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Objective Tiam1, a member of guanine nucleotide exchange factors, plays an important role in tumor invasion and metastasis.This study aimed to construct Tiam1 truncated recombinant plasmids and induce the expression of GST-tagged human Tiam1 fusion proteins in Escherichia coli (E.coli), followed by purification and identification of the GST-Tiam1 fusion protein.Methods The cDNA fragments of Tiam1 C685, C751 and C1199 were amplified by PCR and cloned into the pGEX-4T-1 vector.After verified by DNA sequencing, the recombinant plasmid was transformed into E.coli BL21 (DE3) competent cells, and the expression of the fusion protein was induced by IPTG.The GST-tagged human Tiam1 fusion proteins were purified with glutathione-agarose resin and indentified by Western blot.Results Three Tiam1 truncated recombinant plasmids were constructed successfully.The recombinant fusion proteins GST-Tiam1 C685, GST-Tiam1 C751, and GST-Tiam1 C1199 were expressed mainly in the form of soluble proteins in the cell lysate supernatant with expected relative molecular weight of 100, 108, and 157 kD.Conclusion The recombinant plasmids expressing the bioactive fusion proteins were constructed successfully, which has prepared the ground for the subsequent studies of the Tiam1 protein.
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Purpose To investigate the clinicopathological significance of Tiam1 overexpression in ovarian carcinoma.Methods Tiam1 protein was detected in 98 serous carcinomas,64 ovarian mucinous cystadenocarcinomas and 27 benign ovarian tumors using immunohistochemical of SP two-step staining.Correlations between Tiam1 overexpression and clinicopathological features of ovarian carcinoma were evaluated.Overall survival and disease-free survival rates of ovarian carcinoma patients were calculated by Kaplan-Meier method.Results Tiam1 protein showed a cytoplasmic and nuclear staining pattern in ovarian carcinoma.The positive rate and strongly positive rate of Tiam1 protein were 84.7％ (83/98)and 70.4％ (69/98)in serous carcinoma,the positive rate and strongly positive rate of Tiam1 protein were 87.5％ (56/64) and 73.4％ (47/64) in ovarian mucinous cystadenocarcinoma,both of which were significantly higher than those in benign ovarian tumor (both P ＜ 0.01).High-level expression of Tiam1 was positively correlated with the histological grade,FIGO stage,and metastasis in ovarian carcinoma (both P ＜ 0.05),while independent of age and menopausal status (both P ＞ 0.05).Kaplan-Meier survival analysis showed that overall survival and disease-free survival of ovarian cancer patients with high expression of Tiam1 protein were significantly lower than those of patients with lower expression of Tiam1 protein (P ＜ 0.05),overall survival in metastatic ovarian cancer patients with high expression of Tiam1 protein was significantly lower than those in patients with low Tiam1 protein expression (P ＜ 0.05).Conclusion Tiam1 expression is significantly high in ovarian carcinoma.Tiam1 protein is closely associated with ovarian carcinoma progression,which may be a new potential prognostic biomarker and a therapeutic target for ovarian carcinoma.
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With continuous development of molecular diagnosis,"precision treatment"has entered the therapeutic category for ma-lignant tumors, and targeted gene therapy has been an intense research topic in recent years. Neuroblastoma is the most common ex-tracranial solid tumor that develops during childhood. Studies show that many genes, such as TIAM1, are involved in the development and progression of neuroblastoma. TIAM1 mainly combines with RAC1 to activate downstream factors that mediate differentiation via the TrkA/TIAM1/RAC1 signaling pathway, which is involved in the regulation of neurite. Therefore, further studies and experiments may reveal the specific mechanisms and provide a new direction for the future treatment and development of neuroblastoma.
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Objective To investigate the relationship between the changes of expression of T lymphoma invasion and metastasis inducing factor 1 (Tiam1) mRNA and lymph node micrometastasis and prognosis in patients with node-negative colorectal cancer(CRC).Methods From June 2008 to August 2010,a total of 63 patients with lymph node-negative CRC,20 patients with lymph node-positive CRC and 25 patients with colorectal benign lesions were enrolled.The relative quantity expression (RQ) of Tiam1 mRNA in CRC cancer tissues and colorectal benign lesions were detected by real-time quantitative polymerase chain reaction (PCR).The lymph node micrometastasis was determined by cytokeratin 20 detection in lymph nodes tissues.The five-year postoperative prognosis was evaluated by followed-up in the patients with lymph node-negative CRC.The correlation between the expression of Tiam1 mRNA and lymph node micrometastasis and prognosis was analyzed.The t-test,univariate analysis,multivariate analysis and Log-rank test were performed for statistical analysis.Results Lymphnode micrometastasis occurred in 33 from 63 patients with lymph node-negative CRC.The RQ of Tiam1 mRNA in the patients with lymph node-positive CRC was 9.84±-2.36,which was higher than that of patients with lymph node-negative CRC (5.15±3.58),and the difference was statistically significant (t=5.479,P＜0.01).The RQ of Tiam1 mRNA of these two groups were both higher than that of colorectal benign lesion (0.30± 0.21),and the difference was statistically significant(t=20.169,6.745;both P＜0.01).The RQ of Tiam1 mRNA of positive lymph node micrometastasis group was 6.30±1.95,which was higher than that of negative lymph node micrometastasis group (3.88 ± 1.63),and the difference was statistically significant (t=5.330,P＜0.01).In the patients with lymph node-negative CRC,the lower degree of tumor difference,the deeper invasion,the shorter survival time after surgery and postoperative recurrence and metastasis were related with the higher RQ of Tiam1 mRNA in tumor tissues (t=2.536,3.112,3.213,2.676;all P＜0.05).Univariate analysis revealed that the expression of Tiam1 mRNA in cancer tissues was correlated with lymph node micrometastases(x2=11.878,P =0.001).Multivariate analysis showed that it was an independent risk factors of lymph node micrometastases in CRC (relative risk:9.782).The five-year postoperative cumulative survival rate of high Tiaml mRNA expression group was 75.8 ％,which was lower than that of low expression group (97.1 ％),and the difference was statistically significant (x2 =4.575,P＜0.05).Conclusions Tiam1 may participate in the regulation of lymph node metastasis in CRC.Elevated expression of Tiam1 promotes lymph node metastasis and is closely relatived with poor prognosis of node-negative CRC,and which can be considered as an indicator of prognosis.
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Purpose To screen and identify T cell lymphoma invasion and metastasis 1 (Tiam1) related genes and expression profile during distant metastasis of colorectal cancer. Methods Colorectal cancer( CRC) in a mice model was established by intraperitoneal injection of dimethylhydrazine ( DMH) . Five panels of fresh primary tumor tissues from Tiam1 transgenic mice were collected and com-pared with that from wild type mice. Differentially expressed genes were detected by Affymetrix human genome-wide expression profile chip and verified by real-time quantitative PCR ( qRT-PCR) analysis. Results The genechip result showed that 794 genes were differ-entially expressed by at least 3 folds in Tiam1 transgenic mice with distant metastasis of colorectal cancer tissue, including 400 up-regu-lated and 394 down-regulated ones. Bioinformatics analysis and gene co-expression network building identified 3 genes (PIK3R5, FIGF and RPS6KA6) with specific expression in colorectal cancer tissue with distant metastasis, and this result was confirmed by qRT-PCR. Conclusion A specific Tiam1 gene expression profile related to colorectal cancer distant metastasis has been established through screening and identifying the genes related to Tiam1 by gene chip technique. These findings provide a basis for exploring the molecular markers of colorectal cancer with distant metastasis.
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Objective To explore the effect of Tiam1 overexpression on the biological behaviors of human colorectal cancer cells ( CRC ) . Methods The human CRC lines under the established stable overexpression of Tiam1 were studied. Cell morphology was detected before and after transfection by commassie blue staining and scanning electron microscope. The proliferation in vitro of CRC was tested by cell cycle, MTT and plate colony formation assay, the migration and invasion ability of CRC was tested by Transwell assay. The proliferation ability in vivo was studied by induced subcutaneous tumors of nude mice. Results Compared with HT29/mock cells, HT29/Tiam1 cells formed as spindle, the pseudopodia increased and elongated. The proportion in S phase of HT29/Tiam1 was higher (t=19.546, P=0.000), the proliferation ability enhanced (F=177.125, P=0.000), colonies formation ratio increased ( t = 3 . 222 , P = 0 . 032 ) . The number of HT29/Tiam1 cells acrossing the microporous membrane (t = 4.832, P=0.001)and Matrigel(t=3.779, P = 0.005)all raised. On the fifteenth day, the growth deference between the HT29/Tiam1cells and HT29/mock cells in nude mice in vivo occurred. Till the thirtieth day, the size of the tumors in HT29/Tiam1 cell group were 2.3 times as large as that in HT29/mock cell group (F=53.040, P=0.002). Conclusions Tiam1 stable overexpression can promote the proliferation, migration and invasion ability of CRC which indicates its important role in carcinogenesis and evolution of colorectal cancer. Tiam1 may represent a new therapeutic target for colorectal cancer.
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Objective: Invasion and metastasis are the main causes of carcinoma mortality; hence, the timely blocking of the invasion and metastasis of carcinoma has become a research hotspot. The present study aims to investigate the expression levels of Tiam-1 mRNA and HPA-1 mRNA and their correlation with the invasion and metastasis of hepatocellular carcinoma. Methods: From May 2009 to Jan 2012, 65 hepatocellular carcinoma patients admitted consecutively in our hospital for surgical treatment were included in this study. Real-time fluorescent quantitative reverse transcriptase polymerase chain reaction (RT-PCR) was used to investigate the expression levels of Tiam-1 mRNA and HPA-1 mRNA in hepatocellular carcinoma, paired adjacent hepatocellular carcinoma (2 cm from the carcinoma), and surgical marginal normal hepato mucosa tissues (5 cm from the carcinoma). RT-PCR was also used to analyze their correlation with the clinicopathological characteristics of hepatocellular carcinoma. Results: The expression level of HPA-1 mRNA was significantly higher in hepatocellular carcinoma (43.83±11.62) than in paired adjacent hepatocellular carcinoma (14.82±8.16) and normal hepato mucosa tissues (6.02±5.36) (P<0.001). The expression level of HPA-1 mRNA was higher in paired adjacent hepatocellular carcinoma than in normal hepato mucosa tissues (P<0.05). The expression of Tiam-1 mRNA was higher in hepatocellular carcinoma (35.28±11.81) than in paired adjacent hepatocellular carcinoma (12.94±6.25) and normal hepato mucosa tissues (4.17±3.49) (P<0.05). The expression level of Tiam-1 mRNA was higher in paired adjacent hepatocellular carcinoma than in normal hepato mucosa tissues (P<0.05). The expression levels of Tiam-1 mRNA and HPA-1 mRNA were closely associated with the degree of differentiation, depth of infiltration, lymph node metastasis, vessel metastasis, and TNM (Tumor, Node, Metastasis)staging of gastric carcinoma (P<0.05). Spearman rank correlation analysis demonstrated a significant correlation between Tiam-1 and HPA-1 (OR=0.523, P<0.05). Conclusion: The expression levels of Tiam-1 mRNA and HPA-1 mRNA were high in hepatocellular carcinoma. Meanwhile, the increased ex-pression levels of Tiam-1 and HPA-1 can promote the invasion and metastasis of hepatocellular carcinoma. Moreover, the determination of Tiam-1 and HPA-1 may be valuable for the treatment and prognosis of hepatocellular carcinoma.
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Objective To study the relation between Tiam1 gene(T lymphoma invasion/metastasis 1)and carcinomas of larynx metastasized to lymph node.Method Using reverse transcription polymerase chain (RT-PCR) mRNA overexpression of Tiam1 gene in 30 cases of carcinoma of larynx tissue,12 lymph nodes and 10 cases of normal larynx tissue was studied.Result The frequency of TIAM1 overexpression was 75% (6/8) in primary carcinomas of larynx with metastasis but only 18.7%(4/22) in those without metastasis(P=0.0072).Overexpression of TIAM1 in metastasized lymph nodes was observed in 100% (8/8) of lymph nodes with metastasis but in only 25%(1/4) of the lymph nodes without metastasis of carcinoma(P=0.0182).The frequency of TIAM1 overexpression was 33.3% (10/30) in primary carcinomas of larynx.Conclusion Our data suggest that the overexpression of the TIAM1 gene correlates with lymph node metastasis of carcinomas of larynx.
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As a guanine nucleotide dissociation stimulator (GDS) of Rho-like GTPases, T lymphoma invasion and metastasis inducing factor 1(Tiam 1)possesses such biological functions as regulating reconstruction of cytoskeletal structure and migratory potential of cells. Studies had shown that over-expression of Tiam 1 was able to induce invasion and metastasis of tumor cells, the molecular biological basis of which includes the interactions between Tiam 1 and trans-membrane system of cytoskeleton, adhesion molecules and extra-cellular matrix, the effect on proliferation and apoptosis of tumor cells exposed by Tiam 1, as well as the activities of Tiam 1 regulated by other invasion and meatstasis associated factors. Simultaneously, these experimental results suggested that much more work would be needed in Tiam 1 on the diversity of its activity regulation ,the specificity of its biological effect, the realationship among Tiam 1 with other invasion and metastasis associated factors, as well as their mechanisms of signal transduction.
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Objective To investigate the relationship between the expression of T lymphoma invasion and (metastasis) inducing factor 1 (Tiam1), ras-related C3 botulinum toxin substrate 1 (Rac1) with the invasion and metastasis of gastric cancer. Methods The expressions of Tiam1 and Rac1 proteins in 60 cases of (gastric) cancer and paracarcioma gastric mucosa tissues were detected using Strept Avidin-Biotin Complex(SABC) immunohistochemical method,and the correlation between the expression of Tiam1, Rac1 and (clinicopathological) parameters of gastric cancer were analysed. Results (1)There was negative staining of Tiam1 protein in paracarcioma gastric mucosa tissues, while positive staining was detected in gastric cancer (tissues)(78.33%)(P 0.05). (4) The expression of Rac1 in patients with positive staining of Tiam1 was significantlyhigher than that in patients with negative staining(P