BACKGROUNDIn general, it is very important to understand the state of T cell immune response against tumor cells in leukemia patients and it is especially critical to assess the T cell repertoire of untreated patients. As we know, few studies have dealt with the distribution of oligoclonal T cells in leukemia, so we investigated the distribution and clonality of TCR Vbeta repertoire of T cells in patients with chronic myelogenous leukemia (CML) in chronic phase.

METHODSThe complementarity determining region 3 (CDR3) of TCR Vbeta24 subfamily genes were amplified in peripheral blood mononuclear cells from 27 cases with CML using reverse transcription-polymerase chain reaction (RT-PCR). In order to observe the distribution of TCR Vbeta repertoire, the PCR products were further analyzed by genescan technique to evaluate clonality of the detectable TCR Vbeta T cells. The PCR products of the oligoclonal T cells from three cases were analyzed by direct sequencing to define the sequence of CDR3.

RESULTSThe expression pattern of TCR Vbeta repertoire in different individuals are different. Vbeta2-21 subfamilies could be detected in CML cases. The frequent usage Vbeta repertoire in CML was Vbeta1, Vbeta2 or Vbeta13. Most of the PCR products from 27 patients displayed polyclonality, while a part of the PCR products from 21 out of 27 samples displayed clonal expansion pattern. The clonal expanded T cells in CML could be found in Vbeta16 subfamilies. The frequent usage of Vbeta genes in clonal expansion was Vbeta3, Vbeta13 or Vbeta21. Multiple Vbeta clonal expansion was a general phenomenon in the same patient. The CDR3 sequence of Vbeta21 oligoclonal T cells from 3 cases showed some difference in splice regions and in the usage of J segments.

CONCLUSIONSThese results indicated that clonal expanded T cells could be found in patients with CML and were tendentious in Vbeta3, Vbeta13 and Vbeta21 subfamilies that may be related to the specific immune response for leukemia cell associated antigen.

Clone Cells ; Complementarity Determining Regions ; analysis ; Humans ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; immunology ; Receptors, Antigen, T-Cell, alpha-beta ; analysis ; T-Lymphocytes ; immunology ; pathology

OBJECTIVETo study the effect of DPC4 gene expression on the growth of colon cancer cells and its mechanism.

METHODSExpression plasmid pcDNA3.1-DPC4 was constructed and transfected into the colon cancer cell line SW620 by use of lipofectamine gene transfer technique. DPC4 protein expression was detected by Western blot and immunocytochemistry. The effect of DPC4 gene on the growth of SW620 cells was monitored by population doubling time (PDT) and cloning efficiency. The influence of DPC4 expression on p21WAF1 transcription was investigated by RT-PCR to detect p21WAF1 mRNA.

RESULTSSuccessful expression of DPC4 protein was detected in the transfected SW620 cells. Compared with the control cells, PDT (74 h) of the DPC4 expressing cells was prolonged and the cloning efficiency (21%) decreased. In addition, the mRNA level of p21(WAF1) in DPC4 transfected cells was increased.

CONCLUSIONSOverexpression of DPC4 gene inhibits the growth of colon cancer in vitro, and induction of p21(WAF1) expression may be an important functional aspect of DPC4.

Carcinoma ; genetics ; metabolism ; pathology ; Cell Cycle Proteins ; biosynthesis ; genetics ; Cell Division ; Colorectal Neoplasms ; genetics ; metabolism ; pathology ; Cyclin-Dependent Kinase Inhibitor p21 ; DNA-Binding Proteins ; biosynthesis ; genetics ; Gene Transfer Techniques ; Genes, Tumor Suppressor ; Humans ; Smad4 Protein ; Trans-Activators ; biosynthesis ; genetics ; Transfection ; Tumor Cells, Cultured

OBJECTIVE: To determine the effect of EphA2 protein on the expression of vascular endothelial growth factor (VEGF) and matrix metalloproteinase 9 (MMP9) proteins in HCT116 cells. METHODS: High expression of EphA2 protein in HCT116 cells was confirmed by Western blot. HCT116 cells were transfected with EphA2 antisense oligonucleotide. The expression of the transfection efficiency was analyzed by Western blot. VEGF proteins in the cell supernatants were detected by enzyme linked immunosorbent assay(ELISA), and the expressions of MMP9 in cell supernatants were examined by gelatin zymography. RESULTS: EphA2 antisense oligonucleotide suppressed the expression of VEGF and MMP9 proteins in HCT116 cells. CONCLUSION EphA2 could decrease the invasion and metastasis of HCT116 cells by suppressing the expression of VEGF and MMP9.
HCT116 Cells ; Humans ; Matrix Metalloproteinase 9 ; metabolism ; Oligonucleotides, Antisense ; Receptor, EphA2 ; genetics ; metabolism ; Vascular Endothelial Growth Factor A ; metabolism

OBJECTIVE: To explore the effect of Twist gene on the migration and invasion of human gastric carcinoma cells. METHODS: MKN28 cells, a human gastric carcinoma cell line, were transfected with PcDNA3.1-Twist plasmid by lipofectamine transfecting technique. The transfected cells were selected with geneticin. Expressions of Twist,ecadherin and vimentin protein were detected by Western blot in cells transfected Twist gene. Matrigel invision chambers were performed to analyse the cell migration and invasion. RESULTS: MKN28 cells transfected with PcDNA3.1-Twist plasmid showed stronger intracellular expression of Twist protein than MKN28 cells transfected with PcDNA3.1 and MKN28 cells without transfection. The expression of ecadherin protein in MKN28 cells transfected with PcDNA3.1-Twist plasmid was significantly decreased compared with that in MKN28 cells transfected with PcDNA3.1 and MKN28 cells without the transfection. However, The expression of vimentin protein in MKN28 cells transfected with PcDNA3.1-Twist plasmid was significantly increased compared with that in MKN28 cells transfected with PcDNA3.1 and MKN28 cells without transfection. The migration and invasion ability of Twist+ - MKN28 cells were stronger than that of MKN28 cells transfected with PcDNA3.1 and MKN28 cells without transfection. CONCLUSION Twist gene may promote the migration and invasion ability of gastric carcinoma cells through epithelial mesenchymal transition.
Cadherins ; biosynthesis ; Cell Movement ; genetics ; Humans ; Neoplasm Invasiveness ; Neoplasm Metastasis ; Stomach Neoplasms ; genetics ; metabolism ; pathology ; Transfection ; Tumor Cells, Cultured ; Twist-Related Protein 1 ; biosynthesis ; genetics ; Vimentin ; biosynthesis

The rearrangement segments of TCR Valpha40 gene with Jdelta1, Ddelta3 or psi Jalpha were amplified in genomic DNA from peripheral blood mononuclear cells of 10 normal subjects, sorted CD3(+) cells from peripheral blood of 4 cases and thymocytes from 7 cases, by using nested PCR. Different amounts of DNA from all samples were amplified to estimate the frequency of Valpha40 gene rearrangements. The results indicated that the rearrangements of TCR Valpha40 gene with Jdelta1, Ddelta3 or psi Jalpha could be found respectively in the most samples of peripheral blood T cells or thymocytes. The frequencies of Valpha40 rearrangements were different in peripheral blood T cells and thymocytes by analysis of PCR with different amounts DNA. It is concluded that the TCR V alpha40-psi Jalpha was the most frequent rearrangement in mature and immature T cells, whereas TCR Valpha40-Ddelta3 was more frequently rearranged in immature T cells
Gene Rearrangement ; Humans ; Polymerase Chain Reaction ; Protein Subunits ; genetics ; Receptors, Antigen, T-Cell, alpha-beta ; genetics ; T-Lymphocytes ; physiology

To investigate the distribution and clonal expansion of TCR Vbeta subfamily T cells in patients with B-NHL and T-NHL, the CDR3 of TCR Vbeta 24 subfamily genes was amplified in peripheral blood mononuclear cells from 4 cases with B-NHL and 2 cases with T-NHL using RT-PCR, and to observe the usage of TCR Vbeta repertoire, the PCR products were further labeled with fluorescence and analyzed by genescan technique for the CDR3 size, to evaluating clonality of the detectable TCR Vbeta T cells. The results indicated that only selected expression of 6-12 Vbeta subfamily T cells could be identified in the 6 cases with NHL, and Vbeta1, Vbeta8, Vbeta13 and Vbeta19 were expressed in all samples, Vbeta2 and Vbeta16 could be found in 5 samples, whereas Vbeta4-6, Vbeta10-12, Vbeta15, Vbeta17-18, Vbeta20 and Vbeta22-23 were absent in all samples. Genescan analysis showed that clonal expansion of T cells could be found in 1-3 Vbeta subfamilies from 2 cases with B-NHL and 1 case with T-NHL. In conclusions, the similar selected usage of TCR Vbeta subfamily T cells could be found in peripheral blood from patients with B and T NHL, clonal expansion of T cells which were considered to be related to lymphoma cell antigen could be detected in a part of patients with B or T NHL.
Cell Line ; Clone Cells ; Complementarity Determining Regions ; genetics ; Genes, T-Cell Receptor beta ; genetics ; Humans ; Jurkat Cells ; Lymphoma, Non-Hodgkin ; genetics ; immunology ; RNA, Neoplasm ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; T-Lymphocytes ; metabolism

OBJECTIVETo screen the molecular markers for refractory anemia with excess blasts in transformation (RAEB) in myelodysplastic syndromes (MDS) by serum proteome profiling.

METHODSThe serum protein were isolated from patients with RAEB, acute myeloid leukemia or normal subjects by 2-dimensional electrophoresis (2-DE), and the electrophoresis gels were obtained to identify the differentially reacting protein spots. The replica gels of the differentially reacting proteins were analyzed to locate the matching protein spots, which were identified by peptide mass fingerprint based on matrix-assisted laser desorption/ionization time of-flight mass spectrometry (MALDI-TOF-MS) and database searching.

RESULTSSeven differentially expressed proteins in RAEB were found by 2-DE. Of the 7 proteins, 4 were identified by MALDI-TOF-MS to have significantly differential expression in RAEB, including dipeptidyl peptidase (DPP/CD26), polymerase (DNA directed) kappa, PRO2044 and an albumin-like protein.

CONCLUSION2-DE-based serum proteome profiling helps identify serum proteomic biomarkers related to MDS. DDP/CD26 has increased expression in the serum in RAEB subtype MDS, suggesting its possible role in advanced MDS.

Anemia, Refractory, with Excess of Blasts ; blood ; genetics ; Bone Marrow ; pathology ; DNA-Directed DNA Polymerase ; blood ; Dipeptidyl-Peptidases and Tripeptidyl-Peptidases ; blood ; Female ; Humans ; Male ; Middle Aged ; Myelodysplastic Syndromes ; blood ; classification ; genetics ; Proteomics

OBJECTIVETo evaluate the outcome of mometasone furoate nasal spray (MFNS) used for 3 months on non-allergic rhinitis (NAR).

METHODSIn this multicenter study, NAR patients were enrolled from eight hospitals and received MFNS 200 microgram once daily for 3 months. The patients were followed-up for three times (at baseline, month 1 and month 3) to record the symptom scores and nasal endoscopic appearances. At the same time, the adverse events frequency was recorded and analyzed.

RESULTSA total of 188 NAR cases were enrolled in the study. The total nasal symptom score assessment descended significantly at month 1 (1.70 ± 0.75) and month 3 (0.95 ± 0.79) visits versus at baseline (2.67 ± 0.68, Z value were from -11.603 to -10.491, all P < 0.01). The individual symptoms, including nasal stuffiness, nasal discharge, nasal stuffiness-related dizziness or headache, hyposmia, sleep quality, daily life activity, work or study efficiency, mental status, and whole body fatigue, also showed less scores at month 1 and month 3 visits versus at baseline (Z value were from -11.313 to -6.802, all P < 0.01). At the same time, nasal mucosal appearances assessed by endoscopy had lower scores at month 1 (1.40 ± 0.62) and month 3 (0.75 ± 0.71) visits versus at baseline (2.27 ± 0.73, Z value were from -11.484 to -10.002, all P < 0.01). Additionally, adverse events were only observed in 5.3% cases with light rhinorrhagia and nasal dryness. No other side effect was found.

CONCLUSIONSA 3-months administration of intranasal mometasone can effectively and safely improve NAR patients' clinical symptom and nasal mucosal appearances.

Adolescent ; Adult ; Anti-Allergic Agents ; administration & dosage ; therapeutic use ; Female ; Humans ; Male ; Middle Aged ; Mometasone Furoate ; Nasal Sprays ; Pregnadienediols ; administration & dosage ; therapeutic use ; Rhinitis ; classification ; drug therapy ; Treatment Outcome ; Young Adult

OBJECTIVETo investigate the clonal expansion of T cell receptor (TCR) Vbeta subfamily T cells from cord blood induced by bcr3-abl2 peptide in vitro.

METHODST cells from 3 units of cord blood were amplified by anti-CD(3) monoclonal antibody (McAb) and IL-2 with or without synthetic b3a2 peptide. T cell specific cytotoxicity was analyzed by lactate dehydrogenase (LDH) assay, TCR Vbeta subfamilies by using reverse transcriptase-polymerase chain reaction (RT-PCR) and genescan technique.

RESULTSbcr3-abl2 peptide specific cytotoxicity T cells were successfully induced from the 3 units of cord blood by synthetic b3a2 peptide. Compared with that in CD(3) McAb induced cells, distribution pattern of TCR Vbeta repertoire was different in T cells induced with b3a2 peptide. Oligoclonal and oligoclonal tendency TCR Vbeta subfamily T cells could be identified in cord blood T cells induced by b3a2 peptide in 1 or 2 weeks, whereas those induced by anti-CD(3) McAb and IL-2 were mostly polyclonal.

CONCLUSIONThe cytotoxicity T cells with anti-CML specificity could be induced by b3a2 peptide. The specific anti-CML cytotoxicity may be derived from the clonal expansion TCR Vbeta subfamily T cells.

Antibodies, Monoclonal ; immunology ; CD3 Complex ; immunology ; Fusion Proteins, bcr-abl ; pharmacology ; Genes, T-Cell Receptor beta ; Humans ; Interleukin-2 ; pharmacology ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; immunology ; T-Lymphocytes, Cytotoxic ; immunology

Objective: To clarify the characteristics of the A20 regulatory changes by analyzing mutations in the non-coding region of the A20 gene in patients with T-cell lymphoma leukemia (T-LCL) . Methods: PCR and nucleotide sequence analysis were used to detect mutations in the non-coding region of the A20 gene, and DNA samples from PBMCs of 52 cases of T-LCL and 99 healthy controls. Results: A missense mutation (c.-672T>G) was detected in the A20 gene promoter from one T-LCL patient, which has been registered as a SNP (rs139054966) in gene bank. Meanwhile, a new mutation was detected in the 3' UTR mRNA (3916 (C>G) ) . These two mutations were absent in other T-LCL samples and controls. Conclusion: The rs139054966 (c.-672T>G) and 3916 (C>G) mutations in the A20 gene were detected in T-LCL patients for the first time. There was also rs139054966 located on the binding region of the transcription factor P53, and its significance remained to be further clarified.
3' Untranslated Regions ; Humans ; Leukemia ; Lymphoma, T-Cell ; Mutation ; Promoter Regions, Genetic