OBJECTIVETo explore the effect of Jinwu Jiangu Recipe (JJR) on the expression of synovial cells' nuclear factor-kappaB (NF-kappaB) and serum interleukin 17 (IL-17) in collagen induced arthritis (CIA) rats.

METHODSTotally 60 Wistar rats were randomly divided into 6 groups, i.e., the blank control group, the model group, high, middle, and low dose JJR treatment groups, and the tripterygium control group, 10 in each group. Except rats in the blank control group, CIA model was established in rats of the rest 5 groups. Then they were treated from the 7th day of modeling. After 4 weeks of medication they were sacrificed, serum collected, and synovium of joints were isolated. The expression of serum IL-17 was detected in synovium of joints by enzyme linked immunosorbent assay (ELISA). And the expression of NF-kappaB/P65, Ikappabetaalpha and NF-KappaB/P50 were detected by Western blot.

RESULTSCompared with the blank control group, the serum IL-17 level increased in the model group (P <0. 01). Compared with the model group, the serum IL-17 level obviously decreased in high and middle dose JJR groups and the tripterygium control group (P < 0.01). Results of Western blot showed, when compared with the blank control group, protein activities of NF-kappaB/P65 and NF-kappaB/P50 were significantly enhanced in the model group (P < 0.01). Compared with the model group, protein activities of NF-kappaB/P65 and NF-kappaB/P50 significantly decreased in high and middle dose JJR groups and the tripterygium control group (P < 0.05, P < 0.01). All indices mentioned above were higher in the low dose JJR group than in the tripterygium control group (P < 0.05, P < 0.01).

CONCLUSIONJJR could lower the expression of serum IL-17 in CIA model rats, and inhibit protein activities of NF-kappaB/P65 and NF-kappaB/P50.

Animals ; Arthritis, Experimental ; drug therapy ; metabolism ; Drugs, Chinese Herbal ; chemistry ; Interleukin-17 ; blood ; NF-kappa B ; metabolism ; Random Allocation ; Rats ; Rats, Wistar ; Synovial Membrane ; drug effects ; metabolism ; Tripterygium ; chemistry

Objective To determine whether morphine having the ability to influence the antiviral effect of lamivudine(3TC)in vitro study.Methods MT2 cells were randomly assigned into morphine+3TC treatment group,morphine+naloxone+3TC treatment group,naloxone+3TC treatment group.Both 3TC and virus control groups were set up.The corresponding MT2 cells were treated with opiates antagonist(naloxone)for 0.5 hours before the 24-hours morphine treatment program was implemented while all of the groups were then infected with equal amounts of cell-free HIV-1 ⅢB strain and 3TC.HIV-1 p24 antigen in culture supernatants collected at days 3,4,5 and 6after infection status was tested and the inhibition of 3TC anti-HIV-1 p24 antigen of various treatment groups calculated.Results Inhibition of 3TC anti-HIV-1 p24 antigen of Morphine+3TC treatment group was the lowest when HIV-1 infected cells at 3rd and 4th day and showed significant difierence (P<0.05)when compared to the 3TC control.However,there was no statistically significant difference among them(P>0.05),when virus was infected the cells at 5th and 6th day.The difference of 3TC anti-HIV-1 p24 antigen inhibition between the morphine+naloxone+3TC treatment group and the naloxone+3TC treatment group was not significant(P>0.05).Similar results were obtained when these two groups were compared to the 3TC control group(P>0.05),respectively.The 3TC anti-HIV-1 p24 antigen inhibition of each treatment group reduced as the time of infection prolonged,showing a significant and time-course effbct.Conclusion The 3TC antiviral effect was reduced by morphine in the early stage of infection,and could be blocked by naloxone.

OBJECTIVETo explore the relationship between epidermal growth factor receptor (EGFR) gene expression and radiosensitivity of non-small-cell lung cancer (NSCLC) cells.

METHODSEGFR sequence-specific double-stranded RNA (dsRNA-EGFR) was chemically synthesized. NSCLC cell line SPC-A1 was transfected with dsRNA-EGFR formulated with Lipofectamine 2000. Western blot and real-time PCR were used to determine the EGFR mRNA and protein expression, respectively. Colony inhibition test was adopted to observe the radiosensitizing effect. To establish the nude mouse tumor models, calculate the tumor growth inhibition rate and make the tumor growth curve by measuring its size and weight.

RESULTSEGFR mRNA levels were 1.51 ± 0.22, 1.38 ± 0.15 and 0.45 ± 0.11 in the control group, dsRNA-unrelated group and dsRNA-EGFR group, respectively (F = 482.7, P < 0.01). The contents of EGFR protein were 2340.87 ± 10.99, 2231.85 ± 35.66 and 832.03 ± 39.13 in the control group, dsRNA-unrelated group and dsRNA-EGFR group, respectively (F = 263.3, P < 0.05). Compared with the control group, dsRNA-EGFR sequence specifically decreased the expressions of EGFR mRNA by 70.2% and EGFR protein by 64.5%. The colony inhibition rates of the control group, dsRNA-unrelated combined with radiotherapy group and dsRNA-EGFR combined with radiotherapy group were 9.3%, 12.5% and 65.5%, and the tumor growth inhibition rates were 21.3%, 24.4% and 64.2%, respectively. The combination of dsRNA-EGFR and radiotherapy significantly inhibited the tumor growth in vitro and in vivo.

CONCLUSIONSDsRNA-EGFR shows an apparent inhibitory effect on the expression of EGFR mRNA and protein of NSCLC cells, effectively inhibit the tumor growth in vivo, and enhance the radiosensitivity of NSCLC.

Adenocarcinoma ; metabolism ; pathology ; radiotherapy ; Animals ; Cell Line, Tumor ; Cell Proliferation ; radiation effects ; Humans ; Lung Neoplasms ; metabolism ; pathology ; radiotherapy ; Male ; Mice ; Mice, Nude ; Neoplasm Transplantation ; RNA, Double-Stranded ; genetics ; RNA, Messenger ; metabolism ; Radiation Tolerance ; Random Allocation ; Receptor, Epidermal Growth Factor ; genetics ; metabolism ; Transfection ; Tumor Burden ; radiation effects

OBJECTIVETo evaluate the psychological situations of patients with soft tissue injuries in oral and maxillofacial region by different kinds of suturing.

METHODSA total of 200 patients were selected and randomly divided into two groups. Group A received intradermic suture while group B underwent para-position suture. All patients were evaluated by hospital anxiety and depression (HAD) scales pre-suture, after one week, one month and three months.

RESULTSThe HAD total scores of group B were significantly high compared with group A (P < 0.05) after one week and one month, while there was no difference between group A and group B pre-suture and three months later.

CONCLUSIONSIntradermic suture results in less psychological influence in patients with soft tissue injuries in oral and maxillofacial region.

Adult ; Anxiety ; Depression ; Face ; surgery ; Humans ; Maxillofacial Injuries ; psychology ; surgery ; Soft Tissue Injuries ; psychology ; surgery ; Suture Techniques

OBJECTIVETo evaluate the cytogenetic and molecular genetic features of chronic myeloid leukemia (CML) in Chinese.

METHODSA total of 1193 CML patients were retrospectively studied. Chromosome preparation of bone marrow cells was made using direct and short-term culture. Karyotype and bcr-abl fusion genes were analyzed by R-banding, RT-PCR, respectively.

RESULTSIn the 1193 cases, 98.07% was Ph chromosome positive (Ph+) and 1.93% negative (Ph-). In the Ph+ patients, 95.64% was classical Ph and 4.36% variant rearrangements. Additional genetic changes were demonstrated in 11.88% of classical Ph cases. Cytogenetic clonal evolution was found in 7.94% of patients in chronic phase (CP), 27.78% in accelerated phase (AP), and 49. 04% in blast crisis (BC). Among the classical Ph cases, +Ph, +8, -21 were found in 14.62%, 10.77% and 7.69% of them respectively. In patients in BC and AP, the most common additional chromosome changes were + Ph (28.57%), +8 (16.67%) and +19 (7.14%), while in CP, -21 (10.26%), +Ph (8.97%), and +8 (8.97%). The combination of +Ph and +8 (3.60%) was the most frequent of combination pattern. 524 cases were investigated for bcr-abl fusion gene, and 54.01% was b3a2 (+) and 27.67% b2a2 (+).

CONCLUSIONIn Chinese CML patients seem to have their unique features in terms of cytogenetic clonal evaluation.

Adolescent ; Adult ; Aged ; Child ; Child, Preschool ; Female ; Fusion Proteins, bcr-abl ; genetics ; Humans ; Karyotyping ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; genetics ; Male ; Middle Aged ; Retrospective Studies

To establish lentivirus-mediated system of double suicide genes and explore its killing effects on K562 cells, lentivirus transfer vector for double suicide genes was constructed using molecular methods, three plasmids of lentivirus gene transfer vector system were transferred into packaging cell line 293T using lipofectine method, the transfer effect was observed through fluorescence microscopy, the lentivirus particles were observed by means of electron microscopy. High titer of lentivirus was harvested from the supernatant of virus-producing cell culture and concentrated by high-speed centrifugation with Poly-L-Lysine (PLL). The K562 cells were infected with the concentrated supernatant containing the virus with the double suicide genes. Fluorescence microscopy and RT- PCR confirmed the integration and expression of extraneous gene. The cytotoxicity to these transgenic cells treated with 5-FC and GCV was measured by MTT assays. The growth inhibition ratio (GIR) of cells and inhibition concentration 50 (IC(50)) were counted. After administration of GCV and 5-FC, the changes of those cells were observed through scanning electron microscope. The results showed that lentivirus transfer vector with double suicide genes was constructed successfully. The above-mentioned plasmids were effectively transferred into 293T cells. So much green fluorescence was observed through fluorescence microscope. A lot of lentivirus particles were observed through transmission electron microscope. Double suicide genes mediated by lentivirus were stably integrated and expressed in K562 cells after infection with the concentrated virus using fluorescence microscopy and RT-PCR. The GIR of K562 cells using GCV or 5-FC was 48.73% or 50.69% respectively and it was apparently higher than that of untransfected cells (P < 0.01). When using GCV and 5-FC together, the GIR was 87.69%, which was apparently higher than that of group using GCV or 5-FC alone (P < 0.01). In conclusion, lentivirus-mediated gene transfer system could transfer CD and TK double suicide genes into K562 cells with high efficiency and it had strong killing effects when giving 5-FC and/or GCV. The cytotoxic effects of double suicide genes were superior to that of single suicide gene. The lentivirus-mediated double suicide gene transfer system is a high-efficiency gene transfer vector.
Cytosine Deaminase ; genetics ; Flucytosine ; pharmacology ; Ganciclovir ; pharmacology ; Genetic Therapy ; Genetic Vectors ; genetics ; HIV-1 ; genetics ; Humans ; K562 Cells ; Thymidine Kinase ; genetics

OBJECTIVETo explore the feasibility and efficiency of cytosine deaminase (CD)/thymidine kinase (TK) gene-modified donor T cells used in allogeneic bone marrow transplantation (allo-BMT) as an approach to mitigate GVHD without compromising engraftment.

METHODSThe pseudotyped lentivirus vectors containing CD and TK double suicide genes were transfected with lipofectine to donor T cells. Lethally irradiated 615 leukemia mice were transplanted with BALB/c bone marrow plus CD(+)TK(+)T cells. GVHD prophylaxis was by administration of ganciclovir (GCV) and 5-Fluoride cytosine (5-FC).

RESULTSThe pseudotyped lentivirus-mediated gene transfer system could efficiently transfer CD and TK double suicide genes into donor T cells. Administration of GCV and 5-FC to the mice could markedly potentiate the CFU-S and CFU-GM yields and raise the number of peripheral white blood cells. 1 x 10(7) CD(+)TK(+) allogeneic T cells caused GVHD of a similar magnitude and time course to that of fresh, naive T cells after allo-BMT. Administration of GCV and 5-FC in mice received CD(+)TK(+)T cells reduced the severity of GVHD and resulted in significantly longer survival as compared with non-administration mice, and the effect was stronger than that of administration of GCV or 5-FC alone.

CONCLUSIONAdministration of CD + TK gene-modified donor T cells to recipient in allo-BMT might be an approach to mitigate GVHD without compromising alloengraftment.

Animals ; Body Weight ; Bone Marrow Transplantation ; Cytosine Deaminase ; genetics ; Female ; Genetic Therapy ; Graft vs Host Disease ; pathology ; therapy ; Lentivirus ; genetics ; Mice ; Mice, Inbred BALB C ; Thymidine Kinase ; genetics ; Transplantation, Homologous

OBJECTIVETo study chemical constituents from rhizome of Daphne papyracea var. crassiuscula.

METHODEthyl acetate fraction of 75% ethanol extracts from rhizome of D. papyracea var. crassiuscula, and its strucutre was identified by spectral method.

RESULTNine compounds were separated and identified as daphneticin (1), daphnetin (2), hydrangetin (3), daphnoretin (4), 1-4'-hydroxyphenyl-5-phenyl-2 (E)-en-1-pentanone (5), daphneolon (6), 3beta-O-acetyl-olean-12-en (7), and (+)-usnic acid (8).

CONCLUSIONCompounds 1-8 were separated from D. papyracea var. crassiuscula for the first time. Compound 8 was separated from the genus for first time.

Chromatography, High Pressure Liquid ; Daphne ; chemistry ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Magnetic Resonance Spectroscopy ; Molecular Structure ; Rhizome ; chemistry

OBJECTIVETo construct the multi-probe ribonuclease protection assay (RPA) template set to be used for detecting expression patterns of MD-2, TLR4, CD14 mRNAs in human peripheral blood mononuclear cells.

METHODSThe designed cDNA fragments of the three genes were generated by polymerase chain reaction (PCR) using specific primers and directionally cloned into EcoR I and Hind III sites of expression plasmid pSP72 containing the T7 promoter, the linearized plasmids was used as template to synthesize anti-sense RNA probes. Then we extracted total RNA from peripheral blood mononuclear cells (PBMC) and detected the dynamic expression patterns of the three genes with RPA method.

RESULTSThe proper sequence and orientation of the template set were confirmed by sequencing and the template set was successfully used to assay TLR4, MD-2 and CD14 mRNAs in human PBMC. The results showed that the three detected genes decreased transiently 1-3 hours after 100 ng/ml LPS stimulation.

CONCLUSIONSThese new RPA multi-probe set provided valuable tool for the simultaneous quantitative determination of expression of TLR4, CD14 and MD-2 mRNAs in both constitutive and inducible types.

Antigens, Surface ; analysis ; Base Sequence ; Biological Assay ; Cells, Cultured ; DNA ; analysis ; genetics ; Gene Expression Profiling ; methods ; Humans ; Lipopolysaccharide Receptors ; analysis ; Lymphocyte Antigen 96 ; Membrane Glycoproteins ; analysis ; Molecular Probe Techniques ; Molecular Sequence Data ; Monocytes ; metabolism ; RNA Probes ; analysis ; genetics ; Receptors, Cell Surface ; analysis ; Receptors, Immunologic ; analysis ; Ribonucleases ; Toll-Like Receptor 4 ; Toll-Like Receptors

The aim of study was to investigate the killing effect of double suicide gene system mediated by retroviral vector on K562 cells in vivo and ex vivo. CDglyTK gene was transfected into PA317 cells by using lipofectamine. K562 cells were infected with viral supernatant. K562/CDglyTK cells were treated with 5-fluorocytosine (5-FC) and/or ganciclovir (GCV). Mice were randomly divided into three groups: tumor formation, tumor inhibition and tumor therapy. Each mouse was implanted with K562/CDglyTK cells or K562 cells. The results indicated that the killing effect of 5-FC in combination with GCV on K562/CDglyTK was more significant than using 5-FC or GCV alone. In vivo study showed that after being injected subcutaneously with K562 cells and K562/CDglyTK cells, there was not obvious difference in tumor formation rate of mice, 5-FC + GCV could suppress tumor formation of the K562/CDglyTK cells. After being treated with 5-FC and GCV, the median tumor volume of mice implanted with K562/CDglyTK cells decreased obviously, compared with the control group. Their median survival was significantly prolonged. It is concluded that double suicide genes are more effective for killing effect on K562 cells in vivo and in ex vivo. It may be applicable to clinical gene therapy.
Cytosine Deaminase ; genetics ; Flucytosine ; pharmacology ; Ganciclovir ; pharmacology ; Genes, Transgenic, Suicide ; genetics ; Genetic Therapy ; Genetic Vectors ; genetics ; Humans ; K562 Cells ; Protein-Tyrosine Kinases ; genetics ; Receptor Protein-Tyrosine Kinases ; biosynthesis ; genetics ; Recombinant Fusion Proteins ; genetics ; Recombination, Genetic ; Retroviridae ; genetics