Arthritis, Gouty ; diagnostic imaging ; pathology ; surgery ; Foot Joints ; diagnostic imaging ; pathology ; surgery ; Hand Joints ; diagnostic imaging ; pathology ; surgery ; Humans ; Male ; Middle Aged ; Radiography

Objective To prepare hepatocyte growth factor(HGF) recombinant protein and confirm its activity preliminarily according to building HGF gene prokaryotic expression vector and transforming into E.coli.Methods Clone HGF inserted into the vector pET-26b(+) to construct prokaryotic expression vector pET-26b(+)-HGF and transform into E.coli Rosseta(DE3).The transformed bacteria induced by IPTG was purified through Ni-NTA resin affinity chromatography frozen-drying after renaturation.Results HGF gene recombinant prokaryotic expression vector pET-26b(+)-HGF was constructed successfully.E.coli Rosseta(DE3) which was transformed into pET-26b(+)-HGF expresses the target protein as the form of inclusion bodies,accounting for 38％ of the total bacterial proteins,and confirmed by Western blot.HGF protein which was purified by Ni-NTA resin affinity chromatography,has a purity of about 95％,and can promote proliferation,migration,and inhibition of apoptosis for human non-small cell lung cancer cell line A549 cells after interaction.Conclusion HGF gene recombinant prokaryotic expression vector pET-26b (+)-HGF was constructed and expressed in transformed E.coli Rosseta(DE3) successfully.They resumed their recombinant HGF protein structure after purification and renaturation,and had biological activity confirmed by in vitro studies.

OBJECTIVETo explore the mechanism of miR-124 inhibiting the proliferative activity of prostate cancer PC3 cells.

METHODSLuciferase reporter gene assay was used to examine the specific binding ability of miR-124 to PKM2 mRNA 3'-UTR. After miR-124 was transfected mimic to PC3 cells, the expression levels of PKM2 mRNA and protein were detected by real-time fluorescence quantitative PCR (qRT-PCR) and Western blot, respectively. The effects of miR-124 mimic and PKM2 siRNA on the proliferative activity of the PC3 cells were determined by MTT assay.

RESULTSThe expressions of PKM2 mRNA and protein were upregulated (5.12 +/- 0.35) times and (4.05 +/- 0.20) times respectively in the PC3 cells as compared with those in the RWPE-1 cells (P < 0.05). Luciferase reporter gene assay demonstrated that miR-124 targeted PKM2 3'-UTR. At 24 hours after transfection with miR-124 mimic, the PKM2 protein expression in the PC3 cells was downregulated (0.16 +/- 0.04) times (P < 0.05), while the PKM2 mRNA level was not changed significantly (P > 0.05), as compared with the control group. MTT assay showed that both miRNA-124 mimic and PKM2 siRNA could inhibit the proliferation of the PC3 cells, but the former exhibited a greater inhibitory effect than the latter. After transfection with miR-124 mimic and PKM2 siRNA, the cell growth rates were (66.20 +/- 5.10)% vs (82.10 +/- 6.35)% at 24 hours (P < 0.05) and (49.34 +/- 2.37)% vs (70.10 +/- 5.80)% at 48 hours (P < 0.05).

CONCLUSIONmiR-124 can suppress the proliferation of PC3 cells by regulating the PKM2 gene.

Carrier Proteins ; genetics ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; genetics ; Humans ; Male ; Membrane Proteins ; genetics ; metabolism ; MicroRNAs ; genetics ; Prostatic Neoplasms ; genetics ; metabolism ; pathology ; Thyroid Hormones ; genetics ; metabolism ; Transfection

AIMTo study the antitumor effect of baicalein on human leukemia K562 cell and its mechanism.

METHODSThe IC50 value and cytotoxity of K562 cell were detected by MTT method. The apoptotic cell was analyzed by FCM using Annexin V FITC--PI staining method. Sub-G1 peak was also measured by FCM. Protein expressions of Bcl-2, Fas, Caspase 3 were evaluated with FCM.

RESULTSBaicalein was shown to significantly inhibit the proliferation of K562 cell in a dose-dependent manner and selectively induce apoptosis of human leukemia K562 cells. Flow cytometric analysis showed that baicalein arrested K562 cells in the S phase. In addition, protein expression of Fas, Caspase 3 of K562 cells increased after exposure to baicalein, but Bcl-2 was unchanged.

CONCLUSIONBaicalein can selectively induce apoptosis of human leukemia K562 cell dose and time dependently through up-regulation of caspase-3 and fas gene expression level.

Antineoplastic Agents, Phytogenic ; pharmacology ; Apoptosis ; drug effects ; Caspase 3 ; Caspases ; metabolism ; Cell Cycle ; drug effects ; Flavanones ; Flavonoids ; pharmacology ; Humans ; K562 Cells ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Up-Regulation

OBJECTIVETo detect the expression of epidermal fatty acid-binding protein (E-FABP) and fatty acid synthase (FAS) in human breast cancer and identify the potential markers and therapeutic targets for breast cancer.

METHODSFAS and E-FABP expressions were detected in 76 patients with infiltrating ductal breast carcinoma using RT-PCR, immunohistochemistry and Western blotting. The possible associations of the expression of the two proteins with the major clinicopathological factors were analyzed.

RESULTSE-FABP and FAS expression levels were significantly decreased (P<0.05) in grade III as compared with grades I and II infiltrating ductal breast carcinoma. There was a positive correlation between E-FABP and FAS expressions, but their expressions were not correlated to the clinicopathological factors of the patients except for the tumor grades. High E-FABP expression level in grades I and II tumors were associated with an early increased responsiveness to FAS.

CONCLUSIONThe variation of the E-FABP and FAS expressions in the lesions is associated with increase of the risk for breast cancer, and the results of this study provide evidence for developing new molecular markers of high-risk lesions and identifying new the targets for breast cancer therapy.

Adult ; Aged ; Biomarkers, Tumor ; biosynthesis ; genetics ; Blotting, Western ; Breast Neoplasms ; genetics ; metabolism ; pathology ; Carcinoma, Ductal, Breast ; genetics ; metabolism ; pathology ; Fatty Acid Synthases ; biosynthesis ; genetics ; Fatty Acid-Binding Proteins ; biosynthesis ; genetics ; Female ; Humans ; Immunohistochemistry ; Middle Aged ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo investigate the expression of extracellular matrix metalloproteinase induced (EMMPRIN) in the interface tissue, and explore the role of EMMPRIN in the aseptic loosening of prostheses.

METHODSImmunohistochemistry was performed to characterize the EMMPRIN-expressing cells at sites of interface tissue around aseptic loosened hip prostheses in 16 cases. Reverse transcription-polymerase chain reaction (RT-PCR) was performed to study the existence of EMMPRIN mRNA in interface tissue samples. And it was followed up by computer assisted image analysis in order to detect the A values of their expression. Synovium of hip joint of 8 femoral neck fracture were in control group.

RESULTSStrong immunostaining of EMMPRIN was found in the macrophages and fibroblasts of lining-like layers and vascular endothelium of synovial membrane-like interface tissue around loosened prostheses. Expression of EMMPRIN was significantly higher in interface tissue than the control synovium (z=-3.252, P=0.001). RT-PCR of interface tissue samples disclosed the presence of EMMPRIN mRNA of 14 cases. In interface tissue, the A value of EMMPRIN increased significantly compared to control synovium (P<0.01).

CONCLUSIONOver-expression of EMMPRIN up-regulates the production of matrix metalloproteinase (MMPs) in the interface tissue. And it can promote the bone destruction around prostheses. Thereby it may be one of methods to prevent and treat aseptic loosening of prostheses by repression the biology activity of EMMPRIN.

Adult ; Aged ; Arthroplasty, Replacement, Hip ; Basigin ; genetics ; physiology ; Female ; Hip Prosthesis ; Humans ; Immunohistochemistry ; Male ; Matrix Metalloproteinases ; metabolism ; Middle Aged ; Osteolysis ; physiopathology ; Prosthesis Failure ; RNA, Messenger ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Synovial Membrane ; metabolism

This study was aimed to investigate the expression level of NF-kappaB mRNA in acute myeloid leukemia (AML) using real time fluorescence quantitative polymerase chain reduction (qPCR) detection and to explore the effect of NF-kappaB mRNA in pathogenesis of AML. The fresh bone marrow was collected from 60 newly diagnosed patients with AML, the total RNA was extracted by means of RTIzoL, the cDNA was synthesized, the expression of NF-kappaB mRNA was detected by qPCR using GAPDH as internal reference. 12 normal healthy persons were selected as controls. The results showed that the expression of NF-kappaB mRNA in patients with AML was higher than that in normal healthy persons with significant difference (p<0.05), the expression of NF-kappaB mRNA in patients with AML-M4 and -M5 were higher than that in patients with AML-M1, -M2 and -M3. It is concluded that the expression of NF-kappaB mRNA is higher in patients with AML. The up-regulation of NF-kappaB expression in patients with AML may play an important role in pathogenesis of AML.
Adolescent ; Adult ; Aged ; Bone Marrow ; metabolism ; Case-Control Studies ; Female ; Humans ; Leukemia, Myeloid, Acute ; genetics ; metabolism ; pathology ; Male ; Middle Aged ; NF-kappa B ; genetics ; metabolism ; RNA, Messenger ; genetics ; Young Adult

OBJECTIVETo explore the relationship between hypoxia and the apoptosis of corpus cavernosum smooth muscle cells (CCSMC) in SD rats.

METHODSCCSMCs were cultured in vitro and identified by immunohistochemistry, and then underwent hypoxia interference at the concentration of 1% O2 for 12, 24, 48 and 72 hours, with normal oxygen concentration as the control. Flow cytometry was used to determine the cycles and apoptosis of the cells.

RESULTSThe cultured CCSMCs grew well, positive for anti-smooth muscle alpha-actin monoclonal antibody immunohistochemical staining. Flow cytometry showed that the number of CCSMCs in G0/G1 was gradually increased within 48 hours and then decreased, just opposite to the proportion of the S phase cells. But no regular change was found in the proportion of the cells in the G2/M phase.

CONCLUSIONHypoxia promotes the apoptosis of CCSMCs in a time-dependent manner, to the maximum at 48 hours, and then cell lysis may occur, but with no further apoptosis.

Animals ; Apoptosis ; Cell Hypoxia ; Cells, Cultured ; Male ; Muscle, Smooth, Vascular ; cytology ; Penis ; pathology ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo evaluate the effects of radiotherapy in the treatment of small-cell lung cancer and the optimal radiation doses, irradiation volume and fractionations.

METHODSUsing evidence-based principles to search and evaluate clinical evidence on radiotherapy of small-cell lung cancer and giving grades of recommendation in clinical practice.

RESULTS AND CONCLUSIONThe combination of chemotherapy and thoracic irradiation were the treatment strategy in limited stage small-cell lung cancer (LD SCLC). There were no clear answers on optimal irradiation dose and volume. Early thoracic irradiations were better than later ones. Radiotherapy should be started at the first or second cycle of chemotherapy. Hyperfractionated irradiation may have therapeutic benefit compared with conventional irradiation. Prophylactic cranial irradiation could improve survival for patients with complete response after chemotherapy and radiotherapy.

Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Carcinoma ; drug therapy ; radiotherapy ; Combined Modality Therapy ; Evidence-Based Medicine ; Humans ; Lung Neoplasms ; drug therapy ; radiotherapy ; Radiation Dosage

OBJECTIVETo explore the effects of overexpression of human tissue inhibitors of metalloproteinase-1 (hTIMP-1) on proliferation, invasion, metastasis, angiogenesis, and apoptosis in human hepatocellular carcinoma (HCC) cells in vitro and in vivo.

METHODSRecombinant adenoviral vector containing hTIMP-1 (AdhTIMP-1) was constructed previously. HepG2 cells were infected by AdhTIMP-1 and the changes of cell proliferation and invasion were detected in vitro. The anticancer activity of AdhTIMP-1 was evaluated in BAL B/c mice bearing HCC. Tumor volume and pulmonary metastases were observed. The mechanisms underlying the antitumor effect in vivo were investigated based on detection of microvessel density and apoptosis in tumor tissues.

RESULTSThe resultant AdhTIMP-1 was successfully constructed and the expression of hTIMP-1 was detected by Western blot and RT-PCR. AdTIMP-1 could effectively infect HepG2 cells and significantly inhibit the proliferative activity and invasive ability of the tumor cells. Compared with the controls, pre-infection of HepG2 cells by AdhTIMP-1 resulted in a significant inhibition of tumor formation by 75. 8%. A single local injection of AdhTIMP-1 into pre-established tumors significantly reduced the tumor growth rate by 45.4%, tumor-associated angiogenesis index by 47.8%, lung metastases by 70.4%, and showed a 3-fold increase of apoptotic tumor cells.

CONCLUSIONOur data indicated that AdhTIMP-1 can significantly attenuate tumor proliferation and invasion, reduce metastasis, inhibit angiogenesis, and induce apoptosis in HCC-bearing mice and may pave the way for further liver cancer gene therapy.

Adenoviridae ; genetics ; Animals ; Apoptosis ; Cell Line, Tumor ; Cell Proliferation ; Female ; Genetic Vectors ; Humans ; Liver Neoplasms ; metabolism ; pathology ; Lung Neoplasms ; prevention & control ; secondary ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Neoplasm Transplantation ; Neovascularization, Pathologic ; prevention & control ; Plasmids ; Random Allocation ; Recombinant Proteins ; genetics ; metabolism ; Tissue Inhibitor of Metalloproteinase-1 ; genetics ; metabolism ; Transfection ; Tumor Burden