特异性下调葡萄糖调节蛋白78降低人涎腺腺样囊性癌侵袭和转移的能力

Specific down-regulation of glucose-regulated protein 78 decreases the invasion and metastasis capability of salivary adenoid cystic carcinoma

目的探讨特异性下调葡萄糖调节蛋白78(GRP78)对涎腺腺样囊性癌(ACC)侵袭和转移能力的影响;方法应用siRNA干扰技术,特异性下调ACC-2细胞内GRP78的表达,通过划痕实验、Transwell实验、四甲基偶氮唑盐(MTT)实验和细胞骨架染色,观察下调GRP78表达后细胞的变化,并应用免疫印迹法检测下调GRP78表达对基质金属蛋白酶(MMP)-2、MMP-9、基质金属蛋白酶抑制因子(TIMP)-1和TIMP-2的影响。结果划痕实验显示,下调GRP78的表达能够抑制划痕愈合;Transwell实验发现,下调GRP78在ACC-2的表达明显降低穿过的细胞数;下调GRP78表达后细胞骨架染色发现,骨架纤维明显增粗;MTT实验显示,下调GRP78后细胞的增殖能力降低1/3;免疫印迹法检测结果发现,下调GRP78表达后ACC-2细胞的MMP-2、MMP-9的表达降低,TIMP-1、TIMP-2的表达增高。结论特异性下调GRP78的表达能够降低ACC的侵袭和转移能力,这与GRP78参与调节MMP和TIMP的表达有关。

Objective To explore the effect of specifically down-regulating glucose-regulated protein 78 on invasion and metastasis capability of adenoid cystic carcinoma. Methods Applying small interfering RNA （siRNA） technology to specifically down-regulate the expression of GRP78 and analyzing the difference between ACC-2 and si-GRP78 treated ACC-2 by wound healing, Transwell assay, MTT and cytoskeleton staining. Then we detected the effect of downregulating GRP78 in ACC-2 on matrix metalloproteinases （MMP）-2, M MP-9, tissue inhibitors of metalloproteinase （TIMP）-1 and TIMP-2 by Western blotting technology to compare the ACC-2, control siRNA treated ACC-2 and si-GRP78 treated ACC-2. Results Wound healing assay revealed down-regulating GRP78 in ACC-2 could inhibit the wound healing. From Transwell assay, we found that the penetrated cells number decreased significantly by down-regulating GRP78. Cytoskeleton staining showed the fibers of si-GRP78 treated ACC-2 were thickening and distinct obviously, and the fibers of ACC-2 and control siRNA treated ACC-2 were thin and blurred. MTT assay showed that down-regulating GRP78 could inhibit 1/3 reproductive ability of ACC-2. Western blotting manifested that down-regulating the expression of GRP78 in ACC-2 decreased the expression of MMP-2, MMP-9 and increased the expression of TIMP-1, TIMP-2. Conclusion Specific down-regulation of GRP78 could inhibit ACC invasion and migration and this is due to GRP78 participating in regulating MMP and TIMP.