Objective To evaluate the performance of nested PCR in the detection of different fungi in paraffin wax embedded tissues. Methods Forty-four tissue samples were resected from rats infected with Fonsecaea monophora, patients with chromoblastomycosis, sporotrichosis or penicilliposis marneffei followed by preparation of paraffin wax embedded tissue sections for pathological examination and DNA extraction. Nested PCR was performed by using specific primers targeting the ribosomal DNA of Fonsecaea, Sporothrix and Penicillium marneffei, respectively. The sensitivity and specificity of nested PCR were analyzed and compared with those of pathological examination. Results The nested PCR showed positive results in 8 of 20 samples from rats with chromoblastomycosis, 7 of 10 samples from patients with sporotrichosis and all of the 10 samples from patients with penicilliposis marneffei, but not in the control samples. In the detection of Fonsecaea,Sporothrix schenki and Penicillium marneffei, the sensitivity was 40% ,70% and 100%, respectively, and the specificity was consistently 100%, for the nested PCR. Pathological examination revealed fungal elements in 95%, 70% and 80% of the corresponding samples, respectively. Conclusion Detection of fungal DNA in paraffin wax embedded tissue by nested PCR can be applied to the diagnosis of deep mycosis, especially to the diagnosis of penicilliposis marneffei.

A double targets of high throughput screening model for xanthine oxidase inhibitors and superoxide anion scavengers was established. In the reaction system of xanthine oxidase, WST-1 works as the probe for the ultra oxygen anion generation, and product uric acid works as xanthine oxidase activity indicator. By using SpectraMax M5 continuous spectrum enzyme sign reflectoscope reflector, the changes of these indicators' concentration were observed and the influence factors of this reaction system to establish the high throughput screening model were studied. And the model is confirmed by positive drugs. In the reaction system, the final volume of reaction system is 50 μL and the concentrations of xanthine oxidase is 4 mU x mL(-1), xanthine 250 μmol x L(-1) and WST-1 100 μmol x L(-1), separately. The Z'-factor of model for xanthine oxidase inhibitors is 0.537 4, S/N is 47.519 9; the Z'-factor of model for superoxide anion scavengers is 0.507 4, S/N is 5.388 9. This model for xanthine oxidase inhibitors and superoxide anion scavengers has more common characteristics of the good stability, the fewer reagent types and quantity, the good repeatability, and so on. And it can be widely applied in high-throughput screening research.
Enzyme Inhibitors ; pharmacology ; Free Radical Scavengers ; pharmacology ; High-Throughput Screening Assays ; Superoxides ; Uric Acid ; Xanthine ; Xanthine Oxidase ; antagonists & inhibitors

The chemical structures of P1 A was identified by complete acid hydrolysis, partial acid hydrolysis, periodate oxidation-Smith degradation, methylation analysis, IR and NMR. The results showed that P1 A had a backbone consisting rhamnose, mannose, glucose and galactose. The side chain possessed arabinose and xylose. 1-->, 1-->6 and non-reducing terminal linkages existed in polysaccharide P1A, but there are doubling amount of 1-->2 and 1-->4 linkages. Oxidable linkage of P1 A accounted for 45%, and inoxidable linkage of P1A accounted for 55%. Mannose, glucose and galactose were mainly linked by 1-->2 linkage. Rhamnose, arabinose and xylose were mainly linked by 1-->2 and 1-->4 linkages. PlA contained beta-Glc(1,6)-,beta-Gal(1,3)-,beta-Man(1,4)-beta-Rha,-Glc(1,4)-, Glc(1)-,-Gal(1,4)- and Man(1)-.
Acanthaceae ; chemistry ; Drugs, Chinese Herbal ; chemistry ; Magnetic Resonance Spectroscopy ; Molecular Structure ; Molecular Weight ; Polysaccharides ; chemistry

A GeXP based multiplex PCR assay was developed to simultaneously detect six different chicken respiratory viruses including H5, H7, H9 subtypes of avian influenza virus(AIV), new castle disease virus (NDV), infectious bronchitis virus(IBV) and infectious laryngotracheitis virus(ILTV). According to the conserved sequences of genes of each pathogen, seven pairs of specific primers were designed, and the reaction conditions were optimized. The specificity and accuracy of GeXP were examined using samples of single and mixed infections of virus. The sensitivity was evaluated by performing the assay on serial 10-fold dilutions of cloned plasmids. To further evaluate the reliability, thirty-four clinical samples were detected by GeXP. The corresponding specific fragments of genes were amplified. The detection limit of GeXP was 10(2) copies/microL when all of 7 pre-mixed plasmids containing target genes of six chicken respiratory viruses were present. In the detection of thirty-four clinical samples, the results of GeXP were accorded with the viral isolation completely. In conclusion, this GeXP assay is a rapid, specific, sensitive and high-throughput method for the detection of chicken respiratory virus infections. It can be applied in rapid differential diagnosis for clinical samples, and also provide an effective tool to prevent and control chicken respiratory diseases with similar clinical symptoms.
Animals ; Chickens ; Influenza A virus ; classification ; genetics ; isolation & purification ; physiology ; Influenza in Birds ; diagnosis ; virology ; Multiplex Polymerase Chain Reaction ; methods ; Poultry Diseases ; diagnosis ; virology ; Respiratory Tract Infections ; diagnosis ; veterinary ; virology

OBJECTIVETo observe the effect of carbon disulfide (CS(2)) on the expression of matrix metalloproteinase (MMP)-2, MMP-9 in mouse embryo and uterus tissues and to explore the mechanism of embryo toxicity induced by CS(2).

METHODSAt the phases of follicular development and embryonic implantation which was subdivided into early-implantation phase and late-implantation phase, mice were intraperitoneally exposed to CS(2) (the dosage was 631.4 mg/kg, and the volume was 0.1ml/10 g body weight) for 2 consecutive days. All indicators were got at the ninth day in gestation, and the expression of MMP-2 and MMP-9 in embryo and uterus tissues was analyzed by gelatin zymography.

RESULTSThe number of implanted embryos significantly decreased after exposure at late-implantation phase (16.000 ± 12.166) compared with those of the control (30.700 ± 5.599, P < 0.05). Expression of MMP-2 and MMP-9 in embryos declined obviously at the three reproductive phases (P < 0.01), and the levels of MMP-2 and MMP-9 expression in embryos at the phases of late-implantation phase (0.6837 ± 0.0929, 0.7309 ± 0.0822) and follicular development (0.6222 ± 0.0997, 0.7520 ± 0.1068) were much lower than those of the control (1.0000 ± 0.0710, 1.0000 ± 0.0413, P < 0.01). Expression of MMP-2 and MMP-9 in uterus significantly increased at the phase of late-implantation (1.3153 ± 0.3032, 5.0210 ± 4.0307) compared with those of the control (1.0000 ± 0.1771, 1.0000 ± 0.0996, P < 0.01).

CONCLUSIONEmbryo toxicity of CS(2) is more obvious at the phase of late-implantation. Exposure to CS(2) disturbs expression of MMP-2 and MMP-9 in embryo and uterus tissues, which might be one of the important factors contributed to embryo toxicity induced by CS(2).

Animals ; Carbon Disulfide ; toxicity ; Embryo Implantation ; Embryo, Mammalian ; drug effects ; metabolism ; Female ; Matrix Metalloproteinase 2 ; metabolism ; Matrix Metalloproteinase 9 ; metabolism ; Mice ; Mice, Inbred Strains ; Pregnancy ; Uterus ; drug effects ; metabolism

OBJECTIVETo investigate prevalence of human immunodeficiency virus (HIV) infection and risk factors for its transmission among injection drug users (IDUs) in Liangshan Yizu Autonomous Prefecture of Sichuan Province, China.

METHODSA community-based survey was conducted to investigate demographic characteristics, pattern and frequency of sharing injection equipment, and sexual behaviors in IDUs. Blood samples were also collected from them to detect for antibodies against HIV and syphilis.

RESULTSA total of 379 subjects were recruited with informed consent for study through community outreach and peer recruiting methods. Their prevalence of HIV infection was 11.3% (43/379). Ethnicity, frequency of sharing syringes and cotton swab during the past three months and syphilis infection associated with HIV infection by univariate analysis using chi-square test. Multivariate logistic regression analysis showed odds ratio of frequency of sharing syringes for HIV infection during the past three months was 2.28 (95% CI 1.18 - 4.43), and that for syphilis infection 3.10 (95% CI 1.48 - 6.48).

CONCLUSIONFrequency of sharing syringes during the past three months associated with syphilis and HIV infection.

Adolescent ; Adult ; China ; epidemiology ; Disease Transmission, Infectious ; Female ; HIV Infections ; epidemiology ; transmission ; Humans ; Logistic Models ; Male ; Needle Sharing ; adverse effects ; Prevalence ; Sexual Behavior ; Substance Abuse, Intravenous ; virology

OBJECTIVETo compare the difference in the clinical efficacy on cervical spondylosis between acupuncture at three lines of cervical Jiaji (EX-B 2) and oral administration of jingfukang granules.

METHODSThree hundred cases of cervical spondylosis were divided into an acupuncture group and a medication group, 150 cases in each one. In the acupuncture group, according to the different types of cervical spondylosis, acupuncture was applied at three lines of cervical Jiaji (EX-B 2), once a day. In the medication group, jingfukang granules were prescribed for oral administration, one bag each time, three times a day. The treatment of ten days made one session in the two groups and two sessions were required totally. Before and after two sessions of treatment, the clinical assessment scale for cervical spondylosis (CASCS) was adopted to evaluate the score of subjective symptoms, clinical physical signs and adaptability as well as the total score in the patients of the two groups and the efficacy was compared.

RESULTSThe patients' symptoms and physical signs were alleviated, the adaptability was improved and the score of each item and the total score were increased in the two groups after treatment (all P<0.01). The improvements in the acupuncture group were better than those in the medication group (all P<0.01). The curative and markedly effective rate was 90.7% (136/150) in the acupuncture group, better than 66.0% (99/150) in the medication group (P<0.01).

CONCLUSIONAcupuncture at three lines of cervical Jiaji (EX-B 2) achieves the significant clinical efficacy on cervical spondylosis. This therapy is superior to relieving symptoms and physical signs and recovering adaptability as compared with jingfukang granules.

Acupuncture Points ; Acupuncture Therapy ; Adult ; Female ; Humans ; Male ; Middle Aged ; Spondylosis ; therapy ; Treatment Outcome ; Young Adult

OBJECTIVETo explore the sensitive point of embryotoxicity of carbon disulphide on female mice.

METHODSAt the phases of follicle developing, implanting and post-implantation of blastocyst, female mice were injected intraperitoneally with carbon disulphide 631.4 mg/kg per day for three days while controls with plant oil. All indexes were detected at the fourteenth day of pregnancy.

RESULTS(1) In follicle developing CS(2) exposed group, the weight of embryos fossa [(1.23 +/- 0.36) g was 41% less than that in controls [(2.08 +/- 0.48) g, P = 0.000], and in implanting CS(2) exposed group the weight of embryo fossa, and embryos [(1.27 +/- 0.97) g, and (0.12 +/- 0.09) g respectively] were 39% and 37% less than those in controls [(2.08 +/- 0.48), (0.19 +/- 0.06) g, P = 0.068, P = 0.045]; (2) In both follicle developing and implanting CS(2) exposed group, the weights of uterus and placenta were also less than those in controls (P < 0.01). (3) In post-implantation CS(2) exposed group, the above parameters were not significantly different from those in controls.

CONCLUSIONFollicle developing phase as well as implanting of blastocyst may be sensitive point for embryotoxicity induced by carbon disulphide.

Animals ; Carbon Disulfide ; toxicity ; Embryo Implantation ; drug effects ; Embryonic Development ; drug effects ; Female ; Male ; Mice ; Mice, Inbred Strains ; Pregnancy ; Teratogens ; toxicity

OBJECTIVETo elucidate the effect of angiotensin II (AngII) on the expression of tissue factor (TF) by monocytes and its mechanisms.

METHODSMonocytes were isolated from healthy volunteers by Ficoll-Hypaque gradient and Percoll, and cultured in RPMI-1640. Procoagulant activity (PCA) was determined by one-stage clotting method, TF antigen by ELISA. Reverse transcriptase-polymerase chain reaction (RT-PCR) was used to detect the TF gene mRNA. The levels of IkappaBalpha was detected by Western blot. Electrophoretic mobility shift assays (EMSA) were performed to evaluate the activity of NF-kappaB.

RESULTSAngII (10(-9) - 10(-7) mol/L) significantly increased monocyte PCA, TF antigen and TF mRNA expression in a dose and time dependent manner. Losartan (10(-6) - 10(-5) mol/L) significantly inhibited the effects of AngII on TF activity, antigen and mRNA expression in a dose-dependent effects. Staurosporine (2.5 x 10(-7) mol/L) and genistein (4 x 10(-5) mol/L) lowered TF level of monocytes (P < 0.05). Western blot analysis revealed that after exposure to AngII (10(-7) mol/L), IkappaBalpha level decreased at 15 min, reached nadir at 60 min, and recovered at 180 min. EMSA showed NF-kappaB binding activity increased at 15 min, reached peak at 60 min, and recovered at 180 min. Pyrrolidine dithiocarbamate (PDTC, 10(-4) mol/L), an inhibitor of NF-kappaB, or AT1R antagonist losartan (10(-5)mol/L) inhibited AngII-induced NF-kappaB translocation.

CONCLUSIONSAngII could induce the expression of TF in human monocytes, and this effect was mediated by AT1R. The PKC pathway played the most important role in AngII-induced TF expression. The activation of NF-kappaB was involved in TF expression in monocytes.

Angiotensin II ; pharmacology ; Gene Expression Regulation ; drug effects ; Genistein ; pharmacology ; Humans ; Losartan ; pharmacology ; Monocytes ; drug effects ; metabolism ; NF-kappa B ; metabolism ; Protein Kinase C ; physiology ; RNA, Messenger ; analysis ; Receptor, Angiotensin, Type 1 ; physiology ; Staurosporine ; pharmacology ; Thromboplastin ; genetics

OBJECTIVETo investigate the effects and mechanisms of Curcumol beta-cyclodextrin Compound (CbetaC) on the proliferation and apoptosis of esophageal carcinoma cell line TE-1.

METHODSThe CbetaC was prepared by saturated solution and confirmed by infrared absorption spectroscopy. The effects of CbetaC (at 25, 50, 100 mg/L) on the proliferation of human esophageal carcinoma cell line TE-1 in vitro was analyzed by MTT assay. The cell cycles and apoptosis were detected by flow cytometer. The relative expression of survivin mRNA was detected by real-time fluorescent quantitative PCR and calculated by the 2(-deltaCt) method. The protein expression of survivin was measured by Western blot.

RESULTSCompared with the control group, results of MTT showed that CbetaC at each dose significantly inhibited the proliferation of TE-1 cells in a dose-dependent manner (P < 0.05). The results of flow cytometry showed that CbetaC resulted in the cell cycle arrest at G0/G1 and G2/M phase, and promoted the cell apoptosis. Besides, when compared with the control group, the protein and mRNA expressions of survivin obviously decreased in each CbetaC group (P < 0.05).

CONCLUSIONSCbetaC could inhibit the proliferation of esophageal carcinoma cell TE-1 and promote the apoptosis. Its inhibition on the survivin expression was correlated with its inhibition on the malignant phenotypes of esophageal carcinoma cells.

Apoptosis ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Esophageal Neoplasms ; pathology ; Humans ; Sesquiterpenes ; pharmacology ; beta-Cyclodextrins ; pharmacology