In order to decrease the osmotic damage and chemical cytotoxicity of the cryoprotectant with high concentration, an appropriate scheme of loading and removing the cryoprotective agents is necessary during optimizing the procedure of effective cell and tissue cryopreservation. In this study, the membrane permeability of SD rat cranium parietal bone-derived osteoblasts was studied by microscopy image analysis method. It was found that the osteoblasts isosmotic cell volume Vo of the osteoblast is 2585.22 urn3, and the osmotic behavior of the osteoblasts behaves as ideal osmometers. According to the Boyle van't Hoff plot, the osmotically inactive volume Vb of the osteoblast is 0.335 Vo The found hydraulic conductivities of the osteoblast cell membrane in the presence of cryoprotective agents, dimethyl sulfoxide and 1,2-propanediol, are 1.5× 10-13 and 5×l0-13 m3·N-1·s-1, respectively, and the permeability of the above cryoprotectants are 7.5×10-11 and 120×10-11 mol·N-1·s-1, respectively. The experimental data indicate that the anisotonic condition tolerance of the adherent osteoblasts is lower than that of the osteoblasts in suspension. The study results obtained provide initial biophysical data to characterize osteoblasts and will provide a basis for designing the optimal cryopreservation procedures for engineered bone tissue.%在优化组织工程骨低温保存程序时,需要了解体外培养的种子细胞成骨细胞的渗透特性.尤其对于高浓度的玻璃化保护剂,更有必要设计合适的保护剂导入、洗脱方案来减小高浓度保护剂的渗透效应损伤及其化学毒性作用.利用显微镜图像分析法研究了用于构建组织工程骨的种子细胞—-成骨细胞的渗透特性,其中包括细胞的等渗体积、细胞在高渗及低渗溶液中的平衡体积和细胞的非渗透体积以及细胞膜对水和冷冻保护剂的渗透系数,并比较了悬浮和贴壁的成骨细胞对渗透压的耐受范围.结果表明,在等渗条件下(289 mmol·kg-1),成骨细胞的平均体积为2585.22 μm3；细胞体积随溶液渗透压的变化规律符合理想渗透压变化曲线,即Boyle van’t Hoff关系式,由该关系式得到成骨细胞的渗透体积为0.335 V0;室温下,在2 mol·L-1二甲基亚砜(DMSO)和1,2-丙二醇(PD)中成骨细胞膜对水的水力传导系数分别为1.5×10-13和5×10-13 m3·N-1.s-1,对保护剂的渗透系数分别为7.5×10-11和120× 10-11mol·N-1.s-1；贴壁的成骨细胞对渗透压的耐受范围远低于悬浮细胞,说明了贴壁细胞与悬浮细胞渗透特性的差异.由以上参数结合相关数学模型能够设计和研究冷冻保护剂的导入、洗脱方案,.以使组织工程骨的低温保存获得更好的效果.
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