中文摘要：

目的探讨α1，3-半乳糖基转移酶（α1，3-GT）基因沉默对人自然杀伤（NK）细胞介导的异种细胞毒作用的影响。方法将培养的永生化猪血管内皮细胞系细胞株PED分为3组。转染组：PED转染了特异性α1，3-GT核糖核酸干扰分子（siRNA-1）；错配组：PED转染了错配的SiRNA；空转染组：PED未转染SiRNA。观察各组PED与人NK细胞系细胞株NK92在静止和流动状态下的粘附作用、不同效靶比下的NK92细胞毒作用以及PED和NK92混和培养上清液中γ干扰素（IFN-γ）的水平。结果PED转染后48h，转染组、错配组和空转染组在静止状态下粘附NK92的数量（个）分别为262±26、275±24和252±23；流动状态下分别为132±12、125±15和110±20，无论是静止还是流动状态下，各组PED对NK92的粘附能力的比较，差异均无统计学意义（P〉0．05）。无论PED是否被活化，在相同效靶比下NK92对各组PED的杀伤率的比较，差异均无统计学意义（P〉0．05），但杀伤率与效靶比呈正相关。NK92与PED共孵育早期即有反应性IFN-γ分泌，其分泌水平[（366．9±28．7）ng／L]较其自发分泌水平[（60．1±6．4）ng／L]增加了5倍，在共孵育后12h分泌值最高，但各组间不同时段IFN-γ分泌值的比较，差异均无统计学意义（P〉0．05）。结论利用RNA干扰技术下调PED上的α1，3-半乳糖残基（reGal）的表达，未引起NK92和PED相互作用的明显改变。α-Gal可能并非是NK细胞作用的靶位点，其表达量的高低对NK细胞的活化、粘附及杀伤能力未产生负性影响。

英文摘要：

Objective To evaluate the effect of RNA interference （RNAi） targeting pig alpha 1, 3-galactosyltransferase （α1,3-GT） on the human natural killer cell-induced xeno-cytotoxicity. Methods The immortalized pig endothelial cell line （PED） was subgrouped into transfected （SiRNA-1）, mock and scrambled control groups. The effect of SiRNA-1 on the interaction between PED and human natural killer cell line （NK92） was assessed in terms of adhesion and cytotoxicity, as well as IFN-γ levels of the coculture supernatant of PED and NK92. Results Forty-eight h post-transfection of SiRNA-1 into PED, there was no significant difference in the adhesion of NK 92 to PED under static or dynamic condition among SiRNA-1, scrambled and mock groups. The number of NK 92 adhering to PED was 262 ＋- 26, 275 ＋- 24 and 252 ＋- 23 under static condition and 132 ＋- 12, 125 ＋- 15 and 110 ＋- 20 under dynamic condition in SiRNA-1, scrambled and mock groups, respectively （P〉0. 05）. Direct cytotoxicity of NK 92 towards PED in all groups wag almost the same with or without PED activation and exhibited a positive correlation with the ratio of effector to target cells. There was induced IFN-γ secretion in supernatant at the early period of co-culture [（366. 9 ＋- 28.7）ng/L], resulting in a five-fold rise compared to spontaneous secretion [（60. 1 ＋- 6. 4） ng/L]. The maximum secretion level was at 12 h post-transfection and reduced subsequently. There was no statistically significant difference at various time points among SiRNA-1 transfected, scrambled transfected and mock groups （P〉 0. 05）. Conclusions There was no significant change in the interaction of NK92 and PED after knock-down expression of α-Gal by RNAi, indicating that α-Gal might not the target site of hNK cytotoxicity. The level of α-Gal expression was not related with hNK cells activation, adhesion and cytotoxicity.