BACKGROUND: Both hydroxyapatite (HA) and large diameter TiO2 nanotubes have excellent biocompatibility, but bone-forming ability of nano-HA (nHA) deposited large diameter TiO2 nanotubes is rarely reported.OBJECTIVE: To evaluate the bone-forming ability of nHA/large-diameter TiO2 nanotube composite coating.METHODS: Large-diameter TiO2 nanotubes were prepared by anodic oxidation method, and then nHA was electrochemically deposited on the surface of TiO2 nanotubes. Preosteoblasts MC3T3-E1 were co-cultured with the nHA/large diameter TiO2 nanotube composite, pure titanium and TiO2 nanotube coatings, respectively. At 0.5, 1, 2 hours after culture, the initial cell adhesion was observed. At 1, 3, 5 day after culture, cell proliferation was assessed. At 2 days after culture, cell morphology was observed. At 3 and 7 days after osteogenic induction, intracellular alkaline phosphatase activity was detected. At 14 days after osteogenic induction, mineralization of extracellular matrix was detected.RESULTS AND CONCLUSION: (1) After 2 hours of culture, the number of adherent cells on the composite coating was significantly lower than that on the TiO2 nanotube coating (P < 0.05), but slightly higher than that on the pure titanium coating with no statistical difference. (2) After 1, 3, 5 days of culture, the cell proliferation on the composite coating was significantly lower than that on the TiO2 nanotube coating (P < 0.05), but slightly higher than that on the pure titanium with no statistical difference. (3) The cells on the pure titanium showed a spindle-shape, while those on the TiO2 nanotube coating processed filopodia. The cells on the composite coating showed polygonal shape with a larger number of filopodia. (4) The intracellular alkaline phosphatase activity of the composite coating group was significantly higher than that of the pure titanium group and TiO2 nanotube group. The trend of mineralization of extracellular matrix was ranked from high to low: the composite coating group > TiO2 nanotube group > pure titanium group. To conclude, the nHA/large diameter TiO2 nanotube composite coating not only has good biocompatibility, but also has the ideal ability to promote bone formation.

OBJECTIVETo clone A73 gene of Epstein-Barr virus (EBV) and examine the variation of its coding sequence.

METHODSA73 coding sequence (CDS) amplified from 7 patients in nasophryngeal carcinoma (NPC) biopsies by RT-PCR was cloned into pGEM-T-Easy vector to construct the recombinant plasmid, which was subjected to sequence analysis in Gen Bank database using Blast software.

RESULTSA73 gene from nasophryngeal carcinoma was successfully cloned into pGEM-T Easy vector. A locus with conversion of A-->C in A73 was found at CDS 45 nt, located in the exonVB157154 nt, which did not result in an amino acid replacement, and the variation frequency was 7/7.

CONCLUSIONThere is a point mutation in A73 CDS of EBV isolated from NPC tissue, which might produce NPC-associated polymorphism with possible involvement in the composition of some subtype of EB virus.

Cloning, Molecular ; DNA, Complementary ; chemistry ; genetics ; Herpesvirus 4, Human ; genetics ; Humans ; Molecular Sequence Data ; Nasopharyngeal Neoplasms ; pathology ; virology ; Open Reading Frames ; genetics ; Point Mutation ; Sequence Analysis, DNA ; Viral Proteins ; genetics

OBJECTIVETo analysis the clinic and genotype in two Chinese patients with Dyskeratosis congenita (DC).

METHODSThe two patients were characterized by mucocutaneous abnormalities (abnormal nails, lacey reticular pigmentation, and oral leukoplakia), bone marrow failure. They were diagnosed with DC. DC genes were amplified by polymerase chain reaction (PCR), including DKC1, TERT, TERC, TINF2, NOP10, NHP2, then DNA sequencing was performed for abnormal exons.

RESULTSAn abnormal peak was found in exon 6 of TINF2 gene of the two patients. DNA sequencing showed a 845G→A transition in TINF2 gene in the two patients.

CONCLUSIONWe should think about DC if the young patients with mucocutaneous abnormalities and marrow failure. TINF2 c.845G→A(R282H) does exist in the two patients. It is reported in China for the first time.

Base Sequence ; Child, Preschool ; DNA Mutational Analysis ; Dyskeratosis Congenita ; diagnosis ; genetics ; Exons ; Female ; Humans ; Infant ; Male ; Telomere-Binding Proteins ; genetics

OBJECTIVETo investigate the impact of hepatic steatosis on virologic response in chronic hepatitis B (CHB) patients treated with pegylated interferon-alpha (PEG-IFNa).

METHODSNinety-six naive patients positive for hepatitis B e antigen (HBeAg) and with biopsy-proven CHB were administered PEG-IFNa-2a or PEG-IFNa-2b for 48 weeks. Virologic response (HBeAg clearance and hepatitis B virus (HBV) DNA less than 5 log10 copies/ml) and biochemical response (alanine transaminase (ALT) normalization) were compared between patients with (n=34) and without (n=62) steatosis.

RESULTSThe HBV DNA titer in the steatosis group was significantly lower than that of the non-steatosis group (6.961.27 vs. 7.541.28 log10 copies/ml; t=2.161, P=0.033). After 48 weeks of PEG-IFNa treatments, there was no significant difference in HBeAg seroconversion or the percentage of undetectable HBV DNA (less than 3 log10 copies/ml) between steatosis and non-steatosis patients. However, the steatosis patients presented with a significantly lower complete response rate (virologic response plus biochemical response) compared to non-steatosis patients (26.5% vs. 48.4%; x² =4.373, P=0.037). Of the 45 CHB patients with undetectable HBV DNA after 48 weeks of treatment, seven did not achieve ALT normalization. The rate of patients with non-biochemical response was significantly higher in the steatosis group than in the non-steatosis group (33.3% vs. 6.67%; P=0.032).

CONCLUSIONHepatic steatosis does not affect the virologic response, but does affect the biochemical response in CHB patients treated with PEG-IFNa for 48 weeks.

Adult ; Antiviral Agents ; therapeutic use ; Fatty Liver ; complications ; pathology ; virology ; Female ; Hepatitis B, Chronic ; complications ; drug therapy ; pathology ; Humans ; Interferon-alpha ; therapeutic use ; Liver ; pathology ; Male ; Polyethylene Glycols ; therapeutic use ; Recombinant Proteins ; therapeutic use ; Young Adult

To investigate the effect of a novel p21-modulating protein WISp39 on proliferation, apoptosis and cell cycle of leukemia cells, the plasmid pLenti6/V5-WISp39 was constructed and transfected into the human myelocytic leukemia cell line-U937 cells. The expression of WISp39 was detected by real-time PCR at 48 hours after transfection, proliferation of U937 cells assayed by CCK-8, apoptosis and cell cycle were determined by flow cytometry. The results showed that plasmid pLenti6/V5-WISp39 could readily enhance the expression of WISp39 in U937 cells. A significant growth inhibition (37.6%) was observed in cells tranfected with pLenti6/V5-WISp39, while the control plasmid pLenti6/V5-lacZ showed little effect on U937 growth. Further analysis revealed that pLenti6/V5-WISp39 did not show obvious apoptosis induction effect, but it could really regulate U937 proliferation via cell cycle modulation. Compared with pLenti6/V5-lacZ, pLenti6/V5-WISp39 resulted in increase of cells in G(0)/G(1) phase by 10% at 48 hours after transfection. It is concluded that the WISp39 gene has no significant apoptosis induction effect on leukemic cells, but it can increase cells at G(0)/G(1) phase via effect on cell cycle, thus inhibiting the U937 proliferation. This result means WISp39 gene can act as a negative modulator on tumour cells.
Apoptosis ; genetics ; Cell Cycle ; Cell Proliferation ; Cyclin-Dependent Kinase Inhibitor p21 ; metabolism ; Humans ; Immunophilins ; metabolism ; RNA, Messenger ; metabolism ; Sincalide ; pharmacology ; Transfection ; U937 Cells

CCL23 is a human CC chemokine with potential suppression effects on both human and murine myeloid progenitor cells both in vitro and in vivo, and only expressed and released by dendritic cells differentiated from monocytes in blood cells. However, recent study has shown that CCL23 was over-expressed in bone marrow and peripheral blood cells from pediatric patients with acute myeloid leukemia (AML). In order to investigate the effects of CCL23 on the development, therapy and prognosis of leukemia, the U937 cells, a leukemic cell strain, were adopted and cultured with rhCCL23 for 72 hours. The cell proliferation and apoptosis rate were detected by Cell Counting Kit-8 and FITC-AnnexinV/PI respectively; the morphologic changes and the expression of CCR1 (the only receptor of CCL23 known by now) were observed during the differentiation process. The results showed that no obvious effect on the proliferation, apoptosis and differentiation of U937 was found by using CCL23 alone (P > 0.05), but cultured in combination with CCL23 and PMA, the differentiation of U937 cells were promoted remarkably, during which the CCR1 expression increased (P < 0.05). It is concluded that CCL23 alone did not inhibit the proliferation and differentiation of U937, while its use in combination with PMA may possess synergistic effect on inducting differentiation of U937 through the increase of receptor CCR1 expression.
Apoptosis ; physiology ; Cell Proliferation ; drug effects ; Cell Transformation, Neoplastic ; drug effects ; Chemokines, CC ; pharmacology ; Humans ; U937 Cells

The aim of study was to explore the potential application of targeting at Toll-like receptors (TLRs) in the immunotherapy of acute myelocytic leukemia, and to investigate the expression of TLR and the effects of TLR 8 agonist ssRNA40/LyoVec on proliferation, apoptosis and cell cycle of U937 cells. The expression of TLR 1 - 9 in U937 cells was detected by using reverse transcription polymerase chain reaction (RT-PCR) and the expression of TLR 8 was assayed by flow cytometry (FCM). The effect of TLR 8 agonist, ssRNA40/LyoVec, at different concentrations on U937 cells proliferation was evaluated by CCK-8, apoptosis and cell cycle were detected by FCM. The results showed that U937 cells expressed TLR 1 - 9. TLR 8 agonist ssRNA40/LyoVec could inhibit the growth of U937 cells both in time-and dose-dependent manner and the inhibitory rate could reach 70%. It also increased the percentage of cells in G(0)/G(1) phase. There was no significant difference in percentage of apoptotic cells between control and treated groups. It is concluded that TLRs including TLR 1 - 9 express on U937 cells and TLR 8 agonist ssRNA40/LyoVec may be able to inhibit the growth of U937 cells, arrest the cells in G(0)/G(1) phase, but have no effect of promoting apoptosis.
Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; Dose-Response Relationship, Drug ; Humans ; Leukemia, Myeloid, Acute ; metabolism ; pathology ; Reverse Transcriptase Polymerase Chain Reaction ; Toll-Like Receptor 8 ; agonists ; Toll-Like Receptors ; metabolism ; U937 Cells

To determine whether the microRNAs (miRNAs) contained in cancer-derived microvesicles (MVs) mirror those of the parental tumor cells, we compared the miRNA expression profiles of MVs derived from their parental hepatocellular carcinoma (HCC) cells. The presence and levels of 888 miRNAs from SMMC-7721 cells and MVs were detected by Agilent miRNA microarray analysis. Four selected miRNAs were verified by real time qRT-PCR. Furthermore, the genes of the miRNAs were bioinformatically identified to explore potential roles of the miRNAs in HCC microenvironment. Our results showed that miRNAs expression profiles of MVs derived from HCC were significantly changed. Of all the miRNAs tested, 148 miRNAs were co-expressed in MVs and SMMC-7721 cells, only 121 and 15 miRNAs were detected in MVs and SMMC-7721 cells, respectively. Among the 148 co-expressing miRNAs, 48 miRNAs had the similar expression level and 6 of them were supposed to be oncogenic or suppressive miRNAs. According to the target prediction by Quantile Algorithm method, these miRNAs may regulate 3831 genes which were closely related to cell cycle, apoptosis and oncogenesis, and 78 were known tumor suppressors or oncogenes. Gene ontology (GO) analysis indicated that 3831 genes were mainly associated with nucleic acid binding, cell death, cell adhesion. MVs containing miRNAs, released into the HCC microenvironment, bear the characteristic miRNAs of the original cells and might participate in cancer progression.
Carcinoma, Hepatocellular ; genetics ; Cell Line, Tumor ; Cell Transformation, Neoplastic ; genetics ; Gene Expression Profiling ; Gene Expression Regulation, Neoplastic ; genetics ; Humans ; Liver Neoplasms ; genetics ; MicroRNAs ; genetics ; Neoplasm Proteins ; genetics ; RNA, Neoplasm ; genetics ; Transport Vesicles ; genetics

OBJECTIVESTo screen and clone the genes encoding hepatocellular carcinoma associated tumor antigens.

METHODSA hepatocellular carcinoma cDNA express library was constructed with ZAP vector and analyzed by serological analysis of recombinant cDNA expression library (SEREX) with sera from autologous and allogenous patients. Monoclonalized positive phage clones were converted into pBK-CMV phagemid forms by in vivo excision. The cDNA inserts were determined by restriction endonuclease digestion with EcoR I and Xho I. The cDNA inserts were sequenced and analyzed with bioinformatics. LIMS1 insert was cut from the clone HCL5-70 and constructed into pQE 31 express vector. The recombinant LIMS1 was expressed in M15 and analyzed with SDS-PAGE and Western blot.

RESULTSFourteen genes were cloned from autologous screening and eleven genes were obtained with allogeneous analysis. One gene, kinectin, was identified in both autologous and allogeneous screening. Eight of the total twenty-four genes were unknown for their functions; the other sixteen genes can be classified into eight groups according to their established or putative function. Recombinant LIMS1 was expressed in M15.

CONCLUSIONThe identification of hepatocellular carcinoma associated tumor antigens provides potential targets for immunotherapy of hepatocellular carcinoma patients and will help in the understanding of the carcinogenesis of hepatocellular carcinoma.

Antigens, Neoplasm ; genetics ; immunology ; Carcinoma, Hepatocellular ; genetics ; immunology ; DNA, Complementary ; genetics ; Gene Expression Regulation, Neoplastic ; Genetic Therapy ; Humans ; Liver Neoplasms ; genetics ; immunology

BACKGROUNDCerebral glucose metabolism changes are always observed in patients suffering from malignant tumors. This preliminary study aimed to investigate the brain glucose metabolism changes in patients with lung cancer of different histological types.

METHODSOne hundred and twenty patients with primary untreated lung cancer, who visited People's Hospital of Zhengzhou University from February 2012 to July 2013, were divided into three groups based on histological types confirmed by biopsy or surgical pathology, which included adenocarcinoma (52 cases), squamous cell carcinoma (43 cases), and small-cell carcinoma (25 cases). The whole body 18F-fluorodeoxyglucose (18F-FDG) positron emission tomography (PET)/computed tomography (CT) of these cases was retrospectively studied. The brain PET data of three groups were analyzed individually using statistical parametric maps (SPM) software, with 50 age-matched and gender-matched healthy controls for comparison.

RESULTSThe brain resting glucose metabolism in all three lung cancer groups showed regional cerebral metabolic reduction. The hypo-metabolic cerebral regions were mainly distributed at the left superior and middle frontal, bilateral superior and middle temporal and inferior and middle temporal gyrus. Besides, the hypo-metabolic regions were also found in the right inferior parietal lobule and hippocampus in the small-cell carcinoma group. The area of the total hypo-metabolic cerebral regions in the small-cell carcinoma group (total voxel value 3255) was larger than those in the adenocarcinoma group (total voxel value 1217) and squamous cell carcinoma group (total voxel value 1292).

CONCLUSIONSThe brain resting glucose metabolism in patients with lung cancer shows regional cerebral metabolic reduction and the brain hypo-metabolic changes are related to the histological types of lung cancer.

Adult ; Aged ; Brain ; metabolism ; Female ; Fluorodeoxyglucose F18 ; Glucose ; metabolism ; Humans ; Lung Neoplasms ; metabolism ; Male ; Middle Aged ; Positron-Emission Tomography