[海藻糖对冷冻保存的人皮肤中α-辅肌动蛋白影响的研究]

[Investigation on the effect of trehalose on alpha-actinin in cryopreserved human skin].

作者信息

Jia Xiaoming, Yu Yu, Li Dongjie

机构信息

Burns Institute, First Affiliated Hospital to PLA General Hospital, Beijing, 100037, P. R. China.

出版信息

Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2008 Apr;22(4):446-9.

PMID:18575445

Abstract

OBJECTIVE

To compare the effect of trehalose with that of different traditional cryoprotectants on human skin and to detect the new protection mechanism of trehalose in hypothermia.

METHODS

The skins to be cryopreserved were first treated with DMSO/Propyleneglycol (D/P group), trehalose/DMSO (T/D group), DMSO/serumfree keratinocyte medium (D/K group), DMEM (DMEM group), respectively, so as to be compared with fresh skin (control grouop). Then the histological structure of skin of different groups was observed and analyzed by pathological technology (SP immunohistochemistry, DAB staining). Furthermore, the influence of trehalose on alpha-actinin at gene level with RT-PCR was investigated. The viability of skin in 5 respective groups was evaluated by using succinate dehydrogenase (SDH). The experiments were carried out 14 days after cryopreservation.

RESULTS

The results of immunohistochemistry showed that A values of control group, T/D group, D/P group, D/K group and DMEM group were 27.50 +/- 7.92, 18.40 +/- 5.81, 13.10 +/- 5.11, 11.50 +/- 4.54 and 5.30 +/- 2.14, respectively. There was no significant difference between control group and T/D group (P > 0.05), but control group was significantly different from the other groups (P < 0.05). The results of PCR studies showed that A values of control group, T/D group, D/P group, D/K group and DMEM group were 0.816 +/- 0.134, 0.723 +/- 0.245, 0.564 +/- 0.265, 0.245 +/- 0.071 and 0.148 +/- 0.048, respectively. Control group was not significantly different from T/D group and D/P group (P > 0.05), but was significantly different from D/K group and DMEM group (P < 0.05). The results of SDH showed that A valuse of control group, T/D group, D/P group, D/K group and DMEM group were 18.2 +/- 3.7, 12.3 +/- 3.6, 10.2 +/- 2.4, 7.3 +/- 2.1 and 5.7 +/- 1.5, respectively. There was no significant difference between control group and T/D group (P > 0.05), while control group was significantly different from the other groups (P < 0.05).

CONCLUSION

The results suggest that cryopreservation protocol-trehalose/DMSO is better than the traditional cryoprotectant for cryopreservation on alpha-actinin of human skin.

摘要

目的

比较海藻糖与不同传统冷冻保护剂对人体皮肤的作用，并检测海藻糖在低温状态下的新保护机制。

方法

将待冷冻保存的皮肤分别用二甲基亚砜/丙二醇（D/P组）、海藻糖/二甲基亚砜（T/D组）、二甲基亚砜/无血清角质形成细胞培养基（D/K组）、杜氏改良Eagle培养基（DMEM组）处理，以便与新鲜皮肤（对照组）进行比较。然后采用病理技术（SP免疫组化、DAB染色）观察并分析不同组皮肤的组织结构。此外，运用逆转录聚合酶链反应（RT-PCR）研究海藻糖在基因水平对α-辅肌动蛋白的影响。使用琥珀酸脱氢酶（SDH）评估5个组皮肤的活力。冷冻保存14天后进行实验。

结果

免疫组化结果显示，对照组、T/D组、D/P组、D/K组和DMEM组的A值分别为27.50±7.92、18.40±5.81、13.10±5.11、11.50±4.54和5.30±2.14。对照组与T/D组之间无显著差异（P>0.05），但对照组与其他组有显著差异（P<0.05）。PCR研究结果显示，对照组、T/D组、D/P组、D/K组和DMEM组的A值分别为0.816±0.134、0.723±0.245、0.564±0.265、0.245±0.071和0.148±0.048。对照组与T/D组和D/P组无显著差异（P>0.05），但与D/K组和DMEM组有显著差异（P<0.05）。SDH结果显示，对照组、T/D组、D/P组、D/K组和DMEM组的A值分别为18.2±3.7、12.3±3.6、10.2±2.4、7.3±2.1和5.7±1.5。对照组与T/D组之间无显著差异（P>0.05），而对照组与其他组有显著差异（P<0.05）。