Measurement Of Free Testosterone Research Articles

A-075 Comparability of Free Testosterone Measurements- Results of the Interlaboratory Study

Abstract Background Assessment of free testosterone (FT) has been recommended as a biomarker for diagnosis and management of hypogonadism and female hyperandrogenism. Reliable laboratory measurements are essential for correctly diagnosing and treating these diseases. FT is currently measured using direct commercial immunoassays, assays employing equilibrium dialysis (ED) or other separation techniques, or it is estimated by calculating free testosterone (cFT). There are concerns about the accuracy and reliability of current methods that may impact patient care. To obtain more information about variability of FT measurements used in patient care, CDC’s Clinical Standardization Programs (CDC CSP) conducted an interlaboratory comparison study. Methods CDC CSP developed an accurate, automated method using ED coupled with isotope dilution ultra-high-performance liquid chromatography tandem mass spectrometry (ED-UHPLC-MS/MS). The ED step follows an internationally recognized procedure. The method is calibrated with primary reference material (National Measurement Institute-M914) and was used to assign target values to sera used in the study. The FT concentrations ranged from 3 to 339 pg/mL. In the interlaboratory comparison study single-donor serum samples were provided to participants operating FT methods used in patient care. The participating methods included commercially available platforms utilizing Chemiluminescence Immunoassay or radioimmunoassay techniques, as well as laboratory-developed assays using ED-LC-MS/MS. Results Preliminary findings of the CSP interlaboratory comparison study found large variability among participating assays. The agreement among IA and among ED-LC-MS/MS assays was close. However, FT concentrations obtained by IAs were in average 6 times lower compared to the ED-LC-MS/MS methods. Results estimated using the Vermeulen equation overestimated FT in average by 15% compared to the ED-LC-MS/MS methods and by over 6 times compared to IAs. Conclusions Large variability is observed between FT laboratory developed tests (LDTs) and IAs. As both LDTs and IAs are used in patient care, standardization of FT measurements to assure better patient care is needed. CDC CSP is addressing the need by developing highly accurate and precise FT method and conducting an interlaboratory comparison studies that can aid with improvement of the agreement among assays and identifying potential factors causing differences among assays.

A-078 Harmonization of Immunoassay Reference Intervals with LC-MS/MS for Total and Calculated Free Testosterone Measurement

Abstract Background Management of male hypogonadism relies on accurate serum testosterone testing and reference values. This study aimed to establish harmonized reference ranges for total and calculated free testosterone in adults using the Siemens Atellica immunoassay, validated against a reference method. Methods The Siemens Atellica IM total testosterone assay is a fully automated competitive chemiluminescent immunoassay. We conducted 25 de-identified serum samples. Inter-laboratory correlation studies for total testosterone were performed on in-house Siemens Atellica and liquid chromatography-tandem mass spectrometry (LC-MS/MS) at a reference laboratory. Free testosterone was calculated using the International Society for the Study of the Aging Male (ISSAM) Free Testosterone calculator, which can be accessed from the ISSAM website, http://www.issam.ch/freetesto.htm, using the assayed concentrations of testosterone and Sex Hormone Binding Globulin (SHBG) on the Siemens Atellica IM system. The Siemens Atellica IM SHBG assay is a sandwich monoclonal immunoassay. Calculated free testosterone was compared with LC-MS/MS. Deming and linear regression, and Bland-Altman plot were employed for data analysis. Reference intervals were adopted from the reference laboratory and verified using 20 male and 20 female healthy samples. Total testosterone: Male 240-950 ng/dL, Female 8-60 ng/dL; Free testosterone: Male 2.29-20.7 ng/dL, Female 0-1.08 ng/dL. Results Strong correlations were observed between Siemens Atellica and LC-MS/MS for total testosterone (R^2 = 0.9915, slope = 0.988, intercept = 35) and for calculated free testosterone (R^2 = 0.9645, slope = 0.988, intercept = -0.4). Adopted reference intervals were verified using local healthy samples. Conclusions Harmonized reference intervals for total and calculated free testosterone, validated on Siemens Atellica, offer consistent results comparable to LC-MS/MS. This study demonstrates the feasibility of establishing standardized reference ranges for testosterone assays across laboratories.

Effect of Metformin and Vitamin D Supplementation on Metabolic and Hormonal Profiles in Polycystic Ovary Syndrome Women: A Follow-Up Study

Background: Insulin resistance in association with obesity is implicated in the pathogenesis of polycystic ovarian syndrome and its primary characteristic of hyperandrogenism. Metformin at a dose of (1700-2000) mg/day, with and without lifestyle, is associated with a beneficial enhancement in BMI and menstrual cycles. We aimed to determine the role of free testosterone: dihydrotestosterone ratio, and sex hormone binding globulin in the evaluation of the efficacy of metformin and vitamin D in the treatment of polycystic ovary syndrome. Methods: This cross-sectional study included 50 infertile women who were diagnosed with PCOS with an age range of (18-40 years). They were classified into Group I: Twenty-five women treated with metformin 850 mg/twice daily and followed for 8 weeks; Group II: Twenty-five women treated with vitamin D3 50.000 IU/wk for the same period. Serum investigations included measurements of prolactin, 25 hydroxyvitamin D, LH, FSH, free testosterone, DHT and SHBG by ELISA technique. Results: This study revealed that the mean values of BMI, HOMA-IR, prolactin and free testosterone: dihydrotestosterone ratio were significantly improved (by lowering) after treatment with metformin and vitamin D. Moreover, Polycystic Ovary Syndrome women treated with metformin revealed significant negative association between HOMA-IR with SHBG (r= -0.593, p=0.009), also significant positive association between HOMA-IR with BMI (r= 0.631, p=0.003). Conclusion: Serum measurements of free testosterone: dihydrotestosterone ratio, along with SHBG, are good biochemical markers that reflect the enhancement of insulin resistance and BMI in Polycystic Ovary Syndrome women post metformin treatment or vitamin D supplementation.

A-048 Accurate Measurement of Free Testosterone in Serum using Equilibrium Dialysis Coupled with ID-UHPLC-MS/MS

Abstract Background Assessment of free testosterone (FT) has been recommended in recent clinical practice guidelines as a biomarker for diagnosis and management of testosterone related disorders such as male testosterone deficiency due to hypogonadism and female androgen excess due to polycystic ovary syndrome (PCOS). FT is currently measured using direct commercial immunoassays, assays using equilibrium dialysis (ED) or other separation techniques. Though the ED approach is recognized as the recommended approach, it is laborious and technically challenging for routine clinical use. The CDC Clinical Standardization Program is developing a high throughput automated method using the ED procedure coupled with isotope dilution ultra-high-performance liquid chromatography tandem mass spectrometry (ID-UHPLC-MS/MS) that is suitable for routine patient care and large-scale epidemiologic studies. Methods Serum samples are dialyzed in a custom-designed multi-well plate against a protein-free HEPES buffer (pH 7.4) at 37 °C until equilibrium. After isolating endogenous FT from protein-bound testosterone by ED, isotope-labeled internal standard (13C3-testosterone) was added to the dialysate for quantification. Certified pure primary reference material (National Measurement Institute-M914) was used to prepare calibrators, enabling traceability and ensuring measurement trueness. FT was further isolated from the dialysate matrix using supported liquid extraction and a chromatographic separation from interfering compounds and quantitation by tandem MS. Results The measurement ranges covered 0.2–1000 ng/dL for testosterone, with the bias within ±5% and precision less than 10% CV. A total of 45 samples with a wide range of total testosterone (TT, 21–912 ng/dL) and SHBG (11–129 nmol/L) were analyzed and showed the suitability of this assay to measure the serum free testosterone levels in normal, hypogonadal males as well as in the majority of normal females and females with androgen excess. The application of 48-well format plate and automated liquid handler system significantly improves the throughput of sample preparation. In addition, comparison of the TT results measured with the presented TT method with calculated TT concentrations, suggest that on average calculation overestimates TT concentration by 34%. Conclusion The described high throughput method for FT allows for sufficiently accurate and precise measurement for routine applications including large epidemiologic studies to help with establishing reference intervals.

ODP419 Measurement of Free Testosterone in Serum using Equilibrium Dialysis Coupled with ID-UHPLC-MS/MS: in Comparison to Calculated FT

Abstract Assessment of free testosterone (FT) has been recommended in recent clinical practice guidelines as a biomarker for diagnosis and management of testosterone related disorders such as male testosterone deficiency due to hypogonadism and female androgen excess due to polycystic ovary syndrome (PCOS). FT is currently measured using direct commercial immunoassays, assays using equilibrium dialysis (ED) or other separation techniques, or estimated by calculating free testosterone (cFT) using different formulas. The most widely used formula was developed by Vermeulen et al and uses total testosterone and sex hormone binding globulin (SHBG) concentrations. The ED approach is recognized as the gold standard; however, it is technically challenging for routine clinical use. CDC's Clinical Standardization Program (CDC CSP) has developed a high throughput method using ED coupled with isotope dilution ultra-high-performance liquid chromatography tandem mass spectrometry. The study to evaluate the agreement of this ED-based method to the cFT using different algorithms is being conducted. Serum samples are dialyzed in a custom-designed multi-well plate against a protein-free HEPES buffer (pH 7.4) at 37 °C until equilibrium. After isolating endogenous FT from protein-bound testosterone by ED, isotope-labeled internal standard (13C3-testosterone) was added to the resulting dialysate for quantification. Certified pure primary reference material (National Measurement Institute-M914) was used to prepare calibrators, enabling traceable quantitation and ensuring measurement trueness. FT was further isolated from the dialysate matrix using supported liquid extraction and a chromatographic separation from interfering compounds and quantitation by tandem MS. The assay allowed detection of 2.9 pg/mL FT, with the bias within ± 5% and precision less than 10% CV. A total of 45 samples with a wide range of TT (21-912 ng/dL) and SHBG (11-129 nmol/L) were analyzed. The obtained FT values were compared to the cFT values obtained with the Vermeulen algorithm. The mean difference of 34.1% (95% CI 29.1% to 39.9%) between the measured and calculated FT were observed, with majority of the calculated values overestimating FT concentrations. The described high throughput method for FT allows for sufficiently accurate and precise measurement and can provide more reliable FT measurements compared to cFT (that can lead to overestimation of FT) for routine applications including large epidemiologic studies. Presentation: No date and time listed

PMON248 Predictive value of markers of hyperandrogenism in 1610 women with PCOS

Abstract Polycystic ovary syndrome (PCOS), an endocrine disorder that affects 10% of women worldwide. In 1990, the NIH established the first consensus on diagnostic criteria for PCOS which included chronic anovulation with clinical and/or biochemical hyperandrogenism, after the exclusion of related disorders. Traditionally the assessment of hyperandrogenism included serum studies of total testosterone (TT), free testosterone (FT), and dehydroepiandrosterone sulfate (DHEA-S), and physical exam assessing hirsutism using the modified Ferriman-Gallwey (mFG) score. The most recent 2018 International Guidelines suggest the measurement of DHEA-S can be considered if TT or FT are not elevated, but probably provide limited diagnostic value. We evaluated the diagnostic utility of TT, FT, DHEA-S and hirsutism in a large prospective cohort of PCOS women. Subjects presented to tertiary-care specialty clinics with symptoms of androgen excess in Birmingham, Alabama and Los Angeles, California from 1987-2010. Clinical hyperandrogenism was assessed using the mFG score, and hirsutism was defined as mFG >4. Participants underwent serum studies to assess for hyperandrogenemia and exclude other causes of hyperandrogenism. Fasting blood samples were obtained on cycle days 3 through 8 of a spontaneous or progesterone-induced vaginal bleed for measurement of TT, FT and DHEA-S. Elevated serum androgens were defined as values exceeding the laboratory reference range or greater than the 95th percentile of reported values. Subjects were considered to have oligomenorrhea if they reported >35 day cycles, <10 bleeds per year or reported being eumenorrheic but were confirmed non-ovulatory with a luteal phase progesterone <4 ng/mL. One thousand six hundred and ten women and adolescents (n=1610), aged > 14 years, were diagnosed with PCOS by NIH Criteria and included. Of these 1610 subjects, 39.3% (n=643) were diagnosed with PCOS based upon an elevated TT and FT with oligomenorrhea; 23.9% (n=384) based upon an elevated TT, normal FT and oligomenorrhea; 11.3% (n=182) based upon an elevated FT, normal TT and oligomenorrhea; 1.1% (n=17) based upon oligomenorrhea with normal TT, normal FT and elevated DHEA-S; and 23.9% (n=384) based upon normal serum androgens, oligomenorrhea, and hirsutism (mFG>4). Results suggest that the diagnosis of PCOS is rarely made based upon elevated DHEA-S in the setting of normal TT and FT. Most women had elevated TT and/or FT values, but still a significant proportion (23.9%) of women were diagnosed with PCOS based upon hirsutism in the context of normal serum androgens. Thus, an assessment of hirsutism, which is both timely and inexpensive, is a strong predictor and valuable tool in diagnosing PCOS, reserving DHEAS when better measures of hyperandrogenism are not diagnostic. Presentation: Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.

P-696 Serum androgen levels and risk of endometrial hyperplasia and malignancy in patients with polycystic ovarian syndrome: a cross-sectional analysis

Abstract Study question In patients with polycystic ovarian syndrome (PCOS), are serum androgen levels (total and free testosterone) associated with the risk of endometrial hyperplasia or cancer? Summary answer Patients with PCOS diagnosed with endometrial hyperplasia or cancer had statistically significantly lower serum levels of androgens compared to PCOS patients with a normal endometrium. What is known already PCOS affects as many as 12% of women of reproductive age. Women with PCOS are almost three times as likely to develop endometrial cancer as women without PCOS, and endometrial hyperplasia precedes cancer in 36% of cases. Known risk factors of endometrial hyperplasia include anovulation, obesity, and resultant unopposed estrogen stimulation of the endometrium. A hallmark of PCOS is hyperandrogenism, however the effect of androgens on the endometrium is unclear. Study design, size, duration This was a cross-sectional, retrospective study of patients aged 18-40, being evaluated for oligomenorrhea and/or PCOS in the Reproductive Endocrinology Clinic at Los Angeles County Hospital, California, USA. All patients underwent an endometrial biopsy and serum measurement of free and total testosterone, estradiol, and hemoglobin A1c. Patients diagnosed with PCOS and with an endometrial biopsy diagnosis were included in the study. Patients on any hormonal therapy were excluded. Participants/materials, setting, methods In all patients included in the study, serum androgen levels (free and total testosterone) were measured by mass spectrometry. All endometrial biopsies were analyzed by an experienced histopathology, and all diagnoses of endometrial hyperplasia or cancer were confirmed by histologic studies. Main results and the role of chance Using the Rotterdam criteria, a total of 232 patients were diagnosed with PCOS during the study period. Of these patients, 161 patients (70%) had a normal endometrium, 19 (8%) were diagnosed with endometrial hyperplasia (EH) without atypia, 38 (16%) were diagnosed with EH with atypia, and 14 (6%) were diagnosed with endometrial cancer. Patients with a normal endometrium had a mean total testosterone of 60.9 ng/dL and mean free testosterone of 9.3 ng/dL. Relatively, patients with EH without atypia, EH with atypia, and endometrial cancer had lower mean total testosterone levels of 50.1 ng/dL, 45.5 ng/dL, and 30.1 ng/dL, respectively (p-values 0.04, 0.02, 0.001, respectively), as well as lower mean free testosterone levels of 8.2 pg/mL, 6.7 pg/mL, and 5.5 pg/mL, respectively (p-values 0.04, 0.03, and 0.01, respectively). There was no significant difference in age, parity, BMI, estradiol levels, or hemoglobin A1c between the four groups. Limitations, reasons for caution Limitations of our study include its retrospective nature. Furthermore, hormone levels were only measured in the serum, and not in the endometrial tissue directly, which would give the most insight into their actions on the endometrium. Wider implications of the findings Our findings suggest that androgens have a protective effect on the endometrium against endometrial hyperplasia and cancer. Further studies are needed to evaluate the therapeutic potential role of androgens in the prevention or treatment of these conditions. Trial registration number not applicable

Preliminary comparison of the measurement of free testosterone with a chemiluminescence and a radioimmunoassay method

article: Preliminary comparison of the measurement of free testosterone with a chemiluminescence and a radioimmunoassay method - La Rivista Italiana della Medicina di Laboratorio 2022 Marzo;18(1):37-41 - Minerva Medica - Journals

Detection of Male Hypogonadism in Patients with Post COVID-19 Condition.

The pathogenesis and prognosis of post COVID-19 condition have remained unclear. We set up an outpatient clinic specializing in long COVID in February 2021 and we have been investigating post COVID-19 condition. Based on the results of our earlier study showing that “general fatigue” mimicking myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is the most common symptom in long COVID patients, a retrospective analysis was performed for 39 male patients in whom serum free testosterone (FT) levels were measured out of 61 male patients who visited our clinic. We analyzed the medical records of the patients’ backgrounds, symptoms and laboratory results. Among the 39 patients, 19 patients (48.7%) met the criteria for late-onset hypogonadism (LOH; FT < 8.5 pg/mL: LOH group) and 14 patients were under 50 years of age. A weak negative correlation was found between age and serum FT level (r = −0.301, p = 0.0624). Symptoms including general fatigue, anxiety, cough and hair loss were more frequent in the LOH group than in the non-LOH group (FT ≥ 8.5 pg/mL). Among various laboratory parameters, blood hemoglobin level was slightly, but significantly, lower in the LOH group. Serum level of FT was positively correlated with the levels of blood hemoglobin and serum total protein and albumin in the total population, whereas these interrelationships were blurred in the LOH group. Collectively, the results indicate that the incidence of LOH is relatively high in male patients, even young male patients, with post COVID-19 and that serum FT measurement is useful for revealing occult LOH status in patients with long COVID.

The importance of association between sexsteroids deficiency, reduction of bone mineral density and falling risk in men with implications in medical rehabilitation

Introduction. Endocrino-metabolic rehabilitation represent one of the most complex sector in clinical medicine, regarding functional rehabilitation. Sex hormones deficiency plays an important role in the etiology of osteoporosis in men. At the same time, with age, the trophic role of androgens on muscle decreases and determines an increased frequency of falls. The objective of our study is to determine the association between sexsteroids deficiency, reduction of bone mineral density (BMD) and falling risk in men. Methods. Our retrospective cross-sectional study included 146 men aged between 65–85 years with low BMD (study group) and 121 men with normal BMD (control group). The measurement of Total testosterone (Tt), free testosterone (Tf) and estradiol (E2) serum levels was performed using the immunoassay or the immunoenzymatic methods. Femoral neck and lumbar spine BMD was determined using Dual-energy X-ray absorptiometry (DEXA). The risk of falls was assessed by Tandem Standing, Up &amp; Go, Chair – Rising and walking speed tests. Results. We found a significantly association between Tf and E2 deficiency and low BMD (p=0.007). Also, in men with reduced BMD (study group) we observed significant lower levels of Tf (p&lt;0.001) and E2 (p=0.003) compared to control group. E2 deficiency was associated significantly with low BMD and increased fall risk (p=0.001). At the same time the results highlighted significant lower levels of Tf in patients with BMD reduction and increased risk of falls (p=0.002). Tt deficiency was not associated with BMD reduction (p=0.088) or increased risk of falling (p=0.277). Conclusions. This research revealed a significant association between male sexsteroids deficiency, low BMD and increase of falling risk, with implications in rehabilitation program. The risk of ostoporosis and for falling in man can be estimated by determining serum Tf and E2 levels. Keywords: sexsteroids deficiency; bone mineral density; falling risk,

Semen parameters in Egyptian diabetic patients and its correlation with glycosylated hemoglobin level: A case - control study

Background: Diabetes mellitus (DM) is a well-known metabolic disorder affecting a large population worldwide. Both types of DM could have detrimental effects on semen parameters as well as the hormonal profile of the affected males.Objective: On designing the current study, we sought to evaluate the effect of long-term control of DM by measuring glycosylated hemoglobin (HbA1c) level and detect its possible correlation with semen parameters and hormonal profile of male patients with DM.Patients and Methods: A prospective case–control study was conducted on 60 males with DM and 40 matched controls. All participants were subjected to conventional semen analysis and measurement of free and total testosterone, follicle-stimulating hormone, luteinizing hormone, and prolactin levels by enzyme-linked immunosorbent assay technique. Measurement of HbA1c level was made for all participants.Results: A significant negative correlation was found between HbA1c and free testosterone levels, whereas a significant positive correlation between HbA1c and prolactin levels in patients with DM. No significant correlation was reported between HbA1c and semen parameters in patients with DM.Conclusion: Patients with DM have significantly lower semen parameters and reproductive hormone levels than matched healthy controls which might impair their fecundity and reproductive capacity. We do recommend that all males with abnormal semen analysis should undergo screening for DM.

SAT-LB5 Prevalence of Hypogonadism in Young Obese Males

Ageing, obesity, and chronic illness are major factors affecting serum testosterone (T) levels in men.The magnitude of the impact of ageing on serum T levels is well established, for obesity this is less clear. Severe obesity may lead to isolated hypogonadotropic hypogonadism (IHH). Several explanations have been offered to clarify the presence of reduced T levels in obese men. One relates to the technique that is generally employed to measure serum androgen levels, i.e. measurement of total testosterone (TT) instead of free testosterone (FT). TT represents the sum of FT and T bound to albumin and sex hormone binding globulin (SHBG). A profound reduction in SHBG level is commonly found in obese men, and this is a major factor causing a decrease in TT.Measurement of free testosterone levels may provide a more accurate assessment of androgen status than the (usually preferred) measurement of total testosterone in situations where SHBG levels are outside the reference range. However, reference ranges for free testosterone levels are not well established, especially in older men, and some have argued that the measurement of free testosterone levels merely reintroduces age in a covert form. This is a cross sectional study to estimate prevalence of hypogonadism in young obese males. In this study 147 young obese men participated, of which we confirmed low total testosterone (TT) levels in 35.37% of subjects with a p value of 0.06. Since only Total Testosterone was measured for categorizing subjects with or without hypogonadism, Free Testosterone measurement would be a better indicator for the diagnosis of hypogonadism as in cases where the total testosterone is borderline-low or when SHBG concentrations are abnormal. As such, the study is valuable in the context of the ongoing controversy as to whether testosterone treatment should be limited to men with classical hypogonadism, or be considered for appropriately selected men with functional hypogonadism as well. The principal findings are in general agreement with existing literature reporting correlation between levels of testosterone, body mass index and constitutional symptoms. However, this has never been shown before in context of Indian population. The present study was carried out at Armed Forces Medical College and Command Hospital, Pune between October 2017 to August 2019.We studied to see if there is association between testosterone levels and BMI. In our study we found no statistical association as the p value was 0.26 (>0.05)

Phthalates and Sex Steroid Hormones Among Men From NHANES, 2013-2016.

Phthalates are commonly found in commercial packaging, solvents, vinyl, and personal care products, and there is concern for potential endocrine-disrupting effects in males. The commonly used di-2-ethylhexyl phthalate (DEHP) has progressively been replaced by seldom studied compounds, such as bis-2-ethylhexyl terephthalate and 1,2-cyclohexane dicarboxylic acid di-isononyl ester (DINCH). To investigate the associations between the urinary phthalate metabolites and serum sex steroid hormone concentrations in a nationally representative sample of adult males. This was a cross-sectional analysis of data from the 2013-2016 National Health and Nutrition Examination Survey among 1420 male participants aged ≥20 years. Serum levels of total testosterone, estradiol, SHBG, and derived sex hormone measurements of free testosterone, bioavailable testosterone, and free androgen index were examined as log-transformed continuous variables. Phthalate metabolites were not statistically significantly associated with sex hormone concentrations among all men. However, associations varied by age. High molecular weight phthalates were associated with lower total, free, and bioavailable testosterone among men age ≥60. Specifically, each doubling of ΣDEHP was associated with 7.72% lower total testosterone among older men (95% confidence interval, -12.76% to -2.39%). Low molecular phthalates were associated with lower total, free, and bioavailable testosterone among men age 20 to 39 and ∑DINCH was associated with lower total testosterone among men age ≥40. Our results indicate that males may be vulnerable to different phthalate metabolites in age-specific ways. These results support further investigation into the endocrine-disrupting effects of phthalates.

Screening and Management of the Hyperandrogenic Adolescent: ACOG Committee Opinion Summary, Number 789.

Although androgen excess can manifest in many ways, the most common and recognizable symptoms are hirsutism and acne. Reports of hirsutism and acne should be taken seriously because of their possible association with medical disorders, their substantial effect on self-esteem and quality of life, and the potential for psychosocial morbidity. In patients with symptoms of androgen excess, the differential diagnosis should include physiologic hyperandrogenism of puberty, idiopathic hyperandrogenism, and polycystic ovary syndrome (PCOS). There is a great deal of overlap between the symptoms of PCOS and those of normal puberty, which makes the diagnosis of PCOS in the adolescent difficult. Treatment of acne and hirsutism should not be withheld during the ongoing longitudinal evaluation for possible PCOS. On physical examination, body mass index, blood pressure, and signs of hyperandrogenism, such as acne and hirsutism, should be evaluated. Although guidelines differ on recommended laboratory studies, most include measurement of total testosterone, free testosterone, or both, and screening for nonclassic congenital adrenal hyperplasia with a 17-hydroxyprogesterone test. Elevation of the free or total testosterone level higher than the adult female normative values is a key diagnostic feature of biochemical hyperandrogenism. Because treatment is indicated only when symptoms are distressing to the patient, the degree to which acne or hirsutism bothers the patient should be assessed. Before initiation of any medical therapy, expectations of treatment should be discussed with the patient. Anticipatory guidance is critical to help patients understand the timeline for expected responses to therapy.

Screening and Management of the Hyperandrogenic Adolescent: ACOG Committee Opinion, Number 789.

Although androgen excess can manifest in many ways, the most common and recognizable symptoms are hirsutism and acne. Reports of hirsutism and acne should be taken seriously because of their possible association with medical disorders, their substantial effect on self-esteem and quality of life, and the potential for psychosocial morbidity. In patients with symptoms of androgen excess, the differential diagnosis should include physiologic hyperandrogenism of puberty, idiopathic hyperandrogenism, and polycystic ovary syndrome (PCOS). There is a great deal of overlap between the symptoms of PCOS and those of normal puberty, which makes the diagnosis of PCOS in the adolescent difficult. Treatment of acne and hirsutism should not be withheld during the ongoing longitudinal evaluation for possible PCOS. On physical examination, body mass index, blood pressure, and signs of hyperandrogenism, such as acne and hirsutism, should be evaluated. Although guidelines differ on recommended laboratory studies, most include measurement of total testosterone, free testosterone, or both, and screening for nonclassic congenital adrenal hyperplasia with a 17-hydroxyprogesterone test. Elevation of the free or total testosterone level higher than the adult female normative values is a key diagnostic feature of biochemical hyperandrogenism. Because treatment is indicated only when symptoms are distressing to the patient, the degree to which acne or hirsutism bothers the patient should be assessed. Before initiation of any medical therapy, expectations of treatment should be discussed with the patient. Anticipatory guidance is critical to help patients understand the timeline for expected responses to therapy.

37-Year-Old Man With Fatigue and Erectile Dysfunction

37-Year-Old Man With Fatigue and Erectile Dysfunction

Measurement of free testosterone, androstenedione, and 17-hydroxyprogesterone: Comparison of current laboratory methods

Measurement of free testosterone, androstenedione, and 17-hydroxyprogesterone: Comparison of current laboratory methods

Evaluation of free testosterone measurement on the ids-isys automated platform as compared to RIA

Elevation of blood triglycerides, primarily triglyceride-rich lipoproteins (TGRL), is an independent risk factor for cardiovascular disease and vascular dementia (VaD). Accumulating evidence indicates that both atherosclerosis and VaD are linked to vascular inflammation. However, the role of TGRL in vascular inflammation, which increases risk for VaD, remains largely unknown and its underlying mechanisms are still unclear. We strived to determine the effects of postprandial TGRL exposure on brain microvascular endothelial cells, the potential risk factor of vascular inflammation, resulting in VaD. We showed in Aung et al., J Lipid Res., 2016 that postprandial TGRL lipolysis products (TL) activate mitochondrial reactive oxygen species (ROS) and increase the expression of the stress-responsive protein, activating transcription factor 3 (ATF3), which injures human brain microvascular endothelial cells (HBMECs) in vitro. In this study, we deployed high-throughput sequencing (HTS)-based RNA sequencing methods and mito stress and glycolytic rate assays with an Agilent Seahorse XF analyzer and profiled the differential expression of transcripts, constructed signaling pathways, and measured mitochondrial respiration, ATP production, proton leak, and glycolysis of HBMECs treated with TL. Conclusions: TL potentiate ROS by mitochondria which activate mitochondrial oxidative stress, decrease ATP production, increase mitochondrial proton leak and glycolysis rate, and mitochondria DNA damage. Additionally, CPT1A1 siRNA knockdown suppresses oxidative stress and prevents mitochondrial dysfunction and vascular inflammation in TL treated HBMECs. TL activates ATF3-MAPKinase, TNF, and NRF2 signaling pathways. Furthermore, the NRF2 signaling pathway which is upstream of the ATF3-MAPKinase signaling pathway, is also regulated by the mitochondrial oxidative stress. We are the first to report differential inflammatory characteristics of transcript variants 4 (ATF3-T4) and 5 (ATF3-T5) of the stress responsive gene ATF3 in HBMECs induced by postprandial TL. Specifically, our data indicates that ATF3-T4 predominantly regulates the TL-induced brain microvascular inflammation and TNF signaling. Both siRNAs of ATF3-T4 and ATF3-T5 suppress cells apoptosis and lipotoxic brain microvascular endothelial cells. These novel signaling pathways triggered by oxidative stress-responsive transcript variants, ATF3-T4 and ATF3-T5, in the brain microvascular inflammation induced by TGRL lipolysis products may contribute to pathophysiological processes of vascular dementia.

Validity of free testosterone calculation in pregnant women.

ObjectiveIncreased maternal testosterone concentration during pregnancy may affect the fetus. Therefore it is clinically relevant to have a quick and reliable method to determine free testosterone levels. Current calculators for free testosterone are suspected to perform poorly during pregnancy due to suggested competition between high levels of estradiol and free (bio-active) testosterone for sex hormone-binding globulin (SHBG) binding. Therefore, it is claimed that reliable calculation of free testosterone concentration is not possible. However, recent evidence on SHBG-binding sites questions the estradiol effect on the testosterone-SHBG binding during pregnancy. In this study, we investigated whether the free testosterone concentration can be calculated in pregnant women.Design and methodsFree testosterone was measured with a specially developed equilibrium dialysis method combined with liquid chromatography tandem mass spectrometry (LC-MS/MS). Free testosterone was also calculated with the formulas of Vermeulen et al. and Ross et al. ResultsTotal and free testosterone measured in healthy men and women were in good agreement with earlier reports. In pregnant women, total testosterone values were higher than in non-pregnant women, whereas free testosterone values were comparable. Calculated free testosterone levels in pregnant women were highly correlated, but marginally higher, compared to measured free testosterone levels.ConclusionsWe developed an equilibrium dialysis–LC-MS/MS method for the measurement of free testosterone in the low range of pregnant and non-pregnant women. Although during pregnancy total testosterone is increased, this is not the case for free testosterone. The free testosterone formulas perform well in pregnant women.

Salivary versus Serum Testosterone Levels in Boys with Constitutional Delay of Growth and Puberty

BackgroundSalivary testosterone (Sal-T) has been established as a noninvasive biomarker in the diagnosis of androgen deficiency in men. However, available data on its utility in adolescent boys with constitutional delay of growth and puberty (CDGP) are not conclusive. Our study was designed to compare salivary versus serum measurements of free testosterone (FT) from the samples collected simultaneously and correlated them with the clinical parameters in boys with CDGP.Patients and methodsThe study enrolled 25 adolescent boys with CDGP and 20 healthy controls matched for age and sex. Weight, height, BMI, testicular volume, bone age, serum follicle-stimulating hormone (FSH), and luteinizing hormone (LH) were assessed. Simultaneous morning saliva and serum samples were obtained for FT measurements by radioimmunoassay and Sal-T by the enzyme-linked immunosorbent assay technique.ResultsAdolescent boys with CDGP had significantly lower Sal-T and serum free-T than normal controls (P<0.001). Sal-T positively correlated with BMI, bone age, testicular volume, serum FSH, LH, and FT. With multiple regression analysis BMI, FSH, LH, and serum FT remained independently correlated with Sal-T. Using the receiver operating characteristic curve, Sal-T was found to be more sensitive than serum FT (76 vs. 69%) in the clinical assessment of boys with CDGP.ConclusionOur data shows that Sal-T strongly correlates with FT and provides a sensitive, simple, noninvasive, and diagnostic approach for androgen status in adolescent boys with CDGP.