The expression of cortactin in colorectal cancer and its relationship with clinicopathological and prognostic parameters / 中华肿瘤杂志

Objective: To investigate the expression of cortactin in colorectal cancer and its correlation with clinicopathological parameters and prognosis. Methods: The expressions of cortactin in normal colorectal mucosal tissue and colorectal cancer tissue in paraffin-embedded tissue microarray from 319 patients who were diagnosed as colorectal cancer and treated in Cancer Hospital of Chinese Academy of Medical Sciences from 2006 to 2009 was detected by immunohistochemistry. Kaplan-Meier method and Log rank test were used for survival analysis, and Cox proportional risk regression model was used for multivariate analysis. Results: The positive expression rates of cortactin in colorectal cancer tissue and normal colorectal mucosal tissue were 61.1% (195/319) and 5.6% (18/319, P<0.001), respectively. T-stage, N-stage, American Joint Committee on Cancer (AJCC) stage, degree of tumor differentiation, neural invasion and preoperative carcinoembryonic antigen (CEA) levels were associated with the expression of cortactin (P<0.05). The positive expression of cortactin was associated with poorer disease-free survival (P=0.036) and overall survival (P=0.043), and the effect was more significant in patients with stage Ⅱ to Ⅲ. For patients with stage Ⅱ-Ⅲ colorectal cancer, postoperative adjuvant therapy was associated with disease-free survival (P=0.007) and overall survival (P=0.015). The vascular tumor embolus, pathological type, preoperative CEA level and cortactin expression were independent influencing factors for disease-free survival (P<0.05). The age, AJCC stage, preoperative CEA level and cortactin expression were independent influencing factors for overall survival (P<0.05). Preoperative CEA level and cortactin expression were independent influencing factors for disease-free survival and overall survival (P<0.05). Conclusion: Cortactin is expressed in colorectal cancer and in stage Ⅱ-Ⅲ patients, it is a potential predictor of colorectal cancer prognosis.

Mechanism study of the protective effects of selective cyclooxygenase-2 enzyme inhibitors on the liver of rats with type 2 diabetes mellitus combined with nonalcoholic steatohepatitis via Rho/ROCK pathway / 中华肝脏病杂志

Objective: To investigate whether the selective cyclooxygenase-2 enzyme inhibitors celecoxib has protective effect on the liver of rats with type 2 diabetes mellitus (T2DM) combined with nonalcoholic steatohepatitis (NASH) via inhibiting the expression of Rho/ROCK pathway. Methods: Forty male SD rats were randomly divided into four groups: type 2 diabetes mellitus combined with nonalcoholic steatohepatitis (T2DM-NASH) group, T2DM-NASH + celecoxib group, control group, and control+celecoxib group. The T2DM-NASH and T2DM-NASH + celecoxib groups were fed with high-sugar and fat diet, and the control group and control + celecoxib group were fed with basal diet (25 kJ/kg). Four weeks later, streptozotocin (STZ, 30 mg/kg) was intraperitoneally injected into the NASH group and T2DM-NASH + celecoxib group to induce T2DM model, and the control group and control + celecoxib group were intraperitoneally injected with isovolumic citric acid-sodium citrate buffer. Four weeks after STZ injection, the T2DM-NASH + celecoxib group and the control + celecoxib group were gavaged with celecoxib (10 mg·kg·d) dissolved in normal saline for 4 weeks, and the remaining two groups of rats were gavaged with isovolumic normal saline for 4 weeks. Animals were sacrificed at the end of the 12- weeks, and the liver tissue was collected. Liver pathological changes were observed by HE staining. The expressions of RhoA, RhoA, ROCK1 and ROCK2 proteins in liver were detected by immunohistochemistry and western blot. The expressional condition of RhoA, ROCK1 and ROCK2 mRNA in liver were detected by real-time quantitative PCR. The differences were compared between protein and mRNA expression among the groups by analysis of variance and t-test. Results: Compared with the control group and the control + celecoxib group, the liver tissue of the T2DM-NASH group and the T2DM-NASH + celecoxib group had severe steatosis, and there was partial inflammatory cell infiltration under the light microscope. The expression levels of RhoA, ROCK1 and ROCK2 protein and mRNA were significantly increased (P < 0.05) in each liver tissue, while liver steatosis was reduced to certain extent in T2DM-NASH + celecoxib group than T2DM-NASH group, and the expression levels of RhoA, ROCK1 and ROCK2 protein and mRNA were decreased in each liver tissue of T2DM-NASH group (P < 0.05). Conclusion: The selective cyclooxygenase-2 enzyme inhibitors celecoxib has a protective effect on the liver of rats with T2DM-NASH, and its effect may be achieved by inhibiting the expression of Rho/ROCK pathway.

Effects of adenovirus-mediated shRNA down-regulates PTEN expression on fibril-binding proteins vinculin, filamin A and cortactin in activated hepatic stellate cells / 中华肝脏病杂志

Objective: To investigate the effect of adenovirus-mediated shRNA down-regulating phosphatase and tensin homolog deleted on chromosome 10 (PTEN) expression on vinculin, filamin A, and cortactin in activated hepatic stellate cells (HSCs). Methods: Activated rats hepatic stellate cell line (HSC-T6) was cultured in vitro. Recombinant adenovirus Ad-shRNA/PTEN carrying PTEN targeted RNA interference sequence [short hairpin RNA (shRNA)] and empty control virus Ad-GFP were transfected into HSCs. The PTEN mRNA and protein expression of HSCs in each group were detected by real-time fluorescence quantitative PCR and Western blot. The expressional change of vinculin, filamin A and cortactin in HSCs of each group were detected by confocal laser scanning immunofluorescence microscope. Image-pro plus 6.0 software was used for image analysis and processing. The integrated optical density (IOD) of the fluorescence protein expression was measured. The experiment was divided into three groups: control group (DMEM instead of adenovirus solution in the adenovirus transfection step), Ad-GFP group (transfected with empty virus Ad-GFP only expressing green fluorescent protein), and Ad-shRNA/PTEN group (recombinant adenovirus Ad-shRNA/PTEN carrying shRNA targeting PTEN and expressing green fluorescent protein). One-way analysis of variance was used for comparison of mean value among the three groups, and LSD-test was used for comparison between the groups. Results: shRNA targeted PTEN was successfully transfected and the expression of PTEN mRNA and protein in HSC (P < 0.05) was significantly down-regulated. HSCs vinculin was mainly expressed in the cytoplasm. HSCs vinculin fluorescence IOD in the Ad-shRNA/PTEN group (19 758.83 ± 1 520.60) was higher than control (7 737.16 ± 279.93) and Ad-GFP group (7 725.50 ± 373.03) (P < 0.05), but there was no statistically significant difference between control group and Ad-GFP group (P > 0.05). There was no statistically significant difference in the fluorescence IOD of Filamin A among the three groups (P > 0.05), but the subcellular distribution of Filamin A among the three groups were changed. Filamin A in the Ad-shrNA /PTEN HSC group was mainly distributed in the cytoplasm. Filamin A HSC was mainly located in the nucleus.The filamin A HSC in the control group and Ad-GFP group was mainly located in the nucleus. The nucleocytoplasmic ratio of Filamin A in the AD-shrNA /PTEN group (0.60 ± 0.15) was significantly lower than control group (1.20 ± 0.15) and Ad-GFP group (1.08 ± 0.23), P < 0.05. but there was no statistically significant difference in filamin A nucleocytoplasmic ratio of HSC between the control group and the Ad-GFP group (P > 0.05). Cortactin HSCs in the three groups was mainly distributed in the cytoplasm. The cortactin fluorescence IOD of HSCs in the Ad-shRNA/PTEN group was significantly higher than control group (22 959.94 ± 1 710.42) and the Ad-GFP group (22 547.11 ± 1 588.72 ) (P < 0.05), while there was no statistically significant difference in the IOD of cortactin fluorescence in HSCs between the control group and the Ad-GFP group (P > 0.05). Conclusion: The down-regulation of PTEN expression raises the expression of microfilament-binding protein vinculin and cortactin, and changes the subcellular distribution of another microfilament binding protein filamin A, that is, translocation from nucleus to the cytoplasm in activated HSC in vitro.

Expressions of vascular endothelial growth factor C and cortactin in esophageal squamous cell carcinoma and their clinical significance / 医学研究生学报

Objective Vascular endothelial growth factor C (VEGFC) and cortactin (CTTN) have been found to be closely related to the growth of esophageal squamous cell carcinoma (ESCC), but their specific relationship has not been clearly defined up to the present time. This study aimed to investigate the effects of VEGFC and CTTN on the proliferation and apoptosis of ESCC cells. Methods Human ESCC TE1 cells were treated with normal culture medium (the blank control group), MATE transfection reagent ( the MATE group), negative control RNA and MATE reagent (the negative control group), positive control RNA and MATE reagent (the positive control group), VEGFC siRNA and MATE transfection reagent (the VEGFC siRNA group), and CTTN siRNA and MATE transfection reagent (the CTTN siRNA group). The proliferation of the ESCC TE1 cells in different groups was detected by CCK-8 assay and their apoptosis determined by flow cytometry. Results Compared with the blank control group, the ESCC cells of the VEGFC siRNA and CTTN siRNA groups showed significantly decreased expressions of VEGFC mRNA (1.00 ± 0.00 vs 0.13 ± 0.01, P < 0.05) and CTTN mRNA (1.00 ± 0.00 vs 0.29 ± 0.02, P < 0.05). The proliferation rate of the ESCC cells was remarkably lower in the VEGFC siRNA and CTTN siRNA than in the other groups (P < 0.05), and even lower in the VEGFC siRNA than in the CTTN siRNA group ([31.26 ± 5.25]% vs [46.99 ± 4.82]%, P < 0.05), but their apoptosis rate was markedly higher in the former than in the latter group ([48.41 ± 5.37]% vs [36.78 ± 4.29]%, P < 0.05). Conclusion Interfering with the expressions of VEGFC and CTTN can inhibit the proliferation and promote the apoptosis of ESCC cells, and VEGFC has an even better effect than CTTN.

Effect of Zusanli Point Electroacupuncture on Smooth Muscle Contraction Function and Cortactin Expression in Rats with Irritable Bowel Syndrome / 广州中医药大学学报

Objective To investigate the effect of electroacupuncture at Zusanli (ST36) point on the expression of cortical actin-binding protein (cortactin) in smooth muscle cells of rats with irritable bowel syndrome(IBS). Methods Eighty female Sprague-Dawley(SD) rats aged 3 months were randomly divided into 4 groups: normal group, model group, acupoint group and sham-acupoint group, 20 rats in each group. The diarrhea-predominant IBS (IBS-D) rat model was established by high-lactose feeding combined with restraint stress method. Acupoint group was given electroacupuncture at Zusanli point, and sham-acupoint group was given electroacupuncture at external region of 5 mm beside Zusanli point. After treatment, time for the discharge of active carbon through all of the gastrointestinal duct was measured, the contraction of rat proximal colon smooth muscle strips was tested, the expression level of cortactin and the ratio of G/F-actin in the proximal colon were detected by Western blotting method. Results Compared with the normal group, time for the discharge of active carbon through all of the gastrointestinal duct was shortened, and the systolic basic tension of colonic smooth muscle, the expression level of cortactin, and the ratio of G/F-actin in the model group were increased (P<0.05). After electroacupuncture treatment, the time for the discharge of active carbon through all of the gastrointestinal duct in acupoint group was prolonged, and the systolic basic tension of the intestinal smooth muscle, the expression level of cortactin, and ratio of G/F-actin were decreased (P < 0.05 compared with the model group and non-acupoint group). Conclusion Electroacupuncture at Zusanli point can effectively improve the gastrointestinal function of IBS rats and has regulatory effect on the contraction of colonic smooth muscle.

Cortactin antibody in myasthenia gravis and its clinical significance / 中华神经科杂志

Objective To investigate the clinical significance of cortactin antibody in myasthenia gravis (MG).Methods Cortactin antibody in the serum of 100 MG patients, 40 normal controls and 40 other neuroimmune diseases patients was examined by Western blotting and ELISA using purified recombinant human protein cortactin as antigen .Acetylcholine receptor antibody ( AchR-ab ) and muscle specific kinase antibody (MuSK-ab) were parallely measured by ELISA.Results Antibodies to cortactin were found in nine (9％) serum samples of 100 MG patients.Four of the nine cortactin antibody positive sera were also positive for AChR-ab.The rest five MG patients only had antibodies against cortactin ( no detectable AChR-ab or MuSK-ab).None of the control subjects (including 40 normal controls and 40 other neuroimmune diseases patients ) had cortactin antibodies.Most (7/9) of the cortactin antibody positive MG patients presented with early-onset subgroup.Patients only with cortactin antibodies did not appear to have thymoma.Patients with MG who had both AChR and cortactin antibodies showed maximum involvement of muscles and severe Osserman's classification ( three cases of type ⅡB and one case of type Ⅳ) . Conclusion Cortactin antibody may be a new antibody for MG , which can provide clues for further exploring the potential pathogenic mechanisms of the disease .

Inhibition of invasion and metastasis by imrecoxib and its mechanisms in lung adenocarcinoma A549 cell xenograft in nude mice / 中国癌症杂志

Background and purpose:Cyclooxygenase-2 (COX-2) participates in angiogenesis and lymph node metastasis of lung cancer. COX-2 inhibitors could inhibit invasion and metastasis of lung cancer. This study aimed to investigate the inhibition of invasion and metastasis by COX-2 inhibitor imrecoxib in xenograft tumor of lung adenocar-cinoma A549 cell in nude mice and to explore its possible mechanisms, in addition, to observe the efficacy of imrecoxib combined with lobaplatin.Methods:Thirty male BALB/c nude mice were injected subcutaneously with A549 cells into the right axillary region to establish xenograft models. Twenty-nine successfully modeled mice were randomly divided into four groups: control group (n=7), imrecoxib group (n=8), lobaplatin group (n=7), imrecoxib combined with lobaplatin group (n=7). The control group was treated with the same amount of sterile distilled water and injected with the same amount of 0.9% sodium chloride solution via caudal vein. The treatment group was treated with imrecoxib tablets 40 mg/kg per day through gavage and injected with lobaplatin 7.5 mg/kg per week via caudal vein respectively. The diet, physical activity and other normal conditions of nude mice were observed everyday. After 6 weeks, 29 mice were sacrificed and transplanted tumor tissues were cut off. The expression of PTEN, cortactin protein and mRNA were detected by immunohistochemistry and real-time PCR. The data were analyzed with one-way anova and non-parametric test.Results:In the last week, the diet and physical activity of all nude mice were less than before, and they became thinner, which were more obvious among the mice in lobaplatin group and imrecoxib combined with lobaplatin group. Compared with the control group, the expression of PTEN protein and mRNA were significantly increased in imrecoxib group and imrecoxib combined with lobaplatin group (P<0.001, respectively). Compared with the control group, the expression of cortactin protein and mRNA were significantly decreased in imrecoxib group and imrecoxib combined with lobaplatin group (P<0.001, respectively). PTEN and cortactin protein, PTEN and cortactin mRNA had significantly negative correlation (r=-0.660, -0.983,P<0.001, respectively). Conclusion:Imrecoxib can inhibit non-small cell lung cancer invasion and metastasis which may be involved in upregu-lating PTEN protein and reducing cortactin protein. Imrecoxib could enhance the effect of lobaplatin chemotherapy.

Chromokinesin KIF4A as a tumor suppressor by inhibiting cell invasion / 国际生物医学工程杂志

Objective To explore the role of chromokinesin KIF4A in gastric cancer cell invasion using gastric cancer cells SGC-7901 and chromokinesin KIF4A deficient gastric cancer cells (SGC-shKIF4A).Methods Expression levels of KIF4A in controlling gastric cancer cells (SGC-shNC) and SGC-shKIF4A cells were determined by Western Blot.Invasion of gastric cancer cells were assessed using Transwell invasion assay and the number of cells passing through the matrigel was counted.Changing numbers of cortactin in SGC-7901,SGC-shNC,and SGC-shKIF4A cells were analyzed by immunofluorescence staining.Results Compared to the SGC-shNC cells,invasion ability of SGC-shKIF4A cells was increased.Compared to other cells,the numbers of cortactin in SGC-shKlF4A cells was also increased suggesting invadopodia in these cells was increased(P＜0.01).Conclusions Chromokinesin KIF4A acts as a tumor suppressor by inhibiting gastric cancer cells invasion and the results provides strong evidences for KIF4A serving as one of potent targets for gastric cancer prognostics and treatment in clinic.

Cortactin protein expression and its relationship with cell division and clinical pathology in colorectal cancer / 中华普通外科杂志

Objective To investigate cortactin expression in malignant colorectal tissues and corresponding adjacent non-cancerous colon tissues,precancerous lesions (adenomatous polyps) and the relationship between the expression of cortactin and cell division in colorectal cancer cells.Methods The expression of cortactin was detected by immunohistochemistry in colorectal cancer,colorectal adenomatous polyp (precancerous lesions) and colorectal tissues adjacent to adenocarcinomas (normal tissues).Kaplan-Meier method was employed to compare the survival between the groups.Cortactin expression and cell division were detected by Western blot and immunofluorescence in SW-620 colon cancer cells treated with cortactin siRNA.Results The positive expression rate of cortactin was significantly higher in colorectal cancer tissues than in adenomatous polyp tissues and pericarcinomatous normal tissues.Overexpression of cortactin in colorectal cancer tissues was correlated with poor differentiation (P ＜ 0.01),lymph node metastasis (P =0.006),and TNM stage (P =0.022).The 5 year survival rate of the group of negative/weak positive expression of eortactin was significantly higher than the group of strong positive expression of cortactin.CTTN gene amplification in colorectal cancer tissues was obvious.Cortactin siRNA induction caused a lower cortactin protein expression in colorectal cancer cells.Conclusions It is suggested that the excessive expression of cortactin contributes to the growth of cancer cells in colorectal cancer.

Expression of Cortactin and Focal Adhesion Kinase in Colorectal Adenocarcinoma: Correlation with Clinicopathologic Parameters and Their Prognostic Implication

BACKGROUND: Cortactin and focal adhesion kinase (FAK) are two important components among actin cross-linking proteins that play a central role in cell migration. METHODS: The aims of this study were to evaluate the expression of cortactin and FAK in normal colorectal mucosa and colorectal adenocarcinoma (CRC) using tissue microarray of 2 mm cores to correlate their expression with other clinicopathological factors and, investigate their prognostic significance. RESULTS: Twenty (9%) and 24 cases (11%) of normal colorectal mucosa were immunoreactive for cortactin and FAK. In addition, 184 (84%) and 133 cases (61%) of CRCs were immunoreactive for cortactin and FAK, respectively. Cortactin expression was associated with histologic differentiation and FAK expression. Cortactin, but not FAK expression was also correlated with poor overall and relapse-free survival and served well as an independent prognostic factor for poor survival. CONCLUSIONS: Cortactin expression, in association with FAK expression, may plays an important role in tumor progression. Furthermore, it may also be a satisfactory biomarker to predict tumor progression and survival in CRC patients.

Expressions of E-cadherin, Cortactin and MMP-9 in Pseudoepitheliomatous Hyperplasia and Squamous Cell Carcinoma of the Head and Neck: Their Relationships with Clinicopathologic Factors and Prognostic Implication

BACKGROUND: E-cadherin, cortactin, and matrix metalloproteinase (MMP)-9 have roles in tumor development or progression, but their expression has not been fully investigated in pseudoepitheliomatous hyperplasia (PEH) and squamous cell carcinoma (SCC) of the head and neck. METHODS: We evaluated the immunohistochemical expression of E-cadherin, cortactin, and MMP-9 in 29 cases of PEH and 97 cases of SCC. Additionally, we evaluated their relationship with clinicopathologic factors and prognostic implications in SCC. RESULTS: Thirty-five cases of SCC showed reduced expression of E-cadherin, whereas none of the PEH did. A total of 20 cases and 11 cases of SCC were immunoreactive for cortactin and MMP-9, respectively, whereas none of the PEH did. In SCC, reduced expression of E-cadherin was correlated with cortactin expression and invasion depth. Cortactin expression was correlated with differentiation, T classification, and recurrence and/or metastasis. MMP-9 expression was correlated with invasion depth. Cortactin expression was correlated with poor overall survival and relapse-free survival and it was an independent prognostic factor. CONCLUSIONS: The reduced expression of E-cadherin and the expression of cortactin may be helpful for the differential diagnosis of PEH and SCC. Furthermore, cortactin expression in association with reduced E-cadherin expression is correlated with poor prognosis in SCC.

Significance of cortactin expression in primary liver cancer / 中华消化外科杂志

Objective To investigate the significance of cortactin in primary liver cancer. Methods Fiftythree paraffin embedded primary liver cancer specimens were collected at General Hospital of Air Force of PLA from January 2002 to May 2008. The expression of cortactin was detected by immunohistochemistry, and the relationship between cortactin expression and clinicopathologic parameters was analyzed. All data were analyzed via Wilcoxon rank sum test and Spearman rank correlation analysis. Results There was a significant difference in cortactin expression among tumor capsule integrity, TNM staging, portal vein tumor thrombus and extrahepatic metastasis (u =2. 19, 3. 584, 2. 796, P ＜ 0.05 ). There was a positive correlation between tumor invasion and cortactin expression ( r = 0. 5794, P ＜ 0.05 ). Conclusions Overexpression of cortactin may be one of the factors enhancing the invasion of primary liver cancer. The level of cortactin expression can be used in evaluating the invasive potential of primary liver cancer.

Expression of cortactin in human breast cancer cell lines with different metastasis potentials / 第二军医大学学报

Objective: To screen for human breast cancer sub-cell lines with different metastasis potentials, and to investigate the role of cortactin in proliferation and metastasis of human breast cancer cells. Methods: Human breast cancer sub-cell lines with different metastasis potentials were screened by continuous in vitro invasion assays. Ultra-microstructures of sub-cell lines were observed under transmission electron microscope. The proliferation of sub-cell lines was detected by MTT; the cell cycle was observed by flow cytometry; and the migration ability of sub-cell lines was observed by Transwell assay. The mRNA and protein expression of cortactin was examined by immunofluorescence, RT-PCR and Western blotting assay. Results: Two sub-cell lines with different potentials of metastasis were obtained through continuous in vitro invasion assay, and they showed essentially identical morphology under the transmission electron microscope. The MTT results showed that proliferation of high metastasis cells was faster than that of low metastasis cells (P<0.05). Flow cytometry demonstrated that the proportion of high metastasis cells in G 0/G1 phase was less ([52.67±3.69]% vs [64.46±2.79]%), and that in S phase was more than that of low metastasis cells ([30.53±6.19]% vs [24.63±2.04]%). The proliferation index (PI) of high metastasis cells was higher than that of low metastasis cells ([47.32±3.69]% vs [35.53±2.80]%, P<0.05). Compared with low metastasis cells, the number of high metastasis cells passing through the membrane was significantly more ([61.46± 7.08] vs [25.32±4.87] cells/field, P<0.05). The expression levels of mRNA and protein of cortactin in high metastasis cells was higher than that in low metastasis cells (P<0.05). Conclusion: Over-expression of cortactin is closely related to the proliferation and metastasis of human breast cancer cells.

The correlation between cortactin differential expression and metastasis of colorectal cancer / 中华普通外科杂志

ObjectiveTo explore the correlation between cortactin differential expression and metastasis of colorectal cancer by testing the expression of cortactin in cell sublines with different metastatic ability.MethodHuman colon cancer cell line SW1116, and sublines of fifth subcutaneous ( SW1116_5), first, fifth generation (CRCLM1 、CRCLM3 ) characteristic of different liver metastasis were implanted subcutaneously in BALB/c mouse.Tumors were transplanted into the colon, and liver metastasis was observed, Western blotting and real time-PCR was used to detect the varience of cortactin. Transwell assay was applied to evaluate four cell lines migration and invasion ability. Immunohistochemistry was used to analyze the relationship of cortactin and clinicopathological characterizing in 86 cases of colorectal cancer.ResultCRCLM3( 100％ )has higher ability of metastasis than that of CRCLM1 ( 89％ ), SW1116(40％ )and SW1116_5 (44％) respectively. Expression of cortactin in the SW1116, SW1116_5, CRCLM1,CRCLM3 was gradually increased. There were significant differences between four cell lines by comparison between each others ( P ＜ 0.05 ). Immunohistochemical expression of cortactin in 86 cases was positive in 57 cases(66％ ), and negative in 29 cases (34％).ConclusionsDifferent expression of cortactin with colorectal cancer metastasis and clinical stage was positively correlated. Cortactin is a potential indicator for clinical staging and tumor metastasis of colorectal cancer.

Attenuated endocytosis by down-regulation of cortactin protein in cancer cells / 基础医学与临床

Objective To study cortactin function in cancer cell endocytosis.Methods We applied cortactin siRNA interference to MDA-Mb-231,a human breast adenocarcinoma cell line in which cortactin was over-expressed,and introduced anti-cortactin immunoreagents into the cells with BioPorter system to interfere with cortactin function in vivo.Capture-ELISA assay was used to measure transferrin uptake.We used immunoblot assay to assess the effect of cortactin knock-down and immunoflurescence microscopy to examine the effect of cortactin down-regulation on transferrin uptake.Results Interference of cortactin function in cells resulted in impairment of transferrin endocytosis.Transferrin fluorescent intensity in cytoplasm in cortactin siRNA treated-cells was significantly reduced in comparison to that of mock-treated cells.Less than 50% of cells subjected to cortactin siRNA treatment had normal transferrin uptake.Endocytosis in MDA-Mb-231 cells introduced with cortactin antibodies was impaired as well,showing a 30%～ 60% reduction in transferrin uptake.Conclusion Crtactin,an actin-binding protein,plays an essential role in cell endocytosis.

Effects of silencing cortactin expression by siRNA on the proliferation and invasion in Hep-2 cells / 中国癌症杂志

Background and purpose:The cortical actin-binding protein,cortactin,participates in several functions in the cytoskeleton system,cellular signal transduction and cell adhesion.There is also increasing evidence that it regulates tumor invasion and metastasis.However,the role played by cortactin in laryngeal carcinoma has not been clearly delineated.The purpose of this experiment was to investigate the effect of silencing cortactin expression on the proliferation and invasion in the human laryngeal carcinoma cell line Hep-2.Methods:A plasmid from a siRNA targeting cortactin was constructed and transfected into a Hep-2 cell line.The siRNA interference efficiency of cortactin was determined by Western blot.The proliferation was measured by MTT assay and plate colony formation. The Transwell test was used to detect the migration and invasion ability of the Hep-2 cells.Empty plasmid-transfected Hep-2 and normal Hep-2 were used as control groups.Results:Compared to Hep-2 cells,the cortactin expression of pSilencer3.1-cortactin-siRNA/Hep-2 was 11.22%(P