articles by Skakkebæk, N. E. Related Collections Male Endocrinology