Objective To observe the activation effect of histamine on mast cells in the acupoint area and its analgesic effects similar to acupuncture stimulation,so as to reveal the underlying mechanism of acupoint injection therapy.Methods A total of 60 SD rats were divided into control,model,normal saline(NS),histamine-injection and acupuncture groups(12 rats/group).Acute adjuvant-induced-arthritis(AIA) model was duplicated by injection of complete Freunds' Adjuvant(0.05 mL) into the left ankle articular cavity.For rats of saline and histamine groups,normal saline(50 ?L) and histamine(50 ?L,100 ?g/mL) were injected into the left "Zusanli"(ST 36) acupoint area.For rats of acupuncture group,manual acupuncture(lifting-thrusting and twisting needle intermittently) was applied to ST 36 for 20 min.Paw withdrawal(pain threshold,PT) was detected by radiant heat irradiation of the hindpaw.Skin and muscle tissues of ST 36 area were sampled to be fixed in formalin,then,sliced(5 ?m),and stained with Toluidine Blue,separately,followed by counting the degranulation rate of mast cells under microscope.Results Compared with normal control group and pre-modeling,PT of model,NS,acupuncture and histamine groups decreased significantly(P

The transcription factor of ethylene responsive factor binding protein (ERF) is belonged to AP2/ERF superfamily, which is known to be unique in plants. AP2/ERF proteins have important functions in the transcriptional regulation of a variety of biological processes related to growth and development, as well as various responses to environmental stimuli. An ERF gene from Salvia miltiorrhiza is cloned and divided into ERF gene family group VII of Arabidopsis and Rice. It contains a MCGGAI (I/L) motif referred to as CMVII-1 and a single intron in the 5'-flanking region of the AP2/ERF domain. Sequence analysis reveals that the region of second extron has abundant polymorphism sites. There are 21 single nucleotide polymorphism sites (SNPs) in the 264 bp region, among them, 14 SNPs are synonymous substitutions and 7 SNPs are non-synonymous substitutions. Though analysis of 181 samples from Shandong, Shaanxi and Sichuan Provinces, it reveals that each production area has its own special genotypes, 5 SNPs show significant difference. Cluster based on UPGMA method reveals that different populations from specific province have clustered together. It shows that SmERF gene will be a candidate molecular marker for the identification of Salvia miltiorrhiza from different areas.

ObjectiveTo determine the serum levels of cytokines associated with vascular endothelial cells before and after treatment with either transcatheter arterial chemoembolization (TACE) or partial hepatectomy in patients with hepatocellular carcinoma (HCC).MethodsThere were 30 patients who received partial hepatectomy (the operation group) and 30 patients who received TACE (the TACE group).Cytokines were measured before and after treatment.ResultsThe serum levels of IL-1β,IL-6,IL-8,VEGF and EGF of the post-TACE patients were significantly lower than the pre-TACE patients.The serum levels of IL-10,IFNγ and TNFα of the post-TACE patients were significantly higher than the pre-TACE patients.The serum levels of IL-1β,IL-6.IL-8,VEGF and EGF in the postoperative patients were significantly lower than the post-TACE patients.The serum levels of IL-10,IFNγ and TNFα of the postoperative patients were significantly higher than the post-TACE patients.ConclusionThe results suggested that the serum levels of angiogenic factors in the postTACE patients were significantly higher than the postoperative patients.The serum levels of the inhibitors of vascular endothelial cells of the post-TACE patients were significantly lower than the postoperative patients.

Objective To determine the serum levels of cytokine IL-1α, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, VEGF, IFNγ, EGF, MCP-1 and TNFα in preoperative and postoperative patients with hepatocarcinoma(HCC). Methods 30 patients with hepatocirrhosis were taken as hepatocirrho-sis group; 30 normal health examiners were taken as the normal control group. 30 patients with hepatocarcinoma were taken as HCC group; 30 patients with hepatic hemangioma were taken as the control group. Cytokines had been measured by biochips methods in evidence 180 automatic biochips analyze.Results The serum levels of IL-1β, IL-6, IL-8, VEGF and EGF of the patient with HCC and cirrhosis were significantly higher than those in normal control group. The serum levels of IL-10, IFNγand TNFα of the patient with HCC and cirrhosis were significantly lower than those in normal control group. The serum levels of IL-1β, IL-6, IL-8, VEGF and EGF of the preoperative patient with HCC were significantly higher than those in the postoperative patient with HCC. The serum levels of IL-10,IFNγ and TNFa of the preoperative patient with HCC were significantly lower than those in the postoperative patient with HCC. Conclusions These results suggest that the serum levels of angiogenic factors in HCC were increased. The serum levels of the inhibitors of vascular endothelial cells in HCC were decreased. The serum levels of angiogenic factors in the postoperative patient with HCC were decreased. The serum levels of inhibitors of vascular endothelial cells in the postoperative patient with HCC were increased.

Acetyl-CoA C-acetyltransferase (AACT) is the first enzyme in the terpene synthesis pathway, catalyzed two units of acetyl-CoA to acetoacetyl-CoA. In order to study the tanshinone biosynthesis in Salvia miltiorrhiza, a novel AACT gene, SmAACT, was cloned using cDNA microarray and RACE strategy. The full length cDNA of SmAACT is 1 623 bp (accession No. EF635969), which contained a 1 200 bp open reading frame (ORF) encoding a 399 amino acid protein. Nine introns were found in the genomic sequence. SmAACT was upregulated by YE and Ag+ elicitors both with cDNA microarray and quantitative RT-PCR analyses along with the accumulation of tanshinones. Sequence homology comparison and phylogenetic analysis all suggested that SmAACT belonged to the class of acetyl-CoA C-acetyltransferase. The transcription level of SmAACT was relatively higher in root than that in stem and leaf tissues. SNP analysis revealed that SmAACT was highly variable in the region of 6 to 9 introns with 33 SNPs in the 600 bp region, there are 5 SNPs in the cDNA region while they are all synonymous cSNPs. Some special genotypes were found in Salvia miltiorrhiza from different areas. SmAACT will be an useful gene for further analyze the mechanism of gene regulation among the tanshinones biosynthesis.

OBJECTIVE:To establish a method for determination of the contents of chlorogenic acid and baicalin in Shiwuwei Xiaoyanzhike oral liquid .METHODS:HPLC was used,the mobile phase was acetonitrile - acetic acid,the flow rate was 0.8ml/ min and the detecting wavelengths were 326nm and 274nm.RESULTS: The recoveries of chlorogenic acid and baicalin were 96.8% - 104.5% , 94.9% -97.7% and RSD were 1.09% -3.12% , 1.19% -1.76% (n = 3), respectively .CONCLUSION: The method is simple and feasible with a good reproducibility, the RESULTS: can provide a basis for establishing the quality standard of this oral liquid.

In order to separate chiral D-phenylalanine from L-phenylalanine,the enzyme catalytic method by papain and immobilized papain were used in this investigation.It indicated that the yield rate of N-acetyl-DL-phenylalanine synthesized from DL-phenylalanine was 88.7%.The ionic strength was the strongest influencing factor in the synthesis of N-acetyl-L-phenylalanylaniline by both papain and immobilized papain on sodium alginate-ehitosan(IPSAC).But the reaction temperature was the strongest influencing factor in the synthesis by immobilized papain on nylon cloth(IPN).The ionic strength and the pH were the important influencing factors in the synthesis.The yield rates of the best synthesis system of N-acetyl-L-phenylalanylaniline by papain,IPSAC,IPN were 61.2%,54.7%,36.3%,respectively.The yield rate of L-phenylalanine from N-acetyl-L-phenylalanylaniline hydrolyzation was 59.2%.The optical purity of the product was 96.6%.The vield rate of D-phenylalanine from N-acetyl-D-phenylalanine hydrolyzation was 61.7%.The optical purity was 95.7%.

Objective To understand the characteristics of pathogenic bacteria and drug resistance of ventilator-associated pneu-monia(VAP)in neonatal intensive care unit (NICU),and to explore the corresponding prevention and control measures,and to pro-vide the basis for the VAP antibiotic treatment.Methods A total of 80 children with respiratory failure and ventilator assisted breathing were selected from the NICU as objects in this study.The clinical data,pathogenic bacteria and drug resistance were ana-lyzed retrospectively.Results In 80 cases,the incidence of VAP was 43.75% (35/80),a total of 70 strains of pathogenic bacteria were isolated,gram negative bacilli accounted for the highest proportion,accounting for 81.43% (57/70).Pseudomonas aeruginosa, Klebsiella pneumoniae were the most common gram negative bacilli.Gram positive cocci accounted for 8.57% (13/70),which domi-nated by methicillin resistant Staphylococcus aureus,in addition to susceptible to vancomycin,but resistant to the other antibiotics. Conclusion Gram negative bacillus are the main bacteria in VAP cases,and which are multiple drug-resistant pathogens.

Objective To knockout and identify the Antigen 43 (Ag43) in the Escherichia Coli JM109 .Methods Mutation group Ⅱ introns RNA protein complexes (RNP) gene sequence was obtained by Sigma Company′s TargeTron Gene Knockout Sys‐tem and Ag43 gene specific designed PCR primers amplification ,then ,to acquired Ag43 specific recombinant RNP plasmid pACD4K‐Ag4 ,this gene sequence was inserted into the plasmid pACD4K‐C of RNA′s expression .Finally ,pEGFP‐Ag43 was trans‐formed into JM109 and inserted the group Ⅱ intron into the Ag43′s locus by IPTG inducing expression .Results The best insertion locus was between 1 812 and 1 913 .Through the agarose electrophoresis gel ,the RNP gene sequence was consistent with the expec‐ted value (350 bp) .The pEGFP‐Ag43 vector was correctly constructed which was proofed by endonuclease Nhe Ⅰ and Hind ⅡI di‐gestion as predicted products (3 646 and 4 029 bp;7 000 and 550 bp ,respectively ) .The PCR and gene sequence results indicated that the group Ⅱ intron was inserted into the locus between 1 812 and 1 913 in the Ag43 gene .Conclusion Successful knockout of the Ag43 in Escherichia Coli JM109 found basis to further study the Ag43′s function and regard the coli as host bacteria of Ag43 chimeric protein recombinant .

Three plasmids (pGenesil-P1, pGenesil-P2, pGenesil-P3) with different p27Kip1-shRNA sequences were designed and synthesized. Their effects on the proliferation of bovine corneal endothelial cells (bCEC) were investigated. Plasmid expressing irrelevant shRNA with a random combination was used as negative control (pGenesil-HK). The recombination of four plamids was confirmed by restrictive enzyme digestion and sequence analysis. The expression of mRNA and protein of p27Kip1 was detected by RT-PCR and Western blotting after stable transfection. The expressions of p27Kip1 mRNA and p27Kip1 protein of pGenesil-P1 group, pGenesil-P2 group and pGenesil-P3 group were all lower than those in the pGenesil-HK group and the blank group (non-transfected group). pGenesil-P3 had the strongest inhibitory effect and was selected for the next steps. The proliferation rates of the pGenesil-P3 group, the pGenesil-HK group and the blank group were assessed by MTT. The influence of shRNA-p27Kip1 on bCEC cell cycle was detected by flow cytometry (FCM). Compared with the control groups, the proliferation rate of the pGenesil-P3 group was increased significantly, and the ratio of S-phase also increased. It is concluded that shRNA-p27Kip1 could down-regulate the expression of p27Kip1 effectively and increase the proliferation of bCEC. RNA interference (RNAi) may be an effective means to promote the proliferation of CEC.
Cell Proliferation ; Cornea/cytology ; Cyclin-Dependent Kinase Inhibitor p27/*metabolism ; Endothelial Cells/*cytology ; Endothelial Cells/metabolism ; Gene Expression Regulation ; Models, Biological ; Plasmids/metabolism ; RNA Interference ; RNA, Messenger/metabolism ; RNA, Small Interfering/metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Tetrazolium Salts/pharmacology ; Thiazoles/pharmacology ; Transfection