Brigham Research Assay Core Laboratory, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA.
Research Program in Men's Health: Aging and Metabolism, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA.
J Gerontol A Biol Sci Med Sci. 2022 Apr 1;77(4):763-769. doi: 10.1093/gerona/glab146.
Growth and differentiation factor (GDF)-11 controls embryonic development and has been proposed as an antiaging factor. GDF-8 (myostatin) inhibits skeletal muscle growth. Difficulties in accurately measuring circulating GDF-11 and GDF-8 have generated controversy.
We developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for simultaneous measurement of circulating GDF-8 and GDF-11 that employs denaturation, reduction, and alkylation; cation-exchange solid-phase extraction; tryptic digestion; followed by separation and quantification using 2 signature peptides for multiple reaction monitoring and C-terminal [13C615N4]-Arg peptides as internal standards. We evaluated age trends in serum GDF-11 and GDF-8 concentrations in community-dwelling healthy men, 19 years or older, and determined the effects of graded testosterone doses on GDF-8 and GDF-11 concentrations in healthy men in a randomized trial.
The assay demonstrated linearity over a wide range, lower limit of quantitation 0.5 ng/mL for both proteins, and excellent precision, accuracy, and specificity (no detectable cross-reactivity of GDF-8 in GDF-11 assay or of GDF-11 in GDF-8 assay). Mean ± SD (median ± 1QR) GDF-8 and GDF-11 levels in healthy community-dwelling men, 19 years and older, were 7.2 ± 1.9 (6.8 ± 1.4) ng/mL. Neither GDF-8 nor GDF-11 levels were related to age or body composition. Testosterone treatment significantly increased serum GDF-8 but not GDF-11 levels.
The LC-MS/MS method for the simultaneous measurement of circulating total GDF-8 and GDF-11 demonstrates the characteristics of a valid assay. Testosterone treatment increased GDF-8 levels, but not GDF-11. Increase in GDF-8 levels by testosterone treatment, which increased muscle mass, suggests that GDF-8 acts as a chalone to restrain muscle growth.
生长分化因子 11（GDF-11）控制胚胎发育，被认为是一种抗衰老因子。GDF-8（肌肉生长抑制素）抑制骨骼肌生长。由于难以准确测量循环中的 GDF-11 和 GDF-8，因此产生了争议。
我们开发了一种液相色谱-串联质谱（LC-MS/MS）方法，用于同时测量循环中的 GDF-8 和 GDF-11，该方法采用变性、还原和烷基化；阳离子交换固相萃取；胰蛋白酶消化；然后使用 2 个特征肽进行多重反应监测和 C 末端 [13C615N4]-Arg 肽作为内标进行分离和定量。我们评估了社区居住的健康男性中血清 GDF-11 和 GDF-8 浓度的年龄趋势，年龄在 19 岁及以上，并在一项随机试验中确定了不同剂量睾酮对健康男性中 GDF-8 和 GDF-11 浓度的影响。
该测定法显示出宽线性范围，两种蛋白质的定量下限均为 0.5ng/mL，具有出色的精密度、准确性和特异性（GDF-8 测定中未检测到 GDF-11 的交叉反应，GDF-11 测定中也未检测到 GDF-8 的交叉反应）。19 岁及以上的社区居住健康男性的平均±SD（中位数±1QR）GDF-8 和 GDF-11 水平分别为 7.2±1.9（6.8±1.4）ng/mL。GDF-8 和 GDF-11 水平均与年龄或身体成分无关。睾酮治疗显著增加了血清 GDF-8 但不增加 GDF-11 水平。
用于同时测量循环总 GDF-8 和 GDF-11 的 LC-MS/MS 方法证明了有效测定的特征。睾酮治疗增加了 GDF-8 水平，但没有增加 GDF-11 水平。睾酮治疗增加 GDF-8 水平会增加肌肉质量，这表明 GDF-8 作为一种抑素来抑制肌肉生长。
