【摘要】 目的探讨海藻糖在气管低温保存中的保护作用和机制。方法新鲜配制4组保存液,其中:Ⅰ组 LPD;Ⅱ组LPD+10%DMSO;Ⅲ组LPD+0.15M海藻糖;Ⅳ组LPD+10%DMSO+0.15M海藻糖。切取SD 大鼠气管后立即分别放入含上述4种溶液的冻存管,在程序降温仪降至-80℃后投入液氮中保存。20d后对 气管标本进行HE染色和Bcl 2、Bax蛋白检测(免疫组化),并将其异位移植到Wistar大鼠腹腔,15d后取出 移植气管,观察组织学变化。结果4种保存液低温保存气管20d后,Ⅲ组和Ⅳ组气管结构完整,软骨细胞Bax 蛋白表达较低,尤其Ⅳ组保存效果最好,同种异体异位移植后生长良好。各组Bcl 2蛋白表达无明显区别。 结论在气管低温保存中海藻糖的保护作用主要是通过对软骨细胞的保护实现的,抑制软骨细胞Bax基因的 表达是其中的保护机制之一。海藻糖和DMSO合用保护作用更好。更多还原

【Abstract】 Objective To detect the protective mechanism of trehalose in tracheal cryopreservation.Methods Inbred male Sprague-Dawley (SD) rats were sacrificed with intraperitoneal injection of ketamine(150mg·kg -1).The tracheas were removed and immersed immediately in the freezing medium of low potassium dextran (LPD) solution only(Group Ⅰ) ,containing with 10% dimethylsulfoxide(DMSO)(Group Ⅱ), containing with 0.15mol·L -1 trehalose (Group Ⅲ),and containing with 10% DMSO and 0.15mol·L -1 trehalose (Group Ⅳ) respectively. A sterile plastic tube containing a 1-cm-long trachea was filled with the freezing medium,sealed,and frozen to -80℃ at rate of -1℃ per minute in a programmable freezer.Then the tube was stored in liquid nitrogen(-196℃) for 20 days. Then the specimen was thawed in a 37℃ water bath and rinsed with physiologic saline solution 10 times.Histologic changes before cryopreservation and after thawing were examined in each group. After the specimens were embedded in paraffin,5-(m-thick sections were stained with hematoxylin and eosin.The epithelium and cartilage was assessed. We also observed Bcl-2 and Bax gene expression by immunohistochemistry. At last, some tracheas(SD) after cryopreservation were thawed and transplanted into the abdominal cavity of Wistar rats. The transplanted tracheas were retrieved and assessed histologically.Results Microscopic findings of the tracheas in Group Ⅲ and Group Ⅳ showed their structure were intact and Bax gene expression was lower in cartilage after cryopreservation(20d) compared with other groups,especially in Group Ⅳ.The tracheas in Group Ⅲ and Group Ⅳ grew well after they were transplanted into cavity of Wistar rats heterotopically,too.There were no significant differences among 4 groups in Bcl-2 gene expression.Conclusion In tracheal cryopreservation the trehalose can protect the trachea by protecting the tracheal cartilage.It is one of the protective mechanism that the trehalose inhibit the Bax gene expression of cartilage cells.The concomitant use of trehalose and DMSO has a synergistic effect.更多还原