OBJECTIVETo investigate the effect of protein kinase CK2 gene silencing on the radiosensitization in human nasopharyngeal carcinoma (NPC) cells and its possible mechanism.

METHODSRNA interference (RNAi) technique was used to down-regulate the protein kinase CK2alpha expression in 5-8F cells, and clonogenic assay was employed to observe the changes in the radiosensitivity of the cells. DNA double-strand break was assessed by immunofluorescence staining of gamma-H2AX foci, and the cell apoptosis was examined using Annexin V-FITC/PI double-staining flow cytometry.

RESULTSCK2alpha protein was successfully silenced by siRNA. CK2alpha knockdown significantly decreased the clonogenic activity and increased the radiosensitivity of the NPC cells. After a 15-min exposure of the cells to 1 Gy radiation, significant difference occurred in the gamma-H2AX foci between CK2alpha knockdown cells and the control cells (P<0.01). CK2alpha silencing significantly increased the cell apoptosis after the exposure (P<0.01).

CONCLUSIONSProtein kinase CK2 plays an important role in the radiosensitivity of the NPC cells, and suppression of its expression might be a potential therapeutic approach of cancer.

Animals ; Annexin A5 ; metabolism ; Casein Kinase II ; deficiency ; genetics ; metabolism ; Cell Line, Tumor ; Histones ; genetics ; Humans ; Nasopharyngeal Neoplasms ; genetics ; pathology ; RNA Interference ; RNA, Small Interfering ; genetics ; Radiation Tolerance ; genetics ; Transfection

OBJECTIVETo investigate the expression of casein kinase 2β (ck2β) in colorectal cancer in relation to the metastatic ability of the cancer cells.

METHODSThe expression of ck2β in 46 normal colorectal mucosa, 20 colorectal adenomas and 66 colorectal cancers were detected immunohistochemically. In colorectal cancer cells, Ck2β protein expression was knockdown by RNA interference using ck2β-specific small interfering RNA (siRNA) and the interference efficiency was assessed by Western blotting. The effect of ck2β gene knockdown on the proliferation of the colorectal cancer cells was tested with colony formation assay, and the changes in the invasive ability of the cells were observed using Transwell chamber assay.

RESULTSNegative or weak ck2β expression was detected in normal colorectal mucosa, with nuclear positivity in 8.7% and cytoplasmic positivity in 13.0% of the cases. Colorectal adenomas showed moderate ck2β expression, with 60% cases showing positivity in the cell nuclei and 40% in the cytoplasm. In colorectal cancers, significantly stronger expression of ck2β was found than that in colorectal adenomas and normal colorectal mucosa (P<0.05), and 75.8% cases showed positivity in the cell nuclei and 62.1% showed cytoplasmic positivity; the expression of ck2β protein in colorectal cancers with lymph node metastasis was even higher (P<0.05). Ck2β knockdown obviously inhibited the proliferation and invasiveness of colorectal cancer cells in vitro.

CONCLUSIONThe high expression of ck2β in colorectal cancer is closely correlated to the carcinogenesis and metastasis of the tumor.

Adult ; Aged ; Casein Kinase II ; metabolism ; Cell Proliferation ; Cell Transformation, Neoplastic ; Colorectal Neoplasms ; metabolism ; pathology ; Female ; Humans ; Intestinal Mucosa ; metabolism ; pathology ; Male ; Middle Aged ; Tumor Cells, Cultured ; Young Adult

BACKGROUND: Proximal humeral fracture is a common disease of fall injury in the elderly, because of bone nonunion after treatment with a variety of factors such as senile osteoporosis. Currently, the use of reverse total shoulder arthroplasty has achieved good clinical effect, but has certain limitations. OBJECTIVE: To compare and observe the clinical effects of reverse total shoulder arthroplasty and open reduction and internal plate fixation in the treatment of nonunion of proximal humeral fractures. METHODS: Totally 120 cases of nonunion of proximal humeral fractures were randomly divided into observation group and control group, with 60 cases in each group. The observation group received reverse total shoulder arthroplasty (replacement of artificial shoulder joint). The control group received open reduction and internal plate fixation. RESULTS AND CONCLUSION: (1) Follow-up results: At 3 years after surgery, the pain score was lower in the observation group than that in the control group (P < 0.05). Constant daily activities, range of activities, strength test score, Constant total score, satisfaction and hospitalization expenses were higher in the observation group than in the control group. Functions of flexion, laterotorsion and intorsion were better in the observation group than those in the control group (P < 0.05). (2) Adverse reactions: At 3 years after surgery, 26 and 22 cases had adverse reaction in the observation group and the control group respectively. (3) The results show that the clinical effect of the elders' nonunion of proximal humeral fracture treated with reverse total shoulder arthroplasty is quite good, and the pain degree and shoulder function are obviously improved. The curative effect of reverse total shoulder arthroplasty is better than that of open reduction and internal plate fixation.

OBJECTIVE: To determine the effect of activation of specific anti-tumor cytotoxic T lymphocytes (CTL) and the ability of cross-presentation in vitro by fusion of HLA-A2+ human dendritic cells (DCs) with HLA-A2- melanoma cells. METHODS: The HLA-A2+ human dendritic cells and HLA-A2- melanoma cells were fused by PEG and were cultivated in complete RPMI1640 media containing FCS (10%) and GM-CSF for 24-48 h, and then co-cultured fusion cells with Melan-A specific T cells. HLA-A2- melanoma cells were negative control,While T2 cells and DC+Pts were positive control. The activation of anti-tumor CTL elicited by the fusion cells was detected by intracellular cytokine staining. RESULTS: The immature DC could express CD80, CD83, CD86, HLA-DR, and HLA-ABC,but the mature DC induced by TNF-alpha, PGE-2, and CD40L further highly expressed above molecules. The rate of specific CTL cells primed by the fusion cells was 16.72%+/-4.26%, negative control was 0.21%+/-1.84%,and positive control was 28.60%+/-5.67%. The CTL from vaccine by fusing DC and LAR6 induced lysis of HLA-A2+ LAR1 cells. CONCLUSION The HLA-A2 restricted specific anti-tumor CTL can be induced in vitro by fusion of HLA-A2+ human dendritic cells with HLA-A2- melanoma cells.
Antigen Presentation ; immunology ; Antigens, Neoplasm ; immunology ; Cancer Vaccines ; immunology ; Cell Fusion ; Cell Line, Tumor ; Dendritic Cells ; cytology ; immunology ; HLA-A2 Antigen ; immunology ; Humans ; MART-1 Antigen ; immunology ; Melanoma ; immunology ; pathology ; T-Lymphocytes, Cytotoxic ; immunology

OBJECTIVETo construct a recombinant lentiviral vector for p38 MAPK and establish a human prostatic carcinoma cell line that stably expresses p38 MAPK.

METHODSEGFP/p38 fusion gene was subcloned into the lentiviral vector pTYF- EF1α-IRES-EGFP. The recombinant lentiviral vector pTYF-EF1α-EGFP/p38 was indentified by restriction enzyme digestion, and packaged in HEK 293T cells using lipofectamintm2000 with the packaging plasmid psPAX2 and envelope plasmid pMD2.G. The viral titer was tested according to the expression level of GFP. The resulting recombinant lentiviral vector was transduced into human prostatic carcinoma DU145 cells, and stably transduced cells were selected by limiting dilution analysis. The intracellular expression level of total p38 was detected by Western blotting and the cell growth curve was drawn.

RESULTSDNA restriction enzyme digestion demonstrated that the recombinant lentiviral vector of the fusion gene EGFP/p38 (pTYF-EF1α-EGFP/p38) was constructed successfully. The recombinant lentiviral vector was packaged in 293T with a viral titer of 4.7×10(6) TU/ml. A stable cell line, EGFP/p38-DU145, was established, which stably expressed exogenous EGFP/p38 MAPK fusion protein as detected by Western blotting and showed a lowered growth rate compared to the control cells.

CONCLUSIONWe have successfully constructed a recombinant lentiviral vector of the fusion gene EGFP/p38 and established a stable cell line EGFP/p38-DU145. Overexpression of p38 has a significant inhibitory effect on the proliferation of DU145 cells in vitro.

Cell Line, Tumor ; Cloning, Molecular ; Genetic Vectors ; genetics ; Green Fluorescent Proteins ; genetics ; HEK293 Cells ; Humans ; Lentivirus ; genetics ; metabolism ; Male ; Prostatic Neoplasms ; metabolism ; pathology ; Recombinant Proteins ; biosynthesis ; genetics ; Transfection ; p38 Mitogen-Activated Protein Kinases ; biosynthesis ; genetics

Glioblastoma(GBM)is a lethal cancer with limited therapeutic options.Dendritic cell(DC)-based cancer vaccines provide a promising approach for GBM treatment.Clinical studies suggest that other immu-notherapeutic agents may be combined with DC vaccines to further enhance antitumor activity.Here,we report a GBM case with combination immunotherapy consisting of DC vaccines,anti-programmed death-1(anti-PD-1)and poly I:C as well as the chemotherapeutic agent cyclophosphamide that was integrated with standard chemoradiation therapy,and the patient remained disease-free for 69 months.The patient received DC vaccines loaded with multiple forms of tumor antigens,including mRNA-tumor associated antigens(TAA),mRNA-neoantigens,and hypochlorous acid(HOCl)-oxidized tumor lysates.Furthermore,mRNA-TAAAs were modified with a novel TriVac technology that fuses TAAs with a destabilization domain and inserts TAAs into full-length lysosomal associated membrane protein-1 to enhance major histo-compatibility complex(MHC)class Ⅰ and Ⅱ antigen presentation.The treatment consisted of 42 DC cancer vaccine infusions,26 anti-PD-1 antibody nivolumab administrations and 126 poly I:C injections for DC infusions.The patient also received 28 doses of cyclophosphamide for depletion of regulatory T cells.No immunotherapy-related adverse events were observed during the treatment.Robust antitumor CD4+and CD8+T-cell responses were detected.The patient remains free of disease progression.This is the first case report on the combination of the above three agents to treat glioblastoma patients.Our results suggest that integrated combination immunotherapy is safe and feasible for long-term treatment in this patient.A large-scale trial to validate these findings is warranted.