Objective To prepare huperzine A-carrier protein conjugate and determine its immunogenicity for later production of monoclonal antibody against HupA and establishing an enzyme-linked immunosorbent assay ( ELISA) -based detection method for HupA. Methods HupA was conjugated with bovine serum albumin ( BSA) using glutaraldehyde ( GA) method to obtain the HupA-GA-BSA conjugate,and the hapten number in the conjugate was determined by matrix-assisted laser absorption ionization time-of-flight mass spectrometry ( MALDI-TOF-MS) . BALB /c mice were immunized with HupA-GA-BSA to prepare the antiserum against HupA. The serum titer and specificity of the HupA antibodies were detected by indirect ELISA and competitive ELISA,respectively. Results The conjugation ratio of HupA and the carrier protein BSA was 8∶ 1. The antiserum against HupA with a titer of 1∶ 62 500 was obtained after immunizing the mice with the conjugate,and the antiserum reacted specifically to HupA. Conclusion The synthesized HupA-GA-BSA conjugate possesses good immunogenicity in mice,suggesting the feasibility of preparing the monoclonal antibody against HupA using this conjugate.

Air Pollutants, Occupational ; analysis ; Chromatography, Gas ; methods ; Fluorocarbon Polymers ; analysis ; Workplace

Objective To establish brain hypoxic-ischemic (HI) model using postnatal day 3(P3) SD rats and evaluate the apoptotic neuronal cells in cerebral cortex and neurofunctional development.Methods The P3 rats were randomly divided into HI group (n=35) and control group(n=18).HI was induced in P3 rats with right carotid artery ligation followed by 2.5 h hypoxia in 60 mL?L-1 oxygen at 37 ℃.The injury of neural cells in the cerebral cortex was evaluated by tumor necrosis factor receptor-1(TNF-R1),Caspase-3 immunostaining and Hematoxylin-Eosin (HE) staining in 24 h and 7 d after HI,respectively.Furthermore,the neurofunctional development was evaluated by negative geotaxis reflex and eye opening time.The data were analyzed by SPSS 12.0 software.Results Caspase-3 and TNF-R1 positive cells were abundant in the ipsilateral cortex at 24 h after HI,compared with contralateral part and control group(P

Objective To determine chromosomal localization of the primary gout susceptibility gene in a pedigree.Methods The clinical data and the peripheral blood samples were collected in the pedigree members and the genomic DNA was extracted from peripheral blood.A genome-wide screening was performed using 400 micro-satellite DNA markers in this family,and linkage analysis was used to determine the chromosomal location of the primary gout susceptibility gene.Results Linkage analysis showed that the maximum LOD score reached 1.50 at marker D4S1572 (at recombination fraction?=0.00).Conclusion Since D4S1572 is localized at 4q25,the primary gout susceptibility gene of this pedigree is localized at 4q25.

The medicinal fungi, which are of great importance in traditional medicine, are facing the problems of wild resources scarcity and low concentration of bioactive compounds. Velvet family and LaeA global regulator play a vital role in secondary metabolism and developmental programs, which are found in a wide variety of fungi ranging from Chytridiomycota to Basidiomycota. This review elaborates the structures and functions between Velvet family and LaeA protein. The Velvet family which shares the Velvet protein domain, including VeA (Velvet), VelB (Velvet like B), VosA (viability of spores A) and VelC (Velvet like C), acts on the regulation function is secondary metabolism and developmental programs such as asexual and sexual development. Furthermore, the function is affected by environmental factors such as light and temperature. LaeA protein which owns S-adenosylmethionine-dependent methyltransferase domain, coordinately regulates development and secondary metabolism by regulating and modifying the Velvet proteins. The regulation of LaeA is mediated by light receptor proteins. Therefore, clarifying the mechanism of Velvet and LaeA proteins in medicinal fungi will pave the way for nurturing medicinal fungi and improving production of bioactive compounds.
Fungal Proteins ; metabolism ; Fungi ; chemistry ; Gene Expression Regulation, Fungal ; Genes, Regulator ; Protein Structure, Tertiary ; Secondary Metabolism ; Structure-Activity Relationship

In order to authenticate the components of antler powder in the market, DNA barcoding technology coupled with cloning method were used. Cytochrome c oxidase subunit I (COI) sequences were obtained according to the DNA barcoding standard operation procedure (SOP). For antler powder with possible mixed components, the cloning method was used to get each COI sequence. 65 COI sequences were successfully obtained from commercial antler powders via sequencing PCR products. The results indicates that only 38% of these samples were derived from Cervus nippon Temminck or Cervus elaphus Linnaeus which is recorded in the 2010 edition of "Chinese Pharmacopoeia", while 62% of them were derived from other species. Rangifer tarandus Linnaeus was the most frequent species among the adulterants. Further analysis showed that some samples collected from different regions, companies and prices, contained adulterants. Analysis of 36 COI sequences obtained by the cloning method showed that C. elaphus and C. nippon were main components. In addition, some samples were marked clearly as antler powder on the label, however, C. elaphus or R. tarandus were their main components. In summary, DNA barcoding can accurately and efficiently distinguish the exact content in the commercial antler powder, which provides a new technique to ensure clinical safety and improve quality control of Chinese traditional medicine
Animals ; Antlers ; DNA Barcoding, Taxonomic ; Deer ; Medicine, Chinese Traditional ; Polymerase Chain Reaction ; Powders ; Quality Control

In order to authenticate the components of antler powder in the market, DNA barcoding technology coupled with cloning method were used. Cytochrome c oxidase subunit I (COI) sequences were obtained according to the DNA barcoding standard operation procedure (SOP). For antler powder with possible mixed components, the cloning method was used to get each COI sequence. 65 COI sequences were successfully obtained from commercial antler powders via sequencing PCR products. The results indicates that only 38% of these samples were derived from Cervus nippon Temminck or Cervus elaphus Linnaeus which is recorded in the 2010 edition of "Chinese Pharmacopoeia", while 62% of them were derived from other species. Rangifer tarandus Linnaeus was the most frequent species among the adulterants. Further analysis showed that some samples collected from different regions, companies and prices, contained adulterants. Analysis of 36 COI sequences obtained by the cloning method showed that C. elaphus and C. nippon were main components. In addition, some samples were marked clearly as antler powder on the label, however, C. elaphus or R. tarandus were their main components. In summary, DNA barcoding can accurately and efficiently distinguish the exact content in the commercial antler powder, which provides a new technique to ensure clinical safety and improve quality control of Chinese traditional medicine

BACKGROUND: The association of genetic variation in the IRAK-1 gene with sepsis outcome has been proved. However, few studies have addressed the impact of the IRAK-4 gene variants on sepsis risk. This study aimed to determine whether the polymorphisms in the IRAK-4 gene are associated with susceptibility to and prognosis of severe sepsis in the Chinese Han ethnic population.METHODS: In this case-control study, 192 patients with severe sepsis hospitalized in the emergency department of Zhongshan Hospital from February 2006 to December 2009 and 192 healthy volunteers were enrolled. Exclusion criteria included metastatic tumors, autoimmune diseases, AIDS or treatment with immunosuppressive drugs. This study was approved by the ethical committee of Zhongshan Hospital, Fudan University. Sepsis patients were divided into a survival group (n=124) and a non-survival group (n=68) according to the 30-day mortality. Primer 3 software was used to design PCR and sequencing primers. Genomic DNA was extracted from peripheral blood mononuclear cells. Seven tagSNPs in IRAK-4 were selected according to the data of the Chinese Han population in Beijing from the Hapmap project and genotyped by direct sequencing. The chi-square test was used to evaluate the differences in genotype and allele frequencies between the two groups.RESULTS: The distributions of all tagSNPs were consistent with Hardy-Weinberg equilibrium. The allele and genotype frequencies of rs4251545 (G/A) were significantly different between the severe sepsis and healthy control groups (P=0.015, P=0.035, respectively). Carriers of the rs4251545A had a higher risk for severe sepsis compared with carriers of the rs4251545G (OR=1.69, 95％ CI: 1.10-2.58). The allele and genotype frequencies of all SNPs were not significantly different between the survival group and non-survival group.CONCLUSION: These findings indicate that the variants in IRAK-4 are significantly associated with susceptibility to severe sepsis in the Chinese Han ethnic population.

OBJECTIVETo analysis the changes of two chemical constituents, namely 2, 3-dihydro-3, 5- dihydroxy-6-methyl-4H-pyran-4-one (DDMP) and 5-hydryoxymethyl-furfural (5-HMF) produced in Radix Polygoni Multiflori after processing, with processing time, and to determine the contents of 5-HMF in samples of Radix Polygoni Multiflori and Radix Polygoni Multiflori preparata.

METHODAn HPLC method was applied with a Zobax SB-C18 (3.9 mm x 150 mm, 5 microm) column by a elution using methanol-water (10: 90) as the mobile phase. The detection was set at UV 280 nm.

RESULTThe contents of DDMP were increasing with the processing time until 24 hour, followed by a decrease until 60 hour process. The contents of 5-HMF were increasing gradually throughout the 60 hour steaming process. The contents of 5-HMF in 11 samples of Radix Polygoni Multiflori preparata were from 0.013% to 0.101%, and only one in 4 samples of Radix Polygoni Multiflori containing trace amount of 5-HMF.

CONCLUSIONThe chemical components in Radix Polygoni Multiflori were changed during the processing procedures. Therefore, the processing of Radix Polygoni Multiflori should be controlled and standardized.

Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; chemistry ; Polygonaceae ; chemistry ; Reproducibility of Results

OBJECTIVETo investigate the effect of telomerase reverse transcriptase (TERT) to the proliferation of 5-HT induced pulmonary artery smooth muscle cells (PASMCs).

METHODSThe PASMCs proliferation experiment was performed to detect the effort on PASMCs of 5-HT or ASODN TERT (antisense oligoribonucleotides TERT designed according to the rat TERT mRNA sequence of gene bank). The immunohistochemistry staining experiment and the in situ hybridization experiment were to detect the TERT protein and mRNA expression with 5-HT or ASODN TERT. FITC marked ASODN TERT experiment was done to research the distribution of ASODN TERT in PASMCs.

RESULTS5-HT promoted PASMCs proliferation in a dose-dependent manner (10(-9) - 10(-5) mol/L). 5-HT also significantly increased TERT expression at protein and mRNA levels as shown by immunohistochemistry staining and the in situ hybridization studies. This effect could be blocked by ASODN TERT in a time and dose-dependent manner.

CONCLUSIONSOur experiments show TERT is one of the key factors in the procession of 5-HT induced PASMCs proliferation. ASODN TERT might be a potential therapy agent for pulmonary hypertension.

Animals ; Cell Proliferation ; Cells, Cultured ; Muscle, Smooth, Vascular ; cytology ; Myocytes, Smooth Muscle ; cytology ; Pulmonary Artery ; cytology ; RNA Replicase ; RNA, Messenger ; genetics ; Rats ; Serotonin ; pharmacology ; Telomerase ; pharmacology