Azuma Haruhito, Ehata Shogo, Miyazaki Hideyo, Watabe Tetsuro, Maruyama Osamu, Imamura Takeshi, Sakamoto Takeshi, Kiyama Satoshi, Kiyama Yuko, Ubai Takanobu, Inamoto Teruo, Takahara Shiro, Itoh Yuko, Otsuki Yoshinori, Katsuoka Yoji, Miyazono Kohei, Horie Shigeo
Department of Urology, Osaka Medical College, Takatsuki, Osaka, Japan.
J Natl Cancer Inst. 2005 Dec 7;97(23):1734-46. doi: 10.1093/jnci/dji399.
Transforming growth factor beta (TGF-beta) facilitates metastasis during the advanced stages of cancer. Smad6, Smad7, and c-Ski block signaling by the TGF-beta superfamily proteins through different modes of action. We used adenovirus-mediated gene transfer of these natural inhibitors in a mouse model of breast cancer to examine the roles of TGF-beta superfamily signaling in tumor growth and metastasis.
We systemically administered, by intravenous injection, adenoviruses (AdCMV) containing the mouse cDNAs for Smad7, Smad6, c-Ski, the c-Ski mutant c-Ski (ARPG), or LacZ (control) to nude mice (>19 mice/group) bearing tumors derived from mouse mammary carcinoma JygMC(A) cells, which spontaneously metastasize to lung and liver, and examined their effects on survival and metastasis. High-throughput western blotting analysis was used to examine the expression levels for 47 signal transduction proteins in JygMC(A) cells and primary tumors. We also investigated the proliferation, migration, and invasion of JygMC(A) cells that stably overexpressed Smad6 or Smad7. Nonparametric comparisons were done by Kruskal-Wallis H statistic and Wilcoxon's rank sum tests. Parametric comparisons were done by one-way analysis of variance or two-sided unpaired Student's t tests. All statistical tests were two-sided.
Control mice bearing tumors derived from JygMC(A) cells showed many metastases to the lung and liver; all animals died by 50 days after cell inoculation. By contrast, mice treated with AdCMV-Smad7 or AdCMV-c-Ski demonstrated a dramatic decrease in metastasis and statistically significantly longer survival than control mice (Smad7 versus LacZ: medium survival = 55 days versus 41 days, difference = 14 days [95% confidence interval {CI} = 6 days to 22 days], P < .001), whereas mice treated with AdCMV-Smad6 or AdCMV-c-Ski (ARPG) did not. Expression of Smad7 in JygMC(A) cells was associated with increased expression of major components of adherens and tight junctions, including E-cadherin, decreased expression of N-cadherin, and decreases in the migratory and invasive abilities of the JygMC(A) cells.
Smad7 inhibits metastasis, possibly by regulating cell-cell adhesion. Systemic expression of Smad7 may be a novel strategy for the prevention of metastasis of advanced cancers.
转化生长因子β（TGF-β）在癌症晚期促进转移。Smad6、Smad7和c-Ski通过不同作用方式阻断TGF-β超家族蛋白的信号传导。我们在乳腺癌小鼠模型中利用腺病毒介导这些天然抑制剂的基因转移，以研究TGF-β超家族信号传导在肿瘤生长和转移中的作用。
我们通过静脉注射将含有小鼠Smad7、Smad6、c-Ski、c-Ski突变体c-Ski（ARPG）或LacZ（对照）cDNA的腺病毒（AdCMV）全身给药给携带源自小鼠乳腺癌JygMC(A)细胞的肿瘤的裸鼠（每组>19只小鼠），这些肿瘤会自发转移至肺和肝脏，并检查它们对生存和转移的影响。采用高通量蛋白质印迹分析来检测JygMC(A)细胞和原发性肿瘤中47种信号转导蛋白的表达水平。我们还研究了稳定过表达Smad6或Smad7的JygMC(A)细胞的增殖、迁移和侵袭情况。非参数比较采用Kruskal-Wallis H统计量和Wilcoxon秩和检验。参数比较采用单因素方差分析或双侧非配对学生t检验。所有统计检验均为双侧。
携带源自JygMC(A)细胞肿瘤的对照小鼠出现许多肺和肝转移；所有动物在细胞接种后50天内死亡。相比之下，用AdCMV-Smad7或AdCMV-c-Ski处理的小鼠转移显著减少，且生存期在统计学上显著长于对照小鼠（Smad7与LacZ相比：中位生存期 = 55天对41天，差异 = 14天[95%置信区间{CI}=6天至22天]，P<.001），而用AdCMV-Smad6或AdCMV-c-Ski（ARPG）处理的小鼠则不然。JygMC(A)细胞中Smad7的表达与黏附连接和紧密连接的主要成分（包括E-钙黏蛋白）表达增加、N-钙黏蛋白表达减少以及JygMC(A)细胞迁移和侵袭能力降低有关。
Smad7可能通过调节细胞间黏附来抑制转移。Smad7的全身表达可能是预防晚期癌症转移的一种新策略。
