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 study the clinicopathologic features, diagnosis and differential diagnosis of epithelioid hemangioma.

METHODSThe morphologic features of 7 cases of epithelioid hemangioma of skin, bone and venous vessels were studied.

RESULTSThere were altogether 4 male and 3 female patients (median age = 34 years; age range from 14 to 54 years). The 3 skin cases presented as single or multiple erythematous to bluish nodules or papules, with or without itchiness. The 2 bone cases appeared as osteolytic expansile lesions on radiologic examination. The remaining 2 cases involved medium-sized venous structures and presented as small isolated nodules in soft tissue. Histologically, the lesions were characterized by the presence of exuberant endothelial proliferations with various degree of inflammatory reaction. The neoplastic endothelial cells were plump, eosinophilic and polygonal, forming vascular channels. Occasional solid sheet-like arrangement was demonstrated. Intracytoplasmic vacuoles were commonly identified, indicating formation of primary lumen. The surrounding stroma contained various number of eosinophils and lymphoplasmacytic cells. Immunohistochemical study showed that the tumor cells were positive for endothelial markers (CD31 and CD34) and negative for epithelial marker (cytokeratin). Follow-up information was available in 6 cases. The duration of follow-up ranged from 5 to 36 months (median = 14 months). There was no evidence of recurrence or distant metastasis.

CONCLUSIONSEpithelioid hemangioma is a rare benign curable lesion which can be multifocal, involving skin, soft tissue and bone. It needs to be distinguished from Kimura's disease and epithelioid hemangioendothelioma.

Adolescent ; Adult ; Angiolymphoid Hyperplasia with Eosinophilia ; pathology ; Antigens, CD34 ; metabolism ; Bone Neoplasms ; metabolism ; pathology ; surgery ; Diagnosis, Differential ; Female ; Follow-Up Studies ; Hemangioendothelioma, Epithelioid ; pathology ; Hemangioma ; metabolism ; pathology ; surgery ; Humans ; Male ; Middle Aged ; Platelet Endothelial Cell Adhesion Molecule-1 ; metabolism ; Skin Neoplasms ; metabolism ; pathology ; surgery

OBJECTIVETo observe the peripheral neurotoxicity of 1-bromopropane (1-BP) by developing an animal model of peripheral neuropathy through oral administration of 1-BP.

METHODSForty male Wistar rats were randomly and equally divided into low-dose group (200 mg/kg), medium-dose group (400 mg/kg), high-dose group (800 mg/kg), and control group. The rats in the low-dose, medium-dose, and high-dose groups were orally given 1-BP (dissolved in corn oil), while the rats in the control group were orally given an equal volume of corn oil. The oral administration (0.2 ml/100 g BW) was performed once per day, 5 days per week, for 16 consecutive weeks. Neurobehavioral indices including gait score, hindlimb grip strength, and hindlimb landing foot splay were recorded periodically. Hematological and biochemical parameters were also measured during and after 1-BP exposure.

RESULTSThe gait scores were significantly higher in the high-dose group (after 8 ∼ 16 weeks of 1-BP exposure), medium-dose group (after 14 ∼ 16 weeks of 1-BP exposure), and low-dose group (after 15 ∼ 16 weeks of 1-BP exposure) than in the control group (P < 0.05, P < 0.01). Compared with the control group, the high-dose group showed significantly decreased hindlimb grip strength after 9, 12, and 14 weeks of 1-BP exposure (P < 0.05, P < 0.01), with the hindlimbs paralyzed after 16 weeks of 1-BP exposure. After 16 weeks of 1-BP exposure, the hindlimb grip strengths of rats in the medium-dose and low-dose groups were decreased to 72.6% and 91.2% of the control value (P < 0.01, P < 0.05). Compared with the control group, the high-dose group showed significantly increased hindlimb landing foot splay after 12, 14, and 16 weeks of 1-BP exposure, and the medium-dose group showed significantly increased hindlimb landing foot splay after 14 and 16 weeks of 1-BP exposure (P < 0.05, P < 0.01). The high-dose and medium-dose groups showed significantly higher serum alanine aminotransferase (ALT) activity than the control group after 8 weeks of 1-BP exposure, and so did the low-dose group after 16 weeks of 1-BP exposure (P < 0.01).

CONCLUSIONThe nervous system is sensitive to the toxic effect of 1-BP, and 1-BP exposure can induce peripheral neuropathy in rats.

Animals ; Disease Models, Animal ; Hydrocarbons, Brominated ; administration & dosage ; toxicity ; Male ; Peripheral Nervous System Diseases ; chemically induced ; physiopathology ; Rats ; Rats, Wistar

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 investigate the feasibility of life span extension of transformed human embryonic tendon cells (THETC) by reconstitution of the telomerase activity.

METHODSTHETC were transfected by pGRN145 plasmid containing the human telomerase reverse transcriptase (hTERT) cNDA in vitro by molecular cloning technique. The biological characteristics of transfected cells were detected and compared by morphological observation, plate cloning efficiency, soft agar culture, growth curve of cells cultured in different conditions, immunohistochemistry, telomerase activity assay by telomeric repeat amplification protocol (TRAP).

RESULTSThe THETC transfected by pGRN145 plasmid (telT) could express the telomerase activity with extension of life span. The telT maintained the original characteristics of temperature-dependant and serum-dependant, as well as secretion of type I collagen normally and without tendency of malignant transformation.

CONCLUSIONSThe life span of THETC can be prolonged by reconstitution of telomerase activity, which provides the novel experimental methods to establish the standard cells line.

Cell Line ; Cell Survival ; Embryo, Mammalian ; Humans ; Plasmids ; genetics ; RNA-Directed DNA Polymerase ; Telomerase ; genetics ; metabolism ; Tendons ; cytology ; enzymology ; Transfection