Identification and validation of a new target is one of the most important steps for new antituberculosis (TB) drug discovery. Researches have shown that Mycobacterium tuberculosis (Mtb) encodes 20 CYP450 enzymes which play important roles in the synthesis and metabolism of lipid, cholesterol utilization, and the electron transport of respiratory chain in Mtb. With the critical roles within the organism as well as the protein structures of six Mtb CYP450 enzymes being clarified, some of them have been highlighted as potential anti-tuberculosis targets. In this paper, the phylogenetic analysis, the structural features, and the enzymatic functions of Mtb CYPs, as well as the mechanism of interactions with selective inhibitors such as azole antifungal agents for the CYPs have been reviewed and summarized. The druggability of the CYPs has also been analyzed for their further utility as targets in high throughput screening and rational design of more selective inhibitors.
Antitubercular Agents ; chemistry ; pharmacology ; Azoles ; chemistry ; pharmacology ; Cytochrome P-450 Enzyme Inhibitors ; chemistry ; pharmacology ; Cytochrome P-450 Enzyme System ; genetics ; metabolism ; Drug Delivery Systems ; methods ; Drug Discovery ; Humans ; Mycobacterium tuberculosis ; drug effects ; enzymology ; genetics ; Phylogeny ; Tuberculosis ; drug therapy ; microbiology

Calpain ; genetics ; Diabetes, Gestational ; genetics ; Female ; Genotype ; Haploidy ; Humans ; Polymorphism, Single Nucleotide ; Pregnancy

[Objective] To investigate the pathogen spectrum and characteristics of diarrheal disea-ses among children under five and to provide the basis for prevention and control measures . [ Methods] From February 2012 to January 2013, a total of 619 cases from 2 sentinel hospitals were collected for case survey .From them were collected stool specimens for detection of enteric pathogenic bacteria and virus nu-cleic acids. [Results] The virus detection rate was norovirus (11.95%), rotavirus(7.27%), enteric adenovirus(2.91%), saporovirus(1.45%) and astrovirus(0.97%).The bacteria detection rate was E. coli(5.65%), nontyphoidal salmonella (2.42%), campylobacter (0.81%), aeromonas hydrophila (0.65%),shigella(0.48%) and yersinia enterocolitica (0.32%).The virus detection rate was 27.70%from August to next March, which showed higher than that of other months (18.48%) (χ2 =7.18,P <0.05).The bacteria detection rate was 15.34%from April to October, showing higher than the rate of oth-er months(3.15%)(χ2 =31.22,P<0.05). [Conclusion] Norovirus is the main pathogen for viral diarrheal diseases in children under five and E .coli is the main pathogen of bacterial diarrheal diseases . All study results suggested that summer and autumn should be as the focus time in prevention and control of bacterial diarrhea , while those of viral diarrhea should cover the whole year .

OBJECTIVETo study the effect of fluridone concentration, stimulating period, temperature and salt on the seed germination of three species of Cistanche.

METHODThe seeds were cultured in Petri dish, and the germination percentage was counted.

RESULTThe highest germination percentage was observed in Cistanche tubulosa, C. deserticola, C. sala seeds pre-treated by 0.1 mg x L(-1) fluridone for 24-29 h. The optimal temperature for the seeds germination of three species of Cistanche was at 20-30 degrees C, and the seeds did not germinate at sub-or supraoptimal temperatures (5 and 35 degrees C). The salt tolerance of C. sala seeds was strong, and the critical value of NaCl concentration was 0.04 mol x L(-1). By contrast, C. tubulosa and C. deserticola seeds were more sensitive to the salt stress, the critical value of NaCl concentration was 0.02 mol x L(-1).

CONCLUSIONThe optimal germination condition and the method of testing germination percentage of three species of Cistanche seeds are as follow: the seeds are pre-treated by 0.1 mg x L(-1) fluridone for 24 h and then cultured at 20-30 degrees C in salt solution which concentration is lower than 0.02 mol x L(-1).

Cistanche ; classification ; growth & development ; Germination ; drug effects ; physiology ; Plants, Medicinal ; growth & development ; Pyridones ; pharmacology ; Seeds ; growth & development ; Sodium Chloride ; pharmacology ; Species Specificity ; Temperature

·Familial exudative vitreoretinopathy ( FEVR ) is a hereditary disease associated with abnormal angiogenesis in the pediatric period. The most prominent finding of the disease is avascularity in the peripheral retina. Whereas, the phenotypic features are variable. In some minor cases, missed diagnosis would happened due to asymptom, while, in severe FEVR, neovascularization, retinal exudation, retinal folds, macular heterotopy and retinal detachment may occur and give rise to extremely poor vision or even blindness. Mutations in the FZD4, LRP5, NDP, TSPAN12, ZNF408, and KIF11 genes have been reported to contribute to FEVR with X - linked recessive, autosomal dominant, and autosomal recessive inheritance manners. We have summarized aspects of pathogenesis, clinical features and classification, mutations genes as well as diagnosis and treatment of FEVR in this review.

OBJECTIVETo investigate the effects of formaldehyde inhalation on the morphological damage, and Glu, GABA and NOS contents in olfactory bulb and hippocampus of rats.

METHODSTwenty SD rats were equally divided into two groups: rats in the control group inhaled fresh air, while the animals in experimental group were exposed to the air containing formaldehyde (12.5 mg/m(3), 4 h/d) for 7 days. Then rats were sacrificed and frozen sections of olfactory bulb and hippocampus were prepared. The morphological changes were examined and the Glu, GABA and NOS contents were detected using Nissl-staining, immunohistochemistry and Western blot, respectively.

RESULTCompared with the control group, there was a significant confusion and shrink of neuron morphology in experimental group, the number and staining intensity of Glu and NOS positive cells and protein contents were reduced. The protein expression of GABA was also decreased in the formaldehyde group.

CONCLUSIONFormaldehyde inhalation can cause a severe morphological damage of olfactory bulb and hippocampus in SD rats,which may further impair memory and learning ability through the reduction of Glu, GABA and NOS expression.

Animals ; Formaldehyde ; toxicity ; Glutamic Acid ; metabolism ; Hippocampus ; drug effects ; metabolism ; pathology ; Inhalation Exposure ; Learning ; drug effects ; Neurons ; drug effects ; metabolism ; pathology ; Nitric Oxide Synthase ; metabolism ; Olfactory Bulb ; drug effects ; metabolism ; pathology ; Rats ; Rats, Sprague-Dawley ; gamma-Aminobutyric Acid ; metabolism

Tyrosol, crenulatin and salidroside are the main active constituents of Rhodiola crenulata, with extensive pharmacological activities. In the study, grams of high purity tyrosol, crenulatin and salidroside were simultaneously separated from R. crenulata by the first time. Firstly, R. crenulata was extracted by 70% alcohol. Then, with the yields of three compounds as the index, the macroporous resin was optimized. At last, grams of high purity tyrosol, crenulatin and salidroside were isolated by D-101 macroporousresin, purified by column chromatography. Detected by HPLC, the purity of three compounds were higher than 98%. This method has the advantages of simple process and operation, less dosage of organic solvent, highly yield and reproducibility, suitable for the simultaneously preparation of tyrosol, crenulatin and salidroside.
Chemical Fractionation ; methods ; Chemistry, Pharmaceutical ; Chromatography, High Pressure Liquid ; Coumarins ; analysis ; isolation & purification ; Drugs, Chinese Herbal ; analysis ; isolation & purification ; Glucosides ; analysis ; isolation & purification ; Phenols ; analysis ; isolation & purification ; Phenylethyl Alcohol ; analogs & derivatives ; analysis ; isolation & purification ; Rhodiola ; chemistry

OBJECTIVETo transfect pcDNA3.1/hTM plasmids containing human thrombomodulin (hTM) gene into human umbilical vein endothelial cells (HUVECs), and investigate the expression of hTM and anticoagulating function of transfected HUVECs.

METHODSHUVECs were transfected with pcDNA3.1/hTM by lipofectin. Expression of hTM mRNA was determined by semi-quantitative RT-PCR, hTM antigen on HUVECs membrane by immunohistochemistry, and activated protein C (PC) in HUVECs by chronometry. By using a semiautomatic coagulator, the effect of the reacting liquid from transfected HUVECs mixed with PC from normal peripheral blood was assayed.

RESULTSAbout 10% HUVECs were transfected by pcDNA3.1/hTM with high-level hTM mRNA and protein expression. Activated PC produced by pcDNA3.1/hTM group, pcDNA3.1(+)/neo group and untransfected group was (2.80 +/- 0.43) microg/ml, (0.75 +/- 0.08) microg/ml and (0.85 +/- 0.11) microg/ml, respectively. APTT was (51.68 +/- 2.73) s, (38.38 +/- 2.44) s, (39.65 +/- 2.39) s, (33.93 +/- 1.73) s and (34.60 +/- 1.86) s and PT was (21.89 +/- 1.66) s, (20.56 +/- 1.74) s, (20.42 +/- 2.04) s, (19.57 +/- 1.36) s and (20.16 +/- 1.35) s in pcDNA3.1/hTM group, pcDNA3.1(+)/neo group, untransfected group, inactivating PC group and control, respectively.

CONCLUSIONSThe pcDNA3.1/hTM plasmid could be transfected into endothelial cells and expressed biologically functioning hTM protein on HUVECs membrane. Activated PC could inhibit intrinsic coagulation pathway obviously with slight effect on extrinsic pathway.

Cells, Cultured ; Endothelial Cells ; metabolism ; Endothelium, Vascular ; cytology ; Gene Expression Regulation ; Genes, Transgenic, Suicide ; Humans ; In Vitro Techniques ; Partial Thromboplastin Time ; Plasmids ; Protein C ; Prothrombin Time ; Thrombomodulin ; genetics ; Transfection ; Umbilical Veins ; cytology

OBJECTIVETo construct T vectors based on Xcm I recognition site and optimize the PCR fragments for its ligation.

METHODSWe firstly cloned the human histone H4 cDNA containing one Xcm I recognition site at both its 5' and 3' end into pCDNA 3.0 vector and then generated T vector with pCDNA 3.0 backbone by cutting the recombinant plasmid with Xcm I. To increase the ligation efficiency, the primers were firstly phosphorylated before DNA fragments amplification and then the PCR products were treated with Taq DNA polymerase and dATP after PCR amplification. Two DNA fragments with the length of 312 bp and 1 329 bp were ligated to it and the ligation mixture was transformed into E. coli DH5α competent cells and the positive rates of the transformants were evaluated by PCR and DNA agarose gel electrophoresis.

RESULTSOur results showed that the T vector produced by our method could ligate to the target DNA fragments with high efficiency. Besides, the phosphorylation state of the primers used for PCR amplification is also an important factor determining the cloning efficiency. What's more, as for longer DNA fragments, retreatment with Taq DNA polymerase and dATP after PCR amplification and purification could improve the ligation efficiency significantly.

CONCLUSIONOur protocol may overcome the dependence on blue/white screening to get positive clones and provide a potent way to generate T vectors and ligate them to the target PCR fragment.

Cloning, Molecular ; DNA, Complementary ; genetics ; Escherichia coli ; genetics ; Genetic Vectors ; Histones ; genetics ; Humans ; Polymerase Chain Reaction ; methods

OBJECTIVETo explore the influence on AM fungi infection rate and medicine quality of Pinellia ternate in the condition of three soil impact factors.

METHODSet the orthogonal test of three factors and levels. Determinate the AM fungi infection rate in early stage of mature & stage of mature of P. ternata, and the water content, water soluble extract, butanedioic acid content and alkaloid content of P. ternata tuber that be harvested also had be determinated.

RESULT AND CONCLUSIONWith the P levels to 30 mg x kg(-1) and 90 mg x kg(-1), AM fungi infection was the best when mixed inoculated of EM. Microbial agent inoculated played a decisive role in P. ternata growth and physiological activity, secondary influenced factor was P concentration, and the water stress was the minimal impact. Mixed inoculated of AM fungi and EM treatment with the low P levels (30, 90 mg x kg(-1)) proved better effect on enhancing the water extract content, anedioic acid and alkaloid content, while decreasing the water contents of P. ternata tuber.

Alkaloids ; chemistry ; Drugs, Chinese Herbal ; chemistry ; standards ; Fungi ; Pinellia ; chemistry ; microbiology ; Plant Extracts ; chemistry ; standards ; Soil