Adult ; Aged ; Aged, 80 and over ; Extracorporeal Circulation ; Female ; Follow-Up Studies ; Humans ; Kidney Neoplasms ; pathology ; surgery ; Liver Neoplasms ; pathology ; surgery ; Male ; Middle Aged ; Neoplastic Cells, Circulating ; Survival Rate ; Vena Cava, Inferior ; pathology ; surgery

OBJECTIVETo investigate the therapeutical effect of recombinant plasmid containing vasoactive intestinal peptide gene (pcDNA3.1+/VIP) on collagen-induced arthritis (CIA) in rats.

METHODSThe experimental arthritis was induced by intradermal injection of bovine type II collagen emulsified in Freund's adjuvants in male SD rats. The rats then were given intra-articular injection with recombinant plasmid (pcDNA3.1+/VIP). The levels of serum TNF-alpha, IL-4 and IL-2 were detected by Avidin-Biotin Peroxdase Complex-enzyme-linked immunosorbent assay (ABC-ELISA) and the pathological changes in the joint of rats were observed.

RESULTHistological examination showed massive inflammatory infiltration in the joint with destruction of bone and cartilage, while the severity of pathological changes in synovia of VIP-treated rats was markedly reduced. Compared with normal group, the serum TNF-alpha, IL-2 levels of CIA rats were significantly increased (P <0.05) and IL-4 level was decreased (P<0.05). Compared with control and pcDNA3.1+ -treated CIA rats, serum TNF-alpha and IL-2 levels of pcDNA3.1+/VIP-treated rats were decreased and IL-4 level was increased (P<0.05).

CONCLUSIONRecombinant plasmid containing vasoactive intestinal peptide gene (pcDNA3.1+/VIP) can reduce the clinical and histological severity of established CIA and it might be a promising candidate for treatment of rheumatoid arthritis.

Animals ; Arthritis, Experimental ; therapy ; Arthritis, Rheumatoid ; therapy ; Genetic Therapy ; Injections, Intra-Articular ; Male ; Plasmids ; therapeutic use ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Recombinant Proteins ; therapeutic use ; Vasoactive Intestinal Peptide ; biosynthesis ; genetics ; therapeutic use

OBJECTIVETo explore the expression of tyrosine phosphatase containing C-src homology SH-2 (SHP-1 and SHP-2) in benign prostate hyperplasia.

METHODSWith En Vision two-step method, the expression of SHP-1 and SHP-2 was detected in 10 cases of normal prostate tissue, 30 cases of BPH, 20 cases of PIN, 20 cases of high differential Pca and 20 cases of low differential Pca.

RESULTThe expression of SHP-2 in normal group was mainly distributed in the cytoplasm of secretive cells and basal cells, and a little part in the nucleu. In BPH it was distributed equally in the plasm and nucleu. In PIN, high differential Pca and low differential Pca, SHP-2 expressed mainly in nucleu. The average dyeing index of SHP-2 in each group is 0.4, 1.7, 2.1, 2.2 and 2.6. SHP-1 positive expression in normal prostate, BPH, PIN and high differential Pca showed differentiating layer staining in the cytoplasm of secretive cells and basal cells, while not in low differential Pca. The average dyeing index of SHP-1 in each group is 1.8, 1.8, 1.5, 1.2 and 0.4.

CONCLUSIONThere are transformation in signal transduction relation with SHP-1 and SHP-2 in the progress of prostate cell proliferation, differentiation and malignant. The abnormal activation and distribution of SHP-2 might induce prostate reconstruction and hyperplasia, even carcinoma.

Adult ; Aged ; Cell Nucleus ; enzymology ; Cytoplasm ; enzymology ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Prostatic Hyperplasia ; enzymology ; pathology ; Protein Tyrosine Phosphatase, Non-Receptor Type 11 ; metabolism ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; metabolism ; Protein Tyrosine Phosphatases ; metabolism ; SH2 Domain-Containing Protein Tyrosine Phosphatases ; metabolism ; src-Family Kinases ; metabolism

Leaves of Chrysanthemum morifolium were potential medicinal resource. The present study aims to estimate the main bioactive components: total flavonoids (TF), galuteolin (GA), quercitrin (QU), chlorogenic acid (CA) and 3 ,5-O-caffeoylquinic acid ( CQ), which were considered to be the main effective components, in leaves of C. morfolium cultivars in China. The TF content was estimated hy UV-VIS spectrophotometry, while GA, QU, CA, and CQ were quantitatively determined by HPLC. The highest TF content (7. 13% w/w) was found in cultivar Wan Cong (Shexian county). Cultivar Da Bo ( Bozhou county) had the highest GA content (33. 45 mg - g-1); Cultivar Hong Xin (Sheyang county) contained the highest QU content (29.25 mg · g(-1)); Cultivar Chang Ban (Sheyang county) had the highest CA content (13.14 mg ·(-1)). The maximum CQ content (7.35 mg · g(-1)) was observed in culti- r Da Yang ( Tongxiang county). Different cultivars of C. morfolium had significant difference in components, but the leaf and capitulum of C. morifolium. were found to possess similar chemical compositions. The high content of bioactive components in several cultivars suggested the potential utilization of C. morifolium leaves.
China ; Chromatography, High Pressure Liquid ; Chrysanthemum ; chemistry ; growth & development ; Drugs, Chinese Herbal ; analysis ; Plant Leaves ; chemistry

OBJECTIVETo prepare monoclonal antibodies against oh(8)dG and to evaluate the relationship between Hp infection and oxidative DNA damage by detecting oh8dG in gastric mucosa.

METHODSBALB/C mice were immunized with BSA-oh(8)dG conjugate, monoclonal antibodies were prepared by hybridoma technique, the biological characteristics of antibodies were analysed by competitive ELISA, Western blot and immunohistochemistry.

RESULTSTwo strains of hybridoma cell were obtained. ELISA and Western blot indicated that the antibodies were fairly specific for oh(8)dG. In immunohistochemistry,the positive rate of oh(8)dG expression in Hp positive tissues and Hp negative tissues was 55% and 5%, respectively(P<0.01).

CONCLUSIONThe prepared antibodies can specially recognize oh(8)dG and immunohistochemistry with the monoclonal antibodies showed Hp infection can increase oh(8)dG level in gastric mucosa.

Animals ; Antibodies, Monoclonal ; immunology ; Blotting, Western ; Deoxyguanosine ; analogs & derivatives ; analysis ; immunology ; Enzyme-Linked Immunosorbent Assay ; Female ; Gastric Mucosa ; chemistry ; Helicobacter Infections ; diagnosis ; Helicobacter pylori ; Immunohistochemistry ; Mice ; Mice, Inbred BALB C

The aim of this study is to develop monoclonal antibody against human hepatocyte growth factor activator inhibitor 1 (HAI-1) for future study of HAI-1. The cDNA fragments of human hepatocyte growth factor activator inhibitor 1 (HAI-1) were subcloned to construct GST-HAI-1 fusion protein expression vectors. The vectors were transformed into E. coli and fusion protein expression was induced by IPTG. The GST-HAI-1 fusion proteins were separated on preparative SDS-PAGE and recovered by electroelution, and used to immunize BALB/c mice. Hybridomas producing monoclonal antibodies against human HAI-1 were prepared by cell fusion technique and characterized by ELISA, Western Blot and immunohistochemical staining. One hybridoma cell line, ZMC6, was obtained, which produces specific antibody against the expressed GST-HAI-1 fusion protein. The monoclonal antibody recognizes both the membrane-type and secretory-type HAI-1 proteins of colorectal tissue. The successful development of anti-HAI-1 antibody provides a powerful tool for further investigation on HAI-1's function.
Animals ; Antibodies, Monoclonal ; immunology ; Blotting, Western ; Glutathione Transferase ; genetics ; Immunohistochemistry ; Mice ; Mice, Inbred BALB C ; Proteinase Inhibitory Proteins, Secretory ; analysis ; genetics ; immunology ; Recombinant Fusion Proteins ; biosynthesis ; immunology

OBJECTIVETo prepare monoclonal antibodies (McAbs) against human mesenchymal stem cells (hMSCs) and to study their biological characteristics.

METHODSBALB/C mice were immunized with pooled hMSCs. McAbs were prepared by hybridoma technique and their biological characteristics were analyzed by indirect immunofluorescence, immunohistochemistry and flow cytometry.

RESULTFive hybridoma cell lines were successfully established, which secret McAbs specifically against hMSCs. Investigations showed that all these McAb reacted only to hMSCs and had no cross-reaction to other human cells, the relative affinities of 5 McAbs were 1x10(6) (ZUB1), 1x10(5) (ZUB4), 1x10(6) (ZUC3), ZUE12 (1x10(5)) and 1x10(5) (ZUF10), respectively. Isotype analysis showed that ZUB1, ZUE12, ZUF10 against the same isotype, while ZUC3, ZUB4 against other two different isotypes alone. Flow cytometric analysis showed that the positive expression rate of cultured hMSCs was 87.39% (ZUB1), 88.07% (ZUB4), 88.12% (ZUC3), 69.89% (ZUE12) and 83.67% (ZUF10).

CONCLUSIONThe prepared five McAbs can specifically react against hMSCs, which can be used for selection and study of hMCSs.

Animals ; Antibodies, Monoclonal ; biosynthesis ; Antibody Specificity ; Bone Marrow Cells ; cytology ; immunology ; Fluorescent Antibody Technique ; methods ; HL-60 Cells ; Humans ; Hybridomas ; secretion ; Immunoglobulin G ; immunology ; Immunoglobulin M ; immunology ; Mesenchymal Stromal Cells ; cytology ; immunology ; Mice ; Mice, Inbred BALB C ; Rats

OBJECTIVETo study lysosomes involvement in the degradation of ricin A chain.

METHODSA lysosome-targeted singal KFERQ was added to the C terminus of rRTA by DNA recombinant technology. A pKK223.3 expression system in E. coli was used to produce recombinant ricine A chain (rRTA) and rRTA-KFERQ. Recombinant proteins were purified by affinity chromatography using Blue-Sepharose 6B. The cytotoxicity of recombinant proteins was measured by the MTT method.

RESULTSRecombinant RTA-KFERQ was 49.87%, 54.18% and 88.68% less cytotoxic than RTA itself on the three cell lines HEPG2, Hela and A549, respectively.

CONCLUSIONLysosomes can degrade, but not completely inactivate RTA in different cells, suggesting cells may have other degradation pathways for RTA.

Chromatography, Affinity ; Escherichia coli ; genetics ; metabolism ; HeLa Cells ; Humans ; Lung Neoplasms ; pathology ; Lysosomes ; metabolism ; Recombinant Proteins ; genetics ; isolation & purification ; metabolism ; Ricin ; genetics ; metabolism ; Tumor Cells, Cultured

OBJECTIVETo construct the eukaryotic expression plasmid containing mouse vasoactive intestinal peptide(VIP) gene with biological activities.

METHODSVIP cDNA including the sequences of signal peptide was cloned from mouse thymus by RT-PCR, and then inserted into the mammalian expression vector pcDNA3.1 between Hind III and EcoR I restriction sites. COS-7 cells were transfected with pcDNA3. 1-VIP using liposome, the expression of VIP was identified by Western blot and ELISA. Supernatant of transfected cell culture was added to LPS-stimulated macrophages and the TNF-alpha production in cell medium was observed by ELISA.

RESULTSThe cloned VIP cDNA was confirmed by enzyme digestion and DNA sequencing. The expression of VIP was detected in the pcDNA3. 1-VIP transfected COS-7 cells by Western blot and ELISA. The VIP in culture supernatant potently inhibited TNF-alpha production by LPS-induced Macrophages in vitro.

CONCLUSIONThe eukaryotic expression plasmid that expresses biological active murine VIP has been constructed successfully.

Animals ; Base Sequence ; COS Cells ; Cercopithecus aethiops ; Cloning, Molecular ; DNA, Complementary ; genetics ; Eukaryotic Cells ; metabolism ; Mice ; Molecular Sequence Data ; Plasmids ; Recombinant Proteins ; biosynthesis ; genetics ; Recombination, Genetic ; Reverse Transcriptase Polymerase Chain Reaction ; Vasoactive Intestinal Peptide ; biosynthesis ; genetics

OBJECTIVETo determine the association of CD133 expression with the sensitivity to radiotherapy among rectal cancer patients.

METHODSThe clinical data of 32 rectal cancer patients was retrospectively collected for patients who received a short-term preoperative radiotherapy(5 Gy/d,×5 d) from 2008 to 2010. Pretreatment tumor biopsies were immunostained for CD133 expression. Rectal cancer regression grade (RCRG) was used to evaluate the sensitivity of the rectal cancer to preoperative radiotherapy. The correlation of CD133 expression and sensitivity to radiotherapy was analyzed.

RESULTSCD133 differentially expressed in rectal cancer tissue with 17 high expression and 15 low expression. The expression of CD133 was associated with the differentiation of rectal cancer with higher expression of CD133 among poorly differentiated rectal cancers(P<0.05). Among the CD133-high patients, two patients showed 1st RCRG, five patients showed 2nd RCRG and ten patients showed 3rd RCRG. For the CD133-low patients, there were five 1st RCRG, seven 2nd RCRG and three 3rd RCRG. There was a significant association between CD133 expression and sensitivity to radiotherapy (P=0.037). Multivariate logistic regression analysis showed that the expression level of CD133(P=0.027) and the differentiation of rectal cancer(P=0.046) were independent predictive factors for the sensitivity of rectal cancer to radiotherapy.

CONCLUSIONSCorrelation between CD133 expression and sensitivity to radiotherapy of rectal cancer may exist, which may be helpful in predicting the sensitivity of rectal cancer to preoperative radiotherapy.

AC133 Antigen ; Antigens, CD ; metabolism ; Biomarkers, Tumor ; metabolism ; Biopsy ; Combined Modality Therapy ; Glycoproteins ; metabolism ; Humans ; Peptides ; metabolism ; Rectal Neoplasms ; metabolism ; radiotherapy ; Retrospective Studies