Overexpression of Rho GDP-dissociation inhibitor alpha is associated with tumor progression and poor prognosis of colorectal cancer.

The GDP dissociation inhibitors (GDIs) are pivotal regulators of Rho GTPases, which are essential for tumor progression, particularly in the area of metastasis. One member of GDIs was identified as RhoGDI (Rho GDP-dissociation inhibitor alpha, or RhoGDIalpha), but little is known about this protein in tumors. In this study, we used comparative proteomic analysis to show that RhoGDI is markedly up-regulated in metastatic colorectal cancer (CRC).

[Construction of a lentiviral vector for RNA interference of human CDH22 gene and its gene silencing effect in SW480 cells].

Objective: To construct a lentiviral expression vector for RNA interference of human CDH22 gene, and assess its gene silencing effect in colorectal cancer cells to provide a basis for investigating the role of CDH22 gene in the signaling pathway involved in human colorectal carcinoma metastasis.

Methods: Human CDH22 gene short hairpin RNA (shRNA) sequence was designed using a software available on-line. After synthesis and annealing, the double-stranded oligonucleotides (dsOligoe) were cloned into the pENTR(TM)/U6 plasmid followed by sequence analysis.

[Construction of a lentiviral vector for RNA interference of PRL-3 gene and its stable expression in SW480 cells].

Objective: To construct a lentiviral vector for RNA interference (RNAi) of PRL-3 gene and establish a human colon carcinoma cell line with PRL-3 gene knock-down.

Methods: The plasmids were constructed expressing two different short hairpin RNAs (shRNA) targeting PRL-3 gene under control by the U6 promoter by lentiviral vector. An entry clone was generated in the pENTR/U6 vector.

Silencing of ZNF217 gene influences the biological behavior of a human ovarian cancer cell line.

Zinc-finger protein 217 (ZNF217), a candidate oncogene on 20q13.2, can lead cultured human ovarian and mammary epithelial cells to immortalization, which indicates selective expression of ZNF217 affecting 20q13 amplification during critical early stages of cancer progression. In this study, we tested the hypothesis that ZNF217 is a key factor in regulating ovarian cancer proliferation and progression.

Serum proteome analysis for profiling protein markers associated with carcinogenesis and lymph node metastasis in nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC), one of the most common cancers in population with Chinese or Asian progeny, poses a serious health problem for southern China. It is unfortunate that most NPC victims have had lymph node metastasis (LNM) when first diagnosed. We believe that the 2D based serum proteome analysis can be useful in discovering new biomarkers that may aid in the diagnosis and therapy of NPC patients.

[Expression of COX-2 and VEGF-C in squamous cell carcinoma of the tongue and its correlation to lymph node metastasis].

Objective: To evaluate the coexpression of cyclooxygenase-2 (COX-2) and vascular endothelial growth factor-C (VEGF-C) in human squamous cell carcinoma of the tongue (SCCT) and assess their correlations to neoangiogenesis and lymph node metastasis of the tumor.

Methods: Tissue samples of primary SCCT and the metastatic lymph nodes were obtained from 46 patients undergoing surgical resections of SCCT for immunohistochemical detection of COX-2 and VEGF-C expressions.

Results: The over-expression rates of COX-2 and VEGF-C was 82.

[Expression of the zinc finger transcriptional factor Snail in colorectal carcinoma and its significance].

Objective: To study expression of the zinc finger transcriptional factor Snail in colorectal carcinoma and its significance.

Methods: Expressions of Snail in colorectal carcinoma SW480 and SW620 cells were assayed by immunocytochemistry and immunofluorescent cytochemistry. The paraffin-embedded specimens from 68 cases of colorectal carcinoma and its corresponding adjacent tissues, 33 cases of adenoma and 35 cases of metastatic lymph nodes were also examined for Snail expressions using immunohistochemistry.

[Comparative proteomics of the serum in patients with nasopharyngeal carcinoma: a study with two-dimensional electrophoresis and MALDI-TOF-MS].

Objective: To screen the tumor specific antigens of nasopharyngeal carcinoma (NPC) in the serum of the patients.

Methods: Two-dimensional electrophoresis (2-DE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was employed to screen the specific biomarkers of NPC in the sera from 42 healthy volunteers, 37 NPC patients with lymph node metastasis and 27 NPC patients without lymphatic metastasis.

Results: After pretreatment including albumin and immunoglobulin (IgG) depletion and desalting, the sera were subjected to 2-DE and image analysis.

[Expression of HSP27 in colorectal carcinoma and its relationship with lymphatic metastasis].

Objective: To investigate the relationship between heat shock protein 27 (HSP27) expression and lymphatic metastasis of colorectal carcinoma (CRC).

Methods: Immunohistochemistry was used to detect HSP27 expression in 68 specimens of human CRC. The expression of HSP27 mRNA and protein was also detected in two colorectal carcinoma cell lines with different lymphatic metastasis potentials by RT-PCR, Western blotting and immunohistochemistry.

Well-characterized monoclonal antibodies against cell wall antigen of Aspergillus species improve immunoassay specificity and sensitivity.

The diagnosis of invasive aspergillosis (IA) based on the detection of Aspergillus galactomannan (GM) is complicated by the presence of cross-reactive GM epitopes in patient specimens. We have developed a novel and specific Aspergillus antigen-capture enzyme-linked immunosorbent assay (ELISA) by the selection of two well-characterized monoclonal antibodies from 17 candidate antibodies. The epitopes recognized by the monoclonal antibodies were present on the cell walls of the hyphae and the conidia of Aspergillus species, which were circulating or excreted as immunodominant antigens during the acute phase of IA established in the animal models.

Protein and mRNA characterization in human colorectal carcinoma cell lines with different metastatic potentials.

Metastasis, the important characteristic of malignant tumors, is closely associated with a series of changes in the expressions of genes and proteins. In this study, we compared mRNA and protein expressions in a pair of human colorectal carcinoma cell lines named SW620 and SW480 with different metastatic potentials by suppression subtractive hybridization and 2-dimensional gel electrophoresis combined with the matrix-assisted laser desorption/ionization time-of-flight mass spectrometer. After suppression subtractive hybridization and differential screening, 24 differentially expressed gene fragments were obtained, including 9 known genes and 15 novel genes.

[Effect of Tiam1 overexpression on proliferation and metastatic potential of human colorectal cancer].

Objective: To confirm the role of Tiam1 (T lymphoma invasion and metastasis 1) gene in the proliferation and metastasis of colorectal cancer.

Methods: Proliferative and metastatic abilities of Tiam1 transfectant were investigated by subcutaneous injection of cells and surgical orthotopic transplantation (SOI) in mice.

Results: The expression of Tiam1 led to a pronounced increase in HT29/Tiam1 cell growth starting from day 7, up to 2.

[Association of the coding region single nucleotide polymorphisms in PLUNC gene with nasopharyngeal carcinoma].

Objective: To investigate whether the single nucleotide polymorphism (SNP) site G14595T of PLUNC gene coding region is significantly correlated with nasopharyngeal carcinoma (NPC).

Methods: Peripheral blood samples were collected from 239 NPC patients, 163 males and 76 females, aged 46.9, and 286 sex(-), and age(-)-matched healthy controls in Guangdong, China.

[Clinicopathological implications of positive CK7 expression in colorectal carcinoma].

Objective: To investigate the clinicopathological significance of positive CK7 expression in human colorectal carcinoma (HCC).

Methods: Immunohistochemistry was used to detect CK7 and CK20 protein expressions in 68 cases of HCC, 20 cases of canalicular adenoma (CA), 5 cases of serrated adenoma (SA) and 20 cases of hyperplastic polyps (HP).

Results: The positivity rate of CK20 expression was 89.

[Establishment of a whole-body visualization model of orthotopically implanted colorectal carcinoma and metastasis model in nude mice].

Objective: To establish an animal model visualizing orthotopic growth and metastasis of colorectal cancer.

Methods: pEGFP-N1 plasmid was transfected into human colorectal carcinoma cell line SW480 so that the resultant SW480/EGFP cells emitted fluorescence that could be detected externally by fluorescence stereo microscope. SW480/EGFP cells were inoculated subcutaneously in nude mice, and the orthotopic tumor growth was evaluated in real time.

[Microsatellite analysis and hMLH1/hMSH2 expression detection in young patients with colorectal cancer: value in screening hereditary nonpolyposis colorectal cancer].

Objective: To evaluate the frequency of microsatellite instability (MSI) and absence of hMLH1/hMSH2 expression in young patients with colorectal cancer, and investigate their role in screening hereditary nonpolyposis colorectal cancer (HNPCC).

Method: Seventy-three young patients (below 40 years old) with colorectal cancer were examined for DNA mismatch repair deficiency by microsatellite testing and immunohistochemical detection of hMLH1/hMSH2 gene products.

Results: The frequency of MSI and absence rate of hMLH1/hMSH2 expression are 56.

[Effect of Tiam-l gene silencing on human colorectal carcinoma cell line SW480 metastasis in nude mice observed by real-time whole-body fluorescence imaging].

Objective: To observe the effect of Tiam-l gene silencing on the metastasis of human colorectal carcinoma cell line SW480 in nude mice by real-time whole-body fluorescence imaging.

Methods: Enhanced green fluorescence protein (EGFP)-labeled human colorectal carcinoma cells, SW480/EGFP(+)/Tiam-1(-) and SW480/EGFP(+), were implanted into nude mice via tail vein injection or orthotopic colonal inoculation, and real-time whole-body fluorescence imaging was performed to compare the difference in tumor progression and metastasis between the two cells.

Results: Both SW480/EGFP(+) and SW480/ EGFP(+)/Tiam-1(-) cells stably expressed EGFP, and Tiam1 gene expression was reduced in SW480/EGFP(+)Tiam-1(-) to 30% of the expression level in SW480/EGFP(+) cells.

[Cloning of human PRL-3 gene and construction of its prokaryotic expression vector].

Objective: To obtain the entire coding sequence of human PRL-3 gene and construct its prokaryotic expression vector.

Methods: With total RNA extracted from the human colorectal carcinoma cell line SW480 as the template, PRL-3 gene was amplified by RT-PCR with primers designed according to the published sequence of GenBank, and the product was inserted into pGEM-T Easy vector. The recombinant plasmid pGEM-T-PRL-3 was identified by restriction endonuclease analysis and DNA sequencing.

[Preliminary study of PRL-3 gene promoter binding sites of Snail in SW480 cells].

Objective: To identify the region in PRL-3 gene promoter where the transcriptional factor Snail can bind.

Methods: PRL-3 promoter and the possible binding sites of the transcription factors were analyzed by bioinformatical methods. Chromatin immunoprecipitation and PCR were performed using the antibody specific for Snail to verify the binding of Snail to PRL-3 promoter.

Differential proteomic analysis of human colorectal carcinoma cell lines metastasis-associated proteins.

Purpose: Metastasis is a common phenomenon and the major lethal cause of colorectal carcinoma (CRC). To better comprehend the mechanism underlying CRC metastasis and to search for potential markers for predicting CRC metastasis, two CRC cell lines with different metastatic potentials, SW480 and SW620, were investigated by phenotypic analyses and proteomics technologies.

Methods: The surgical orthotopic implantation (SOI) technique was originally used to develop a reproducible colorectal cancer model in nude mice with stable tumor growth and metastasizing course.

Proteomic analysis of Tiam1-mediated metastasis in colorectal cancer.

Tiam1 (T lymphoma invasion and metastasis 1), a guanine nucleotide exchange factor that activates Rac, was recently identified as a novel colorectal cancer metastasis-related gene. To better understand the mechanism underlying Tiam1-mediated metastasis, we applied two-dimensional polyacrylamide gel electrophoresis (2-DE) and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis to identify differentially expressed proteins between Tiam1 transfected and mock transfected colorectal cancer HT29 cells. Eleven differentially expressed proteins were identified and further validated by Western blot and/or real-time PCR.

[Monitoring changes of serum protein markers in metastatic colorectal carcinoma model].

Objective: To investigate the changes of several protein markers in a metastatic colorectal carcinoma model by serum proteomic analysis.

Methods: The pEGFP-N1 plasmid with enhanced expression of green fluorescence protein (EGFP) was transfected into human colon carcinoma cell line SW480 to obtain a stable SW480-EGFP cell line, the SW480-EGFP cells were then injected subcutaneously into nude mice. The harvested tumor cells were implanted orthotopically into the colon of the nude mice.