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

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

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

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

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

OBJECTIVETo identify protein markers for the early diagnosis of pancreatic cancer by a comparative proteomic method.

METHODSComparative analysis on the pancreatic peripheral blood protein profiling from 20 pancreatic cancer patients, 10 chronic pancreatitis patients and 20 cancer-free controls from May 2007 to September 2008 was carried out by two-dimensional fluorescence electrophoresis (2D-DIGE). Differentially expressed proteins were identified by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS). The significance difference proteins were confirmed by Western-blot.

RESULTSA differentially expressed proteins: complement 3 (C3) was identified. The gray level of C3 in pancreatic cancer tissue, chronic pancreatitis, and normal control group were 1.63 ± 0.28, 0.65 ± 0.13 (t = 11.81, P = 0.00) and 0.88 ± 0.19 (t = 9.93, P = 0.00), respectively. C3 was high expression in pancreatic cancer group compared with normal control group. The expression of C3 was higher in pancreatic cancer group than in chronic pancreatitis group. The high expression of C3 in pancreatic carcinoma was confirmed by Western blot.

CONCLUSIONS2D-DIGE and MALDI-TOF-MS technology is a quick, easy and practical method to screen for specific biomarkers in serum of patients with pancreatic carcinoma. The identified protein C3 in this study may be as specific serum biomarkers of pancreatic carcinoma.

Adult ; Aged ; Aged, 80 and over ; Biomarkers, Tumor ; blood ; Case-Control Studies ; Complement C3 ; analysis ; Early Diagnosis ; Female ; Humans ; Male ; Middle Aged ; Pancreatic Neoplasms ; blood ; diagnosis ; Pancreatitis, Chronic ; blood ; Proteomics ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Two-Dimensional Difference Gel Electrophoresis

OBJECTIVETo isolate and characterize human rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLSs).

METHODSThe synovial membrane tissues were obtained from 4 RA patients, 1 chondroma patient and 1 healthy subject and FLS were isolated by means of tissue culture. The cell morphology was observed by phase-contrast microscope and the cell surface markers were detected by flow cytometry.

RESULTSThe FLSs were successfully cultured from the synovial membrane tissues with good cell homogeneity after the third passage. The FLSs of the 3rd to 7th passages were stable and proliferated actively, followed by slow proliferation and aging since the 8th passage. Flow cytometry showed that the 4th-passage FLSs from the RA patients contained 99.04% CD90(+) cells, 2.73% CD3(+) cells, 0.29% CD3(-)CD19(+) cells, 2.81% CD3(-)CD16(+)CD56(+) cells, 5.89% CD14(+) cells, and 54.17% CD55(+) cells. The presence of interleukin-1 receptor type I (IL-1RI, 158.63-/+20.32 pg/ml) and IL-1beta (4.67-/+0.82 pg/ml) were detected in the cell culture supernatant of the 4th-passage FLSs from the RA patients by enzyme-linked immunosorbent assay ELISA.

CONCLUSIONFLSs from RA patients can be effectively culture by means of tissue culture, and the cultured FLSs show high expressions of CD90, IL-1RI and IL-1beta.

Adult ; Aged ; Arthritis, Rheumatoid ; pathology ; Cell Proliferation ; Cell Separation ; Cells, Cultured ; Female ; Fibroblasts ; pathology ; Humans ; Interleukin-1beta ; metabolism ; Male ; Middle Aged ; Receptors, Interleukin-1 Type I ; metabolism ; Synovial Membrane ; cytology ; pathology ; Thy-1 Antigens ; metabolism

The automatic cleaning machine we have developed, adopts a SCM system in automatic cleaning. The machine has five functions: ultrasonic cleaning, cold or hot water spraying, drying and greasing. The clinical applications show that the machine with a good effectiveness is suitable for the cleaning of many surgical instruments. It also raises working efficiency, cuts down on the cost of repair and maintenance and reduces the injury and infection to nurses caused by manual cleaning, satisfying the needs of clinical applications.
Automation ; instrumentation ; Disinfection ; instrumentation ; Equipment Design ; Surgical Instruments ; standards ; Ultrasonics ; instrumentation

OBJECTIVETo study the influence of distal tibiofibular synostosis on ankle function.

METHODSFrom October 1998 to October 2004, a total of 281 consecutive patients underwent operations because of ankle fractures or distal fractures of the tibia and fibula. Distal tibiofibular synostosis occurred after operation in 8 patients. The duration of follow-up averaged 20.6 months (14-44 months). The ankle function was assessed on the basis of functional rating system described by Mazur.(1)

RESULTSAccording to Mazur's ankle evaluation system, 4 patients achieved an excellent result, 2 a good result and 2 a fair result. The dorsiflexion of the synostosis ankle reduced by 8.26 degrees as compared with that of the contralateral ankle, and there was little influence on the plantar flexion. All the patients had a normal gait.

CONCLUSIONThe distal tibiofibular synostosis after the operation of ankle fractures or distal fractures of the tibia and fibula usually gives rise to few symptoms and needs no specific treatment.

Adult ; Ankle Injuries ; physiopathology ; Ankle Joint ; physiopathology ; Female ; Fracture Healing ; Humans ; Male ; Middle Aged ; Range of Motion, Articular ; Retrospective Studies ; Young Adult