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Testosterone Replacement, Low T, HCG, & Beyond
Testosterone and Men's Health Articles
What's Coming? Accurate measurement of total and free testosterone levels for the diagnosis of androgen disorders
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<blockquote data-quote="madman" data-source="post: 228759" data-attributes="member: 13851"><p>Critical points for everyone to keep in mind:</p><p></p><p><em><strong>*SHBG circulates as a homodimer with a single binding site on each of the two monomers. Recent studies using modern biophysical techniques and molecular modeling have shown that the binding of testosterone, as well as estradiol to SHBG, is a complex, multi-step, dynamic process that involves inter-monomeric allostery such that the <u>binding affinities, conformations, and energy states of the two monomers are not identical in unbound or even fully bound states</u>.7,8 </strong></em></p><p><em><strong></strong></em></p><p><em><strong><em><strong>*The binding of testosterone to human serum albumin also is far more complex than had been recognized previously; our recent studies show that there are multiple, allosterically-coupled binding sites for testosterone on human serum albumin. 9</strong> <strong>Testosterone shares these binding sites on human serum albumin with free fatty acids and many commonly used drugs such as ibuprofen and coumadin, which could displace testosterone from its binding sites under various physiological states or disease conditions, affecting its bioavailability </strong></em></strong></em></p><p><em><strong><em><strong></strong></em></strong></em></p><p><em><strong><em><strong>*<em><strong> It is generally believed that testosterone can rapidly dissociate from one or more binding sites on human serum albumin and become "bioavailable" in some target organs, especially in target organs with long transit times such as the liver and the brain; this premise has not been fully substantiated. The characteristics of testosterone binding to CBG and orosomucoid remain incompletely understood</strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong>*<em><strong> It has been postulated that the HSA-bound testosterone is so loosely bound that it can dissociate in the tissue capillaries and essentially become “free.”14,15 There is also a question of whether SHBG-bound testosterone can be internalized into the cell mediated by the membrane protein, megalin.16 The tertiary complex is delivered to the lysosomal compartments where the testosterone dissociates from the binding proteins</strong></em></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong>*<em><strong>Polymorphisms in the SHBG gene can cause elevated or decreased SHBG levels. Increased SHBG levels are associated with the variants rs6258 and rs12150660 and decreased SHBG levels are associated with the variants rs6257, rs6259, rs727428, rs1799941.38-40 Some of the SHBG polymorphisms can also affect its binding to testosterone while others can affect its clearance and dimerization</strong></em></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong><em><strong></strong></em></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong><em><strong>*<strong><em>Free testosterone concentration should be measured directly, preferably using an equilibrium dialysis assay in a reliable laboratory. 6 Lack of standardization of the equilibrium dialysis method has impeded efforts to generate harmonized reference ranges for free testosterone levels.6 </em></strong></strong></em></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong>*The lack of standardization of the equilibrium dialysis method among laboratories has been a barrier to the generation of a harmonized reference range for free testosterone levels; until such rigorously-derived harmonized reference ranges become available, the clinicians currently must rely on reference ranges provided by a laboratory6 or those published from the analyses of large epidemiologic studies.7</strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong><em><strong><em><strong><em><strong>*Free androgen index does not provide an accurate or rational estimate of free testosterone concentration and its use is not recommended</strong></em></strong></em></strong></em></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong><em><strong><em><strong><em><strong></strong></em></strong></em></strong></em></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong><em><strong><em><strong><em><strong>*Tracer analog methods have been shown to be inaccurate and should be avoided</strong></em></strong></em></strong></em></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong><em><strong><em><strong><em><strong></strong></em></strong></em></strong></em></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong><em><strong><em><strong><em><strong>*<em><strong>Two categories of algorithms to calculate FT from the TT, SHBG, and albumin levels have been published; the linear equations that are based on the assumption of a simple linear model of testosterone's binding to SHBG and human serum albumin with a single fixed Kd, and those that are empirically derived from regression analyses of the relation between free testosterone and total testosterone and SHBG levels. The linear equations for FT estimation are based on the assumption that each SHBG dimer binds two testosterone molecules and that the two binding sites on SHBG have similar binding affinity; <u>these assumptions are not supported by our current understanding of the dynamics of testosterone's binding to SHBG. cFT values obtained using a multi-step, allosteric ensemble model are much closer to those obtained using equilibrium dialysis</u>. 7,49</strong></em></strong></em></strong></em></strong></em></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong><em><strong></strong></em></strong></em></strong></em></strong></em></p><p><em><strong><em><strong><em><strong><em><strong>*<strong><em>Testosterone levels should be measured preferably in a CDCcertified laboratory using validated assays; in cases of equivocal TT concentration and/or abnormal SHBG levels, free testosterone levels should be measured using equilibrium dialysis or <u>calculated using an equation based on our current understanding of the dynamics of testosterone binding, such as the ensemble allostery model</u></em></strong></strong></em></strong></em></strong></em></strong></em></p></blockquote><p></p>
[QUOTE="madman, post: 228759, member: 13851"] Critical points for everyone to keep in mind: [I][B]*SHBG circulates as a homodimer with a single binding site on each of the two monomers. Recent studies using modern biophysical techniques and molecular modeling have shown that the binding of testosterone, as well as estradiol to SHBG, is a complex, multi-step, dynamic process that involves inter-monomeric allostery such that the [U]binding affinities, conformations, and energy states of the two monomers are not identical in unbound or even fully bound states[/U].7,8 [I][B]*The binding of testosterone to human serum albumin also is far more complex than had been recognized previously; our recent studies show that there are multiple, allosterically-coupled binding sites for testosterone on human serum albumin. 9[/B] [B]Testosterone shares these binding sites on human serum albumin with free fatty acids and many commonly used drugs such as ibuprofen and coumadin, which could displace testosterone from its binding sites under various physiological states or disease conditions, affecting its bioavailability *[I][B] It is generally believed that testosterone can rapidly dissociate from one or more binding sites on human serum albumin and become "bioavailable" in some target organs, especially in target organs with long transit times such as the liver and the brain; this premise has not been fully substantiated. The characteristics of testosterone binding to CBG and orosomucoid remain incompletely understood *[I][B] It has been postulated that the HSA-bound testosterone is so loosely bound that it can dissociate in the tissue capillaries and essentially become “free.”14,15 There is also a question of whether SHBG-bound testosterone can be internalized into the cell mediated by the membrane protein, megalin.16 The tertiary complex is delivered to the lysosomal compartments where the testosterone dissociates from the binding proteins[/B][/I] *[I][B]Polymorphisms in the SHBG gene can cause elevated or decreased SHBG levels. Increased SHBG levels are associated with the variants rs6258 and rs12150660 and decreased SHBG levels are associated with the variants rs6257, rs6259, rs727428, rs1799941.38-40 Some of the SHBG polymorphisms can also affect its binding to testosterone while others can affect its clearance and dimerization *[B][I]Free testosterone concentration should be measured directly, preferably using an equilibrium dialysis assay in a reliable laboratory. 6 Lack of standardization of the equilibrium dialysis method has impeded efforts to generate harmonized reference ranges for free testosterone levels.6 [/I][/B][/B][/I] *The lack of standardization of the equilibrium dialysis method among laboratories has been a barrier to the generation of a harmonized reference range for free testosterone levels; until such rigorously-derived harmonized reference ranges become available, the clinicians currently must rely on reference ranges provided by a laboratory6 or those published from the analyses of large epidemiologic studies.7 [I][B][I][B][I][B]*Free androgen index does not provide an accurate or rational estimate of free testosterone concentration and its use is not recommended *Tracer analog methods have been shown to be inaccurate and should be avoided *[I][B]Two categories of algorithms to calculate FT from the TT, SHBG, and albumin levels have been published; the linear equations that are based on the assumption of a simple linear model of testosterone's binding to SHBG and human serum albumin with a single fixed Kd, and those that are empirically derived from regression analyses of the relation between free testosterone and total testosterone and SHBG levels. The linear equations for FT estimation are based on the assumption that each SHBG dimer binds two testosterone molecules and that the two binding sites on SHBG have similar binding affinity; [U]these assumptions are not supported by our current understanding of the dynamics of testosterone's binding to SHBG. cFT values obtained using a multi-step, allosteric ensemble model are much closer to those obtained using equilibrium dialysis[/U]. 7,49[/B][/I][/B][/I][/B][/I] *[B][I]Testosterone levels should be measured preferably in a CDCcertified laboratory using validated assays; in cases of equivocal TT concentration and/or abnormal SHBG levels, free testosterone levels should be measured using equilibrium dialysis or [U]calculated using an equation based on our current understanding of the dynamics of testosterone binding, such as the ensemble allostery model[/U][/I][/B][/B][/I][/B][/I][/B][/I][/B][/I] [/QUOTE]
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Testosterone Replacement, Low T, HCG, & Beyond
Testosterone and Men's Health Articles
What's Coming? Accurate measurement of total and free testosterone levels for the diagnosis of androgen disorders
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