LC-MS/MS-based in vitro and in vivo investigation of blood-brain barrier integrity by simultaneous quantitation of mannitol and sucrose.
LC-MS/MS-based in vitro and in vivo investigation of blood-brain barrier integrity by simultaneous quantitation of mannitol and sucrose.
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通过同时定量甘露醇和蔗糖进行的基于LC-MS/MS的血脑屏障完整性的体外和体内研究。
DOI：
10.1186/s12987-020-00224-1
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发表时间：
2020-10-14
影响因子：
7.3
通讯作者：
Bickel U
中科院分区：
文献类型：
--
作者：
Noorani B;Chowdhury EA;Alqahtani F;Ahn Y;Patel D;Al-Ahmad A;Mehvar R;Bickel U
了解血脑屏障（BBB）的病理生理学在疾病的诊断和治疗中起着至关重要的作用。应用灵敏和特异的LC-MS/MS技术以高精度测量BBB完整性，我们最近引入了非放射性[13 C12]蔗糖作为上级标记物。比较具有不同分子量但在其他方面具有相似理化性质的渗透性标志物，可以提供对BBB处的摄取机制的见解。甘露聚糖是一种亲水性的不带电小分子，其大小是蔗糖的一半。以前仅使用放射性[3 H]甘露醇或[14 C]甘露醇来测量BBB完整性。建立了一种同时分析稳定同位素标记蔗糖和甘露醇的超高效液相色谱-串联质谱法。在小鼠中测量[13 C6]甘露醇和[13 C12]蔗糖的体内BBB渗透性，使用[13 C6]蔗糖作为血管标记物以校正脑血管内含量。此外，使用具有诱导的多能干细胞衍生的脑内皮细胞的Transwell模型在对照和受损（在IL-1β的存在下）条件下测量蔗糖和甘露醇的体外渗透系数。我们发现甘露醇和蔗糖在体外（渗透系数分别为4.99 ± 0.152 × 10−7和3.12 ± 0.176 × 10−7 cm/s）和体内（PS产物分别为0.267 ± 0.021和0.126 ± 0.025 µl g−1 min−1）的渗透率值较低。此外，在IL-1β存在下，两种标记物的体外渗透性显著增加。甘露醇（0.071 ± 0.007和0.065 ± 0.009%注射剂量/g）或蔗糖（0.035 ± 0.003和0.037 ± 0.005%注射剂量/g）的校正脑浓度（Cbr）（通过洗脱与血管标记物校正获得）无显著差异。这些数据还表明甘露醇的Cbr和PS乘积值约为蔗糖相应值的两倍。我们建立了一种高度敏感，特异性和可重复的方法，同时测量两个经典的低分子量，亲水性标记物的稳定同位素标记的格式的血脑屏障通透性。这种方法现在可作为一种工具，在体外和体内不同的疾病模型中量化BBB渗透性，以及监测治疗结果。
Understanding the pathophysiology of the blood brain–barrier (BBB) plays a critical role in diagnosis and treatment of disease conditions. Applying a sensitive and specific LC–MS/MS technique for the measurement of BBB integrity with high precision, we have recently introduced non-radioactive [13C12]sucrose as a superior marker substance. Comparison of permeability markers with different molecular weight, but otherwise similar physicochemical properties, can provide insights into the uptake mechanism at the BBB. Mannitol is a small hydrophilic, uncharged molecule that is half the size of sucrose. Previously only radioactive [3H]mannitol or [14C]mannitol has been used to measure BBB integrity. We developed a UPLC–MS/MS method for simultaneous analysis of stable isotope-labeled sucrose and mannitol. The in vivo BBB permeability of [13C6]mannitol and [13C12]sucrose was measured in mice, using [13C6]sucrose as a vascular marker to correct for brain intravascular content. Moreover, a Transwell model with induced pluripotent stem cell-derived brain endothelial cells was used to measure the permeability coefficient of sucrose and mannitol in vitro both under control and compromised (in the presence of IL-1β) conditions. We found low permeability values for both mannitol and sucrose in vitro (permeability coefficients of 4.99 ± 0.152 × 10−7 and 3.12 ± 0.176 × 10−7 cm/s, respectively) and in vivo (PS products of 0.267 ± 0.021 and 0.126 ± 0.025 µl g−1 min−1, respectively). Further, the in vitro permeability of both markers substantially increased in the presence of IL-1β. Corrected brain concentrations (Cbr), obtained by washout vs. vascular marker correction, were not significantly different for either mannitol (0.071 ± 0.007 and 0.065 ± 0.009 percent injected dose per g) or sucrose (0.035 ± 0.003 and 0.037 ± 0.005 percent injected dose per g). These data also indicate that Cbr and PS product values of mannitol were about twice the corresponding values of sucrose. We established a highly sensitive, specific and reproducible approach to simultaneously measure the BBB permeability of two classical low molecular weight, hydrophilic markers in a stable isotope labeled format. This method is now available as a tool to quantify BBB permeability in vitro and in vivo in different disease models, as well as for monitoring treatment outcomes.
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影响因子：
4.6
作者：
Gervasoni, Jacopo;Primiano, Aniello;Persichilli, Silvia
通讯作者：
Persichilli, Silvia
DOI：
10.1111/j.1748-1716.1980.tb06678.x
发表时间：
1980-01-01
期刊：
ACTA PHYSIOLOGICA SCANDINAVICA
影响因子：
--
作者：
AMTORP, O
通讯作者：
AMTORP, O
DOI：
10.1152/ajpheart.2000.279.3.h1022
发表时间：
2000-09-01
影响因子：
4.8
作者：
Murakami, H;Takanaga, H;Sawada, Y
通讯作者：
Sawada, Y
影响因子：
17.1
作者：
Iliff JJ;Wang M;Liao Y;Plogg BA;Peng W;Gundersen GA;Benveniste H;Vates GE;Deane R;Goldman SA;Nagelhus EA;Nedergaard M
通讯作者：
Nedergaard M
影响因子：
4.7
作者：
Gustafsson, Sofia;Lindstrom, Veronica;Syvanen, Stina
通讯作者：
Syvanen, Stina