High prevalence of subnormal testosterone in obese adolescent males: reversal with bariatric surgery

Introduction

We have previously reported that adolescent males with obesity have markedly lower total and free testosterone concentrations when compared with boys of normal weight (1). These boys may suffer from hypogonadism and infertility in the long term. Male adults with obesity also have lower testosterone concentrations than lean males. Approximately a quarter of men with obesity or metabolic syndrome, and a third of men with type 2 diabetes, have subnormal free testosterone concentrations (2, 3, 4). The gonadotropin concentrations in these males are inappropriately normal or low and thus they have hypogonadotropic hypogonadism. The hypogonadal state is more frequent and more profound in severely obese individuals. In fact, testosterone concentrations are inversely related to BMI (4). Following bariatric surgery in adult males with obesity, testosterone concentrations improve markedly (5, 6). It is, therefore, possible that a similar increase may be observed in adolescents with severe obesity. This potential benefit could prevent a lifelong risk of infertility.

The Teen-Longitudinal Assessment of Bariatric Surgery (Teen-LABS) is the first large study to systematically document the outcome of metabolic bariatric surgery for the treatment of adolescents with severe obesity in the USA (7). As expected, it demonstrated that with marked weight loss, the reversal of diabetes, remission of dyslipidemia and reductions in blood pressure followed (7). In view of these findings, we hypothesized that bariatric surgery in adolescents with severe obesity will also induce an increase in plasma testosterone concentrations. Such an increase would be potentially beneficial beyond improvements in sexual function since testosterone has been shown to exert insulin-sensitizing and anti-inflammatory effects (8).

Subjects and methods

Teen-LABS is a prospective, multicenter, observational study that enrolled consecutive adolescents (≤19 years of age) who were undergoing any bariatric surgical procedure from March 2007 through February 2012. The details of the study design have previously been reported (9). The study was approved by the Institutional Review Board of the participating sites (Cincinnati Children’s Hospital Medical Center, Nationwide Children’s Hospital, The University of Alabama at Birmingham, University of Pittsburgh Medical Center and Texas Children’s Hospital/Baylor College of Medicine). After a full explanation of the purpose and nature of all procedures used in the study, all participants provided their written informed consent. The Teen-LABS study enrolled 242 participants, of which 59 were males. Participants were followed up for 5 years following bariatric surgery. We are presenting results from an ancillary study in which serum from male study participants was used to measure testosterone, estradiol, sex hormone-binding globulin (SHBG) and gonadotropins (luteinizing hormone (LH) and follicle-stimulating hormone (FSH)). Serum samples were collected from participants in the Teen-LABS study at baseline (pre-surgery) and at 6, 12, 24, 36, 48 and 60 months. Baseline serum samples were available only in 34 males. Herein, we report the change in gonadal hormones in sequential samples obtained from these 34 participants. Follow-up samples were available at 6, 12, 24, 36, 48 and 60 months in 29, 27, 24, 21, 15 and 13 participants, respectively.

Total testosterone and estradiol were measured by liquid chromatography–tandem mass spectrometry as previously described (10, 11). The sensitivity of the testosterone assay (limit of quantification (LOQ)) set at a coefficient of variation (CV) of ≤20% is 0.01 nmol/L. The intraassay CV ranges from 7.6 to 10.8% and interassay CV ranges from 9.8 to 13.4% at total testosterone concentrations between 0.4 and 41.7 nmol/L. The normal range for total testosterone on this assay is 9.2–31.8 nmol/L in healthy non-obese men (12). The interassay precision (CV) of the assay is 4.06% at 73 pmol/L of estradiol, 1.42% at 734 pmol/L and 1.88% at 2936 pmol/L. The LOQ for estradiol in this assay is 7 pmol/L. The reference range for men is ≤106 pmol/L, established using healthy donors. The performing laboratory is certified by Clinical Laboratory Improvement Amendments and is a participant in the Hormone Standardization program of the Centers for Disease Control and Prevention.

Obesity is associated with low SHBG concentrations and thus there is a physiological lowering of total testosterone concentrations in obese men (4). An evaluation of free testosterone concentrations is, therefore, essential to accurately assess the gonadal status in obese males. Free testosterone and estradiol were calculated using concentrations of total testosterone, total estradiol, SHBG and albumin by the formula of Sodergard et al. and Vermeulen et al. (13, 14, 15, 16). The normal reference interval for free testosterone is 0.23–0.87 nmol/L. Large studies have not been published on reference intervals for free testosterone in children. In a prior study, the fifth percentile of the calculated free testosterone (using similar methodology as used in the current study) in lean males between 14 and 20 years of age was 0.23 nmol/L (1). We used this as a cut-off to define subnormal free testosterone in the results below. The normal reference interval for calculated free estradiol is 1.10–4.77 pmol/L. This is based on measurements obtained from 207 healthy men (15).

Results

The baseline characteristics of the study participants are shown in Table 1. Most (73%) of the study participants were ≥16 years of age; therefore they had most likely completed their pubertal development prior to surgery. Consistent with this, we did not see any relationship between age and total (r= −0.21, P = 0.22) or free testosterone (r = 0.07, P = 0.70) at baseline. The percent change in free testosterone from baseline was also found to be similar in males <16 years or >16 years of age (P = 0.65). Maximum BMI loss was achieved by 12 months, with slight regain noted between 24 and 36 months (P = 0.032) (Table 2). By 5 years, the mean percent BMI change from baseline was −28.7% (95% CI: −33.0 to −24.4).

Figure 1 shows the total and free testosterone concentrations by study visit. Baseline total testosterone concentration was 6.7 nmol/L (95% CI: 4.7 to 8.8), significantly increasing to 17.6 (95% CI: 15.3 to 19.9) at 2 years (P  < 0.001), with values remaining comparable through 2–5 years (Table 2). Baseline free testosterone concentrations increased from 0.17 (95% CI: 0.13 to 0.20) to 0.34 (95% CI: 0.30 to 0.38) nmol/L by 2 years (P  < 0.001). Following this, the concentrations declined at 3 years (P = 0.014), then remained similar thereafter.

Percent change in total testosterone (black circles) and free testosterone (white circles) at baseline and following bariatric surgery. Data are modeled estimates with 95% CIs. Samples were available at 0, 0.5, 1, 2, 3, 4 and 5 years in 29, 27, 24, 21, 15 and 13 participants, respectively.

Citation: European Journal of Endocrinology 186, 3; 10.1530/EJE-21-0545

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Percent change in total testosterone (black circles) and free testosterone (white circles) at baseline and following bariatric surgery. Data are modeled estimates with 95% CIs. Samples were available at 0, 0.5, 1, 2, 3, 4 and 5 years in 29, 27, 24, 21, 15 and 13 participants, respectively.

Citation: European Journal of Endocrinology 186, 3; 10.1530/EJE-21-0545

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Percent change in total testosterone (black circles) and free testosterone (white circles) at baseline and following bariatric surgery. Data are modeled estimates with 95% CIs. Samples were available at 0, 0.5, 1, 2, 3, 4 and 5 years in 29, 27, 24, 21, 15 and 13 participants, respectively.

Citation: European Journal of Endocrinology 186, 3; 10.1530/EJE-21-0545

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Prior to surgery, 79.4% (95% CI: 67.5 to 93.5) of the participants had subnormal total testosterone concentrations (<9.2 nmol/L), with prevalence significantly declining to 21.6% (95% CI: 9.6 to 48.5) at 5 years post-op (P  < 0.001). Following a similar pattern, 72.7% (95% CI: 58.7 to 59.9) at baseline had subnormal free testosterone (<0.23 nmol/L) concentrations, significantly falling to 33.1% (95% CI: 17.8 to 61.7) at 5 years. By 5 years, 71.4% (95% CI: 51.1 to 100.0) of those with low baseline total testosterone achieved a normal concentration. Over half (54.6% (95% CI: 31.5 to 94.5)) of the participants who had subnormal free testosterone concentrations at baseline were restored to normal free testosterone concentrations by 5 years. Over the 5-year post-operative period, each 10% improvement in BMI change from baseline was associated with 24% odds of normalization of free testosterone (relative risk (RR): 1.24; 95% CI: 1.02 to 1.51; 0.034)

In crude linear mixed models, HOMA2-IR, BMI and CRP were inversely associated with both total and free testosterone (P  < 0.01 for all). We then utilized linear mixed modeling to evaluate the total and free testosterone concentrations over time, adjusting for covariates. Both BMI and CRP were inversely correlated to total testosterone (each P  < 0.01); that is, as each of these metrics decreased, total testosterone concentration increased. HOMA2-IR was evaluated for inclusion in the final model but was not found to be an important predictor of total testosterone in the presence of other terms (P = 0.26). Only CRP was found to have a significant independent association with free testosterone concentration (P = 0.001). Neither BMI (P = 0.27) nor HOMA2-IR (P = 0.07) was found to be important independent predictors of free testosterone in the final model.

Participants predominantly reached nadir weight at 1 (60%) and 2 years (24%) following surgery. At nadir, the mean percent weight change from baseline was −38.3 ± 11.1%. Weight regain was assessed by the percentage of regain from maximum weight lost. One year following nadir, the median percent regain was 11.4% (25th, 75th percentile: 6.3, 18.6); 2 years after nadir, the mean percent regain was 26.7% (9.6, 37.1). Percent change in free testosterone was modeled as a function of percent regain over the first 2 post-nadir years. After adjustment, increased percent regain was associated with a decline in percent free testosterone change from baseline (Beta: −1.87; 95%CI: −3.30 to −0.43; P = 0.015). In Fig. 2, a visual representation of the effect of weight regain on free testosterone concentrations is provided. No significant difference (P = 0.17) in weight regain was noted between participants undergoing Roux-en-Y gastric bypass or sleeve gastrectomy over the first 2 post-nadir years.

Effect of weight regain on change in free testosterone concentrations. Black circles: median percent change in free testosterone and weight in participants who regained ≥10% of their body weight following nadir (regainers, n  = 10). Black squares: median percent change in free testosterone and weight in participants who maintained their weight or regained <10% (maintainers, n  = 20). Follow-up data beyond 2 years were not available for four participants. Left Y axis: % change in free testosterone (black lines). Right Y axis: % change in weight (gray dotted lines). Error bars are 25th and 75th percentile.

Citation: European Journal of Endocrinology 186, 3; 10.1530/EJE-21-0545

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Effect of weight regain on change in free testosterone concentrations. Black circles: median percent change in free testosterone and weight in participants who regained ≥10% of their body weight following nadir (regainers, n  = 10). Black squares: median percent change in free testosterone and weight in participants who maintained their weight or regained <10% (maintainers, n  = 20). Follow-up data beyond 2 years were not available for four participants. Left Y axis: % change in free testosterone (black lines). Right Y axis: % change in weight (gray dotted lines). Error bars are 25th and 75th percentile.

Citation: European Journal of Endocrinology 186, 3; 10.1530/EJE-21-0545

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Effect of weight regain on change in free testosterone concentrations. Black circles: median percent change in free testosterone and weight in participants who regained ≥10% of their body weight following nadir (regainers, n  = 10). Black squares: median percent change in free testosterone and weight in participants who maintained their weight or regained <10% (maintainers, n  = 20). Follow-up data beyond 2 years were not available for four participants. Left Y axis: % change in free testosterone (black lines). Right Y axis: % change in weight (gray dotted lines). Error bars are 25th and 75th percentile.

Citation: European Journal of Endocrinology 186, 3; 10.1530/EJE-21-0545

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Total estradiol concentrations did not differ across study visits in this male cohort (P = 0.47). However, both BMI and testosterone values were positively associated with total estradiol (each P  < 0.001). Free estradiol values were found to significantly decline by 3 years post-op (P = 0.010). However, by 5 years, the values increased and were comparable to baseline (P = 0.15). In parallel with total estradiol, both BMI and testosterone were positively associated with free estradiol (each P  < 0.001).

LH and FSH concentrations were normal in all participants except one, who had elevated LH and FSH. FSH concentrations were higher than baseline at 2 years (P = 0.03) and tended to be higher (P = 0.07) at years 1 and 4. There was a trend toward an increase in LH concentrations (P = 0.08) at 2 and 4 years. The change in LH and FSH correlated with each other (r= 0.27, P = 0.02) but did not relate to changes in weight, CRP or HOMA2-IR. The participant with high gonadotropins was an 18-year-old male with a BMI of 43 kg/m². His total testosterone was 5 nmol/L at baseline and 7.6 nmol/L at 4 years (at his last available sample). His gonadotropins remained elevated throughout the follow-up.

Discussion

Bariatric surgery and weight loss in adolescent males with severe obesity led to an increase in total and free testosterone concentrations. There was also an increase in SHBG concentration and, therefore, the magnitude of increase in free testosterone concentration was smaller than that of the total testosterone concentration. There was a progressive increase for 2 years in total and free testosterone concentrations following the surgery, after which the concentrations tended to decline. This decrease may have been the result of weight regain that followed.

The restoration of normal testosterone concentrations is important since it is a major mediator of sexual function. A recent study found that penis length was 11% lower in obese boys than lean boys at the completion of puberty. Penile length was directly related to the testosterone concentrations (17). It is of interest that the magnitude of increase in free testosterone concentrations (~100%) in our study was much higher than the increases reported in adult males (~50%) (18). This may be reflective of a more responsive hypothalamo–hypophyseal–gonadal axis in adolescent males than adults(19, 20). An intact hypothalamo–pituitary axis stimulates the testes physiologically via LH and FSH, enhancing both testosterone production and spermatogenesis. Starting in the fourth decade of life, men have a 1% annual decline in serum testosterone concentrations. This is largely due to decreased Leydig cell function (21). Thus, it is possible that Leydig cells in young males are more responsive to the stimulation by gonadotropins. LH and FSH concentrations in our study were normal at baseline and tended to increase after bariatric surgery. This is consistent with data from bariatric studies in adults (5, 18). In addition, testosterone is an insulin sensitizer that leads to loss of adiposity and a gain of muscle and lean body mass (2). In view of the effect of testosterone on adiposity, patients with weight regain must be assessed again for their testosterone status so that the systemic benefits are not lost.

One prior study has assessed total testosterone levels in 16 adolescents following bariatric surgery (22). The total testosterone showed a trend toward a rise (P = 0.07) over 2 years after the surgery. The small sample size likely precluded a significant effect of surgery on weight loss. However, BMI and testosterone were negatively related at year 2 (P = 0.003). Free testosterone was not reported. There was no change in LH or FSH. An increase in testosterone concentrations in adults has been described after bariatric surgery. Testosterone concentrations were evaluated over 5 years after bariatric surgery in 24 middle-aged men with type 2 diabetes in Surgical Therapy And Medications Potentially Eradicate Diabetes Efficiently trial (18). Total and free concentrations increased by 80 and 50%, respectively, at 5 years as compared to baseline. LH concentrations also increased but there was no change in estradiol concentrations. A meta-analysis of 439 men in 16 studies of bariatric surgery showed that total testosterone and SHBG concentrations increased by 8.1 and 22 nmol/L, respectively (5). Eight studies calculated free testosterone concentrations in a total of 259 men and showed an increase of 0.08 nmol/L on average. LH and FSH concentrations increased by 1.0 and 1.8 IU/L, respectively. Estradiol concentrations decreased by an average of 18.4 pmol/L.

We found that estradiol concentrations did not change after bariatric surgery in our study. Testosterone in males can be converted to estradiol through the action of aromatase in the mesenchymal cells and pre-adipocytes of adipose tissue. In fact, it has been suggested that excessive estrogen production due to aromatase activity in males with obesity may potentially suppress the hypothalamic secretion of gonadotropin-releasing hormone (GnRH) (23). However, we have previously shown that this widely believed presumption is not true in adolescents or adults with hypogonadism related to obesity (1, 24). Total and free estradiol concentrations in males with subnormal testosterone were significantly lower than in those with normal testosterone concentrations. Population-based studies such as the European Male Aging Study also showed that estradiol concentrations were lower in hypogonadal men as compared to eugonadal men, regardless of whether the hypogonadism was primary or secondary (21). Thus, an increase in testosterone following bariatric surgery should also increase estradiol concentrations. However, fat loss would reduce the amount of aromatase enzyme and reduce the formation of estradiol. Our data from Teen-LABS above show that following bariatric surgery, the change in free estradiol was related to the rise in both free testosterone and weight. Since weight decreased but testosterone concentrations increased, there was no change in estradiol concentrations.

Bariatric surgery in our participants led to a marked decrease in insulin resistance and CRP concentrations. While HOMA2-IR was related to the change in free testosterone on univariate analysis, we found that after fully adjusting for covariates, a fall in CRP was the strongest predictor of the rise in free testosterone concentrations. It is believed that inflammatory mediators cause a suppression of the hypothalamo–pituitary axis. Tumor necrosis factor α and interleukin-1β have been shown to suppress hypothalamic GnRH and LH secretion in experimental animals and in vitro(25). We have previously shown that CRP concentrations are higher in adolescent males with subnormal free testosterone concentrations than in those with normal testosterone (1). CRP concentrations are also higher in hypogonadal middle-aged men than in eugonadal males (26, 27). It is possible that the reduction in inflammation following bariatric surgery may be responsible for the restoration of hypothalamo–pituitary–gonadal axis. It should be noted that the changes in free testosterone, BMI, CRP and HOMA2-IR occurred simultaneously in our study participants. Hence, it is not possible to be completely certain which factor (or factors) is the cause of an increase in free testosterone following bariatric surgery. Other factors that have been implicated in obesity-related hypogonadism are leptin resistance and insulin resistance in the kisspeptin neurons (2). It is possible that bariatric surgery may restore neuronal signaling of these mediators and enhance GnRH production. Additionally, it is not known if the change in incretin hormones or bile acid profiles after bariatric surgery influences testosterone concentrations.

Our study has some limitations. Firstly, we did not have information on the pubertal status of these participants. Obese males achieve Tanner stage 4 at a median age of 13.5 years and tanner stage 5 at 15.5 years (28). An increase in testosterone is expected with age in younger participants. However, we did not find an effect of age on free testosterone concentrations either at baseline or during the follow-up. The change in free testosterone concentrations following bariatric surgery in our study participants was not different in those who were younger or older than 16 years of age. Thus, it is not likely that our results were affected by the age of the participants. The suppressive effect of severe obesity most likely overwhelms any effect of age. Second, the current ‘gold standard method’ of measuring free testosterone involves the separation of free testosterone by equilibrium dialysis and measurement by mass spectrometry (29). We did not have a sufficient volume of serum to perform equilibrium dialysis on the samples. Free testosterone is commonly calculated from SHBG, albumin and total testosterone concentrations. Although the calculation equation continues to be refined, this calculated free testosterone has been shown to correlate very well with the directly measured free testosterone and is well suited for epidemiological and clinical studies, as well as in clinical practice (29, 30). Third, we do not have a comparator arm of males who did not undergo bariatric surgery. However, it is well known that conventional non-surgical methods such as diet and exercise rarely lead to sustained weight loss. Furthermore, the magnitude of weight loss is much less with medical interventions than with bariatric surgery and does not lead to a change in testosterone concentrations in adults (18, 31). Lastly, we could not collect data on symptoms or signs of hypogonadism in the Teen-LABS study since ours was an ancillary study conceived after the completion and the publication of the original study. We cannot comment on the causes of weight regain since objective physical activity data or detailed diet information during follow-up were not available.

In conclusion, severe obesity is associated with subnormal testosterone concentrations in approximately three-quarters of adolescent males. Bariatric surgery resulted in a doubling of free testosterone concentrations within 2 years of surgery. Weight regain predisposes to relapse subnormal testosterone concentrations. Further studies are needed to investigate the factors contributing to the increase in testosterone concentration following bariatric surgery.

Declaration of interest

S D: Bayer, Clarus Therapeutics, Acerus Pharma (consultant). H G: none. F S: Bayer (consultant). P D: Research Support: National Institutes of Health; JDRF, ADA; Novo Nordisk; Bristol Meyer Squibb; AbbVie Pharmaceuticals; Astra Zeneca, Boehringer Ingelheim Pharmaceuticals. Honorarium: Eli Lilly; Novartis; GlaxoSmithKline; Merck; Novo Nordisk; Takeda; Sanofi-Aventis.

Funding

The NIH (National Institute of Diabetes and Digestive and Kidney Diseases) provided grant funding for the Teen-LABS study through grants UM1DK072493 (University of Colorado) and UM1DK095710 (University of Cincinnati). Funding for this ancillary study was provided by Divisions of Endocrinology of University at Buffalo and Saint Louis University.

Author contribution statement

P D put forth the hypothesis, planned and interpreted the study, and wrote the manuscript. S D, T J and H G executed the study, analyzed data, and wrote the manuscript. H G, A G, Z W and M M analyzed samples. T I, C H and F S critically reviewed the manuscript. P D, S D, T J and H G are the guarantors of this work and, as such, have full access to all the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis.