Guo Kun, Liu Yinkun, Zhou Haijun, Dai Zhi, Zhang Jubo, Sun Ruixia, Chen Jie, Sun Qianglin, Lu Wenjing, Kang Xiaonan, Chen Pei
Liver Cancer Institute, Zhongshan Hospital, Fudan University, No. 180 Fenglin Road, Shanghai 200032, China.
Cancer Sci. 2008 Mar;99(3):486-96. doi: 10.1111/j.1349-7006.2007.00702.x. Epub 2007 Dec 27.
To understand the molecular mechanism that underlies the role of various prominent signal pathways in hepatocellular carcinoma (HCC) metastasis, a human signal transduction oligonucleotide microarray analysis was carried out in cultured HCC cell models with increasing spontaneous metastatic potential (MHCC97L, MHCC97H, and HCCLM6). The results revealed that the mitogen-activated protein kinase (MAPK) pathway is the prominently upregulated pathway in HCC metastasis. Further study showed that basal phosphorylated levels of extracellular signal-regulating kinase (ERK)(1/2) and p38 MAPK consecutively increased from MHCC97L to MHCC97H to HCCLM6 cells, but not c-Jun N-terminal kinase. The phosphorylation of ERK(1/2) and p38 MAPK was regulated by upregulated protein kinase C beta (PKC beta) in HCC cells through the integrated use of PKC beta RNA interference, the PKC beta specific inhibitor enzastaurin and a PKC activator phorbol-12-myristate-13-acetate. Heat shock protein 27 (HSP27) was also verified as a downstream common activated protein of PKC beta-ERK(1/2) and PKC beta-p38 MAPK. In vitro migration and invasion assay further showed that the depletion of PKC beta or inhibition of PKC beta activation effectively decreased HCC cell motility and invasion. Moreover, the motility and invasion of phorbol-12-myristate-13-acetate-stimulated PKC beta-mediated HCC cells was significantly negated by an ERK inhibitor, 1.4-diamino-2.3-dicyano-1.4-bis[2-aminophenylthio] butadiene, or a p38 MAPK inhibitor, 4-(4-Fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole. It also showed that HSP27 is critical in PKC beta-mediated HCC cell motility and invasion. Taken together, this study reveals the important role of this PKC beta-ERK(1/2)/p38MAPK-HSP27 pathway, which was verified for the first time, in modulating HCC cell motility and invasion.
为了解各种重要信号通路在肝细胞癌（HCC）转移中所起作用的分子机制，我们利用具有逐渐增强的自发转移潜能的培养HCC细胞模型（MHCC97L、MHCC97H和HCCLM6）进行了一次人类信号转导寡核苷酸微阵列分析。结果显示，丝裂原活化蛋白激酶（MAPK）通路是HCC转移中显著上调的通路。进一步研究表明，细胞外信号调节激酶（ERK）（1/2）和p38 MAPK的基础磷酸化水平从MHCC97L细胞到MHCC97H细胞再到HCCLM6细胞依次升高，但c-Jun氨基末端激酶并非如此。通过综合运用PKCβRNA干扰、PKCβ特异性抑制剂恩杂鲁胺和PKC激活剂佛波醇-12-肉豆蔻酸酯-13-乙酸酯，发现HCC细胞中ERK（1/2）和p38 MAPK的磷酸化受上调的蛋白激酶Cβ（PKCβ）调控。热休克蛋白27（HSP27）也被证实为PKCβ-ERK（1/2）和PKCβ-p38 MAPK下游共同激活的蛋白。体外迁移和侵袭试验进一步表明，PKCβ的缺失或PKCβ激活的抑制有效降低了HCC细胞的运动性和侵袭能力。此外，ERK抑制剂1,4-二氨基-2,3-二氰基-1,4-双[2-氨基苯硫基]丁二烯或p38 MAPK抑制剂4-（4-氟苯基）-2-（4-甲亚磺酰基苯基）-5-（4-吡啶基）-1H-咪唑显著抑制了佛波醇-12-肉豆蔻酸酯-13-乙酸酯刺激的PKCβ介导的HCC细胞的运动性和侵袭能力。研究还表明，HSP27在PKCβ介导的HCC细胞运动性和侵袭中起关键作用。综上所述，本研究首次证实了PKCβ-ERK（1/2）/p38 MAPK-HSP27通路在调节HCC细胞运动性和侵袭中的重要作用。
