MiR-210 regulates cell cycle in nasopharyngeal carcinoma cell line （CNE-1） under hypoxic condition by reducing the expression of cyclin DI

Objective： The aim of our study was to determine the underlying mechanism of miR-210 on regulation of the cell cycle in nasopharyngeal carcinoma cell line CNE-1, particularly through regulation of cyclin D1, under hypoxic conditions. Methods： The CNE-1 cell line was induced with hypoxia, and the expression levels of endogenic miR-210 and cyclin D1 were detected by real-time PCR and Western blotting. Next, the luciferase assay was used to confirm that cyclin D1 is a target gene for miR-210. Cell cycle and cell proliferation were detected in CNE-1 cells that were cultured under hypoxic conditions with either overexpression or knockout of miR-210 using flow cytometry and MTT assay, respectively. Results： Hypoxia induced the expression of miR-210, resulting in reduced mRNA and protein levels of cyclin D1 and repression of cyclin D1 in CNE-1 cells. Further analysis indicated that miR-210 directly binded to the 3＇UTR of the cyclin D1 gene, thus regulated the expression of cyclin DI. The flow cytometry assay showed that, under hypoxic conditions, miR-210 blocked CNE-1 cells in the G1 phase, and miR-210 also inhibited the proliferation of CNE-1 cells. Conclusion： Under hypoxic conditions, miR-210 directly reduced the expression of cyclin D1, leading to CNE-1 cells blocked in G1 phase.     

Inhibition of proliferation of human breast cancer MCF-7 cells by small interference RNA against LRP16 gene

Objective: Our previous studies have firstly demonstrated that 17β-E2 up-regulates LRP16 gene expression in human breast cancer MCF-7 cells, and ectopic expression of the LRP16 gene promotes MCF-7 cells proliferation. Here, the effects of the LRP16 gene expression on growth of MCF-7 human breast cancer cells and the mechanism were further studied by establishing two stably LRP16-inhibitory MCR-7 cell lines. Methods: Hairpin small interference RNA (siRNA)strategy, bY which hairpin siRNA was released by U6 promoter and was mediated by pLPC-based retroviralvector, was adopted to knockdown endogenous LRP16 level in MCF-7 cells. And the hairpin siRNA against green fluorescence protein (GFP) was used as the negative control. The suppressant efficiency of the LRP16 gene expression was confirmed by Nothern blot.Cell proliferation assay and soft agar colony formation assay were used to determine the status of the cell sproliferation. Cell cycle checkpoints including cyclin E and cyclin D1 were examined by Western blot. Results:The results from cell proliferation assays suggested that down-regulation of LRP16 gene expression is capable of inhibiting MCF-7 breast cancer cell growth and down-regulation of the LRP16 gene expression is able to inhibit anchorage-independent growth of breast cancer cells in soft agar. We also demonstrated that cyclin E and cyclin D1 proteins were much lower in the LRP16-inhibitory cells than in the control cells.Conclusion: These data suggest that LRP16 gene play an important role in MCF-7 cells proliferation by regulating the pathway of the G1/S transition and may function as an important modulator in regulating the process of tumorigenesis in human breast.     

肝癌细胞系BEL-7402中siRNA表达载体介导靶向c-myc基因的RNA干涉实验研究

Objective： To explore the inhibition of the expression of c-Myc in human hepatocellular carcinoma via vector-based RNA interference （RNAi）. Methods： Two RNA interference DNA templates targeting c-Myc oncogene were designed via online software and synthesized. By ligation, the fragments were inserted into pSilencer 1.0-U6 to construct the recombinant siRNA expressing plasmids. The identified recombinants were introduced into BEL-7402 cells with Lipofactamine. The inhibition of c-Myc expression, together with the expression of CDK4, hTERT and Gadd45β in c-Myc down-regulated BEL-7402 cells, were analyzed by semi-quantitative RT-PCR. Results： Two recombinant plasmids pSic-myc-1 and pSic-myc-2, which direct the yields of siRNAs targeting c-Myc in cells, were constructed. Among which, pSic-myc-2 was shown to trigger a RNAi-mediated inhibition of expression of c-Myc in BEL-7402 by up to 90%. In c-Myc knockdown BEL-7402 cells, the expression of CDK4 and hTERT were down-regulated with a ratio of 85% and 57%, respectively, while the expression of Gadd45β was up-regulated by up to 110%. Conclusion： The expression of c-Myc in BEL-7402 could be suppressed by vector-based RNA interference successfully. The knockdown of c-Myc in turn resulted in the changes of expression of genes related to cell proliferation and apoptosis. Thus, our study provided a preliminary data in searching of a c-Myc-targeted RNAi therapy of human hepatocellular carcinoma.     

转染AP-2α基因对结肠癌SW620细胞增殖及凋亡的影响(英文)

Objective:We investigated the effects of exogenous AP-2α gene on SW620 cell cycle, apoptosis and proliferation. Methods:The Plasmid pcDNA3.1(+)-AP-2α was transfected into colorectal carcinoma SW620 cells line by liposome mediation for transient expression. After AP-2α transfected SW620 cells, the exogenous AP-2α mRNA and protein express were determined by the method of Real-time PCR and Western blot. In order to elucidate the effect of expression of exogenous AP-2α gene on the colorectal cancer cell SW620, ...     

转染外源CC10对肺腺癌细胞系A549的CyclinD1蛋白及mRNA的影响

Objective： To investigate the effects of exogenous CC10 gene transfection on cell cycle and the expression of cyclinD1 protein and mRNA in A549 cells. Methods： A549 cells in all test groups （group A to E） and control group （group F） were transfected with exogenous CC10 gene by liposome for 8, 16, 24, 36, 48 and 0 h respectively. CC10 protein expression was detected in A549 cells by Western blot. The growth inhibitory rate was detected by MTT method. Flow cyometry analysis （FCS） and AnnexinV-PI staining were used to determine the changes of cell cycle progression and apoptosis rate in all groups. CyclinD1 protein and mRNA expression in A549 cells was detected by the methods of immunocytochemistry and RT-PCR. Results： Exogenous CC10 gene could inhibit the growth of A549 cells, and the growth inhibitory rates in all test groups （from group A to E） were 24.7%, 33.1%, 44.3%, 61.7% and 74.2% respectively, and that in group F was 6.24%. CC10 blocked the cell cycle progression at G0/G1 and induced apoptosis gradually. In A549 cells of test groups, the expression of cyclinD1 protein and mRNA was significantly decreased. Conclusion： The inhibitory effects of the transfection of exogenous CC10 gene on G0/G1 cycle of lung cancer cells might be related with the down-regulation of cyclinD1 gene.     

新辅助化疗对骨肉瘤细胞增殖、凋亡及耐药性的影响及其机制

Objective： To study the influence of neoadjuvant chemotherapy on the expression of cyclin D1, Bcl-2, PCNA and P-gp in osteosarcoma cells and the relationship between the expression and tumor cell necrosis rate （TCNR） after chemotherapy. Methods： By using immunohistochemistry, the expression of cyclin D1, Bcl-2, PCNA and P-gp was detected in 23 cases of osteosarcoma and TCNR were calculated. Results： The pre-chemotherapy positive expression rate of cyclin D1, Bcl-2, PCNA and P-gp was 73.9%, 69.6%, 91.3% and 21.7%, respectively, and that post-chemotherapy positive expression rate was 52.1%, 34.8%, 43.5% and 56.5%, respectively. The positive expression rate of Bcl-2 and PCNA after chemotherapy was much lower than that before chemotherapy （P=0.039, 0.034）. After chemotherapy, the expression rate of P-gp was higher （P=0.021） and the expression of cyclin D1 had no statistically significant difference （P=0.180） comparing with that before chemotherapy. No correlation existed between the expression of cyclin D1, Bcl-2, PCNA, P-gp and TCNR before chemotherapy （P=0.155, 0.371, 1.000 and 0.640）. There was a negative correlation between the expression of Bcl-2, PCNA, P-gp and TCNR （P=0.009, 0.012 and 0.015）, but no relationship existed between the cyclin D1 and TCNR （P=-0.100） after chemotherapy. Conclusion： Chemotherapy could inhibit proliferation and induce apoptosis of osteosarcoma cells. At the same time, due to the overexpression of the P-gp, the drug resistance of the osteosarcoma cells was increased. The detection of cyclin D1, Bcl-2, PCNA and P-gp in osteosarcoma samples before chemotherapy might not be used to predict the curative effect of the chemotherapy.     

DJ-1 promotes proliferation and epithelial-mesenchymal transition of colorectal cancer cells by regulating p53 signaling pathway北大核心CSCD

Objective: To investigate the effects of up-regulating or silencing DJ-1 gene expression on the apoptosis, migration and invasion of colorectal cancer (CRC), and to explore the possible molecular mechanism. Methods: The expression level of DJ-1 in CRC tissues and cells was detected by immunohistochemistry, Western blotting and real-time fluorescent quantitative PCR, respectively. The SW480 and HCT116 cells were transfected with the recombinant lentiviral vector carrying human DJ-1 gene to obtain DJ-1 overexpressed SW480/OE-DJ-1 and HCT116/OE-DJ-1 cells, while the cells transfected with the empty vector was as the negative control (OE-NC). The SW480 and HCT116 cells were transfected with the recombinant lentiviral vector carrying the specific shRNA targeting DJ-1 gene to generate the SW480/shDJ-1 and HCT116/sh-DJ-1 cells with stable knockdown of DJ-1, while the cells transfected with the empty vector was as the negative control (sh-NC). Subsequently, the expressions of DJ-1 and p53 protein and mRNA were detected by immunohistochemistry and real-time fluorescent quantitative PCR, respectively; and their relationship was analyzed. The expressions of p53 and its downstream apoptosis-related proteins Bax and Bcl-2 in SW480 and HCT116 cells with DJ-1 over-expression or knockdown were detected by Western blotting. The effects of overexpressing and silencing DJ-1 gene expression on the invasion and migration abilities of SW480 and HCT116 cells were detected by Transwell chamber assay. The epithelial-mesenchymal transition (EMT) of CRC cells was induced by transforming growth factor-β1 (TGF-β1), then the expression levels of DJ-1 and EMT-related markers (N-cadherin, β-catenin, vimentin, E-cadherin) were analyzed by Western blotting. Results: DJ-1 was highly expressed in 34 CRC tissues (24/34, 70.59%) (P < 0.001). The overall survival time of the patients with DJ-1 high expression was significantly shorter than that of the patients with DJ-1 low expression (P < 0.001). The high expression of DJ-1 was correlated with TNM stage, tumor location, lymph node metastasis, and degree of differentiation (all P < 0.05). There was a negative correlation between DJ-1 and p53 expressions (r =-0.428, P = 0.015). Silencing DJ-1 increased the expression level of p53 and its downstream apoptotic protein Bax, decreased the expression of anti-apoptotic protein Bcl-2 (all P < 0.05), and decreased the invasion and migration capacities of SW480 and HCT116 cells (both P < 0.01); Conversely, overexpressing DJ-1 decreased the expression level of p53 and Bax, increased the expression of Bcl-2 (all P < 0.05), and increased the invasion and migration capacities of SW480 and HCT116 cells (both P < 0.01). Overexpression of DJ-1 induced by TGF-β1 increased the expressions of N-cadherin, β-catenin and vimentin, and decreased the expression of E-cadherin in the process of EMT (P < 0.05). Conclusion: DJ-1 promotes the apoptosis and invasion of CRC cells by negatively regulating the p53 signaling pathway. © 2019 by TUMOR All rights reserved.     

EXPRESSION OF FLIP IN HUMAN COLON CARCINOMAS： A NEW MECHANISM OF IMMUNE EVASION

Objective： It has been proposed that Fas ligand （FasL） may play an important role in immune escape of tumors and FLIP is an important mediator of Fas/FasL pathway. In this study, the expression of FLIP was determined in human colon carcinoma cell lines and tissue to investigate the new mechanism of immune evasion of human colon carcinomas. Methods： RT-PCR and immunohistochemistry （IHC） were performed to investigate the expression of FLIP in human colon carcinoma cell lines SW480, LS174 and twenty human primary colon carcinoma specimens. Results： It was shown that SW480 cells, LS174 cells and primary colon carcinoma specimen constitutively expressed FLIP at the mRNA and protein level. The expression of FLIP was not found in the epithelial cells of normal colon mucosa. Conclusion： FLIP was expressed in human primary colon carcinoma specimens but not in the normal counterpart. It suggested that the expression of FLIP may occur during the malignant transformation from normal colon epithelial cells to colon carcinoma cells. Tumor cells might obtain the ability to resist the Fas-mediated apoptosis by expressing FLIP. The expression of FLIP might contribute to the formation of colon carcinomas.     

Expression of Cyclin E and Its Relationship with the Prognosis of Patients with Breast Cancer

Objective： To investigate the expression of cyclin E in breast cancer tissues and its relationship with prognosis of the patients with breast cancer. Methods： The expression of cyclin E, HER-2/neu, nm23-H1 and actin was detected in 80 breast cancer tissues and 18 benign breast tumor tissues by immunohistochemical methods. The relationship between cyclin E and the remaining genes or the clinical data of the patients with breast cancer was analyzed. Results： The over expression rate of cyclin E in malignant tissues was obviously higher than that in benign tumor tissues （P〈0.01）. The over expression of cyclin E in later stage of disease was higher than that in early stage of disease （P〈0.05）. The expression of cyclin E in ER positive tissues was lower than that in ER negative tissues （P〈0.05）. The expression of cyclin E in PR positive tissues and PR negative tissues had no significant difference （P〉0.05）. The expression of cyclin E in HER-2/neu positive tissues was higher than that in HER-2/neu negative tissues （P〈0.05）. And the expression of cyclin E in ER, PR and HER-2/neu all positive tissues was much higher （P〈0.01）. There was no significant difference in the expression of cyclin E between nm23-H1 positive tissues and nm23-H1 negative tissues （P〉0.05）. The expression of cyclin E in actin positive and continuous distribution tissues was lower than that in actin negative or discontinuous distribution tissues （P〈0.05）. Conclusion： The expression of cyclin E has a strong correlation to the prognosis of the patients with breast cancer.     

hL RH-1 调控结肠腺癌细胞SW480 增殖的分子机制

 目的 初步探讨人核受体hLRH-1在结肠癌发生发展中可能的生物学功能。方法 真核表达质粒pcDNA3-hLRH-1经脂质体介导转入SW480细胞中，RT-PCR和Western blot法分别检测外源基因mRNA及蛋白水平的表达；M1vr法分析hLRH-1过表达对SW480细胞生长增殖的影响，同时行实时定量RT-PCR法分析细胞生长增殖相关基因CyclinE1和CyclinD1、以及凋亡调控相关基因Pten和Rb1的表达。结果 hLRH-1过表达的SW480细胞增殖速度明显加快，同时其CyclinE1基因的表达显著增高，而Pten和Rb1在hLRH-1过表达的SW480细胞中均呈明显的表达下调。结论 外源hLRH-1的表达既通过上调CyclinE1基因的表达而促进SW480细胞增殖，还可能通过调节Pten和Rb1基因的表达而参与细胞凋亡的调控。     

Inhibition of p38 MAPK increases the sensitivity of 5-fluorouracil-resistant SW480 human colon cancer cells to noscapine

A major cause of treatment failure in advanced colon cancer is resistance to chemotherapy. p38 mitogen-activated protein kinase (MAPK) has been associated with cellular apoptosis and plays an important role in multidrug resistance (MDR) in cancer cells. In the present study the effect of p38 MAPK on the sensitivity of 5-fluorouracil (5-FU)-resistant SW480 (SW480/5-FU) human colon cancer cells to noscapine was investigated. Following p38 MAPK interference, the inhibitory effect of noscapine on cell viability and proliferation was increased in the SW480/5-FU cells and there was also a decrease in the expression level of minichromosome maintenance proteins, recombinant Ki-67 and proliferating cell nuclear antigen. Inhibition of p38 MAPK also enhanced noscapine-induced G₁-phase cell cycle arrest in the SW480/5-FU cells and there was also a decrease in the protein and mRNA expression level of cyclin D, cyclin E and cyclin-dependent kinase 2, and an increase in the expression level of P57. Furthermore, p38 MAPK interference increased noscapine-induced apoptosis of the SW480/5-FU cells and there was an increase in the protein and mRNA expression level of caspases-3 and 8 and Bax, and decreased Bcl-2 expression level. The sensitivity of the SW480/5-FU cells to noscapine was also increased following p38 MAPK interference, as demonstrated by MDR inhibition via decreased Akt activity and reduced protein expression level of the MDR proteins P-glycoprotein, multidrug resistance protein 1 and ATP-binding cassette G2. These observations indicated that inhibition of p38 MAPK increased the sensitivity of the SW480/5-FU cells to noscapine by suppressing proliferation, induction of cell cycle arrest and apoptosis, and reversal of MDR in the SW480/5-FU cells.     

DNA-PK／AKT／GSK3β通路与结肠癌细胞的放射抗拒性

目的探索结肠癌细胞株SW480在长期分割放疗下获得放射抗拒性的可能机制。方法经不同剂量0Gy、2Gy、5Gy、10Gy单次x射线照射后,以RT．PCR法检测结肠癌亲本细胞SW480和放射抗拒性细胞SW480-R中CCNDlmRNA的表达;Westernblot法检测DNA．PK／AKT／GSK3[3通路中两种细胞株cyclinD1、CDK4、Rb、P—Rb．Ser795、AKT、p-AKT-ser473、GSK313、p-GSK3β-Ser9、DNA．PKcs、P—DNA．PKcs蛋白的表达。结果经单次x射线照射0Gy、2Gy、5Gy、10Gv后,RT—PCR法检测显示SW480．R细胞中CCNDlmRNA的表达明显低于SW480的表达（P〈0．05）,其相对表达水平分别为0．31±0．02、0．32±0．03、0．34＋0．05、0．44±0．04;Westernblot法检测显示SW480和SW480．R中的cyclinDI、CDK4、Rb、P—Rb．Ser795、AKT、P—AKT．Ser473、p-GSK3β-Ser9、DNA—PKcs及p-DNA-PKcs蛋白的表达含量在两种细胞中显著不同,SW480．R的蛋白表达明显高于SW480（P〈0．05）,而GSK313蛋白的表达含量SW480．R却低于SW480（P〈0．05）。结论经过长期分割放疗的结肠癌细胞获得的放射抗拒性可能与激活DNA—PK／AKT／GSK313通路中cyclinD1蛋白过表达介导的DNA损伤反应（DDR）的改变有关。     

慢病毒介导的RNA干扰沉默KIF14表达对结肠癌细胞生长和转移的影响

目的:探讨慢病毒介导RNA干扰沉默KIF14表达对结肠癌SW480细胞生长和转移的影响。方法:采用脂质体法将构建的慢病毒靶向siRNA-KIF14干扰载体转入SW480细胞后,采用RT-PCR和Western blot检测KIF14 mRNA和蛋白的表达,MTT法、流式细胞仪和Transwell小室实验分别检测SW480细胞增殖、凋亡和侵袭,Western blot检测上皮-间质转化相关蛋白N-cadherin、E-cadherin和Vimentin的表达。结果:转染SW480细胞后成功下调KIF14 mRNA和蛋白的表达。与未转染细胞相比,下调KIF14的表达可抑制SW480细胞增殖、侵袭和上皮-间质转化,并促进细胞凋亡（P＜0.05）。结论:下调KIF14表达可抑制结肠癌SW480细胞生长和转移。     

The mechanisms of 5-FU-PLA-O-CMC-NPS-mediated inhibition of the proliferation of colorectal cancer cell line SW480

We aimed to investigate how 5-FU-PLA-O-CMC-NP (5-FPOCN) inhibits the proliferation of the SW480 colon cancer cell line. Following the treatment of cell line SW480 with 0.1, 1, 10 or 100 μg/ml 5-FPOCN or 5-fluorouracil (fluorouracil, 5-Fu) for 0, 24, 48, or 72, the rate of cell was tested by the tetrazolium assay (MTT). After the SW480 cells were treated with 5-FPOCN or 5-FU for 72 h, the growth rate and apoptosis were detected. After the SW480 cells were treated with 5-FPOCN or 5-FU for 24, 48, 72, or 120, flow cytometry (FCM) was used to determine the cell cycle distribution. The changes in the expression of P21, CyclinD1 and Rb were detected by Western blotting and real-time PCR. We found that different doses of 5-FPOCN can significantly inhibit the growth rate of SW480 cells, and this effect is dose and time dependent. However, there is no significant difference from 72 to 120 h (P?>?0.05). After 5-FPOCN treatment for 72 h, there is a negative correlation between the concentration of 5-FPOCN and the activity of SW480 cells and a positive correlation between the concentration of 5-FPOCN and SW480 cell apoptosis. G1 phase was significantly increased, and S phase was significantly decreased in 5-FPOCN-treated SW480 cells at 72 h compared to the control group (P?<?0.05); there was a positive correlation between the concentration of 5-FPOCN and the above changes. It was suggested that 5-FPOCN can delay G1/S phase and that this is a dose-dependent effect. The expression of P21 protein and messenger RNA (mRNA) and Rb protein and mRNA was significantly increased in 5-FPOCN-treated SW480 cells at 72 h compared to the control group, and this was a dose- and time-dependent effect. CyclinD1 protein and mRNA expression was reduced as the dose increased, and its expression was negatively associated with the increased expression of P21. We concluded that 5-FPOCN can significantly inhibit the growth of colon cancer SW480 cells. 5-FPOCN increased P21 expression and decreased cyclin family and pRb expression to promote cell cycle delay and apoptosis.     

香加皮杠柳苷通过抑制Wnt／β-catenin信号通路诱导结肠癌细胞SW480凋亡

背景与目的：Wnt／β-catenin信号通路在人类肿瘤尤其在大肠癌的发生发展中起着重要作用。本研究分析了香加皮杠柳苷（periplocin extracted fromcortex periplocae,CPP）对人结肠癌细胞SW480增殖的抑制作用,及对Wnt／β-catenin信号通路的调控作用。方法：MTT法检测CPP对SW480细胞增殖的影响,流式细胞技术检测细胞周期的变化和凋亡。Western blot法检测CPP处理组与对照组细胞总蛋白、细胞浆蛋白及细胞核蛋白中β-catenin表达变化,电泳迁移率改变法分析CPP作用后SW480细胞核TCF复合物与其特异性DNA结合序列结合能力变化。半定量RT—PCR法检测CPP作用后细胞中β-catenin、survivin、c—myc和cyclin D1mRNA的表达。结果：CPP明显抑制SW480细胞增殖（P〈0．01）,并呈时间和浓度依赖性;0．5μg／mLCPP可将SW480细胞阻滞于G0／G1期,并诱导细胞凋亡（P〈0．05）。CPP作用后SW480细胞总蛋白、胞浆蛋白及细胞核蛋白中的B—catenin表达均明显降低（P〈0．01）,细胞核中TCF复合物与其特异性DNA结合序列结合能力受到抑制,其下游靶基因mRNA表达水平下降（P〈0．01）,而β-cateninmRNA表达未见明显改变。结论：CPP可明显抑制SW480细胞增殖并诱导细胞凋亡,其作用机制与抑制细胞Wnt／β-catenin信号转导通路有关。     

Beta-Hydroxybutyrate Promotes Proliferation,Migration and Stemness in a Subpopulation of 5FU Treated SW480 Cells: Evidence for Metabolic Plasticity in Colon Cancer

Background: Beta-hydroxybutyrate (BHB) as a ketone body is the metabolic fuel in oxidative phosphorylationpathway. So far the effects of BHB on the biology of tumor cells is contradictory. Therefore, we investigated the effect ofBHB on viability, metabolism, proliferation and migration of 5FU treated SW480 colon cancer cell line. Methods: wetreated the SW480 cells with IC50 dose of 5-fluorouracil (5FU) for 72 h to isolate a subpopulation of 5FU treated cellsthat were resistant to it. Effects of BHB on cell viability was investigated by MTT assay. Measurement of oxygenconsumption rate (OCR) in parallel with extracellular acidification rate (ECAR) upon BHB treatment was used fordetermination of metabolic profile of these cells. Investigating the relationship between metabolic phenotype andthe status of differentiation and stemness was done by analyzing the expression of PGC-1α, c-MYC, NANOG, ALPiand KRT20 genes by qRT-PCR. Clonogenic and scratch assay were performed to determine the proliferation andmigration abilities of incubated with BHB compared to untreated cells. Results: BHB increased cell viability in SW480and 5FU treated SW480 cells. The results showed a significantly decreased ECAR and increased OCR in both celltypes following BHB treatment reflecting the superiority of oxidative phosphorylation profile compared to glycolysisin both cell types. Also, treatment with BHB increases the expression of genes normally associated with stemnessand mitochondrial biogenesis and decreases the expression of genes related to glycolytic program and differentiationin 5FU treated cells. Self-renewal and migration potential of BHB treated cells increased significantly. Conclusion:These findings suggest that BHB utilization via oxidative mitochondrial metabolism can fuel proliferation, migrationand stemness in 5FU treated SW480 colon cancer cells.     

细丝蛋白A抑制荷人结肠腺癌细胞SW480裸鼠移植瘤的生长

目的:探讨细丝蛋白A(fi lamin A,FLNa)基因对人结肠腺癌SW480细胞在裸鼠体内成瘤的影响及其可能的机制。方法:将重组载体质粒pcDNA3.1/V5-His-TOPO/FLNa通过脂质体介导的方式转染到SW480细胞中,RT-PCR和蛋白质印迹法检测SW480/FLNa细胞中FLNa mRNA和蛋白的表达,MTT法检测细胞的体外增殖能力。将SW480/FLNa、SW480/pcDNA3.1和SW480细胞接种于裸鼠皮下,观察移植瘤的生长情况,计算抑瘤率,HE染色观察移植瘤组织的形态学变化,RT-PCR和蛋白质印迹法检测移植瘤组织中基质金属蛋白酶-9(matrix metalloproteinases-9,MMP-9)mRNA和蛋白的表达水平。结果:FLNa基因在SW480细胞中获得稳定表达,SW480/FLNa、SW480/pcDNA3.1和SW480细胞的体外增殖能力相似;SW480/FLNa组移植瘤的生长速度和质量低于SW480组,抑瘤率为(88.7±3.5)%(P=0.000);SW480/FLNa组移植瘤组织出现退变坏死,MMP-9mRNA和蛋白的表达水平(0.11±0.02和0.02±0.00)均低于SW480组(1.12±0.02和1.25±0.05),差异均有统计学意义(P=0.012,P=0.025)。结论:FLNa基因具有抑制人结肠腺癌SW480细胞在裸鼠体内的生长作用,其作用机制可能与下调MMP-9的表达有关。     

The leukocyte protein L-plastin induces proliferation, invasion and loss of E-cadherin expression in colon cancer cells

L-plastin, a gene that codes for an actin-bundling protein, is upregulated in the metastatic colon cancer cell line SW620, when compared to its premetastatic counterpart SW480. The aim of our study was to characterise the effect of L-plastin overexpression on SW480 cells in the context of the acquisition of a metastatic phenotype. SW480 cell lines overexpressing L-plastin were established (SW480-LPL). Analysis of these cell lines revealed significantly higher rates of proliferation and invasion than the control cell line (SW480-Ctrl). In addition, the expression of E-cadherin was lost from SW480-LPL cells. Treatment of SW480-LPL cells with cytochalasin B, an inhibitor of endocytosis, attenuated the loss of E-cadherin expression in these cells. The association of L-plastin overexpression with an increased rate of proliferation and invasion, and loss of E-cadherin expression in the SW480 colon cancer cell line indicates that L-plastin plays an important mechanistic role in colorectal cancer metastasis (supplementary material for this article can be found on the International Journal of Cancer website at http://www.interscience.wiley.com/jpages/0020-7136/suppmat/index.html).