Downregulation of Talin1 promotes hepatocellular carcinoma progression through activation of the ERK1/2 pathway

Talin1 is an adaptor protein that conjugates integrins to the cytoskeleton and regulates integrins and focal adhesion signaling. Several studies have found that Talin1 is overexpressed in several tumor types and promotes tumor progression. However, the explicit role of Talin1 in hepatocellular carcinoma (HCC) progression is still unclear and its functional mechanism remains largely unknown. In this study, we showed a trend of gradually decreasing expression of Talin1 from normal liver tissues to hepatocirrhosis, liver hyperplasia, the corresponding adjacent non‐tumor, primary HCC, and eventually metastatic foci, indicating that Talin1 may correlate with HCC initiation to progression. Talin1 was significantly downregulated in HCC tissues compared with adjacent non‐tumor tissues and low Talin1 expression was associated with HCC progression and poor prognosis. Furthermore, Talin1 knockdown induced epithelial–mesenchymal transition and promoted migration and invasion in SK‐Hep‐1 cells and HepG2 cells. Mechanistically, we found that the ERK pathway was responsible for these promoting effects of Talin1 knockdown in HCC cells. The promoting effects of Talin1 knockdown on epithelial–mesenchymal transition, migration, and invasion were reversed by U0126, a specific ERK1/2 inhibitor. Taken together, our results suggested that Talin1 might serve as a tumor suppressor in HCC and a potential prognostic biomarker for HCC patients.     

Cancer stem/progenitor cells are highly enriched in CD133+CD44+ population in hepatocellular carcinoma

Both our previous study and other reports have suggested that CD133, originally classified as a hematopoietic stem cell marker, could be used for enrichment of cancer stem cells (CSCs) in human hepatocellular carcinoma (HCC). It was also noted that not all of CD133⁺ cells were representative of CSCs. Further identification and characterization of CSCs or tumor‐initiating cells in HCC are necessary to better understand hepatocarcinogenesis. In present study, we demonstrated that CSC phenotype could be precisely defined by co‐expression of CD133 and CD44 cell surface markers. CD133⁺CD44⁺ HCC cells showed stem cell properties, including extensive proliferation, self‐renewal, and differentiation into the bulk of cancer cells. In vivo xenograft experiments revealed that, actually, the highly tumorigenic capacity of CD133⁺ cells as previously described was primarily attributed to CD133⁺CD44⁺ cell subpopulation, instead of their CD133⁺CD44^? counterparts. Moreover, cells double‐positive for CD133 and CD44 exhibited preferential expression of some stem cell‐associated genes and were more resistant to chemotherapeutic agents due to the upregulation of ATP‐binding cassette (ABC) superfamily transporters, including ABCB1, ABCC1, and ABCG2, further supporting these cells as HCC cell origin. Our findings suggest that CD133⁺CD44⁺ cells might represent true cancer stem/progenitor cells in HCC, which could allow a better understanding of HCC initiation and progression, as well as establish a precise target for the development of more effective therapies.     

ERK2和p-ERK1/2蛋白在宫颈癌中的表达和意义

目的 研究细胞外信号调节激酶2(extracellular regulated kinase 2,ERK2)及其磷酸化状态(p-ERK1/2)在宫颈癌的表达情况,探讨二者与宫颈癌侵袭、转移的关系.方法 采用免疫组化(SP)法,检测80例宫颈癌组织、50例CIN组织及30例正常宫颈组织中ERK2和p-ERK1/2的表达.结果 ERK2和p-ERK1/2在宫颈癌中阳性表达率为75%和52.5 %,明显高于CIN组的34%和28%,以及正常宫颈组的13.33%和20%(P<0.01); ERK2和p-ERK1/2阳性表达率随临床分期的增高而增加(P<0.05),病理分级的降低而增加(P<0.01),有淋巴结转移者ERK2和p-ERK1/2阳性表达率明显高于无转移者(P<0.01);ERK2及p-ERK1/2在宫颈癌中的表达呈正相关(r=0.61,P<0.01).结论 ERK2及p-ERK1/2的表达与宫颈癌的生长、浸润、转移等生物学行为关系密切,在宫颈癌的发生发展过程中发挥重要作用.     

ERK2和p-ERK1/2蛋白在宫颈癌中的表达和意义

目的：研究细胞外信号调节激酶2（extracellular regulated kinase 2,ERK2）及其磷酸化状态（p-ERK1/2）在宫颈癌的表达情况,探讨二者与宫颈癌侵袭、转移的关系。方法：采用免疫组化（SP）法,检测80例宫颈癌组织、50例CIN组织及30例正常宫颈组织中ERK2和p-ERK1/2的表达。结果：ERK2和p-ERK1/2在宫颈癌中阳性表达率为75%和52.5%,明显高于CIN组的3 4%和2 8%,以及正常宫颈组的13.33%和20%（P〈0.01）;ERK2和p-ERK1/2阳性表达率随临床分期的增高而增加（P〈0.05）,病理分级的降低而增加（P〈0.01）,有淋巴结转移者ERK2和p-ERK1/2阳性表达率明显高于无转移者（P〈0.01）;ERK2及p-ERK1/2在宫颈癌中的表达呈正相关（r=0.61,P〈0.01）。结论：ERK2及p-ERK1/2的表达与宫颈癌的生长、浸润、转移等生物学行为关系密切,在宫颈癌的发生发展过程中发挥重要作用。     

肝细胞癌组织CD44v6的检测及其临床意义

目的　探讨原发性肝细胞癌患者术后组织切片中CD44v6的表达及其临床意义。方法　通过免疫组化方法 ,用CD44v6的单克隆抗体 ,以S -P染色法检测CD44v6的表达。结果　CD44v6在癌组织中的表达显著高于癌旁组织 ,有肝外转移的患者其CD44v6的表达也明显高于没有转移的患者 ,而CD44v6的表达在肿瘤大小、分化程度、门静脉癌栓、两年内复发等方面没有显著性差异。结论　CD44v6与肝细胞癌的细胞恶变的发生有关 ,而且CD44v6在有肝外转移患者中的高表达说明它在肝癌发生转移中发挥重要作用。而CD44v6的表达与肿瘤大小、分化程度、门静脉癌栓及两年内复发无关。     

CD44在肝细胞癌及癌周组织中的表达及其意义

 目的 研究CD44s及CD44v6蛋白在人肝细胞癌组织中表达及其意义。方法 应用免疫组化方法检测42例人肝细胞癌及其癌周组织与10例正常肝组织标本的CD44s及CD44v6表达情况。结果 CD44s及CD44v6在肝癌组织中表达明显高于癌周及正常肝组织，与门静脉内癌栓、包膜浸润及分化程度密切相关（分别为P＜0.01，P＜0.05，P＜0.05）。生存时间2年及2年以上组CD44s及CD44v6阳性表达分别为17.6 %（3／17），5.8 %（1／17），生存时间短于2年分别为52.6 %（10／19），42.2 %（8／19）（P＜0.05）。结论 肝细胞癌组织中CD44的表达与细胞分化、浸润转移密切相关，其阳性表达有助于判断预后。     

Insulin receptor tyrosine kinase substrate activates EGFR/ERK signalling pathway and promotes cell proliferation of hepatocellular carcinoma

Insulin receptor tyrosine kinase substrate (IRTKS) is closely associated with actin remodelling and membrane protrusion, but its role in the pathogenesis of malignant tumours, including hepatocellular carcinoma (HCC), is still unknown. In this study, we showed that IRTKS was frequently upregulated in HCC samples, and its expression level was significantly associated with tumour size. Enforced expression of IRTKS in human HCC cell lines significantly promoted their proliferation and colony formation in vitro, and their capacity to develop tumour xenografts in vivo, whereas knockdown of IRTKS resulted in the opposite effects. Furthermore, the bromodeoxyuridine (BrdU) incorporation analyses and propidium iodide staining indicated that IRTKS can promote the entry into S phase of cell cycle progression. Significantly, IRTKS can interact with epidermal growth factor receptor (EGFR), results in the phosphorylation of extracellular signal-regulated kinase (ERK). By contrast, inhibition of ERK activation can attenuate the effects of IRTKS overexpression on cellular proliferation. Taken together, these data demonstrate that IRTKS promotes the proliferation of HCC cells by enhancing EGFR–ERK signalling pathway.     

CD44,CD90的表达与肝癌肝移植术后复发转移的关系

目的:探讨CD44,CD90的表达与肝细胞肝癌肝移植患者预后的关系.方法:收集福州总医院2002至2012年行经典原位肝移植治疗的105例原发性肝细胞癌患者的石蜡包埋标本.应用免疫组化染色法检测肿瘤组织中CD90,CD44的表达,并对临床病理特征及术后随访资料进行统计学分析.结果:CD44和CD90的表达相关关系经Spearman相关分析,两者呈正相关(r=0.5,P＜0.001).COX多因素分析显示,患者5年无复发生存率与CD90高表达(HR=2.765,P=0.002)显著相关.Kaplan-Meier分析显示CD90高表达的患者与CD90低表达患者的肝切除术后5年无复发生存率、术后5年总生存率均有显著差异(40.4％ vs57.6％)(P＜0.05)、(12.7％vs 61.1％) (P＜0.001).COX多因素分析显示,患者5年总生存率与肿瘤大小(HR=1.743;P=0.037)、AFP(HR=2.291;P=0.004)、肿瘤分化(HR=0.283;P＜0.001)、CD44(HR=1.977;P =0.029)、CD90(HR=1.883;P=0.041)表达显著相关.Kaplan-Meier分析显示CD44高表达的患者与CD44低表达患者的肝切除术后5年总生存率有显著差异(16.6％vs 62.7％) (P＜0.001),CD44和CD90共同高表达的患者比CD44或CD90低表达患者的术后5年总生存率显著低(10.7％vs 54.5％)(P ＜0.001).结论:CD90与CD44的高表达相关,CD90与肝癌肝移植术后复发转移有关,二者均影响肝癌肝移植患者预后.     

The iron chelator Dp44mT inhibits hepatocellular carcinoma metastasis via N-Myc downstream-regulated gene 2 (NDRG2)/gp130/STAT3 pathway

Here we showed that hepatocellular carcinoma (HCC) cell lines with high metastatic potential had low levels of NDRG2. The iron chelator Dp44mT up-regulated NDRG2, suppressed epithelial-mesenchymal transition (EMT) and inhibited tumor metastasis in HCC having high metastatic potential. Also Dp44mT attenuated the TGF-β1-induced EMT in HCC having low metastatic potential. In agreement, silencing endogenous NDRG2 with shNDRG2 in HCC cells attenuated the effect of Dp44mT. We showed that the NDRG2/gp130/STAT3 pathway can mediate Dp44mT effects. In agreement, we found that a combination of NDRG2 expression and p-STAT3 levels is a strong predictor of prognosis in HCC patients. We suggest that up-regulation of NDRG2 by Dp44mT is a promising therapeutic approach in HCC.     

CAAP1抑制肝癌HepG2细胞凋亡而促进其增殖、迁移和侵袭

目的:探讨CAAP1对肝癌HepG2细胞凋亡、增殖、迁移和侵袭的影响及其作用机制.方法:构建CAAP1过表达载体pcDNA3/CAAP1和敲降载体pSilencer 2.l-U6 neo/shR-CAAPl,转染肝癌HepG2细胞后,qPCR和WB实验分别检测CAAP1mRNA和蛋白的表达水平.实验分为过表达对照组(p...     

Glypican-6在胃肠道间质瘤组织中的表达及其通过ERK1/2通路促进细胞的增殖、迁移及侵袭

目的:探究Glypican-6对胃肠道间质瘤的作用及其可能的作用机制.方法:收集胃肠道间质瘤组织及其癌旁组织27例,RT-PCR和Western blot检测组织中Glypican-6的表达.选取胃肠道间质瘤细胞系GIST-T1进行体外研究.RT-PCR实验检测细胞中Glypican-6 mRNA的表达;Western...     

HECA通过Wnt通路抑制肝癌细胞增殖、迁移和侵袭

目的:拟在通过影响肝细胞癌HepG2细胞株中HECA基因表达变化,观察与HECA基因相关联的Wnt通路上重要调节分子的变化,讨论此通路影响肝癌细胞增殖能力状况及其相关作用机理。方法:HECA慢病毒感染肝癌细胞株HepG2,细胞株中稳定上调HECA,用HECA siRNA转染HepG2细胞株下调HECA,在HECA表达上调的细胞株中用qPCR检测Wnt通路中β-catenin、TCF4、CDK2、CDK9、Cyclin A和Cyclin K的mRNA表达水平的变化,用Western blot检测qPCR结果中差异明显的β-catenin、TCF4蛋白表达水平的变化,应用FH535抑制剂处理HECA过表达的HepG2细胞,进一步验证HECA基因与Wnt通路的关系,利用Western blot检测HECA的表达水平,用MTT、划痕、克隆形成以及Transwell实验检测对肝癌细胞侵袭、迁移、增殖能力的影响。结果:qPCR法以及Western blot法对其相关基因表达水平进行测定,结果显示在HepG2细胞株内HECA过表达后,与HECA相关联的Wnt通路重要调节分子中β-catenin、TCF4的表达水平显著降低;FH535抑制剂处理HECA过表达的HepG2细胞,Western blot实验结果显示FH535处理过的过表达组HECA水平更高,细胞功能实验也表明FH535处理过的过表达HECA组,其细胞增殖及侵袭能力下降最明显。结论:HECA基因抑制肝细胞癌增殖及侵袭的分子机制可能为:HECA通过Wnt/β-catenin这一经典通路内β-catenin与TCF4的结合活性降低而发挥作用。     

Shp2 SUMOylation promotes ERK activation and hepatocellular carcinoma development

Shp2, an ubiquitously expressed protein tyrosine phosphatase, is essential for regulation of Ras/ERK signaling pathway and tumorigenesis. Here we report that Shp2 is modified by SUMO1 at lysine residue 590 (K⁵⁹⁰) in its C-terminus, which is reduced by SUMO1-specific protease SENP1. Analysis of wild-type Shp2 and SUMOylation-defective Shp2^K590R mutant reveals that SUMOylation of Shp2 promotes EGF-stimulated ERK signaling pathway and increases anchorage-independent cell growth and xenografted tumor growth of hepatocellular carcinoma (HCC) cell lines. Furthermore, we find that mutant Shp2^K590R reduces its binding with the scaffolding protein Gab1, and consistent with this, knockdown of SENP1 increased the interaction between Shp2 and Gab1. More surprisingly, we show that human Shp2 (hShp2) and mouse Shp2 (mShp2) have differential effects on ERK activation as a result of different SUMOylation level, which is due to the event of K⁵⁹⁰ at hShp2 substituted by R⁵⁹⁴ at mShp2. In summary, our data demonstrate that SUMOylation of Shp2 promotes ERK activation via facilitating the formation of Shp2-Gab1 complex and thereby accelerates HCC cell and tumor growth, which presents a novel regulatory mechanism underlying Shp2 in regulation of HCC development.     

DDR2 facilitates hepatocellular carcinoma invasion and metastasis via activating ERK signaling and stabilizing SNAIL1

Background

Several studies have found that DDR2 is up-regulated in many tumor types and facilitates tumor progression. However, the role of DDR2 in hepatocellular carcinoma (HCC) progression and its downstream signaling pathways remain unclear.

Methods

DDR2 expression was assessed in several cell lines and 112 pairs of HCC and matched adjacent noncancerous liver tissues. Clinical significance of DDR2 in HCC was analyzed. Phosphorylated DDR2 (p-DDR2) expression was detected by immunoblotting to evaluate its correlation with DDR2. The effect of DDR2 on HCC cell migration and invasion were examined. Cycloheximide chase experiments were performed to detect the half-life of SNAIL1. Moreover, DDR2 expression was detected by immunohistochemistry to evaluate its correlation with SNAIL1. The regulatory effect of DDR2 on ERK signaling, SNAIL1, EMT, MT1-MMP and MMP2 was confirmed by immunoblotting. The effect of type I collagen on DDR2/ERK2/SNAIL1 signaling was assessed.

Results

DDR2 was more highly expressed in HCC than in non-HCC tissues. DDR2 overexpression was correlated with clinicopathological features of poor prognosis. Clinical analysis revealed that DDR2 is an independent prognostic marker for predicting overall survival and disease free survival of HCC patients. Overexpression of DDR2 is associated with p-DDR2 amplification. In vitro studies showed that DDR2 facilitates HCC cell invasion, migration and EMT via activating ERK2 and stabilizing SNAIL1. DDR2 can up-regulate MT1-MMP and MMP2 expression through ERK2/SNAIL1 signaling in HCC. Additionally, collagen I can induce DDR2/ERK2/SNAIL1 signaling activation in HCC cells.

Conclusions

Our findings suggest that DDR2 plays an important role in promoting HCC cell invasion and migration, and may serve as a novel therapeutic target in HCC.     

整合素α5β1与CD44s在非小细胞肺癌中表达的意义

目的:研究整合素α5β1与CD44s在NSCLC中表达的意义。方法:应用免疫组织化学SP法检测53例非小细胞肺癌标本中整合素α5β1和CD44s的表达情况,用卡方检验对各指标表达情况与临床病理参数的关系进行分析,用Spearman分析探讨NSCLC组织中整合素α5β1和CD44s表达强度的相关性。结果:有淋巴结转移组和无淋巴结转移组的NSCLC中整合素α5β1的表达阳性率分别为67.50%和30.77%,差异有显著性(P<0.05);CD44s在鳞癌组织中的表达强度明显高于腺癌,表达率分别为46.87%和14.29%,P<0.05;有淋巴结转移组与无淋巴结转移组的NSCLC组织中CD44s的表达率分别为54.55%和23.00%,两者有显著性差异(P<0.05)。NSCLC组织中整合素α5β1与CD44s表达程度呈正相关(rs=0.502,P<0.001)。结论:整合素α5β1和CD44s在NSCLC的表达呈正相关,是预测NSCLC病人预后的有意义的指标。     

人参皂苷Rh2通过Egr-1/TRL4/mTOR信号通路抑制食管癌细胞Eca-109增殖、迁移和EMT

目的:研究人参皂苷Rh2对食管癌细胞Eca-109增殖、迁移和上皮间质转化（epithelial-mesenchymal transition,EMT）的作用以及作用机制。方法:CCK-8法检测人参皂苷Rh2对食管癌细胞Eca-109增殖的影响;细胞划痕实验检测人参皂苷Rh2对食管癌Eca-109细胞迁移的影响;Western blot检测EMT相关蛋白E-cadherin、Vimentin和Slug的蛋白表达水平。结果:人参皂苷Rh2能够显著抑制Eca-109细胞的增殖,且呈剂量依赖性;此外,人参皂苷Rh2显著抑制E-cadherin、Vimentin和Slug的蛋白表达,并抑制Eca-109细胞迁移;人参皂苷Rh2显著抑制Egr-1、TRL4和mTOR的蛋白表达;进一步的研究结果表明人参皂苷Rh2通过抑制Egr-1/TRL4/mTOR信号通路抑制食管癌细胞Eca-109增殖、迁移和EMT。结论:人参皂苷Rh2能够抑制食管癌细胞Eca-109的增殖、迁移和EMT,其作用机制是通过介导Egr-1/TRL4/mTOR信号通路来实现的。这一结果能够为治疗食管癌的进一步研究提供分子基础。