Carcinoma ; metabolism ; Cell Differentiation ; genetics ; Gallbladder Neoplasms ; genetics ; metabolism ; Gene Expression Regulation, Neoplastic ; Humans ; Kangai-1 Protein ; genetics ; metabolism ; NM23 Nucleoside Diphosphate Kinases ; genetics ; metabolism

OBJECTIVETo study the expression of p16 and nm23 genes in salivary gland tumors and the relation of P16 and nm23 proteins with fumorigenesis of salivary gland tumors.

METHODSExpression of P16 and nm23 proteins was examined by SABC immunohistochemical method in 39 cases of paraffin blocks of normal salivary gland tissues and salivary gland tumors.

RESULTSP16 and nm23 protein positive staining were mainly found in the cytoplasm and cytoblast of all salivary gland tissues. Positive rate of P16 protein expression was 76.9% (10/13) and 40.9% (9/22) in benign and malignant salivary gland tumors, respectively. There was significant difference between P16 protein expression of benign and malignant tumors by chi 2 test (P < 0.05). mm23 protein positive staining was found in 84.6% (11/13) and 45.5% (10/22) of benign and malignant tumors respectively. The expression of nm23 protein in benign and malignant tumors was significantly different (P < 0.05). There was no correlation of the expression of P16 and nm23 in salivary gland tumors was found (P > 0.05).

CONCLUSIONp16 and nm23 genes may play an important role in different sides in salivary gland tumorigenesis and the reduce of the expression of p16 and nm23 genes may contribute to the generation of malignant salivary gland tumors.

Adenoma, Pleomorphic ; genetics ; metabolism ; Carcinoma, Mucoepidermoid ; genetics ; metabolism ; Cyclin-Dependent Kinase Inhibitor p16 ; biosynthesis ; genetics ; Humans ; Immunohistochemistry ; NM23 Nucleoside Diphosphate Kinases ; Nucleoside-Diphosphate Kinase ; Protein Biosynthesis ; Proteins ; genetics ; Salivary Gland Neoplasms ; genetics ; metabolism ; Salivary Glands ; metabolism

OBJECTIVETo transfect nm23-H1 into the BcaCD885 cell lines in order to get safe high-efficiency and low-toxicity, and to find out whether nm23-H1 could affect the invasion and metastases ability of BcaCD885 cell lines.

METHODSLipofect was used to transfect nm23-H1 into BcaCD885 cell lines; immunohistochemistry was used to detect the difference expression of nm23-H1 between transfected and non-transfected cell lines; then transwell-room and wash way were used to detect the difference of invasion and metastases ability between transfected and non-transfected cell lines.

RESULTSPCMV-NEO-BAM system gave the stability expression of nm23-H1; there was significant different NDPKA expression between transfected and non-transfected BcaCD885 cell lines; the invasion and metastases ability of transfected BcaCD885 cell lines decreased obviously.

CONCLUSIONnm23-H1 can inhibit the metastases of BcaCD885 cell lines significantly.

Cell Adhesion ; physiology ; Cell Movement ; physiology ; Genetic Vectors ; genetics ; Humans ; Monomeric GTP-Binding Proteins ; genetics ; metabolism ; Mouth Neoplasms ; genetics ; pathology ; physiopathology ; NM23 Nucleoside Diphosphate Kinases ; Nucleoside-Diphosphate Kinase ; Transcription Factors ; genetics ; metabolism ; Transfection ; Tumor Cells, Cultured

OBJECTIVETo identify molecular markers of lung squamous cell carcinoma by cDNA microarray technique.

METHODScDNA expression profiles were examined by microarrays of 6 surgical specimens of stage I lung squamous cell carcinomas. Those genes, either up-regulated or down-regulated in every specimen studied, were identified. The expression levels of nm23 and BRCA2 by the squamous cell carcinoma of the lung were further examined by immunohistochemical techniques.

RESULTSA total of 107 genes were identified, of which 26 were up-regulated and 81 were down-regulated in all six specimens. Immunohistochemical staining showed that, compared with normal lung tissues, the intensity of nm23 expression by the squamous cell carcinoma of lung was significantly increased while that of BRCA-2 was decreased.

CONCLUSIONcDNA microarrays can be used to identify gene expression profile of lung cancer, some of which may be used as markers of lung squamous cell carcinoma.

BRCA2 Protein ; metabolism ; Biomarkers, Tumor ; Carcinoma, Squamous Cell ; genetics ; metabolism ; Gene Expression Profiling ; Humans ; Lung Neoplasms ; genetics ; metabolism ; Male ; NM23 Nucleoside Diphosphate Kinases ; Nucleoside-Diphosphate Kinase ; metabolism ; Oligonucleotide Array Sequence Analysis

OBJECTIVE: To explore the relationship between the expressions of PTEN and the metastasis of the gallbladder cancer. METHODS: The expression of PTEN and nm23 were detected by immunohistochemical staining in 32 cases of gallbladder cancer with metastasis and the staining intensity was scored semi-quantitatively, compared with the cases without metastasis. RESULTS: The intensity score of PTEN and nm23 in gallbladder cancer with metastasis was 8.9947+/-4.5590 and 10.2003+/-3.9031, respectively, which was lower than that in those without metastasis (12.9433+/-4.7618 and 15.8436+/-5.6917 respectively, P < 0.01 ). The expression of PTEN was correlative with that of nm23 ( Pearson = 0.370, P < 0.05). CONCLUSION The lower expressions of PTEN and nm23 are related to the metastasis of gallbladder cancer.
Female ; Gallbladder Neoplasms ; metabolism ; Humans ; Liver Neoplasms ; metabolism ; secondary ; Lymphatic Metastasis ; Male ; NM23 Nucleoside Diphosphate Kinases ; Nucleoside-Diphosphate Kinase ; biosynthesis ; genetics ; PTEN Phosphohydrolase ; biosynthesis ; genetics ; Tumor Suppressor Proteins ; biosynthesis ; genetics

To purify recombinant human nucleoside diphosphate kinase A (rhNDPK-A) efficiently in pilot scale, cells of rhNDPK-A producing E. coli were homogenized by high pressure under 4 degrees C, 950 Pa. The insoluble debris was removed by microfiltration and the soluble portion was concentrated by ultrafiltration. The resulted crude sample was loaded on DEAE-sepharose Fast Flow. The target fraction was collected and then load on Cibacron Blue 3GA Sepharose CL-4B. Eluted with buffer containing ATP from the AC column, rhNDPK-A was polished with ultrafiltration. The results showed that after homogenized 2 rounds, 1500g cells of E. coli brought crude sample containing 47.6g NDPK-A. Treated with microfiltration and ultrafiltration, 27.3g of NDPK-A were recovered from this bacteria homogenate. After 2-step purification with column chromatography and then polished with ultrafiltration, 17.2 g rhNDPK-A were collected with purity of 96.3%. The recovery of the whole purification process was 36.2%, and the productivity of rhNDPK-A was 1.15 g per 100 g wet cells. Comparing the recovery of each purification step, it was found that the recovery of polish is higher than that of affinity chromatography, which is higher than that of ion exchange chromatography. The limit step was the process of sample pretreatment among the 4 purification steps. Combine with the fermentation results reported before, it was deduced that the productivity of rhNDPK-A was 510 mg/L. In conclusion, an easily controlled purification condition with high yield provides material for the translation researches of NDPK; In addition, it was suggested the crucial step determine the recovery of non-secretive recombinant proteins might be the process of sample pretreatment, not be the process of column chromatography.
Chromatography, Affinity ; Chromatography, Ion Exchange ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli ; genetics ; Humans ; NM23 Nucleoside Diphosphate Kinases ; genetics ; metabolism ; Pilot Projects ; Recombinant Proteins ; isolation & purification ; metabolism ; Ultrafiltration

This experimental study sought to find out the inhibitory effects of Ad-GFP-nm23-H1 on proliferation and metastasis of human colorectal carcinoma cell line Lovo, and, further, to gain an insight into some theoretical and methodical basis for instituting nm23-H1 gene therapy of cancers. MTT assay and Transwell chamber were used to detect the rates of proliferation and invasion as well as the adhesion of Lovo cells in vitro. The results demonstrated that the proliferation inhibition rates of Lovo cells treated with Ad-GFP-nm23-H1 of 10(10) PFU/ml, 10(9) PFU/ml and 10(8) PFU/ml were 84.9% +/- 1.51%, 48.5% +/- 7.23% and 22.5% +/- 5.47%, that the adherence inhibition rates of Lovo cells treated with Ad-GFP-nm23-H1 of 10(10) PFU/ml, 10(9) PFU/ml and 10(8) PFU/ml were 70.3% +/- 2.40%, 60.1% +/- 5.68% and 18.5% +/- 3.61%, and that the invasiveness inhibition rates of Lovo cells treated with Ad-GFP-nm23-H1 of 10(10) PFU/ml, 10(9) PFU/ml and 10(8) PFU/ml were 83.2% +/- 5.71%, 52.2% +/- 6.94% and 28.1% +/- 8.21%. These data suggested that Ad-GFP-nm23-H1 exerted significant inhibitory effects on the proliferation and metastasis of human colorectal carcinoma cell line Lovo in a dose-dependent way.
Adenoviridae ; genetics ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; Colorectal Neoplasms ; genetics ; pathology ; Gene Expression Regulation, Neoplastic ; Genetic Therapy ; Green Fluorescent Proteins ; genetics ; metabolism ; Humans ; NM23 Nucleoside Diphosphate Kinases ; genetics ; metabolism

OBJECTIVETo study the prognostic predictor for nasopharyngeal carcinoma (NPC).

METHODSThe expressions of nm23-H1 and vessel endothelium growth factor (VEGF) protein were examined by immunohistochemistry S-P staining in 108 NPC tissues, the expression of nm23-H1 and VEGF protein in NPC tissues with clinical stage of NPC, radiosensitivity of tumor, survival rate of patients, relapse and metastasis of carcinoma were studied.

RESULTSThe positive rate of nm23-H1 and VEGF was 48.1% and 59.3% respectively. The clinical staging, metastatic potential of lymph nodes were correlated with low-level expression of nm23-H1 protein. The patients with negative nm23-H1 expression had worse prognosis than those with positive nm23-H1 expression. The clinical staging, metastatic potential and poor sensitivity of radiotherapy were correlated with high level expression of VEGF protein. The patients with positive VEGF expression had worse prognosis than those with negative VEGF expression. The expression of nm23-H1 protein was negatively correlated with the expression of VEGF protein (r = -0.577, P < 0.05).

CONCLUSIONSThe low level expression of nm23-H1 protein and the high level expression of VEGF protein may be associated with the development and poor prognosis of NPC.

Adolescent ; Adult ; Aged ; Carcinoma, Squamous Cell ; metabolism ; pathology ; Female ; Humans ; Lymphatic Metastasis ; Middle Aged ; NM23 Nucleoside Diphosphate Kinases ; genetics ; metabolism ; Nasopharyngeal Neoplasms ; metabolism ; pathology ; Neoplasm Staging ; Prognosis ; Survival Rate ; Vascular Endothelial Growth Factor A ; genetics ; metabolism ; Young Adult

A single-stranded oligonucleotides containing a 6 histidine sequence, a hydroxylamine cleavage site, a thrombin cleavage site, and stop codon TAA were inserted into the polylinker's downstream of prokaryotic expression vector pBV220 between BamHI and PstI. The resultant vector is named pBV223. Proteins expressed in this vector will have a 6 histidine tail as affinity handy fused to their C terminus and can be quickly purified by one step immobilized metal affinity chromatography (IMAC). This plasmid is verified by restriction map and DNA sequencing. Subsequently, the metastasis suppressor gene nm23-H1 cDNA (without the stop codon) was cloned into vector PBV223 in frame with the 6-histidine sequence, hydroxylamine and thrombin cleavage sites. The soluble nm23-H1 fusion protein was successfully induced in the bacterial DH5a and easily purified with Ni chromatograph. These results indicated that the strategy to clone the single-stranded oligonucleotides directly into the restriction sties between BamH I and Pst I in the pBV220 vector is the simplest and cost-effective method.
Base Sequence ; Chromatography, Affinity ; methods ; Cloning, Molecular ; DNA, Complementary ; genetics ; Escherichia coli ; genetics ; metabolism ; Genetic Vectors ; genetics ; Molecular Sequence Data ; Mutation ; NM23 Nucleoside Diphosphate Kinases ; biosynthesis ; genetics ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; isolation & purification

NM23 is a family of structurally and functionally conserved proteins known as nucleoside diphosphate kinases (NDPK). There is abundant mRNA expression of NM23-H1, NM23-H2, or a read through transcript (NM23-LV) in the primary sites of hepatocellular carcinoma (HCC). Although the NM23-H1 protein is implicated as a metastasis suppressor, the role of NM23-H2 appears to be less understood. Thus, the aim of this study was to examine whether NM23-H2 is associated with hepatocarcinogenesis. The level of NM23-H2 expression in tumor tissues and the surrounding matrix appeared to be independent of etiology and tumor differentiation. Its subcellular localization was confined to mainly the cytoplasm and to a lesser extent in the nucleus. Ectopic expression of NM23-H2 in NIH3T3 fibroblasts and HLK3 hepatocytes showed a transformed morphology, enhanced focus formation, and allowed anchorage-independent growth. Finally, NIH3T3 fibroblasts and HLK3 hepatocytes stably expressing NM23-H2 produced tumors in athymic mice and showed c-Myc over-expression. In addition, NF-kappaB and cyclin D1 expression were also increased by NM23-H2. Lentiviral delivery of NM23-H2 shRNA inhibited tumor growth of xenotransplanted tumors produced from HLK3 cells stably expressing NM23-H2. Collectively, these results indicate that NM23-H2 may be pro-oncogenic in hepatocarcinogenesis.
Animals ; Carcinoma, Hepatocellular/*enzymology/genetics/pathology ; Cell Line ; Cell Line, Tumor ; *Gene Expression Regulation, Neoplastic ; Humans ; Liver/*enzymology/metabolism/pathology ; Liver Neoplasms/*enzymology/genetics/pathology ; Mice ; Mice, Nude ; NIH 3T3 Cells ; NM23 Nucleoside Diphosphate Kinases/*genetics/metabolism