The changes of bioactive constituents were analyzed for Puhuang-Wulingzhi before and after compatibility and the antiplatelet and antithrombin activitives were evaluated in order to elucidate the scientific and reasonable of Puhuang-Wulingzhi compatibility. UPLC-QTOF-MA-Markerlynx, principal component analysis (PCA) and orthogonal partial least-squares discriminant analysis were used for data analysis and tracking changes of chemical composition during the decocting process. In vitro platelet aggregation induced by ADP, thrombin time(TT) and prothrombin time (PT) were investigated for Puhuang-Wulingzhi before and after compatibility. The results showed that significant differences were found between the mixed decoction and codecoction of Wulingzhi and Puhuang. Five compounds changed obviously were identified as typhaneoside, naringenin, isorhamnetin-3-O-ruinoside, quercetin-3-O-neohesperidoside, kaempferol-3-O-neohesperidoside. The codecoction, comparing with the single decoction, was more significant in antiplatelet aggregation and could prolong thrombin time. In the same crude drug dose, the thrombin time (TT) elongation were greater. These data could provide references for elucidation of bioactive components for this herb pair.
Animals ; Antithrombins ; chemistry ; pharmacology ; Blood Platelets ; drug effects ; physiology ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Female ; Humans ; Molecular Structure ; Platelet Aggregation ; drug effects ; Rabbits ; Thrombin Time

Objectives To study the level and development of serum specific antibody against severe acute respiratory syndrome coronavirus(SARS-CoV)of different populations in SARS pestilence district after SARS outbreak in a general hospital.Discuss SARS sub-clinical infection and protective action of the IgG antibody.Methods Seroepidemiology method,enzyme-linked immunosorbent assay (ELISA)and indirect immunfluorescence assay(IFA)were employed to investigate the changing level of serum antibody to SARS-associated coronavirus in non-SARS population in SARS pestilence district during and after SARS outbreak.The development of IgM and IgG antibody in patients with SARS in 6 weeks after the onset of SARS was studied qualitatively.The level changing of IgG antibody in con- valescent patients with SARS in 82 weeks after the onset was observed dynamically.Results The ELISA test outcome of IgG antibody was negative in 200 non-SARS people who were random samples of normal mass in SARS pestilence district and common community.The positive rate was 0.41% in 487 SARS high risk population tested by ELISA,but showed negative when retested by IFA.The A value level of IgG antibody existed significant difference in non SARS mass during and after SARS outbreak and the later's was higher them the former's(P

In order to evaluate the effect and mechanism of the mulberry leaf alkaloid, flavones, and polysaccharide intervention on diabetes, the overall metabolite profiling characteristics for the plasma of diabetic mouse was performed by using an ultra-performance liquid chromatography/electrospray-tandem mass spectrometry (UPLC-ESI-MS). The 8 potential biomarkers were found in diabetic mice plasma based on the data of MS/MS characteristics obtained from the UPLC-OrbitrapMS analysis, which mainly involved in sphingolipids, amino acid metabolic pathway. The principal component analysis showed that the normal group and model group were obviously distinguished and implied that metabolic disturbance was happened in diabetic mice plasma. The extracts of mulberry leaf flavonoids, polysaccharide, alkaloid had exhibited the effects of callback function for diabetic mice through regulating the amino acid metabolism and sphingolipid metabolism.
Alkaloids ; chemistry ; Amino Acids ; metabolism ; Animals ; Biomarkers ; blood ; Chromatography, High Pressure Liquid ; Diabetes Mellitus, Experimental ; drug therapy ; Flavones ; chemistry ; Flavonoids ; chemistry ; Metabolic Networks and Pathways ; Metabolomics ; Mice ; Morus ; chemistry ; Plant Leaves ; chemistry ; Sphingolipids ; metabolism ; Tandem Mass Spectrometry

OBJECTIVETo explore the myelo-protection effect of mdr1 transfected cord blood cells (CBMNCs) graft against high-dose homoharringtonine leukemia-bearing severe combined immunodeficient (SCID) mice model.

METHODSMultidrug resistant (mdr1)gene was transferred into CBMNCs by a retrovirus vector, containing full-length cDNA of human mdr1 gene. CBMNCs and high-titer retrovirus supernatant were cocultured with cytokine combinations for 5 - 6 days. The SCID mouse models bearing human HL-60 cell leukemia were divided into three groups. Group A received tail vein injection of 2 x 10(6) mdr1 gene transduced CBMNCs at day 1 and 3, groups B and C 2 x 10(6) un-transduced CBMNCs and same volume of normal saline, respectively. The 3 groups of the mouse model were treated with weekly escalated doses of homoharringtonine. The peripheral white blood cell (WBC) counts, the human leukemia cells percentage in peripheral blood, the histological findings of main organs were assayed. The CD33 positive HL-60 cells in bone marrow were determined by flow cytometry. The function and expression of mdr1 gene were examined by PCR, immunochemistry (IC) and DNR extrusion test in vivo.

RESULTS(1) mdr1 gene was transferred into CBMNCs successfully and the transfection frequency was 30%. (2) Leukemia SCID mice were xenotransplanted with mdr1-transfected BMMNCs by a programmed procedure and could be used as a valuable model for in vivo evaluating myelo-protection effects. (3) The transfected mice could tolerate homoharringtonine 5 approximately 6 folds higher than conventional dose and kept peripheral WBC count at a mean of 3 x 10(9)/L, with the peripheral human myeloid leukemia cells percentage decreasing to less than 5%. Histological examination showed that there was no leukemia infiltration in the main organs, the CD33 positive HL-60 cells in bone marrow were less than 5%. (4) The repopulation frequency of the transfected CBMNs in marrow were 9.13%. DNR extrusion test confirmed that the P-gp product maintained its biological function in the marrow.

CONCLUSIONmdr1 transferred-human CBMNC can xenotransplanted and repopulated in leukemia-bearing SCID mouse and are protected from chemotherapy-induced myelosuppression.

ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; metabolism ; Animals ; Antineoplastic Agents, Phytogenic ; administration & dosage ; adverse effects ; therapeutic use ; Cord Blood Stem Cell Transplantation ; methods ; Female ; Fetal Blood ; cytology ; Genetic Vectors ; HL-60 Cells ; Harringtonines ; administration & dosage ; adverse effects ; therapeutic use ; Humans ; Leukemia, Promyelocytic, Acute ; drug therapy ; pathology ; surgery ; Leukocytes, Mononuclear ; cytology ; metabolism ; transplantation ; Male ; Mice ; Mice, SCID ; Random Allocation ; Retroviridae ; genetics ; Transfection ; Treatment Outcome ; Xenograft Model Antitumor Assays

AIMTo establish reference range of blood theology for different gender by sitting and lying in healthy populations.

METHODSIn 31 volunteers blood were drawed in the postures of sitting and lying to measure blood rheology. Data (between July 2002 and July 2006) of 32854 samples (23436 check-up, 6553 out-patient,2865 in-patient) were collected. Differences between different postures and different gender were compared by Independ-Sample t-test with SPSS 13.0 software and each reference range of blood rheology was established.

RESULTSIn sitting and lying postures 12 parameters of blood rheology showed great significance (P < 0.01). 8 parameters such as whole blood viscosity, blood reduce viscosity, plasma viscosity, hematocrit, were decreased by 9.33% on average while posture was changed from sitting to lying. But index of erythrocyte deformability, erythrocyte electrophoresis presented inverse correlation and were increased by 6.49% on average with the same posture change.Various parameters of blood theology in different gender showed great significance (P < 0.01).

CONCLUSIONThere is significant difference in various parameters of blood rheology in the posture change from sitting to lying. Reference ranges of blood theology of different postures are established which may be used to decrease the rate of misdiagnosis by 8-10 percent.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Blood Viscosity ; Child ; China ; Erythrocyte Aggregation ; Erythrocyte Deformability ; Female ; Hemorheology ; Humans ; Male ; Middle Aged ; Posture ; physiology ; Reference Values ; Young Adult

To investigate the optimum harvesting time and utilization of mulberry leaves during different growth periods based on the content of alkaloids and flavonoids, 88 samples of 11 species of mulberry leaves were collected and analyzed. UPLC-TQ/MS method was applied and the results showed that the ingredients of alkaloids and flavonoids in mulberry leaves are quite different in different growth periods and different species. There was a sharp decline of the average content of alkaloids in all samples from October, while the content of flavonoids dropped either from October but with less volatile. The content of flavonoids in M. atropurpurea was much higher than alkaloids, while M. australis was opposite completely. There was a sharp decline of alkaloids in M. cathayana and M. mongolica from Tuly to August, however, the content of alkaloids and flavonoids in M. alba is neither too high nor too low. In summary, it is more suitable to harvest tender mulberry leaves harvested from the end of September to beginning of October that provide a scientific evidence for rational harvest and comprehensive utilization of mulberry leaves.
Alkaloids ; analysis ; isolation & purification ; Chromatography, High Pressure Liquid ; Drug Stability ; Drugs, Chinese Herbal ; analysis ; isolation & purification ; Flavonoids ; analysis ; isolation & purification ; Mass Spectrometry ; Medicine, Chinese Traditional ; Morus ; chemistry ; Plant Leaves ; chemistry ; Plants, Medicinal ; Reproducibility of Results ; Seasons

In order to establish HPLC fingerprint of Liuwei Dihuang soft capsule, and to provide certain reference for an quality control of it, the HPLC method was performed on an Agilent C18 (4.6 mm x 250 mm, 5 μm) column with acetonitrile-0.02% trifluoroacetic acid as mobile phase, gradient elution volume flow of 1.0 mL x min(-1), column temperature was 30 degrees C, detection wavelength: 0-60 min, 238 nm, 60-70 min, 210 nm. The software for chromatographic fingerprint was applied to analysis different batches of Liuwei Dihuang soft capsule samples. Sixteen mutual peaks were selected as the fingerprint peaks in 12 samples with loganin as the reference peak, and all of the detected peaks were separated effectively. Cluster analysis (HCA) and similarity analysis (SA) were done based on data of 12 samples clustering analysis of 12 batches of samples were divided into 2 categories. Including 7 for the first class, the rest was second, similarities calculated by SA were all above 0.92, indicating a good similarity between the reference and twelve batches of samples, also, the analysis results of HCA and SA basically the same. This method is simple with good precision, repeatability and stability, and provides the basis for Liuwei Dihuang soft capsule quality control.
Capsules ; analysis ; Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; analysis ; Quality Control

OBJECTIVERotavirus is the single most common cause of severe dehydrating diarrhea in young children worldwide, but the pathogenesis and immunity against this disease are not completely understood. A prospective study was conducted to assess gene expression of toll-like receptors (TLR) in children with acute rotavirus diarrhea.

METHODSSeventy-five children with acute rotavirus diarrhea and 38 control children were enrolled in this study from Sep. 2004 to Jan. 2005. All the 75 patients had detailed records of clinical characteristics. Rotavirus antigen was detected by ELISA from stools. Peripheral blood mononuclear cells (PBMC) were separated by Ficoll reagent and RNA was extracted by Trizol. The levels of mRNA for five TLRs in PBMC were examined by fluorescent quantitative RT-PCR.

RESULTSPatients with acute rotavirus infection had elevated mean levels of TLR 2, 3, 4, 7, 8 mRNA expressions in PBMC within 3 days since onset of the disease, P less than 0.05. But only TLR 2, 3, 8 mRNA levels remained increased in patients within 7 or 14 days since onset (P less than 0.05). Mean levels of mRNA for TLR 4 in PBMC was higher in patients with more severe diarrhea including longer duration of diarrhea, more episodes of diarrhea per day and higher severity scores (P less than 0.05).

CONCLUSIONManifold TLR may play roles in the start-up and regulation of immune responses in children with acute rotavirus diarrhea. These findings will be helpful to further recognize immune response in Chinese children with rotavirus diarrhea and, consequently, may provide directions and insights that could prove critical to the prevention or treatment of this important disease.

Acute Disease ; Antigens, Viral ; analysis ; Child, Preschool ; Diarrhea ; genetics ; virology ; Enzyme-Linked Immunosorbent Assay ; Feces ; virology ; Gene Expression ; Host-Pathogen Interactions ; Humans ; Infant ; Leukocytes, Mononuclear ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Rotavirus ; immunology ; physiology ; Rotavirus Infections ; genetics ; virology ; Toll-Like Receptors ; genetics

OBJECTIVETo investigate the influence of three pontic types on alveolar ridge mucosal microecosystem.

METHODSSixty patients ready to accept three unit metal ceramic bridges were selected. The bacterial type and the cultivable flora were counted and the proportions of bacteria detected on the top of alveolar ridge mucosal contact area before tooth preparation and three months after bridge insertion.

RESULTSType and CFU of bacteria on the alveolar ridge mucosa under modified base-type pontics and modified ridge lap pontics increased significantly (P < 0.05); while there was no significant change under the ovata pontics (P > 0.05). Before tooth preparation and 3 months after fixed prosthesis insertion, the percentages of oral Streptococci and Neisseriae changed significantly (P < 0.05).

CONCLUSIONSThe ovata pontic had less influence on mucosal microecosystem than the other two pontics and is the appropriate pontic design for clinical dentist.

Adult ; Alveolar Process ; microbiology ; Denture Design ; Denture, Partial, Fixed ; microbiology ; Humans ; Middle Aged ; Mouth Mucosa ; microbiology

Recent studies have shown that epidermal growth factor receptor (EGFR) is an important target for cancer therapy. The present study prepared single chain Fv (scFv) directed against EGFR. Balb/c mice were immunized by human carcinoma A431 cells, and total RNA of the splenic cells was extracted. VH and VL gene fragments were amplified by RT-PCR and further joined into scFv gene with a linker, then scFv gene fragments were ligated into the phagemid vector pCANTAB 5E. The phagemid containing scFv were transformed into electro-competent E. coli TG1 cells. The recombinant phage antibody library was constructed through rescuing the transformed cells with help phage M13K07. The specified recombinant phages were enriched through 5 rounds of affinity panning and the anti-EGFR phage scFv clones were screened and identified with ELISA. A total of 48 clones from the library were selected randomly and 45 clones were identified positive. After infecting E. coli HB2151 cells with one positive clone, soluble recombinant antibodies about 27 kD were produced and located in the periplasm and the supernatant. The result of sequencing showed that the scFv gene was 768 bp, which encoded 256 amino acid residues. VH and VL including 3 CDRs and 4 FRs, respectively, were all homologous to mouse Ig. The soluble scFv showed the specific binding activity to purified EGFR and EGFR located in carcinoma cell membrane. The successful preparation of anti-EGFR scFv will provide an EGFR targeted molecule for the development of antibody-based drugs and biological therapy of cancer.
Animals ; Antibodies, Monoclonal ; Cell Line, Tumor ; Humans ; Immunoglobulin Light Chains ; genetics ; immunology ; Male ; Mice ; Mice, Inbred BALB C ; Peptide Library ; Receptor, Epidermal Growth Factor ; genetics ; immunology ; Single-Chain Antibodies ; genetics ; immunology