Hematopoietic Stem Cell Transplantation ; Humans ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; therapy ; Male ; Middle Aged ; Splenectomy ; adverse effects

OBJECTIVETo achieve high expression of human renal cell carcinoma-associated antigen G250 in Escherichia coli.

METHODSThe gene fragments encoding the protein obtained by PCR was cloned into prokaryotic expression vector pET32a(+) and expressed in E. coli Rosseta. The immunogenicity of the recombinant protein was evaluated by Western blotting.

RESULTSThe plasmid pET32a(+)/G250 was constructed and expressed in E. coli Rosseta successfully. Western blot analysis showed that the recombinant protein could be specifically recognized by monoclonal antibody M75.

CONCLUSIONEfficient G250 expression is achieved in prokaryotic expression system, which may facilitate further functional study of the protein and its monoclonal antibody preparation.

Antibodies, Monoclonal ; immunology ; Antibody Specificity ; immunology ; Antigens, Neoplasm ; genetics ; immunology ; metabolism ; Biomarkers, Tumor ; genetics ; immunology ; metabolism ; Blotting, Western ; Carbonic Anhydrase IX ; Carbonic Anhydrases ; genetics ; immunology ; metabolism ; Cloning, Molecular ; Escherichia coli ; genetics ; Gene Expression ; Humans ; Recombinant Fusion Proteins ; genetics ; immunology ; metabolism

OBJECTIVETo evaluate the effect of early application of high-volume hemofiltration treatment (HVHF) on the levels of lactic acid, pro-inflammatory cytokines and C-reactive protein (CRP) in plasma, as well as APACHE II score in patients suffering from severe sepsis.

METHODSThirty patients meeting the diagnosis of severe sepsis were enrolled in the trial within 24 hours of insults. The level of lactic acid, interleukin-6 (IL-6) and CRP in plasma were measured before HVHF and at 24, 48 or 72 h following HVHF treatment.

RESULTThe plasma levels of lactic acid and IL-6 decreased significantly at 24 h, 48 h, 72 h after HVHF (P <0.05), while, IL-10 did not differ significantly following HVHF (P>0.05), when compared with that before HVHF.

CONCLUSIONThe early application of HVHF could clear the plasma lactic acid and pro-inflammatory cytokines, and improve the tissue oxygenation in severe sepsis.

APACHE ; Adult ; C-Reactive Protein ; analysis ; Female ; Hemofiltration ; methods ; Humans ; Interleukin-10 ; blood ; Interleukin-6 ; blood ; Lactic Acid ; blood ; Male ; Middle Aged ; Sepsis ; blood ; therapy ; Treatment Outcome ; Young Adult

The Pichia pastoris strain GS115-PreS could produce a high expression level of full-length PreS protein that secreted to the supernatant after methanol induction in the fermentation. The Western blot analysis showed a single band with expected molecular mass of 48kD and that the major component of the particles was the full-length PreS protein (PreS1 + PreS2 + S) and small envelope protein (S) of 48 and 28 kD, respectively. Electron microscopy image showed PreS particles with 30 nm in diameter. The supernatants of the fermentation were desalted and concentrated. Purified PreS protein was obtained by DEAE-SFF anion exchange column chromatography and the PreS particles were obtained by ultracentrifugation and sucrose density gradient. The ELISA assay results proved that both full-length PreS protein and particles showed high immunogenicity and specificity. P/N ratio further demonstrated that the immunogenicity of the particles is higher than the full-length PreS protein.
Hepatitis B Surface Antigens ; biosynthesis ; genetics ; Hepatitis B virus ; immunology ; Humans ; Pichia ; genetics ; metabolism ; Protein Precursors ; biosynthesis ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; immunology ; isolation & purification ; Viral Envelope Proteins ; biosynthesis ; genetics

OBJECTIVE: To construct the recombinant lentivirus RNAi vector, and to determine whether the lentivirus mediated short hairpin RNA (shRNA) can inhibit the tissue factor (TF) expression in endothelial cells. METHODS: Two short hairpin RNAs targeting to human TF were cloned into pENTRTM/U6 plasmid to obtain an entry clone, and the positive clones were verified by sequencing. A recombination reaction was performed between the pENTR/U6 entry construction and pLenti6/BLOCKiTTM-DEST vector, and then the positive clones were confirmed by sequencing. The 293FT cell line was transfected by the above recombined plasmid and lentivirus packing materials, the culture supernatant was harvested, and the virus titer was determined. RT-PCR and ELISA were used to observe the inhibition of TF gene expression after the lentivirus transduction in human umbilical vein endothelial cells. RESULTS: The shRNA sequences targeting to human TF were cloned into the vectors, and an entry clone and an expression clone were constructed successfully, which were proved by sequence determination. Viral particles were packaged in the 293FT cell line, all virus stocks were collected, and the transfection titer was 5*10(5)/transduced unit. RT-PCR and enzyme linked immunosorbent assay demonstrated that the lentivirus stocks could suppress the TF expression in endothelial cells remarkably. CONCLUSION Lentivirus RNAi vectors containing human TF gene are successfully constructed, and lentivirus mediated shRNA can inhibit the TF expression in endothelial cells, which may provide a highly effective method for the prevention and treatment of thrombo-embolic diseases.
Base Sequence ; Down-Regulation ; Endothelial Cells ; cytology ; metabolism ; Genetic Vectors ; genetics ; Humans ; Lentivirus ; genetics ; Molecular Sequence Data ; RNA Interference ; RNA, Messenger ; biosynthesis ; genetics ; RNA, Small Interfering ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; Thromboplastin ; biosynthesis ; genetics ; Umbilical Veins ; cytology

BACKGROUNDAll-trans retinoic acid (ATRA) can influence the tumor cell proliferation cycle, and some chemotherapeutic drugs are cycle specific. In this study, we hypothesize that ATRA can enhance chemotherapeutic drug sensitivity by affecting the cell cycle of tumor cells.

METHODSThe cell cycle of LoVo cells was evaluated using flow cytometry (FCM). Cell viability was analyzed using the MTT assay. The morphologic changes in the treated LoVo cells were measured with acridine orange (AO)/ethidium bromide (EB) staining. Expression of survivin in LoVo cells was analyzed by immunofluorescence assay.

RESULTSAfter LoVo cells were treated with ATRA, the G0/G1 ratio of the tumor cells increased and the cell ratio of S- and G2/M-phase decreased. Viability of the cells decreased significantly after combined treatment with ATRA and 5-fluorouracil (5-FU) or mitomycin c (MMC) and was evaluated by fluorescence microscopy. Expression level of survivin in the tumor cells decreased after ATRA combination treatment.

CONCLUSIONSATRA enhances drug sensitivity of the LoVo cell line to cell cycle-specific agents and inhibits the expression of survivin in LoVo cells. The combination of ATRA and 5-FU or MMC promoted cell apoptosis, and the mechanism involved in apoptosis may be related to inhibition of survivin gene expression.

Cell Line, Tumor ; Colonic Neoplasms ; drug therapy ; metabolism ; Drug Resistance, Neoplasm ; Flow Cytometry ; Gene Expression Regulation, Neoplastic ; drug effects ; Humans ; Inhibitor of Apoptosis Proteins ; Microtubule-Associated Proteins ; genetics ; Neoplasm Proteins ; genetics ; Tretinoin ; pharmacology

OBJECTIVETo investigate the expression and its significance of retinoic acid receptor-beta (RAR-beta) in colorectal cancer.

METHODSRAR-beta was detected by immunohistochemistry methods and carcino-embryonic antigen (CEA) was tested by chemiluminescence immunoassay methods in normal tissues, paracancerous tissues and colorectal cancer tissues of 60 patients with colorectal cancer treated from January 2006 to January 2007. Above-mentioned data, together with the clinicopathological data of these 60 patients, were analyzed to figure out the expression and its significance of RAR-beta in colorectal cancer.

RESULTSThe expression rate of RAR-beta in tumor tissues (48%) was significantly lower than those in both normal tissues (87%) and paracancerous tissues (87%) (P < 0.05). And its expression was also significant lower in patients with lymph node metastasis (32%) and patients with advanced cancer (TNM stage III and IV) (29%) than in those without lymph nodes metastasis (60%) and those with early stage cancer (stage I and II) (69%). There was no significant differences among well, mildly and poorly differentiated cancer tissues. The CEA level rose in 20 patients, and its rising rate was remarkably higher in patients with lymph node metastasis (48%) and in patients with advanced cancer (52%) than those without lymph node metastasis (23%) and in early stage(14%).

CONCLUSIONSThe expression of RAR-beta decreases significantly in cancer tissues in patients with colorectal cancer, which may be related to the carcinogenesis of colorectal cancer; and its decreasing degree is correlated negatively with the lymph node metastasis and advanced clinicopathological stage. The expression level of RAR-beta may be a new prognostic indication of patients with colorectal cancer.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Colorectal Neoplasms ; metabolism ; pathology ; Female ; Humans ; Male ; Middle Aged ; Receptors, Retinoic Acid ; metabolism ; Young Adult

BACKGROUNDBcl-2, the anti-apoptotic protein is overexpressed in the majority of gastric cancers and associated with its pathogenesis. To better understanding of the role of Bcl-2, RNA interference (RNAi) was used to inhibit Bcl-2 expression in the human gastric cancer cells in vitro and in vivo.

METHODSBcl-2 small interfering RNA (siRNA) was transfected into human gastric cancer cells SGC-7901, and Bcl-2 expression was monitored by real-time polymerase chain reaction (PCR) and Western blot. Cell proliferation, apoptosis, and telomerase activity were examined by MTT, flow cytometry, and TRAP assay, respectively. Gastric cancer cells treated with 100 nmol/L Bcl-2 siRNA were subcutaneously transplanted into nude mice and tumor growth was assessed.

RESULTSBcl-2 siRNA significantly inhibited the expression of Bcl-2 in human gastric cancer cells at both mRNA and protein levels in a time- and dose-dependent manner. Bcl-2 siRNA also decreased telomerase activity (by 78.76%) and increased the rate of apoptosis (by 37.47%). SGC-7901 cell growth was also significantly suppressed in vivo and in vitro.

CONCLUSIONSBcl-2 expression knockdown suppressed the growth of gastric cancer cells. Thus, Bcl-2 may play a very important role in carcinogenesis of gastric cancer and its knockdown may offer a new potential gene therapy approach for human gastric cancer in future.

Animals ; Apoptosis ; Cell Line, Tumor ; Cell Proliferation ; Humans ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Proto-Oncogene Proteins c-bcl-2 ; antagonists & inhibitors ; genetics ; RNA Interference ; RNA, Small Interfering ; genetics ; Stomach Neoplasms ; pathology ; therapy ; Transfection

BACKGROUNDLow back pain (LBP) is a major medical and social problem among working populations and is associated with high medical expense, loss of productivity, and disability. The aim of this study is to investigate the prevalence of LBP among soldiers and evaluate the possible causative factors in military training. The results may provide an insight into changes needed in military training that will reduce the occurrence of LBP among soldiers.

METHODSA cross-sectional survey was conducted in a group of young soldiers in China to estimate the prevalence of LBP and evaluate possible causative factors in military training.

RESULTSThe survey was distributed to 1659 soldiers, of whom 1624 responded. LBP was reported by 425 of the 1624 (26.2%) soldiers. The prevalence of LBP was higher in the armored force (51.3%) than in the artillery (27.5%) or infantry (11.9%). A multivariate logical regression analysis identified night training, 5 km cross-country race, and grenade-throwing training as military training risk factors for LBP.

CONCLUSIONSThe relatively high incidence of LBP among soldiers was related to night training, 5 km racing, and grenade throwing. Modifications in these training methods should enhance the health of recruits and lower the incidence of LBP.

Adolescent ; Adult ; Cross-Sectional Studies ; Humans ; Low Back Pain ; epidemiology ; Male ; Military Personnel ; Prevalence ; Risk Factors ; Surveys and Questionnaires ; Young Adult

OBJECTIVETo evaluate the feasibility, safety and short-term outcomes of laparoscopy-assisted distal gastrectomy for advanced gastric cancer.

METHODSFrom January 2007 to June 2008, 135 patients with advanced gastric cancer in the lower or middle stomach were operated, of whom 66 underwent laparoscopy-assisted distal gastrectomy(LADG) with D2 dissection of lymph nodes and 69 received conventional open D2 distal gastrectomy(ODG). Clinical data were recorded and compared between the two groups.

RESULTSThere were no significant differences in age, gender, and TNM staging between LADG and ODG(all P>0.05). All the patients in the LADG group underwent gastrectomy and lymph nodes dissection successfully without conversion to open surgery and no operative deaths occurred. The operative time was significantly longer for the LADG group than for the ODG group[(266.1±55.1) min vs. (223.8±26.8) min)]. The patients in the laparoscopic surgery group had less blood loss[(131.9±88.7) ml vs.(342.3±178.7) ml], earlier recovery of bowel activity[(3.18±1.22) d vs.(4.50±1.59) d], and shorter hospitalization time[(9.20±3.39) d vs. (11.35±4.61) d]. No significant differences were found in the total number of retrieved lymph nodes(25.81±12.53 vs. 27.47±10.28). The morbidity of complications was comparable between two groups(6.1% vs. 15.94%). No mortality and recurrence were observed during a follow-up period of 1-19 months.

CONCLUSIONSLADG with D2 lymph node dissection is a safe and feasible procedure with adequate lymphadenectomy for advanced gastric cancer.

Female ; Gastrectomy ; methods ; Humans ; Laparoscopy ; Laparotomy ; Male ; Middle Aged ; Retrospective Studies ; Stomach Neoplasms ; surgery ; Treatment Outcome