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首页> 美国卫生研究院文献>Biopreservation and Biobanking >A Microfluidic Study of Megakaryocytes Membrane Transport Properties to Water and Dimethyl Sulfoxide at Suprazero and Subzero Temperatures
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A Microfluidic Study of Megakaryocytes Membrane Transport Properties to Water and Dimethyl Sulfoxide at Suprazero and Subzero Temperatures

机译：在超零温度和零以下温度下巨核细胞膜向水和二甲基亚砜转运特性的微流控研究

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Megakaryocytes (MKs) are the precursor cells of platelets. Cryopreservation of MKs is critical for facilitating research investigations about the biology of this important cell and may help for scaling-up ex-vivo production of platelets from MKs for clinical transfusion. Determining membrane transport properties of MKs to water and cryoprotectant agents (CPAs) is essential for developing optimal conditions for cryopreserving MKs. To obtain these unknown parameters, membrane transport properties of the human UT-7/TPO megakaryocytic cell line were investigated using a microfluidic perfusion system. UT-7/TPO cells were immobilized in a microfluidic system on poly-D-lysine-coated glass substrate and perfused with various hyper-osmotic salt and CPA solutions at suprazero and subzero temperatures. The kinetics of cell volume changes under various extracellular conditions were monitored by a video camera and the information was processed and analyzed using the Kedem–Katchalsky model to determine the membrane transport properties. The osmotically inactive cell volume (Vb=0.15), the permeability coefficient to water (Lp) at 37°C, 22°C, 12°C, 0°C, −5°C, −10°C, and −20°C, and dimethyl sulfoxide (DMSO; Ps) at 22, 12, 0, −10, −20, as well as associated activation energies of water and DMSO at different temperature regions were obtained. We found that MKs have relatively higher membrane permeability to water (Lp=2.62 μm/min/atm at 22°C) and DMSO (Ps=1.8×10−3 cm/min at 22°C) than most other common mammalian cell types, such as lymphocytes (Lp=0.46 μm/min/atm at 25°C). This information could suggest a higher optimal cooling rate for MKs cryopreservation. The discontinuity effect was also found on activation energy at 0°C–12°C in the Arrhenius plots of membrane permeability by evaluating the slope of linear regression at each temperature region. This phenomenon may imply the occurrence of cell membrane lipid phase transition.
机译：巨核细胞（MKs）是血小板的前体细胞。 MK的超低温保存对于促进有关该重要细胞生物学的研究至关重要，并且可能有助于扩大MK的血小板在体外的临床生产规模。确定MK向水和冷冻保护剂（CPA）的膜运输特性对于开发冷冻保存MK的最佳条件至关重要。为了获得这些未知参数，使用微流灌注系统研究了人UT-7 / TPO巨核细胞系的膜转运特性。将UT-7 / TPO细胞固定在微流体系统中的聚-D-赖氨酸涂层玻璃基板上，并在零下和零下温度下灌注各种高渗透盐和CPA溶液。通过摄像机监控各种细胞外条件下细胞体积变化的动力学，并使用Kedem-Katchalsky模型处理和分析信息，以确定膜的转运特性。在37°C，22°C，12°C，0°C，-5°C，-10°C和-20°C时无渗透活性的细胞体积（Vb = 0.15），对水的渗透系数（Lp） C，22、12、0，-10，-20的二甲基亚砜（DMSO; Ps），以及水和DMSO在不同温度区域的相关活化能。我们发现，MKs对水的膜渗透性相对较高（在22°C下为Lp = 2.62μm/ min / atm）和DMSO（在22°C下为Ps = 1.8×10 -3 cm / min）比大多数其他常见的哺乳动物细胞类型（例如淋巴细胞）（在25°C时Lp = 0.46μm/ min / atm）高。该信息可能表明，MK冷冻保存的最佳最佳冷却速率更高。通过评估每个温度区域的线性回归斜率，在膜渗透性的Arrhenius图中还发现了在0°C–12°C下对活化能的不连续性影响。这种现象可能暗示细胞膜脂质相变的发生。

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