Dendritic cell (DC)s are protessional antigen presenting cells and they have been used for antitumor immunotherapy or cell vaccines. However therapy using DC is restricted because the number of DC available from tissue is very low. Flt3 ligand (FL) has been known as a hematopoietic growth factor that increases proliferation of hematopoietic stem cells and progenitor cells, and recently it showed inducibility of dendritic cell (DC)s and signiticant antitumor effects in vivo. Thus FL will be frequently used for DC induction and antitumor immunotherapy in future. Here we constructed FL plasmid and studied its in vivo effect. FL plasmids were made by cloning of partial FL cDNA into pcDNA3 plasmid, and gene expression and protein producibility of FL plasmid were confirmed in Renca cells transfected with FL plasmid. Mice were injected with FL plasmid (100ug/mouse) three times and 20 days later mouse spleens were harvested for staining and RT-PCR. There were lots of blastogenic cells in the spleen of mice treated with FL plasmid. FL plasmid also induced DEC205, IL-12 and GM-CSF gene expression in mouse splenocyte. All these data suggest FL plasmid may be used for induction of DC and antitumor therapy as DNA adjuvant.
Mice ; Animals

In order to develop an experimental DNA vaccine for the prevention and treatment of hepatitis B virus infection, hepatitis B virus surface antigen (HBsAg) DNA was subcloned into an E. coli-eukaryotic cell shuttle vector and was expressed in the Baculovirus expression system. Intramuscular, intradermal, and intraperitoneal injections of 30 microg of the plasmid DNA expressing HBsAg induced humoral and cellular immune responses in ICR mice. The first IgG antibodies were detected after ten days and specific IgG antibody titers peaked after two months of a single intramuscular DNA injection. Anti-HBs antibody titers gradually increased and peaked at four months following intradermal DNA injection, and in case of intraperitoneal injection they peaked at seven months. Generation of HBs-specific helper T lymphocytes was also investigated through the production of interleukin-2 by T helper cells. Boosting effects of HBs DNA were investigated without much results. In general, DNA-mediated HBs immunization induced humoral and cellular immune responses in mice that appears to simulate immune responses in human during the course of HBV vaccination.
Animal ; Cloning, Molecular ; DNA, Viral/immunology* ; Hepatitis B/prevention & control* ; Hepatitis B Antibodies/immunology ; Hepatitis B Surface Antigens/genetics* ; Hepatitis B Vaccines/immunology* ; Human ; Interleukin-2/biosynthesis ; Male ; Mice ; Mice, Inbred ICR ; Plasmids/immunology* ; Spleen/immunology ; Spleen/cytology ; Vaccination ; Vaccines, DNA/immunology*

CD44 is a glycoprotein expressed in a wide variety of cell types. Recently expression of some alternatively-spliced variants of CD44 transcripts (CD44v) has been suggested to play a potential role in tumor metastasis and the detection of CD44v containing exon 6 to 11 may be helpful for the diagnosis of cancers. Expressions of CD44v containing exon 6 to 11 were investigated in 20 human colorectal cancer samples, peripheral blood leukocytes isolated from colorectal cancer patients, and 4 colorectal cancer cell lines using reverse transcription-polymerase chain reaction and Southern blot analysis. The standard form of CD44 transcripts was expressed in all samples tested. CD44v containing exon 6 to 11 was expressed in 18 cases of colorectal cancers (sensitivity = 90%), 3 out of 4 cell lines, and one normal tissue (specificity = 95%). These results suggest that the expression of CD44v containing exon 6 to 11 can be regarded as tumor specific and that this marker may be helpful for the early diagnosis of colon cancers, if specimens from the early stage are available.
Adenocarcinoma/*genetics ; Adult ; Aged ; Antigens, CD44/*genetics ; Base Sequence ; Blotting, Southern ; Colorectal Neoplasms/diagnosis/*genetics ; DNA Primers ; Electrophoresis, Agar Gel ; Feces/chemistry/cytology ; Female ; Gene Expression Regulation, Neoplastic/genetics ; Human ; Male ; Middle Age ; Molecular Sequence Data ; Polymerase Chain Reaction ; RNA Splicing/*physiology ; RNA, Messenger/analysis ; Tumor Cells, Cultured/*physiology ; Tumor Markers, Biological

Coptis chinensis (CC) is one of the traditional herbs used in Oriental medicine for the treatment of gastrointestinal disorders, anxiety, and insomnia. In this study, neurotrophic and neuritogenic effects of CC on rat pheochromocytoma (PC12) cells were evaluated. Pretreatment of PC12 cells with water extracts of CC (120microgram/ml) produced considerable outgrowth of neurites that is comparable to the effect of nerve growth factor (NGF). Therefore, neurite outgrowth was quantified and expression of NGF mRNA was examined. Furthermore, characteristics of neurites were immunocytochemically confirmed using axon and dendrite-specific antibodies. These results suggest that water extracts of CC contain components that have neurotrophic and neuritogenic properties.
Animals ; Antibodies ; Anxiety Disorders ; Axons ; Coptis* ; Medicine, East Asian Traditional ; Nerve Growth Factor ; Neurites ; PC12 Cells* ; Pheochromocytoma ; Rats ; RNA, Messenger ; Sleep Initiation and Maintenance Disorders ; Water*

No abstract available.
Biopsy* ; Brain* ; Protons*

BACKGROUND: PM is known to induce various pulmonary diseases, including asthma, cancer, fibrosis and chronic bronchitis. Despite the epidemiological evidence the pathogenesis of PM-related pulmonary diseases is unclear. METHODS: This study examined the effects of PM exposure on the secretion of TNF-alpha and IL-1beta in the cultured alveolar macrophages. The cultured primary alveolar macrophages were treated with the medium, PM (5~20 microgram/cm2), LPS (5ng/ml), and PM with LPS for 24h and 48h respectively. ELISA was used to assay the secreted TNF-alpha and IL-beta in the culture medium. Western blotting was used to identify and determine the level of proteins isolated from the culture cells. The cells cultured in the Lab-Tek(R) chamber slides were stained with immunocytochemical stains. RESULTS: PM induced TNF-alpha and IL-1beta secretion in the culturing alveolar macrophages, collected from the SPF and inflammatory rats. However, the effects were only dose-dependent in the inflammatory macrophages. When the cells were co-treated with PM and LPS, there was a significant synergistic effect compared with the LPS in the both cell types. CONCLUSION: PM might be play an important role in the induction and/or potentiation of various lung diseases by oversecretion of TNF-alpha and IL-1beta.
Animals ; Asthma ; Blotting, Western ; Bronchitis, Chronic ; Coloring Agents ; Enzyme-Linked Immunosorbent Assay ; Fibrosis ; Lung Diseases ; Macrophages ; Macrophages, Alveolar* ; Rats ; Tumor Necrosis Factor-alpha*

Urolithiasis and calcium oxalate crystal deposition diseases are still significant medical problems. In the course of nephrocalcin cDNA cloning, we have identified FKBP-12 as an inhibitory molecule of calcium oxalate crystal growth. lambdagt 11 cDNA libraries were constructed from renal carcinoma tissues and screened for nephrocalcin cDNA clones using anti-nephrocalcin antibody as a probe. Clones expressing recombinant proteins, which appeared to be antigenically cross-reactive to nephrocalcin, were isolated and their DNA sequences and inhibitory activities on the calcium oxalate crystal growth were determined. One of the clone lambdagt 11 #31-1 had a partial fragment (80 bp) of FKBP-12 cDNA as an insert. Therefore, a full-length FKBP-12 cDNA was PCR-cloned from the lambdagt 11 renal carcinoma cDNA library and was subcloned into an expression vector. The resultant recombinant FKBP-12 exhibited an inhibitory activity on the calcium oxalate crystal growth (Kd=10(-7) M). Physiological effect of the extracellular FKBP-12 was investigated in terms of macrophage activation and proinflammatory cytokine gene induction. Extracellular FKBP-12 failed to activate macrophages even at high concentrations. FKBP-12 seems an anti-stone molecule for the oxalate crystal deposition disease and recurrent stone diseases.
Animals ; Base Sequence ; Calcium Oxalate/*antagonists & inhibitors ; Carcinoma, Renal Cell ; Crystallization ; DNA, Complementary ; Extracellular Space ; Glycoproteins/genetics ; Humans ; Kidney Calculi/*prevention & control ; Kidney Neoplasms ; Male ; Mice ; Mice, Inbred ICR ; Molecular Sequence Data ; Recombinant Fusion Proteins/genetics/metabolism ; Tacrolimus Binding Protein 1A/genetics/*metabolism

The mechanism by which propofol exerts its action is poorly understood, but may involve a non-specific effect on lipid membrane and has been shown to potentiate GABA-mediated synaptic inhibition. And, midazolam also acts through GABA receptor mediated increased chloride conductance. The aim of this study was to evaluate the dose response of midazolam, propofol and combination of these drugs, and determine possible interaction between two drugs in patients. The effect of propofol on the dose response curve for midazolam was studied in 260 nonpremedicated ASA physical status I or II female patients who were scheduled for elective operation. The response to the verbal command was used as an end-point of hypnosis. Dose response curves for midazolam, propofol, and their type of interaction was determined using Instat software package, nonlinear regression analysis, and algebraic(fractional) analysis of interaction. ED of midazolam and propofol was 0.11 mg/kg, 1.13 mg/kg and ED95 was 0.18 mg/kg, 1.67 mg/kg respectively. ED50 of combined drug(midazolam+propofol) in comhined dose response curve was 29% of each drug and the type of interaction between two drugs was found to be synergistic.
Anesthesia* ; Female ; Humans ; Hypnosis ; Membranes ; Midazolam* ; Propofol* ; Receptors, GABA

In order to study the functions of migration inhibitory factor (MIF) as macrophage activating cytokine and to investigate the possibility of MIF cDNA as gene therapeutic agent or adjuvant, we produced recombinant MIF (rMIF), anti-MIF antibody and pcDNA I plasmid containing mMIF cDNA (mMIF plasmid). We have investigated the effects of recombinant mMIF or mMIF plasmid on the expression of immune response-related gene in the mouse peritoneal macrophage or splenocyte. Recombinant mMIF produced by Baculovirus expression system was biologically active; it increased mRNA expression of tumor necrosis factor (TNF)-a, Interleukin (IL)-1, IL-6, granulocyte monocyte-colony stimulating factor (GM-CSF), nitric oxide synthase (NOS), Fas and Bcl-x when applied to the cultures of mouse peritoneal macrophage. Anti-mMIF antibody blocked these effects of mMIF on macrophage. Plasmid DNA carrying MIF cDNA inoculated into mouse peritoneal cavity also increased mRNA transcriptions from TNF, IL-1, IL-6, IL-12, GM-CSF, NOS genes of peritoneal macrophage. It enhanced proliferation of splenocyte stimulated with phorbol myristate acetate and IL-2 mRNA expression of splenocytes. Frorn these results, we conclude that rMIF is a strong macrophage activating factor and especially MIF plasmid can be used as an immune potentiating DNA drug in gene therapy for cancer or DNA adjuvant in vaccination in future.
Animals ; Baculoviridae ; DNA ; DNA, Complementary* ; Genetic Therapy ; Granulocyte-Macrophage Colony-Stimulating Factor ; Granulocytes ; Interleukin-1 ; Interleukin-12 ; Interleukin-2 ; Interleukin-6 ; Interleukins ; Macrophages* ; Macrophages, Peritoneal ; Mice ; Nitric Oxide Synthase ; Peritoneal Cavity ; Plasmids ; RNA, Messenger* ; Tetradecanoylphorbol Acetate ; Tumor Necrosis Factor-alpha ; Vaccination

OBJECTIVES: This 6-week, open label randomized, multicenter study was conducted to evaluate the antidepressant effect and safety of milnacipran and fluoxetine in patients with major depression. METHODS: The study was done in patients with major depression diagnosed by DSM-IV who score > or =17 in 17 items Hamilton Rating Scale for Depression (17-item HAM-D) and score > or =25 in Montgomery and Asberg Depression Rating Scale (MADRS). A total of 87 patients were randomized to milnacipran group and fluoxetine group. In cases of the patients taking other antidepressants, 6 weeks of each medication was administered after 7 days of drug excretion period. The evaluation was done using 17 item HAM-D, MADRS, Clinical Global Impression Scale (CGI), and COVI scale after baseline, 1 week, 2 weeks, 4 weeks, and 6 weeks. The side effects that had occurred during the period of our study were put in records by developed/disappeared time, severities, incidences, managements and results. RESULTS: A total of 87 patients were enrolled. 70 (milnacipran group 39;fluoxetine group 31) of them were included for the 6 weeks of research and 17 of them dropped out within the first week, not due to adverse reactions or deficiency of effects. Total 17 item HAM-D scores, total points of MADRS, and CGI showed significant decrease after 1 week in each treatment group and continued decrease after 2 weeks and 4, 6 weeks. But there was no difference between milnacipran group and fluoxetine group in the antidepressant effect. There were no significant changes in vital sign, CBC, chemistry, and EKG in each treatment group. The commonly reported side effects of minlacipran were nausea (25.0%), headache (10.7%), vomiting (7.1%), constipation (7.1%), dizziness (7.1%) and those of fluoxetine were GI trouble (11.1%), diarrhea (11.1%), insomnia (11.1%), agitation (5.6%), and dizziness (5.6%). CONCLUSION: Milnacipran was effective for the improvement of depressive symptoms and was well tolerated and safe in patients with depression.
Antidepressive Agents ; Chemistry ; Constipation ; Depression* ; Diagnostic and Statistical Manual of Mental Disorders ; Diarrhea ; Dihydroergotamine ; Dizziness ; Electrocardiography ; Fluoxetine* ; Headache ; Humans ; Incidence ; Nausea ; Sleep Initiation and Maintenance Disorders ; Vital Signs ; Vomiting