转染FRZB基因对人胃癌SGC-7901细胞生长和转移的影响

Effects of FRZB on Growth and Metastasis of Gastric Cancer Cell Line SGC-7901

背景与目的:FRZB(frizzled motif associated with bone development)是sFRP(secreted frizzled related protein)家族的成员,在胚胎发育过程中起重要作用。有文献报道FRZB的表达能抑制基质金属蛋白酶-2(matrix metalloproteinase 2,MMP-2)和MMP-9表达而与前列腺癌的侵袭能力相关。本研究拟探讨FRZB基因对人胃癌SGC-7901细胞成瘤性和侵袭能力的影响,及其可能的机制。方法:构建FRZB真核表达载体并转染SGC-7901细胞,经G418筛选获得稳定表达克隆。用MTT法检测并绘制细胞增殖曲线,软琼脂培养和裸鼠皮下注射的体外和体内实验检测细胞成瘤性变化。用细胞免疫化学检测转染FRZB后细胞中MMP-2、MMP-7、MMP-9的表达变化,体外侵袭和粘附实验检测细胞侵袭能力变化。结果:筛选稳定表达细胞株经定量PCR检测,FRZB表达显著提高。体内外实验证实,转染FRZB的细胞(SGC-7901/FRZB)生长抑制率约为60%,克隆形成能力和成瘤性减弱。免疫细胞化学显示MMP-2、MMP-7、MMP-9在细胞浆内的阳性表达降低或接近阴性,对细胞外基质成分(CollagenI、IV、Vitronectin、Fibronectin、Laminin)粘附能力均增高,比空载体转染对照组(SGC-7901/vector)增加12.8%、19.8%、59.8%、26.7%、15.2%,差异具有显著性(P<0.05),而空载体转染对照组与亲本细胞间差异无统计学意义(P>0.05)。SGC-7901、SGC-7901/vector和SGC-7901/FRZB细胞侵袭能力分别是每高倍视野55.90±5.68、54.80±6.97、6.60±2.63,差异具有统计学意义(P<0.05)。结论:FRZB高表达在体内体外均可抑制SGC-7901细胞的增殖、成瘤性和侵袭能力,具有肿瘤抑制基因的功能。FRZB抑制肿瘤细胞的生长和抑制MMP-2、MMP-7、MMP-9的表达为抑癌功能相关机制之一。

BACKGROUND ＆ OBJECTIVE： FRZB （frizzled motif associated with bone development）, a member of the secreted frizzled related protein （sFRP） family, plays an important role in embryonic development. The expression of FRZB can suppress the invasion ability of prostate cancer by inhibiting the expression of matrix metalloproteinase-2 （MMP-2） and MMP-9. This study was to investigate the effects of FRZB on tumorigenicity and metastasis of gastric cancer cell line SGC-7901. METHODS. FRZB expression vector and empty vector were constructed and transfected into SGC-7901 cells to obtain stable clones （SGC-7901/FRZB and SGC-7901/vector）. Cell proliferation was detected by MTT assay. The tumorigenicity was investigated by soft agar colony formation test （in vitro） and xenograft mouse model （in vivo）. The adhesive and invasive abilities of SGC-7901/FRZB cells were determined by Boyden chambers. The expression of MMPs in SGC-7901 cells was tested by immunocytochemistry. RESULTS. The expression of FRZB was increased in SGC-7901/FRZB cells. Growth rate and colony formation rate of SGC-7901/FRZB cells were decreased to about 60% and 15% as compared with those of SGC-7901/vector cells, respectively. The rates of adhesion to collagen I , collagen IV, vitronectin, fibronectin and laminin were increased in SGC-7901/FRZB cells by 12.8%, 19.8%, 59.8%, 26.7% and 15.2% as compared with those in SGC-7901/ vector cells, respectively. The positive dyeing of MMP-2, MMP-7 and MMP-9 in SGC-7901/FRZB cells was reduced, The numbers of invasive SGC-7901, SGC-7901/vector and SGC-7901/FRZB cells were 55.90±5.68, 54.80±6.97, 6.60±2.63, respectively. CONCLUSION. FRZB exhibits antitumor ability in gastric cancer cell line SGC-7901 in vitro and in vivo, and decreases the expression of MMP-2, MMP-7 and MMP-9.