Molecular Surgeon Research Center, Michael E. DeBakey Department of Surgery, Baylor College of Medicine, Houston, Texas, USA.
Mol Cancer. 2011 Aug 26;10:102. doi: 10.1186/1476-4598-10-102.
Cyclophilin A (CypA) expression is associated with malignant phenotypes in many cancers. However, the role and mechanisms of CypA in liver fluke-associated cholangiocarcinoma (CCA) are not presently known. In this study, we investigated the expression of CypA in CCA tumor tissues and CCA cell lines as well as regulation mechanisms of CypA in tumor growth using CCA cell lines.
CypA expression was determined by real time RT-PCR, Western blot or immunohistochemistry. CypA silence or overexpression in CCA cells was achieved using gene delivery techniques. Cell proliferation was assessed using MTS assay or Ki-67 staining. The effect of silencing CypA on CCA tumor growth was determined in nude mice. The effect of CypA knockdown on ERK1/2 activation was assessed by Western blot.
CypA was upregulated in 68% of CCA tumor tissues. Silencing CypA significantly suppressed cell proliferation in several CCA cell lines. Likewise, inhibition of CypA peptidyl-prolyl cis-trans isomerase (PPIase) activity using cyclosporin A (CsA) decreased cell proliferation. In contrast, overexpression of CypA resulted in 30% to 35% increases in proliferation of CCA cell lines. Interestingly, neither silence nor overexpression of CypA affected cell proliferation of a non-tumor human cholangiocyte cell line, MMNK1. Suppression of CypA expression attenuated ERK1/2 activity in CCA M139 cells by using both transient and stable knockdown methods. In the in vivo study, there was a 43% reduction in weight of tumors derived from CypA-silenced CCA cell lines compared with control vector CCA tumors in mice; these tumors with stable CypA silencing showed a reduced cell proliferation.
CypA is upregulated in majority of CCA patients' tissues and confers a significant growth advantage in CCA cells. Suppression of CypA expression decreases proliferation of CCA cell lines in vitro and reduces tumor growth in the nude mouse model. Inhibition of CypA activity also reduces CCA cell proliferation. The ERK1/2 pathway may be involved in the CypA-mediated CCA cell proliferation. Thus, CypA may represent an important new therapeutic target for liver fluke-associated CCA.
亲环素 A（CypA）的表达与许多癌症的恶性表型有关。然而，CypA 在肝吸虫相关胆管癌（CCA）中的作用和机制目前尚不清楚。在这项研究中，我们通过实时 RT-PCR、Western blot 或免疫组织化学检测了 CypA 在 CCA 肿瘤组织和 CCA 细胞系中的表达，并通过 CCA 细胞系研究了 CypA 对肿瘤生长的调控机制。
采用实时 RT-PCR、Western blot 或免疫组化检测 CypA 的表达。采用基因传递技术沉默或过表达 CCA 细胞中的 CypA。采用 MTS 法或 Ki-67 染色法检测细胞增殖。在裸鼠中检测 CypA 沉默对 CCA 肿瘤生长的影响。Western blot 检测 CypA 敲低对 ERK1/2 激活的影响。
68%的 CCA 肿瘤组织中 CypA 上调。沉默 CypA 可显著抑制几种 CCA 细胞系的增殖。同样，用环孢菌素 A（CsA）抑制 CypA 肽脯氨酰顺反异构酶（PPIase）活性也降低了细胞增殖。相反，过表达 CypA 可使 CCA 细胞系的增殖增加 30%至 35%。有趣的是，CypA 的沉默或过表达均不影响非肿瘤性人类胆管细胞系 MMNK1 的细胞增殖。用瞬时和稳定敲低方法抑制 CypA 表达可减弱 CCA M139 细胞中 ERK1/2 的活性。在体内研究中，与对照组载体 CCA 肿瘤相比，沉默 CypA 的 CCA 细胞系来源的肿瘤重量减轻了 43%；这些稳定沉默 CypA 的肿瘤显示出细胞增殖减少。
CypA 在大多数 CCA 患者的组织中上调，并赋予 CCA 细胞显著的生长优势。抑制 CypA 表达可减少体外 CCA 细胞系的增殖，并减少裸鼠模型中的肿瘤生长。抑制 CypA 活性也可减少 CCA 细胞的增殖。ERK1/2 通路可能参与 CypA 介导的 CCA 细胞增殖。因此，CypA 可能成为肝吸虫相关 CCA 的一个重要新的治疗靶点。
