Huang F, Newman E, Theodorescu D, Kerbel R S, Friedman E
Department of Medicine, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
Cell Growth Differ. 1995 Dec;6(12):1635-42.
A transforming growth factor beta1 (TGF beta1) antisense expression plasmid under constitutive control of the Rous sarcoma virus promoter was introduced into the highly tumorigenic and invasive colon carcinoma U9A cell line, which uses its autocrine TGF beta1 as a growth-stimulating factor. Stable transfectants were infrequent, and only the K6 transfectant exhibited 39 and 33%, respectively, of the levels of TGF beta1 mRNA and active, secreted TGF beta1 protein of the parental line. K6 exhibited no change in TGF beta2 expression, and TGF beta3 expression was not detected in either parental or transfectant cells. Compared to the parental line, the K6 antisense transfectant exhibited a 3-fold increase in lag time in anchorage-dependent colony formation. The parental line was 44 times as invasive through a collagen l-coated polycarbonate membrane in vitro as K6 cells and, after s.c. injection at low-cell inocula, U9A cells induced tumors 75 times as large in vivo as did the K6 antisense transfectant. The decreases in in vitro invasion and anchorage-dependent colony formation seen in K6 cells were largely reversed by the addition of TGF beta1. Tumors that did arise from the K6 antisense transfectant cells had lost antisense TGF beta1 expression and expressed the same TGF beta1 mRNA levels as controls. U9A cells were more metastatic to the liver after intrasplenic injection than K6 cells. These findings demonstrate a role for autocrine TGE beta1 in colon cancer tumorigenicity and invasion. They also show that a relatively small decrease in TGF beta1 levels was enough to markedly decrease colon carcinoma cell aggressiveness. This is not unprecedented, as we had found in an earlier study that a small, 2-4-fold increase in TGF beta1 protein levels in human colon cancers correlated with disease progression to metastases (E. Friedman et al., Cancer Epidemiol, Biomarkers & Prev., 4:549-554, 1995).
将受劳斯肉瘤病毒启动子组成型控制的转化生长因子β1（TGFβ1）反义表达质粒导入具有高致瘤性和侵袭性的结肠癌U9A细胞系，该细胞系利用其自分泌的TGFβ1作为生长刺激因子。稳定转染子很少见，只有K6转染子分别表现出亲代细胞系TGFβ1 mRNA水平和活性分泌型TGFβ1蛋白水平的39%和33%。K6细胞的TGFβ2表达没有变化，在亲代细胞和转染细胞中均未检测到TGFβ3表达。与亲代细胞系相比，K6反义转染子在依赖贴壁的集落形成中的延迟时间增加了3倍。亲代细胞系在体外通过胶原蛋白I包被的聚碳酸酯膜的侵袭能力是K6细胞的44倍，在低细胞接种量皮下注射后，U9A细胞在体内诱导产生的肿瘤大小是K6反义转染子的75倍。添加TGFβ1后，K6细胞中观察到的体外侵袭和依赖贴壁的集落形成的降低在很大程度上得到了逆转。由K6反义转染子细胞产生的肿瘤失去了反义TGFβ1表达，并且表达的TGFβ1 mRNA水平与对照相同。脾内注射后，U9A细胞比K6细胞更易转移至肝脏。这些发现证明了自分泌TGFβ1在结肠癌致瘤性和侵袭中的作用。它们还表明，TGFβ1水平相对较小的降低就足以显著降低结肠癌细胞的侵袭性。这并非没有先例，因为我们在早期研究中发现，人类结肠癌中TGFβ1蛋白水平小幅度的2至4倍增加与疾病进展至转移相关（E. Friedman等人，《癌症流行病学、生物标志物与预防》，4:549 - 554，1995）。
