Establishment of sandwich ELISA for soluble alpha-Klotho measurement: Age-dependent change of soluble alpha-Klotho levels in healthy subjects.
Establishment of sandwich ELISA for soluble alpha-Klotho measurement: Age-dependent change of soluble alpha-Klotho levels in healthy subjects.
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DOI：
10.1016/j.bbrc.2010.06.110
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发表时间：
2010-07-30
影响因子：
3.1
通讯作者：
Nabeshima, Yo-ichi
中科院分区：
文献类型：
--
作者：
Yamazaki, Yuji;Imura, Akihiro;Urakawa, Itaru;Shimada, Takashi;Murakami, Junko;Aono, Yukiko;Hasegawa, Hisashi;Yamashita, Takeyoshi;Nakatani, Kimihiko;Saito, Yoshihiko;Okamoto, Nozomi;Kurumatani, Norio;Namba, Noriyuki;Kitaoka, Taichi;Ozono, Keiichi;Sakai, Tomoyuki;Hataya, Hiroshi;Ichikawa, Shoji;Imel, Erik A.;Econs, Michael J.;Nabeshima, Yo-ichi
α-Klotho （αKl）调节循环中钙离子（Ca2+）和无机磷酸盐（Pi）等矿物质代谢。小鼠的缺陷导致了类似于人类衰老中发现的疾病的临床特征。尽管跨膜型αKl的重要性已被证实，但关于循环中可溶性型αKl （s - αKl）的生理重要性尚不清楚。本研究的目的是：1)建立血清sαKl夹心ELISA检测系统；2)确定健康人群血清sαKl水平参考值及其与肾功能、矿物质代谢、年龄、性别等指标的关系。我们相继建立了一种ELISA方法来检测年龄（61.1±18.5岁）的健康志愿者（n=142，男性66）血清sαKl。这些健康对照成人的水平（平均±SD）如下：总钙（Ca; 9.46±0.41 mg/dL）， Pi(3.63±0.51 mg/dL)，血尿素氮（BUN; 15.7±4.3 mg/dL），肌酐（Cre; 0.69±0.14 mg/dL）， 1,25二羟基维生素D (1,25(OH)2D；54.8±17.7 pg/mL)，完整甲状旁腺激素（iPTH; 49.2±20.6 pg/mL），降钙素（26.0±12.3 pg/mL）和完整成纤维细胞生长因子（FGF23; 43.8±17.6 pg/mL）。正常成人血清sαKl水平为239 ~ 1266 pg/mL（平均±SD； 562±146 pg/mL）。虽然sαKl水平不受性别和矿物质代谢指标的影响，但与Cre和年龄呈负相关。然而，正常儿童（n=39，男性23，平均±SD； 7.1±4.8岁）的sαKl水平（平均±SD； 952±282 pg/mL）明显高于成人（平均±SD； 562±146，P<0.001）。本研究对儿童和成人进行多元线性回归分析，结果表明sαKl与年龄、Ca呈负相关，与Pi呈正相关。最后，我们检测了一名严重肿瘤性钙血症患者的血清s - α kl，该患者源于α-klotho基因的纯合子错义突变。患者s - α kl水平明显低于同龄对照组。首次建立了测定人血清s - α kl的检测系统。正常人群中年龄、钙和Pi似乎影响血清s - α kl水平。该检测系统将成为研究s - α kl在矿物质代谢中的功能的良好工具。
α-Klotho (αKl) regulates mineral metabolism such as calcium ion (Ca2+) and inorganic phosphate (Pi) in circulation. Defects in mice result in clinical features resembling disorders found in human aging. Although the importance of transmembrane-type αKl has been demonstrated, less is known regarding the physiological importance of soluble-type αKl (sαKl) in circulation. The aims of this study were: 1) to establish a sandwich ELISA system enabling detection of circulating serum sαKl, and 2) to determine reference values for sαKl serum levels and relationship to indices of renal function, mineral metabolism, age and sex in healthy subjects. We successively developed an ELISA to measure serum sαKl in healthy volunteers (n=142, males 66) of ages (61.1 ± 18.5 yr). The levels (mean ± SD) in these healthy control adults were as follows: total calcium (Ca; 9.46 ± 0.41 mg/dL), Pi (3.63 ± 0.51 mg/dL), Blood urea nitrogen (BUN; 15.7 ± 4.3 mg/dL), creatinine (Cre; 0.69 ± 0.14 mg/dL), 1,25 dihydroxyvitamin D (1,25(OH)2D; 54.8 ± 17.7 pg/mL), intact parathyroid hormone (iPTH; 49.2 ± 20.6 pg/mL), calcitonin (26.0 ± 12.3 pg/mL) and intact Fibroblast growth factor (FGF23; 43.8 ± 17.6 pg/mL). Serum levels of sαKl ranged from 239 to 1266 pg/mL (mean ± SD; 562 ± 146 pg/mL) in normal adults. Although sαKl levels were not modified by gender or indices of mineral metabolism, sαKl levels were inversely related to Cre and age. However, sαKl levels in normal children (n=39, males 23, mean ± SD; 7.1 ± 4.8 years) were significantly higher (mean ± SD; 952 ± 282 pg/mL) than those in adults (mean ± SD; 562 ± 146, P<0.001). A multivariate linear regression analysis including children and adults in this study demonstrated that sαKl correlated negatively with age and Ca, and positively with Pi. Finally, we measured a serum sαKl from a patient with severe tumoral calcinosis derived from a homozygous missense mutation of α-klotho gene. In this patient, sαKl level was notably lower than those of age matched controls. We established a detection system to measure human serum sαKl for the first time. Age, Ca and Pi seem to influence serum sαKl levels in a normal population. This detection system should be an excellent tool for investigating sαKl functions in mineral metabolism.
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DOI：
10.1006/bbrc.2000.4226
发表时间：
2001-02-02
影响因子：
3.1
作者：
Koh, N;Fujimori, T;Nabeshima, Y
通讯作者：
Nabeshima, Y
DOI：
10.1073/pnas.0913986107
发表时间：
2010-01-26
影响因子：
11.1
作者：
Tomiyama, Ken-ichi;Maeda, Ryota;Nabeshima, Yo-ichi
通讯作者：
Nabeshima, Yo-ichi
影响因子：
15.9
作者：
Ichikawa, Shoji;Lmel, Erik A.;Econs, Michael J.
通讯作者：
Econs, Michael J.
影响因子：
5.8
作者：
Yamazaki, Y;Okazaki, R;Fukumoto, S
通讯作者：
Fukumoto, S
DOI：
10.1073/pnas.101545198
发表时间：
2001-05-22
影响因子：
11.1
作者：
Shimada, T;Mizutani, S;Yamashita, T
通讯作者：
Yamashita, T