Expressing trend of NME3 protein in acute myeloid leukemia HL-60 cells and patients' bone marrow / 中国实验血液学杂志

To verify the differential expression of non-metastasis cell 3 (NME3) protein in HL-60 cells when they were induced to differentiate into monocyte and granulocyte like cells, and study its value in diagnosis of acute myeloid leukemia, all-trans retinoic acid (ATRA) and a new steroidal drug NSC67657 were employed to induce acute myeloid leukemia HL-60 cells into monocyte and granulocyte like cells. Then the cell differentiating direction was observed by chemical staining, the degree of differentiation was determined by surface antigen CD11b/CD14 detection, and the apoptosis was excluded by phosphatidylserine valgus analysis, by which cellular differentiating model was constructed. Furthermore, RT-PCR and Western blot were employed to verify the differentially expression of NME3 before and after differentiation of HL-60 cells. At last, samples from bone marrow nucleated cells of 26 patients with myeloid leukemia, which were diagnosed definitely by clinical doctors, and 5 normal people were chosen. Then the expressing trend of NME3 protein in these testing groups was analyzed by means of comparison. The results showed that ATRA (2 µmol/L for 5 d) and NSC67657 (10 µmol/L for 5 d) could induce HL-60 cells to differentiate into monocyte and granulocyte like cells above 90% without cell apoptosis. The expression of NME3 gene and protein were down-regulated by the inducers, which was accorded with the screening results that was got using proteomics technology in the former research. The expression of NME3 protein in bone marrow from acute myeloid leukemia patients was elevated significantly as compared to normal persons. It is concluded that the expression level of NME3 protein is down-regulated after cellular differentiation, according with the changing trend in leukemia patients, which imply that NME3 protein may be a potential biomarker for diagnosis of acute myeloid leukemia.

Expression and clinical significance of vascular endothelial growth factor-C and nm23-H1 in stage II and III colorectal carcinomas / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To discuss the expression and clinical significance of VEGF-C and nm23-H1 in stage II and III colorectal carcinomas.</p><p><b>METHODS</b>SP immunohistochemical staining was employed to determine the expression of vascular endothelial growth factor-C (VEGF-C) and nm23-H1 in the tumor tissues of 110 cases of stage II and III colorectal carcinomas and in the adjacent mucosal tissues of 53 cases as control, and analyze their correlation with cliniopathological features and prognosis.</p><p><b>RESULTS</b>The positive expression of VEGF-C in the carcinoma tissues was 71.8%, significantly higher than that in the adjacent mucosal tissues (22.6%, P < 0.001). The positive expression of nm23-H1 in the carcinoma tissues was 57.3%, significantly lower than that in the adjacent mucosal tissues (90.6%, P < 0.001). The expression of VEGF-C was significantly correlated with lymph node metastasis (P < 0.05), and the nm23-H1 expression was significantly correlated with lymph node metastasis and pathological type (P < 0.05). The expression of VEGF-C and nm23-H1 did not show a significant correlation with age, gender, primary tumor site, tumor size and depth of invasion (P > 0.05). The VEGF-C expression was negatively related with nm23-H1 expression in colorectal carcinoma (r = -0.361, P < 0.001). The median overall survival (MOS) and median disease free survival (MDFS) of 110 patients with colorectal carcinoma were 55 and 48 months, respectively. The colorectal patients with different VEGF-C and nm23-H1 expression showed significant differences in the 5-year OS rate and 5-year DFS rate (P < 0.001). The patients with negative VEGF-C expression and positive nm23-H1 expression had a better prognosis.</p><p><b>CONCLUSIONS</b>The joint detection of VEGF-C and nm23-H1 expression is very promising in prediction of the prognosis of patients with stage II and III colorectal carcinoma. However, whether it can be used as a marker in prognosis judgment needs further investigation.</p>

Pro-oncogenic potential of NM23-H2 in hepatocellular carcinoma

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.

Significance of TIMP-1 and nm-23 expressions in HER-2-positive breast cancer / 中华肿瘤杂志

<p><b>OBJECTIVE</b>Trastuzmab combined therapy has been recommended as standard therapy for HER-2-positive breast cancer. However, more than 70% HER-2-positive cases do not recur and might thus be with overtreatment risk. The aim of this study was to explore the significance of TIMP-1 and nm-23 expression in identifying the good prognosis subtype in HER-2-positive breast cancer.</p><p><b>METHODS</b>One hundred and sixty-five cases of HER-2-positive breast cancer treated in Tianjin Medical University Cancer Hospital from Jan. 1987 to Dec. 1988 were studied. Expressions of TIMP-1 and nm-23 were detected by immunohistochemistry and the correlation with clinicopathologic characteristics and long-term outcome was analyzed.</p><p><b>RESULTS</b>53.3% (88/165) of TIMP-1 and 72.7% (120/165) of nm-23 expression were detected in the patients. The TIMP-1-negative patients classified as good prognosis group had a 10-year metastasis free survival (MFS) of 58.4% and 10-year overall survival (OS) of 68.8%, compared with the 10-year MFS of 43.2% (P = 0.041) and 10-year OS of 52.0% in the positive group (P = 0.020). The nm-23-positive patients classified as good prognosis group had a 10-year MFS of 54.2% and 10-year OS of 65.7%, compared with the 10-year MFS of 40.0% (P = 0.049) and 44.1% (P = 0.015) in the negative group. The multivariate analysis showed that expressions of TIMP-1 and nm-23, tumor size and lymph node involvement were independent prognostic factors.</p><p><b>CONCLUSIONS</b>In HER-2-positive breast cancer, TIMP-1 and nm-23 can be used to further distinguish between low-risk and high-risk subgroups, and they are independent factors in prognosis.</p>

nm23-H1-siRNA enhances the chemosensitivity to liposome-encapsulated paclitaxel in lung adenocarcinoma cells in vitro / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To study the chemosensitivity of lung adenocarcinoma cell line A549 cells to liposome-encapsulated paclitaxel after treatment by nm23-H1-small interference RNA (nm23-H1-siRNA) in vitro.</p><p><b>METHODS</b>The A549 cells were divided into two groups: non-transfected group and nm23-H1-siRNA-transfected group. Western blot analysis was used to detect the expression of nm23-H1. MTT and flow cytometry were used to determine the cell mortality rate, apoptosis rate and cell cycle after liposome-encapsulated paclitaxel treatment in both groups.</p><p><b>RESULTS</b>The expression of nm23-H1 in A549 cells was significantly decreased after transfection with nm23-H1-siRNA. After treatment for 48 hours with liposome-encapsulated paclitaxel, the cell mortality rate was increased with the increasing concentration of liposome-encapsulated paclitaxel in both groups, but increased higher in the nm23-H1-siRNA-transfected group. When the concentration of liposome-encapsulated paclitaxel was above 5 µg/ml, the cell mortality rate was significantly higher than that in the non-transfected group (P < 0.05). The proportion of apoptotic cells also increased in the nm23-H1-siRNA-transfected group, compared with that of the non-transfected group (t = 3.812, P < 0.05), while the proportion of cells at S and G(2)/M phase decreased after transfection with nm23-H1-siRNA (S phase:t = 8.356, P < 0.05; G(2)/M phase:t = 7.256, P < 0.05).</p><p><b>CONCLUSIONS</b>Nm23-H1 is related with the chemoresistance to liposome-encapsulated paclitaxel in lung adenocarcinoma cell line A549 cells. Inhibition of the expression of nm23-H1 by nm23-H1-siRNA can improve the in vitro chemosensitivity of A549 cells to liposome-encapsulated paclitaxel.</p>

Proteomic analysis of human ovarian cancer paclitaxel-resistant cell lines / 中南大学学报(医学版)

OBJECTIVE@#To study the proteins related to paclitaxel-resistant of ovarian cancer cell line.@*METHODS@#The total proteins of paclitaxel-resistant and paclitaxel-sensitive human ovarian cancer cell lines were separated by 2-dimensional gel electrophoresis (2-DE). The differentially expressed proteins were analyzed using image analysis software. The differential proteins were identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Western blot was used to determine the differential expression levels of the 2 proteins.@*RESULTS@#Forty differentially expressed proteins were found by image analysis software, and 24 differential proteins were identified by mass spectrometry. These proteins included proliferation cell nuclear antigen (PCNA), nm23, prohibitin (PHB), annexin, alpha-enolase, heat shock protein (HSP), and so on.@*CONCLUSION@#Twenty-four proteins in human ovarian cancer cell lines of paclitaxel-resistant and paclitaxel-sensitive are found by proteomic techniques, which may be involved in the paclitaxel-resistance of human ovarian cancer cells.

Expressions of nucleoside diphosphate kinase (nm23) in tumor tissues are related with metastasis and length of survival of patients with hepatocellular carcinoma / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To evaluate the relationship of expressions of nucleoside diphosphate kinase (nm23) and proliferating cell nuclear antigen (PCNA), as well as apoptosis, with the prognosis of HCC patients by analyzing their pathological and clinical data.</p><p><b>METHODS</b>The expressions of nm23 and PCNA were analyzed by immunohistochemistry and the apoptotic phenomena were detected by TUNEL technique in the liver samples from 43 HCC tissues, 39 para-neoplastic tissues, and 10 normal tissues. The mean apoptosis index (AI) and proliferative index (PI) in individual sample were calculated.</p><p><b>RESULTS</b>As shown by the detection, 32.6% of carcinomas had negative nm23 signal in tumor tissues, whereas all para-neoplastic and normal tissues had positive nm23. The AI in nm23 positive HCC was significantly higher than that in nm23 negative one, with statistical difference (P<0.05). Furthermore, the expressions of nm23, and the values of AI and PI were contrastively analyzed with some main pathological and clinical data of HCC. It revealed that HCC with extrahepatic metastasis showed remarkable correlation with the negative nm23 (P=0.013) and higher PI values of HCC (P=0.015). The disease-free survival in HCC patients with negative nm23 expression was significantly poorer than that in patients with positive nm23 expression.</p><p><b>CONCLUSIONS</b>These data suggest that expressions of nm23 protein in tumor tissues are correlated with occurrences of metastasis and length of survival of the HCC patients, which may be an indicator for their prognosis.</p>

Expressions of surviving, MMP2, TIMP1, CD44 and nm23 of two tumors originating from different immunosurveillance in the same patient / 南方医科大学学报

<p><b>OBJECTIVE</b>To explore the differences in the survival, adhesion and diffusion capacity between different carcinomas originating from different immunosurveillance in the same patient.</p><p><b>METHODS</b>The expressions of survivin, MMP2, TIMP1, CD44, and nm23 proteins were detected immunohistochemically in a patient with acute lymphoblastic leukemia and lymphoma after allogeneic blood stem cell transplantation.</p><p><b>RESULTS</b>Survivin, MMP2, TIMP1, CD44, and nm23 proteins were positive in acute lymphoblastic leukemia samples obtained before transplantation and negative in the lymphoma tissue occurring after the transplantation.</p><p><b>CONCLUSION</b>The expressions of survivin, MMP2, TIMP1, CD44, and nm23 proteins vary between the two carcinomas originating from different immunosurveillance in the same patient.</p>

Adenovirus-mediated delivery of nm23-H1 gene inhibits growth of colorectal carcinoma cell line Lovo / 生物医学工程学杂志

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.

Correlation of polymorphism of Nme1-1465 T>C and TGFβ1-509 T>C with genetic susceptibility of gastric carcinoma / 中华病理学杂志

<p><b>OBJECTIVE</b>To explore the correlation of functional genetic variants in Nme1-509 C>T and TGFβ1-1465 T>C genes to the genetic susceptibility of gastric carcinoma in Fujian province, China.</p><p><b>METHODS</b>A case-control study was conducted in a population in Fujian province. The polymorphism of TGFβ1-509 C>T (rs1800469), Nme1-1465 T>C (rs16949649) in 273 gastric carcinoma patients and 277 cancer-free controls, frequency-matched by age and sex, were analysed by using Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry (MALDI-TOF-MS). Adjusted odds ratios (OR) and 95% confidence evaluation intervals (95%CI) measured by multivariate Logistic regression analysis were adopted in studying the correlation of the gene polymorphism with the susceptibility of gastric cancer.</p><p><b>RESULTS</b>After the adjustment using Logistic regression or the potential confounding effects of gender and age, as compared with TT+CT genotype gastric carcinoma patients, the homozygous Nme1-1465CC genotype carriers had a significantly higher risk in lymph node metastasis, with the OR of 2.5 (95%CI 0.08-2.10; P=0.029). There was no association obtained between TGFβ1-509 T>C genotype with the tumor size, cell differentiation, tumor invasion and lymph node metastasis in gastric carcinoma. In the intestinal type gastric carcinoma group, when compared with the wild homozygous Nme1 TT* TGFβ1 CC, Nme1 TC* TGFβ1 TC, Nme1 TC* TGFβ1 TT and Nme1 CC* TGFβ1 TC genotype carriers, there was a significantly decrease of risk in gastric carcinogenesis of 0.42 fold (95%CI 0.54-0.94, P=0.022), 0.32 fold (95%CI 0.42-0.97, P=0.013) and 0.26 fold (95%CI 0.42-0.97, P=0.008), respectively.</p><p><b>CONCLUSIONS</b>There is a significant relationship between polymorphism of Nme1-1465 T>C and the prognosis of carcinoma of stomach. It also demonstrates that coexistence of Nme1-1465 T>C and TGFβ1-509 T>C genes may provide a synergistic effect of increasing the susceptibility of gastric carcinogenesis.</p>

Quantity and distribution of CD44+/CD24- cells in breast cancer tissue and the cell lines / 中华病理学杂志

<p><b>OBJECTIVE</b>To study the distribution and quantity of CD44+/CD24- cells in breast cancer tissue and the cell lines, and as well as its correlation with the expression of various breast cancer markers and molecular subtyping of breast carcinoma.</p><p><b>METHODS</b>The expression of CD44/CD24, estrogen receptor, progesterone receptor, HER2, human estrogen-induced protein PS2, bcl-2 and nm23 in 60 cases of invasive ductal carcinoma of breast were studied by either single or double immunohistochemical staining. The co-expression of CD44 and CD24 in 3 breast cancer cell lines (MCF-7, MDA-MB-468, and MDA-MB-231) was also examined.</p><p><b>RESULTS</b>The quantity and distribution of CD44+/CD24- cells varied greatly and no specific patterns were identified. The percentage of CD44+/CD24- in breast cancer was 65%. The amount of CD44+/CD24- cells did not correlate with the age of patients, lymph node metastasis, tumor size, molecular subtypes and expression of various breast cancer markers in breast carcinoma. The proportion of CD44+/CD24- cells in MCF-7, MDA-MB-468, and MDA-MB-231 cell lines was <1%, 5% and >80%, respectively.</p><p><b>CONCLUSIONS</b>CD44+/CD24- cells are demonstrated in certain breast cancer tissues and cell lines. However, there is no relationship obtained between the quantity or the distribution of these cells and the molecular subtyping or the clinicopathologic parameters in breast cancer.</p>

Expression of nm23 H(1) gene in childhood acute lymphoblastic leukemia and the relationship between nm23 H(1) expression and immunophenotype / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To study the expression of nm23-H(1) gene in children with acute lymphoblastic leukemia (ALL) and the relationship between nm23-H(1) expression and immunophenotype.</p><p><b>METHODS</b>nm23-H(1) expression was measured by semiquantitative RT-PCR in children with ALL (newly diagnosed, n = 40; remission, n = 32; relapse, n = 16; refractory, n = 3). Twenty normal children served as the control group. The relationship between nm23-H(1) expression and immunophenotype was evaluated.</p><p><b>RESULTS</b>The expression of nm23-H(1) in the newly diagnosed ALL group was significantly higher than that in the control (p<0.01) and the remission groups (p<0.01). There was no difference in the nm23-H(1) expression between the remission and the control groups. The expression of nm23-H(1) in the relapse group was significantly higher than that in the control (p<0.01) and the remission groups (p<0.01), and similar to that in the newly diagnosed ALL group. The three children with refractory ALL had higher nm23-H(1) expression. Both the positive rate and expression of nm23-H(1) in children with T-lineage ALL were higher than in children with B-lineage ALL (p<0.05).</p><p><b>CONCLUSIONS</b>The expression level of nm23-H(1) varies with the stages of ALL. Newly diagnosed, relapsed and refractory ALL children have higher nm23-H(1) expression. High nm23-H(1) expression may be associated with a poor prognosis and relapse. A higher expression of nm23-H(1) in children with T-ALL may be contributed to a low remission rate and a poor prognosis.</p>

[Expression of Nm23 marker in benign and malignant thyroid neoplasms immunohistochemical]

Several prognostic factors affect the outcome of thyroid carcinomas including tumor stage and distant metastases. Nm23 is a metastasis suppressor gene and has a crucial role in the control of metastatic potential of several carcinomas. The aim of our study is to evaluate expression of nm23 marker in benign and malignant thyroid neoplasms using the immunohistochemistry method and to elucidate its relationship with tumor size, vascular or capsular invasion and lymph node involvement. In a descriptive study, 200 paraffin blocks comprising of 38 benign and 162 malignant thyroid neoplasms stained with nm23 marker were studied. Cytoplasmic staining in more than 10% of cells was considered as positive. The relationship between nm23 and tumor size, vascular or capsular invasion, lymph node involvement was analysed using SPSS 11.5 software [p=0.05]. There was 40% positive incidence of nm23 in follicular adenoma, 87.5% in hurthle cell adenoma, 67.2% in papillary carcinoma, 66.7% in follicular carcinoma, and 64.7% in medullary carcinoma. In follicular adenoma, frequency of nm23 positive tumors was directly correlated to tumor size [p=0.04]. There are no statistically significant correlation between nm23 and tumor size, vascular or capsular invasion or lymph node involvement in malignant thyroid neoplasms. In papillary and medullary carcinoma, negative predictive value of nm23 for lymphnode involvement was over 80%. Also in follicular carcinoma, sensitivity and negative predictive value of nm23 for vascular invasion were approximately 90%. Lack of significant correlation between nm23 and tumor invasiveness [and probably metastasis] factors, demonstrate that although nm23 is a potentially metastasis suppressor gene, whereas in many other tumors it may play a different role in thyroid neoplasms, a role which necessitates further studies to be conducted

Expression and implication of CD44 and nm23-H1 proteins in laryngeal carcinoma / 临床耳鼻咽喉头颈外科杂志

OBJECTIVE@#To study the expression of CD44 and nm23-H1 gene proteins and their clinical significance in laryngeal carcinoma.@*METHOD@#The expression of CD44 and nm23-H1 proteins were detected by immunohistochemistry method in 40 cases with laryngeal carcinoma, 20 adjacent carcinoma tissues and 12 cases normal laryngeal mucosa tissues.@*RESULT@#The expression of CD44 and nm23-H1 proteins in laryngeal carcinoma were much higher than that in normal laryngeal mucosa. The expression of CD44 protein in laryngeal carcinoma with metastatic lymph node was higher than that in laryngeal carcinoma without metastatic lymph node, but nm23-H1 protein lower. The expression of CD44 protein was positively correlated with the metastasis, clinical staging and pathological classification but not correlated with T classification of laryngeal carcinoma. The expression of nm23-H1 protein was negative correlation with the metastasis and clinical staging of laryngeal carcinoma.@*CONCLUSION@#CD44 and nm23-H1 gene proteins play an important coordinated regulation role in the carcinogenesis, development and metastasis of laryngeal carcinoma and will probably become the key biological marks in the judging and evaluating prognosis of laryngeal carcinoma.

Construction and application of a new prokaryotic expression vector derivative of pBV220 / 生物工程学报

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.

Study of increased sensitivity on Tca8113 cell line to cisplatin by nm23-H1 in vitro / 华西口腔医学杂志

<p><b>OBJECTIVE</b>To study the mechanism of sensitivity variation to cisplatin caused by nm23-H1.</p><p><b>METHODS</b>The samles was divided into two groups: Tca8113 group and Tca8113/nm23-H1 group. Using MTT and flow cytometer, the changes of cell mortality rate, apoptosis and mitochondrial membrane potential were detected. By VG PQ Excell, the changes of the intracellular platinum were detected.</p><p><b>RESULTS</b>In vitro the cell mortality rate and apoptosis were increased in Tca8113/nm23-H1 group, comparing with Tea8113 group. Mitochondrial membrane potential was decreased in Tca8113/nm23-H1 group. The intracellular platinum was increased significantly in Tca8113/nm23-H1 group. This effect could be inhibited by oubain which was an inhibitor of Na+/K+-ATP.</p><p><b>CONCLUSION</b>nm23-H1 can increase the sensitivity of cisplatin on Tca8113 cell line. The mitochondrial membrane potential was decreased by nm23-H1 so that intracellular platinum was increased and finally increased the apoptosis or necrosis.</p>

Study on the relationship between microsatellite alterations of RASSF1A gene and the development of cervical carcinoma / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To explore the relationship between microsatellite alterations of RASSF1A gene and the development of cervical carcinoma, and HPV16 infection.</p><p><b>METHODS</b>Two sites of microsatellite polymorphism of RASSF1A gene were selected, we used polymerase chain reaction (PCR) technique to detect the LOH and MSI of cervical tissues, and to detect the infection state of HPV16.</p><p><b>RESULTS</b>There were significant differences of LOH rates at the two sites between clinical stage and pathological grade (P < 0.05). Significant differences were noted between the cervical carcinomas with lymph node metastasis and those without lymph node metastasis in regard to their LOH and MSI at the two sites ( P < 0.05). The incidence of LOH of RASSF1A gene was higher in HPV16(+) than that in HPV16(-) ( P < 0.05).</p><p><b>CONCLUSION</b>The change of RASSF1A gene is a relatively late event in cervical carcinomas. The detection of the LOH and MSI of RASSF1A gene might be helpful to the early diagnosis and the screening of cervical carcinoma. It might also be useful for predicting the prognosis of cervical carcinoma. Infection of HPV16 and LOH of RASSF1A gene had reacted together in the development of cervical carcinoma.</p>

Influence of nm23-H1 gene silence in K562 cell on its differentiation toward megakaryocyte / 中华血液学杂志

<p><b>OBJECTIVE</b>To construct a stable nm23-H1-knock-down cell model with K562 cell line and study its differentiation toward megakaryocyte.</p><p><b>METHODS</b>Eukaryotic expression vector pSilencer 4.1-CMV-sinm23 expressing siRNA targeting nm23-H1 was transfected into K562 cells with lipofectamine2000. Cells with stably nm23-H1 silence were screened out by G418. Real-time quantitative PCR, immunocytochemistry, western blot were used to confirm the nm23-H1-knock-down K562 model. Cell differentiation capacity was detected by NBT reduction assay. Surface antigen Gp IIb-IIIa (CD41) of knock-down cells treated with phorbol 12-myristate 13-acetate was analyzed by flow cytometry. Western blot was used to detect the ERK1/2 signal pathway after the stimulation of phorbol 12-myristate 13-acetate.</p><p><b>RESULTS</b>Endogenous nm23-H1 was silenced by pSilencer 4.1-CMV-sinm23 and the silence efficiency was up to 75% and 70% in mRNA and protein levels respectively compared with the mock cells. Under phorbol 12-myristate 13-acetate treatment, the knock-down cells displayed a significantly increased differentiation ability toward megakaryocyte compared with control. The NBT reduction values were (0.31 +/- 0.07) and (0.23 +/- 0.05) respectively. Further results revealed that nm23-H1 gene regulating the megakaryocytic differentiation was due in part to the increased ERK1/2 phosphorylation.</p><p><b>CONCLUSIONS</b>A stable nm23-H1-knock-down K562 cell model is successfully constructed. nm23-H1 involves in regulating the megakaryocytic differentiation of K562 cell line.</p>

Expression of nm23 and KAI1 and their clinical significance in gallbladder adenocarcinoma / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To investigate the expression of two tumor metastasis suppressor genes nm23 and KAI1 in gallbladder adenocarcinoma, and their clinicopathological significance.</p><p><b>METHODS</b>Specimens and clinical data from 31 gallbladder adenocarcinoma patients were collected. Histopathological grading and the expression of nm23 and KAI1 were detected by HE and immunohistochemical staining, respectively. All cases were followed up for at least three years.</p><p><b>RESULTS</b>Immunohistochemical staining showed that the positive rate of nm23 and KAI1 proteins was 71.0% (22/31) and 61.3% (19/31), respectively. The positive expression rates of nm23 and KAI1 proteins in the early stage carcinomas were significantly higher than those in the moderate and advanced stage ones (P exact = 0.0051 and P exact = 0.0084), and both had an negative correlation with clinicopathologic stage (P trend = 0.0047 and P trend = 0.0058). There was a significant difference in the expression of nm23 and KAI1 proteins among well, moderately and poorly differentiated carcinomas (P exact = 0.0328 and P exact = 0.0020). The expression of nm23 and KAI1 was positively correlated with histopathological grade (P trend = 0.0086 and P trend = 0.0006). There was also a significant difference in the expression of nm23 and KAI1 proteins between 17 survival and 14 dead patients (P exact = 0.0038 and P exact = 0.0001 ). A synergistic effect of nm23 and KAI1 protein on the survival was observed , and seemed to be more important than any individual gene alone (P exact = 0.0005).</p><p><b>CONCLUSION</b>The expressions of nm23 and KAI1 proteins are negatively correlated with clinical stage, but positively with histopathological grade in gallbladder adenocarcinoma. These two tumor metastasis suppressor genes may act synergistically to inhibit the tumor metastasis.</p>

Efficient purification of recombinant human NDPK-A in pilot-scale / 生物工程学报

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.