Dr. Ron’s Research Review – May 1, 2019

©

This week’s research review focuses on free and bioavailable hormones

Hormones are bound to carrier proteins in serum. Carrier proteins include: albumin, thyroid binding globulin, sex-hormone binding globulin, cortisol binding globulin and vitamin D binding protein. Free hormone concentrations can be measured or calculated.
Hormones are weakly bound to albumin. Hence, the bioavailable concentration includes the fraction bound to albumin. Bioavailable = free + albumin-bound.
Disparity in calculated free hormones reflects the competition effect between E2, E1 and T for the SHBG binding sites. (Mazer, 2009)

T4

Only a tiny percentage (approximately 0.01%) of the total T4 present in blood is free, primarily because of the very high affinity of the major carrier protein (thyroid binding globulin or TBG). Approximately 70% of the total T4 is bound to TBG; approximately 30% is bound equally to transthyretin and albumin and a small percentage is bound to a variety of apolipoproteins. (Berrino et al., 2001)

Testosterone

Like T4, testosterone is poorly soluble in blood. In both men and women, approximately 98% of the total testosterone in plasma is protein-bound, leaving only approximately 2% free. Testosterone is bound to both sex-hormone binding globulin (SHBG) and albumin. (Berrino et al., 2001)
In healthy adult men, ~44% of the circulating testosterone is specifically bound to sex hormone–binding globulin (SHBG), 50% is nonspecifically bound to albumin, and 3.5% is bound to cortisol-binding globulin, indicating that only 2%–3% is unbound or free. Bioavailable testosterone (bioT) = free + albumin-bound testosterone. (de Ronde et al., 2006)

Estradiol

The two proteins that bind E2 are SHBG and albumin. The calculation of free E2 has been compared to measured values, and found to be satisfactory, in only a small sample of postmenopausal patients; the authors caution against using the calculated value in other populations. Moreover, it should be recalled that E2 is bound to SHBG 2–3-fold less tightly to SHBG than is testosterone, rendering the assumption of a fixed value for albumin unacceptable until the appropriate calculations and experiments are at hand. (Rosner, 2015)

Dr. Ron

Articles

Reducing bioavailable sex hormones through a comprehensive change in diet: the diet and androgens (DIANA) randomized trial.
            (Berrino et al., 2001)  Download
High serum levels of testosterone and estradiol, the bioavailability of which may be increased by Western dietary habits, seem to be important risk factors for postmenopausal breast cancer. We hypothesized that an ad libitum diet low in animal fat and refined carbohydrates and rich in low-glycemic-index foods, monounsaturated and n-3 polyunsaturated fatty acids, and phytoestrogens, might favorably modify the hormonal profile of postmenopausal women. One hundred and four postmenopausal women selected from 312 healthy volunteers on the basis of high serum testosterone levels were randomized to dietary intervention or control. The intervention included intensive dietary counseling and specially prepared group meals twice a week over 4.5 months. Changes in serum levels of testosterone, estradiol, and sex hormone-binding globulin were the main outcome measures. In the intervention group, sex hormone-binding globulin increased significantly (from 36.0 to 45.1 nmol/liter) compared with the control group (25 versus 4%,; P < 0.0001) and serum testosterone decreased (from 0.41 to 0.33 ng/ml; -20 versus -7% in control group; P = 0.0038). Serum estradiol also decreased, but the change was not significant. The dietary intervention group also significantly decreased body weight (4.06 kg versus 0.54 kg in the control group), waist:hip ratio, total cholesterol, fasting glucose level, and area under insulin curve after oral glucose tolerance test. A radical modification in diet designed to reduce insulin resistance and also involving increased phytoestrogen intake decreases the bioavailability of serum sex hormones in hyperandrogenic postmenopausal women. Additional studies are needed to determine whether such effects can reduce the risk of developing breast cancer.

Calculation of bioavailable and free testosterone in men: a comparison of 5 published algorithms.
            (de Ronde et al., 2006)  Download
BACKGROUND:  Estimation of serum concentrations of free testosterone (FT) and bioavailable testosterone (bioT) by calculation is an inexpensive and uncomplicated method. We compared results obtained with 5 different algorithms. METHODS:  We used 5 different published algorithms [described by Sodergard et al. (bioTS and FTS), Vermeulen et al. (bioTV and FTV), Emadi-Konjin et al. (bioTE), Morris et al. (bioTM), and Ly et al. (FTL)] to estimate bioT and FT concentrations in samples obtained from 399 independently living men (ages 40-80 years) participating in a cross-sectional, single-center study. RESULTS:  Mean bioT was highest for bioTS (10.4 nmol/L) and lowest for bioT(E) (3.87 nmol/L). Mean FT was highest for FTS (0.41 nmol/L), followed by FTV (0.35 nmol/L), and FTL (0.29 nmol/L). For bioT concentrations, the Pearson correlation coefficient was highest for the association between bioTS and bioTV (r = 0.98) and lowest between bioTM and bioTE (r = 0.66). FTL was significantly associated with both FTS (r = 0.96) and FTV (r = 0.88). The Pearson correlation coefficient for the association between FTL and bioTM almost reached 1.0. Bland-Altman analysis showed large differences between the results of different algorithms. BioTM, bioTE, bioTV, and FTL were all significantly associated with sex hormone binding globulin (SHBG) concentrations. CONCLUSION:  Algorithms to calculate FT and bioT must be revalidated in the local setting, otherwise over- or underestimation of FT and bioT concentrations can occur. Additionally, confounding of the results by SHBG concentrations may be introduced.

A novel spreadsheet method for calculating the free serum concentrations of testosterone, dihydrotestosterone, estradiol, estrone and cortisol: with illustrative examples from male and female populations.
            (Mazer, 2009)  Download
In humans, testosterone (T), dihydrotestosterone (DHT), estradiol (E2), estrone (E1) and cortisol (C) bind to the serum proteins sex hormone-binding globulin (SHBG), albumin (Alb) and corticosteroid-binding globulin (CBG). Equilibrium dialysis is considered to be the "gold standard" for measuring the free concentrations of these steroids but is technically difficult and not widely available. Based on a mathematical model of the 5-ligand/3-protein binding equilibria, we developed a novel spreadsheet method for calculating the free and bioavailable (free+Alb-bound) concentrations of each steroid in terms of the total steroid and protein concentrations. The model uses 15 association constants K(SHBG-X), K(Alb-X), and K(CBG-X) (X=T, DHT, E2, E1 and C) that have been estimated from a systematic review of published binding studies. The computation of the free and bioavailable concentrations uses an iterative numerical method that can be readily programmed on a spreadsheet. The method is illustrated with six examples corresponding to young men (YM), old men (OM), obese men (Ob M), young women (YM), pregnant women in the 3rd trimester (Preg T3) and oophorectomized women on oral conjugated equine estrogens (CEE). The resulting free hormone concentrations for YM and YW fall within the normal references ranges obtained by equilibrium dialysis for all five hormones. The model also accounts for the competitive binding effects of high estrogen levels on the free T levels in Preg T3. This novel spreadsheet method provides a "user-friendly" approach for estimating the free concentrations of circulating sex hormones and cortisol in men and women.


 

Free estradiol and sex hormone-binding globulin.
            (Rosner, 2015)  Download
SHBG is a plasma protein that participates in the regulation of free estradiol and free testosterone in plasma. We discuss the concept of the nature of a free estradiol and how best to ascertain its value. It can be measured or calculated; the ways in which this can be done are explored along with the advantages and disadvantages of each.

 

References

Berrino, F, et al. (2001), ‘Reducing bioavailable sex hormones through a comprehensive change in diet: the diet and androgens (DIANA) randomized trial.’, Cancer Epidemiol Biomarkers Prev, 10 (1), 25-33. PubMed: 11205485
de Ronde, W, et al. (2006), ‘Calculation of bioavailable and free testosterone in men: a comparison of 5 published algorithms.’, Clin Chem, 52 (9), 1777-84. PubMed: 16793931
Mazer, NA (2009), ‘A novel spreadsheet method for calculating the free serum concentrations of testosterone, dihydrotestosterone, estradiol, estrone and cortisol: with illustrative examples from male and female populations.’, Steroids, 74 (6), 512-19. PubMed: 19321131
Rosner, W (2015), ‘Free estradiol and sex hormone-binding globulin.’, Steroids, 99 (Pt A), 113-16. PubMed: 25453337