To obtain chemical constituent information of rat plasma after oral administration of Huang-Lian-Jie-Du Decoction (HLJDD), a LC-FT-ICR-MS method has been established, and both positive and negative ions scan modes were include in the analysis. By comparing their retention time, high resolution mass data of HLJDD extracts, blank plasma and dosed plasma, 38 constituents, including 22 prototype compounds and 16 metabolites, were detected in rat plasma after oral administration of HLJDD. In the 22 prototype compounds, 16 constituents were determined unambiguously by comparing with references. In the analysis of metabolites, phase II reactions like glucuronidation and sulfation were the major biotransformation pathways of HLJDD. M11 was observed as the only phase I metabolite in present experiment. The results will be beneficial for the further pharmacokinetics and pharmacological evaluations of HLJDD.
Administration, Oral ; Alkaloids ; blood ; Animals ; Biotransformation ; Chromatography, High Pressure Liquid ; Drug Combinations ; Drugs, Chinese Herbal ; administration & dosage ; isolation & purification ; metabolism ; Flavonoids ; blood ; Iridoids ; blood ; Male ; Plants, Medicinal ; chemistry ; Rats ; Rats, Sprague-Dawley ; Spectrometry, Mass, Electrospray Ionization ; Tandem Mass Spectrometry

Objective To explore the effect of aminolevulinic acid-based photodynamic therapy(ALA-PDT) on alloreactived peripheral blood mononuclear cells(PBMCs).Methods Human PBMCs from different healthy donor were collected and mixed in the one-way mixed lymphocyte culture(MLC) for 5 days. The cells were harvested and aminolevulinic acid(ALA) were added into ALA group and ALA+Light group with ultimate concentrations of 0.5 mmol/L,1.0 mmol/L,1.5 mmol/L,2.0 mmol/L and 2.5 mmol/L.After cultured for 2 hours, 4 hours and 6 hours respectively in 37 ℃ 5% carbon dioxide incubator,Light group and ALA+Light group were irradiated by light of 410 nm wavelength for 1 hour.The MLC cells were treated with the former stimulator cells for 48 hours.The survival of stimulator cells were detected using MTT colorimetric assay and the kill rates of treated cells were calculated.Results The kill rate of ALA+Light group on stimulators was apparently lower than those of Light group, ALA group and control group, (33.0?26.5)% vs (87.1?2.2)%,(89.2?2.5)%,(90.3?1.9)%(All P

OBJECTIVEThis study investigated the hypothesis that 5-aminolevulinic acid-mediated photodynamic therapy (ALA-PDT) might alleviate acute graft-versus-host disease (GVHD) following allogenic bone marrow transplantation (allo-BMT) in mice.

METHODSAcute GVHD model following allo-BMT was established in 40 recipient BALB/C mice. Fifty C57BL/6J mice were used as donors and another 10 BALB/C mice as blank control without any intervention. Recipients received a lethal dose of 8.5 Gy (60)Co radiation for 10 minutes before transplantation and then were randomly divided into four groups of 10 mice (A-D). Group A was injected with normal saline injection and served as controls. Group B received pure donor bone marrow and spleen cell infusion. Group C received donor bone marrow and mixed donor-recipient spleen cell infusion. Group D was administered with an infusion of donor bone marrow cells and mixed donor-recipient spleen cells treated with ALA-PDT. The 28th day survival rate, incidence of acute GVHD and hematological and pathological changes after transplantation were examined.

RESULTSAll the mice from the Blank control group survived. The survival rates for Groups A-D on the 28th day were 0, 0, 10% and 60% respectively. Group D showed a significantly higher survival rate than the other three groups (P < 0.01). Most of the mice in Groups B and C developed GVHD but only two developed in Group D. Moreover Group D had less severe hematological and pathological changes when compared with Groups B and C.

CONCLUSIONSALA-PDT significantly alleviated GVHD and increased the 28th day survival rate for allo-BMT mice. ALA-PDT may be a promising therapy for GVHD following allo-BMT. Future studies should focus on the underlying mechanism of its therapeutic effect.

Aminolevulinic Acid ; therapeutic use ; Animals ; Bone Marrow Transplantation ; adverse effects ; mortality ; Female ; Graft vs Host Disease ; drug therapy ; Leukocyte Count ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Photochemotherapy ; Transplantation, Homologous

OBJECTIVETo study the effect and potential mechanism of expression of c-jun N-terminal kinase (JNK) signal pathway on neuron autophagy after diffuse brain injury (DBI).

METHODSMale Sprague Dawley rats (n = 216) were randomly divided into four groups: DBI group (n = 54), SP600125 intervene group (n = 54), DMSO group (n = 54) and sham operation group (n = 54). DBI rat model was established according to the description of Marmarou DBI. At different time points (1, 6, 12, 24, 48 and 72 h) after operation, the histopathologic changes of neurons in cortex were observed by HE staining method; The expression of p-JNK, p-P53, DRAM and Beclin-1 were detected by Western blot and immunohistochemistry.

RESULTSThe results showed that under light microscope degenerated and necrotic neurons were observed to be scattered in cortex at 6 h after operation in DBI group, but these changes were low in SP600125 intervene group. Compared with SP600125 intervene group, the expression of p-JNK in DBI group were enhanced obviously at 6, 12 and 24 h (F = 17.902, P < 0.05); the expression of p-P53 in DBI group were enhanced obviously at 12, 24, 48 and 72 h (F = 7.107, P < 0.05); the expression of DRAM in DBI group were enhanced obviously at 6, 12, 24, 48 and 72 h (F = 15.455, P < 0.05); the expression of Beclin-1 in DBI group were enhanced obviously at 6, 12, 24, 48 and 72 h (F = 11.517, P < 0.05). Compared with DBI group, the expression of p-JNK, p-P53, DRAM and Beclin-1 in DMSO group were similar at 1, 6, 12, 24, 48 and 72 h (F = 1.509, P > 0.05).

CONCLUSIONSThe present results indicate that SP600125 can dramatically improve trauma brain injury from autophagy after DBI and the molecular mechanism is related to the modulation of JNK signal pathway following DBI, while it measures the neuron autophagy by means of intervening JNK signal pathway.

Animals ; Anthracenes ; pharmacology ; Autophagy ; Brain Injuries ; metabolism ; pathology ; Disease Models, Animal ; JNK Mitogen-Activated Protein Kinases ; metabolism ; Male ; Neurons ; pathology ; Rats ; Rats, Sprague-Dawley

The purpose of this study is to evaluate the effects and the underline mechanisms of berberine on the cardiac function and left ventricular remodeling in rats with renovascular hypertension. The renovascular hypertensive model was established by the two-kidney, two-clip (2K2C) method in Sprague-Dawley (SD) rats. Two weeks after surgery, all the operated SD rats were randomly assigned into four groups: (1) renovascular hypertensive model group; (2) berberine 5 mg x kg(-1) group; (3) berberine 10 mg x kg(-1) group; (4) captopril 45 mg x kg(-1) group; and the sham operated rats were used as control. Four weeks after the drugs were administered, the cardiac function was assessed. The ratios of heart weight to body weight (HW/BW), left ventricular weight to body weight (LVW/BW) and right ventricular weight to body weight (RVW/BW) were compared between groups. Coronal sections of the left ventricular tissue (LV) were prepared for paraffin sections, picrosirius red and HE staining was performed. The left ventricular wall thickness (LVWT), interventricular septal thickness (IVST), the parameters of myocardial fibrosis indicated by interstitial collagen volume fraction (ICVF) and perivascular collagen area (PVCA) were assessed. Nitric oxide (NO), adenosine cyclophosphate (cAMP) and guanosine cyclophosphate (cGMP) concentrations of left ventricular tissue were measured. Berberine 5 mg x kg(-1) and 10 mg x kg(-1) increased the left ventricular +/- dp/dt(max) and HR. Berberine 10 mg x kg(-1) decreased HW/BW and LVW/BW. The image analysis showed that both 5 and 10 mg x kg(-1) of berberine decreased LVWT, ICVF and PVCA, while increased the NO and cAMP contents in left ventricular tissue. Berberine could improve cardiac contractility of 2K2C model rats, and inhibit left ventricular remodeling especially myocardial fibrosis in renovascular hypertension rats. And such effects may partially associate with the increased NO and cAMP content in left ventricular tissue.
Animals ; Berberine ; pharmacology ; Collagen ; metabolism ; Cyclic AMP ; metabolism ; Cyclic GMP ; metabolism ; Hypertension, Renovascular ; metabolism ; physiopathology ; Male ; Myocardium ; metabolism ; pathology ; Nitric Oxide ; metabolism ; Organ Size ; drug effects ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Ventricular Function, Left ; drug effects ; Ventricular Remodeling ; drug effects

AIM:To investigate the effects of evodiamine on the growth and apoptosis of human hepatocellular carcinoma Huh7 cells,and to illustrate the molecular mechanism that evodiamine enhances antitumor activity of tumors nec -rosis factor-related apoptosis-inducing ligand(TRAIL)in Huh7 cells.METHODS: The cell viability was measured by MTT assay.The cell cycle distribution was analyzed by flow cytometry.The apoptosis rate was determined by TUNEL stai-ning.The protein levels of cell cycle-and apoptosis-related proteins were detected by Western blot analysis.RESULTS:Treatment of Huh7 cells with evodiamine reduced the cell viability(P<0.05).Evodiamine induced cell cycle arrest in G2/M phase by upregulation of p27,cyclin B1, cell division cycle protein 2(Cdc2)and p-Cdc2.Evodiamine triggered apoptosis accompanied by cleavage of caspase-3 and poly(ADP-ribose)polymerase(PARP).Combination of evodiamine with TRAIL significantly reduced the cell viability and increased cleavage of caspase -3 and PARP as compared with the use of each agent alone.Moreover,evodiamine increased the expression of death receptor 5(DR5)in the Huh7 cells.CON-CLUSION:Evodiamine inhibits the cell growth by reducing the cell viability and inducing cell cycle arrest.Evodiamine also triggers cell apoptosis and enhances the sensitivity of Huh 7 cells to TRAIL by upregulating the expression of DR5.

Objective To explore the approach and early effects of endovascular stent-graft deployment in the treatment of portal stenosis of cancerous thrombus.Methods Six cases with portal vein stenosis of cancerous thrombus,which caused by primary hepatic carcinoma(5 cases)and eholangiocarcinoma(1 case)and the severity of stenosis showed on contrast enhanced CT were more than 75% or occluded,were performed percutaneous transhepatie or transsplenic portography.FLUENCY~(TM) endovascular stent-graft(10 mm diameter)was placed at the position of stenosis after gastroesophageal varices embolization.Portal pressure was measured pre-and post-deployment.Results Stents were successfully placed in all patients.The average portal pressure decreased from 50.7 cm H_2O(1 cm H_2O = 0.098 kPa)to 41.3 cm H_2O after endovascular stent-graft deployment.The restenosis were found in 2 cases after one month.Haematemesis and refractory aseites appeared in one case respectively,the other 4 cases showed no significant symptoms above caused by portal hypertension.Conclusion It is safe and feasible for endovaseular stent-graft deployment in the treatment of portal stenosis of cancerous thrombus.Selecting the suitable indications,the symptoms of portal hypertension can be controlled effectively.

BACKGROUNDAlthough DNA vaccine is considered as the next generation of vaccine, most DNA vaccine candidates are still suffering from the relatively weak immunogenicity despite the increased dosage of plasmid DNA administered. In order to enhance the immune responses elicited by a codon-optimized HIV gag DNA vaccine, a modified plasmid vector pDRVI1.0 and a booster immunization with replicating Tiantan vaccinia (RTV) strain expressing the same gene were employed.

METHODSVector pDRVI1.0 was constructed through inserting the 72-bp element from the SV40 enhancer, which was reported promoting nuclear transport of plasmid DNA, to the upstream of cytomegalovirus enhancer/promoter region of the plasmid vector pVR1012. Gene expression levels from expression plasmids based on pDRVI1.0 and pVR1012 were tested. Humoral and cellular immune responses induced by DNA vaccine alone or DNA prime-RTV boost regimen were determined in mice.

RESULTSIt was shown that the 72-bp element significantly enhanced the gene expression level in non-dividing cells. gag-specific humoral and cellular immune responses induced by DNA vaccination were both significantly improved, while the Th1/Th2 balance was not obviously affected by the 72-bp element. RTV boosting further significantly enhanced DNA vaccine-primed antibody and T cell responses in a Th1-biased manner.

CONCLUSIONSThe 72-bp SV40 enhancer element should be included in the DNA vaccine vector and RTV strain is a very efficient live vector for boosting immunization.

AIDS Vaccines ; immunology ; Amino Acid Sequence ; Animals ; Blotting, Western ; CD8-Positive T-Lymphocytes ; immunology ; Enhancer Elements, Genetic ; Female ; Gene Products, gag ; immunology ; HIV Antibodies ; blood ; Immunoglobulin G ; blood ; Mice ; Mice, Inbred BALB C ; Molecular Sequence Data ; Plasmids ; Simian virus 40 ; genetics ; Vaccination ; Vaccines, DNA ; immunology ; Vaccinia ; immunology

OBJECTIVETo evaluate the quantitative and functional changes of T helper (Th) cell subsets in the bone marrow of severe aplastic anemia (SAA) patients and the relationship between these changes and the patients hematopoietic function.

METHODSBy FACS, the quantity and ratio of Th1 and Th2 cells, the percentage of CD3(+)CD8(+) cells in the bone marrow were detected in 24 patients with SAA at active phase, 15 patients with SAA at recovery phase, and 16 normal controls. By radioimmunoassay, the serum levels of TNF-alpha, or IL-4 in 20 SAA patients at active phase, 12 at recovery phase and 16 normal controls were measured. The relationships between CD3(+)CD8(+) cells, TNF-alpha and Ret, ANC; and between Th1 cells and CD3(+)CD8(+) cells, TNF-alpha or Ret, ANC; between IL-4, balance of Th1/Th2 and Ret, ANC were evaluated.

RESULTSThe percentages of Th1 and Th2 cells, and ratio of Th1/Th2 in bone marrow of SAA patients at active phase were (4.87 +/- 2.64)%, (0.41 +/- 0.26)% and 21.22 +/- 5.07, respectively, being higher than those of normal controls [(0.42 +/- 0.30)% (P < 0.01), (0.24 +/- 0.17)% (P < 0.05) and (1.57 +/- 0.93) (P < 0.01), respectively] and all of them reduced to normal levels of SAA at recovery phase (P > 0.05). The percentage of CD3(+)CD8(+) cells significantly decreased from (32.32 +/- 8.69)% at active phase to (13.76 +/- 2.96)% at recovery phase (P < 0.01). The serum levels of TNF-alpha and IL-4 at active phase was (4.29 +/- 3.15) microg/L and (1.24 +/- 0.73) microg/L, respectively, being higher than those of normal controls (1.21 +/- 1.16) microg/L, (1.18 +/- 0.97) microg/L, but only the difference of TNF-alpha was statistically significant (P < 0.01). In recovery SAA patients, the serum levels of TNF-alpha significantly decreased to (1.46 +/- 1.41) microg/L (P < 0.01), and the levels of IL-4 increased markedly to (3.05 +/- 1.94) microg/L. The CD3(+)CD8(+) cells and TNF-alpha of patients negatively correlated with Ret (P < 0.05; P < 0.05) and ANC (P < 0.05; P < 0.05), Th1 cells correlated with CD3(+)CD8(+) cells and TNF-alpha positively (P < 0.01; P < 0.05), the Ret and ANC negatively (P < 0.01; P < 0.01), IL-4 and the balance of Th1/Th2 positively correlated with Ret and ANC (P < 0.05, P < 0.01; P < 0.01, P < 0.01).

CONCLUSIONThe bone marrow failure in SAA might be caused not only by the increase of Th1 cells, Th1 type effector cells and cytokines, but also by insufficient compensation of Th2 cells and Th2 type cytokines, which shifted the balance of Th1/Th2 favorable to Th1.

Adolescent ; Adult ; Anemia, Aplastic ; blood ; pathology ; physiopathology ; Bone Marrow ; metabolism ; pathology ; CD3 Complex ; blood ; CD8 Antigens ; blood ; Child ; Female ; Hematopoietic System ; metabolism ; pathology ; physiopathology ; Humans ; Interleukin-4 ; blood ; Male ; Middle Aged ; Radioimmunoassay ; T-Lymphocytes, Helper-Inducer ; metabolism ; pathology ; Th1 Cells ; metabolism ; pathology ; Th2 Cells ; metabolism ; pathology ; Tumor Necrosis Factor-alpha ; blood ; Young Adult

OBJECTIVETo measure the subsets of dendritic cells 1 (DC1) in the bone marrow of severe aplastic anemia (SAA) patients and evaluate the relationships between the CD11c+CD83+ cells and Th1 cells, CD3+CD8+ cells or hematopoietic function and explore the role of DC1 in the pathogenesis of SAA.

METHODSBy FACS, the quantities and ratios of CD11c+CD1a+ cells, CD11c+CD83+ cells, Th1 cells, and CD3+CD8+ cells in the bone marrow of SAA patients and normal controls were detected respectively. The relationships between CD3+CD8+ cells and reticulocyte absolute value (Ret) or neutrophil absolute value (ANC), between Th1 cells and CD3+CD8+ cells, Ret or ANC, between CD11c+CD83+ cells, and Th1 cells, CD3+CD8+ cells, Ret or ANC were evaluated.

RESULTSIn normal controls' bone marrow, the percentages of Th1 cells, CD11c+CD1a+ cells, CD11c+CD83+ cells and the ratio of CD11c+CD83+/CD11c+CD1a+ were (0.42 +/- 0.30)%, (0.38 +/- 0.29)%, (0.37 +/- 0.32)% and 1.07 +/- 0.10, respectively. In untreated SAA patients, they were (4.87 +/- 0.54)%, (1.73 +/- 0.24)%, (3.38 +/- 0.56)% and 2.21 +/- 0.32 respectively, which were higher than that in normal controls (P < 0.01). In recovering SAA patients, the percentages of Th1 cells, CD11c+CD1a+ cells and CD11c+CD83+ cells decreased significantly to (0.53 +/- 0.22)%, (0.61 +/- 0.23)%, (0.65 +/- 0.22)%, respectively (P < 0.01). The ratio of CD11c+CD83+/CD11c+ CD1a+ in recovering SAA patients decreased to 1.37 +/- 0.25, which was similar to that in normal controls (P > 0.05). The percentage of CD3+CD8+ cells in untreated SAA patients was (32.32 +/- 10.22)%, and in recovering SAA patients decreased to (13.67 +/- 5.24)% (P < 0.01). The percentage of CD3+CD8+ cells in SAA patients was negatively correlated with their Ret and ANC (P < 0.05), while their Th1 cell percentages were positively correlated with their CD3+CD8+ cells (P < 0.01), and negatively correlated with their Ret and ANC (P < 0.01). SAA patient's CD11c+CD83+ cell percentages were positively correlated with their Th1 cell and CD3+CD8 cells (P < 0.01, P < 0.05), but negatively with their Ret and ANC (P < 0.01).

CONCLUSIONBoth immature DC1 and activated DC1 increased in the bone marrow of SAA patients, and the balance of DC1 subsets shifted from stable form to active one, which might promote Th0 cells to polarize to Th1 cells, and cause the over-function of T lymphocytes and hematopoiesis failure in SAA.

Adolescent ; Adult ; Anemia, Aplastic ; immunology ; Antigens, CD ; immunology ; Antigens, CD1 ; immunology ; Bone Marrow ; immunology ; CD11c Antigen ; immunology ; CD8-Positive T-Lymphocytes ; immunology ; Child ; Dendritic Cells ; immunology ; Female ; Humans ; Immunoglobulins ; immunology ; Male ; Membrane Glycoproteins ; immunology ; Th1 Cells ; immunology ; Young Adult