In order to develop a rapid detection kit for novel avian influenza virus (AIV) subtype H7N9, two sets of specific primers and probes were designed based on the nucleotide sequences of hemagglutinin antigen (HA) and neuraminidase antigen (NA) of novel H7N9 virus (2013) available in GenBank to establish the method of TaqMan probe-based multiplex real-time RT-PCR for rapid detection of AIV subtype H7N9. The primer and probe of HA were for all H7 subtype AIVs, while the primer and probe of NA were only for novel N9 subtype AIVs. The results showed that this method had high sensitivity and specificity. This method was applicable to the testing of positive standard sample with a minimum concentration of 10 copies/microL; it not only distinguished H7 subtype from H1, H3, H5, H6, and H9 subtypes, but also distinguished novel N9 subtype from traditional N9 subtype. A total of 2700 samples from Zhuhai, China were tested by this method, and the results were as expected. For the advantages of sensitivity and specificity, the method holds promise for wide application.
Animals ; Birds ; virology ; Influenza A Virus, H7N9 Subtype ; genetics ; isolation & purification ; physiology ; Influenza in Birds ; prevention & control ; virology ; Real-Time Polymerase Chain Reaction ; methods ; Species Specificity ; Taq Polymerase ; metabolism ; Time Factors

OBJECTIVETo investigate nuclear factor kappa B (NF-kappaB) activation induced by lipopolysaccharide (LPS) in rat peritoneal macrophages (PMAs) and the inhibitory effect of resveratrol on NF-kappaB activation.

METHODSPMAS from normal SD rats were randomly divided into 7 groups, including a control group, a LPS group and 5 resveratrol groups (I-V). PMAs of the control group were incubated in DMEM, and those in LPS group in DMEM containing LPS (10 microg/ml). PMAS of resveratrol groups I-V were incubated in DMEM containing LPS (10 microg/ml) and different concentrations of resveratrol. After 24 h of incubation, NF-kappaB activation in the PMAs was determined, and the expression levels of tumor necrosis factor alpha (TNF-alpha), interleukin-1 (IL-1) and nitric oxide (NO) in the culture medium were measured.

RESULTSExposure to LPS resulted in an excessive enhancement of cytokine and NO expressions in the PMAs. Resveratrol at 1.25-10 microg/ml produced a dose- dependent inhibition of cytokine and NO expressions and on NF-kappaB activation in LPS-stimulated PMAs.

CONCLUSIONResveratrol can inhibit LPS-induced NF-kappaB activation in rat PMAs and subsequently suppress the expressions of TNF-alpha, IL-1 and NO.

Animals ; Cytokines ; metabolism ; Lipopolysaccharides ; pharmacology ; Macrophage Activation ; drug effects ; Macrophages, Peritoneal ; drug effects ; immunology ; metabolism ; Male ; NF-kappa B ; metabolism ; Nitric Oxide ; metabolism ; Rats ; Rats, Sprague-Dawley ; Stilbenes ; pharmacology

In order to study the proliferation inhibition effect of H5N1 subtype avian influenza virus (AIV) with small interfere RNA (siRNA), a total of 4 siRNAs were designed in accordance with the NP and PA genes of H5N1 subtype AIV, the siRNAs were then transfected to chicken embryo fibroblast(CEF), CEF was infected with H5N1 subtype AIV after 6 hrs. Virus titer of cell supernatant was tested at 16-56hrs post infection, and pathological changes of the cells was observed; mRNA levels of NP, PA, HA and p13-actin gene were tested at 36hrs post infection. The results showed that these 4 siRNAs could inhibit the prolif-eration of H5N1 subtype AIV in CEF in varying degrees, and one siRNA targeting PA was best per-formed. The experimental results also showed that the inhibition effect was decreased with the time prolonged. This research provides a basis for further studying RNAi on AIV prevention and control.
Actins ; genetics ; Animals ; Chick Embryo ; DNA Primers ; genetics ; Fibroblasts ; virology ; Hemagglutination ; Hemagglutinin Glycoproteins, Influenza Virus ; genetics ; Hemagglutinins ; genetics ; Humans ; Influenza A Virus, H5N1 Subtype ; genetics ; growth & development ; physiology ; RNA Interference ; RNA Replicase ; genetics ; RNA, Small Interfering ; chemical synthesis ; genetics ; RNA-Binding Proteins ; genetics ; Real-Time Polymerase Chain Reaction ; Specific Pathogen-Free Organisms ; Transfection ; Viral Core Proteins ; genetics ; Viral Proteins ; genetics ; Virus Replication

OBJECTIVETo investigate the mutation characteristics of DJ1 gene in Chinese patients with autosomal recessive early-onset Parkinsonism (AR-EP).

METHODSMutations of DJ1 gene were screened by polymerase chain reaction combined with DNA direct sequencing in index patients with AR-EP from 11 unrelated families.

RESULTSNo pathogenetic mutations in the DJ1 gene were detected in this group. Six intronic DJ1 polymorphisms (IVS1-15T-->C, IVS4+30T-->G, IVS4+45G-->A, IVS4+46G-->A, IVS5+31G-->A, g.168-185del) were found. Three of them (IVS1-15T-->C, IVS4+45G-->A, IVS4+46G-->A) were not reported previously.

CONCLUSIONDJ1 mutations were rare in Chinese patients with autosomal recessive early-onset Parkinsonism.

Adolescent ; Adult ; Age of Onset ; Base Sequence ; China ; epidemiology ; DNA Mutational Analysis ; methods ; Female ; Humans ; Intracellular Signaling Peptides and Proteins ; genetics ; Mutation ; Oncogene Proteins ; genetics ; Parkinsonian Disorders ; epidemiology ; genetics ; Polymerase Chain Reaction ; Protein Deglycase DJ-1 ; Young Adult

OBJECTIVEThis study sought to isolate and identify the proteins that interact with ataxin-3, to confirm the interacted domain, and to provide new clues for exploring the function of ataxin-3 and the pathogenesis of spinocerebellar ataxia type 3 and Machado-Joseph disease (SCA3/MJD).

METHODSYeast two-hybrid screen (MATCHMAKER GAL4 Two-Hybrid System 3) and regular molecular biologic techniques were undertaken to screen human brain cDNA library with mutant ataxin-3 bait. Two baits from both normal and mutant C-terminus of ataxin-3 were created by subcloned methods to determine which domain of ataxin-3 interacts with the putative associated proteins and to find out optimal candidate proteins that interact with C-terminus of ataxin-3. Confocal microscope was used to observe whether ataxin-3 co-localized with the obtained interacting proteins in mammalian cells.

RESULTSFive novel ataxin-3 interacting proteins were obtained, among which were three known proteins, namely human rhodopsin guanosine diphosphate dissociation inhibitor alpha, small ubiquitin-like modifier 1, and human neuronal amiloride-sensitive cation channel 2; the other two were unknown. Interacting domain analysis revealed that an unknown protein interacted with the C-terminus near the polyglutamine tract of ataxin-3, the other four all interacted with the N-terminus. In the nucleus of SH-SY5Y cell, small ubiquitin-like modifier 1 co-localized with the wild-type ataxin-3 and with the intranuclear aggregates formed by the mutant ataxin-3.

CONCLUSIONAn unknown protein probably interacting with C-terminus of ataxin-3 is firstly discovered, and the initiative findings suggest first that the interaction of small ubiquitin-like modifier 1 with N-terminus of ataxin-3 and the relevant sumoylation probably participate in the post-translation modifying of ataxin-3 and in the pathogenesis of SCA3/MJD.

Acid Sensing Ion Channels ; Ataxin-3 ; Cell Line, Tumor ; Green Fluorescent Proteins ; genetics ; metabolism ; Humans ; Microscopy, Confocal ; Mutation ; Nerve Tissue Proteins ; genetics ; metabolism ; Nuclear Proteins ; genetics ; metabolism ; Plasmids ; genetics ; Protein Binding ; Recombinant Fusion Proteins ; genetics ; metabolism ; Repressor Proteins ; genetics ; metabolism ; SUMO-1 Protein ; genetics ; metabolism ; Sodium Channels ; genetics ; metabolism ; Transfection ; Two-Hybrid System Techniques

OBJECTIVETo study the characteristics of the mutation of small heat-shock protein 22 (HSP22) gene in Chinese patients with Charcot-Marie-Tooth (CMT) disease.

METHODSA CMT2L proband with 423(G--> T) mutation in HSP22 gene had been studied and reported by the present authors. In this study, mutation analysis of HSP22 gene was performed using polymerase chain reaction and DNA direct sequencing in 114 CMT probands.

RESULTSIn the 114 CMT probands, a 582(C--> T)(T194T)samesense mutation was found in two unrelated families.

CONCLUSIONThe rate of HSP22 gene mutation in Chinese patients with CMT is as low as 0.87%(1/115).

Asian Continental Ancestry Group ; genetics ; Charcot-Marie-Tooth Disease ; ethnology ; genetics ; China ; DNA Mutational Analysis ; Heat-Shock Proteins, Small ; genetics ; Humans ; Mutation ; Polymerase Chain Reaction

OBJECTIVETo make technical standard of acupuncture manipulation for acupuncture treatment of heroin withdrawal syndrome.

METHODSTwo hundred and twenty cases of heroin withdrawal syndrome were randomly divided into an acupuncture group of 111 cases and a control group of 109 cases. They were respectively treated with acupuncture and oral administration of lofexidine hydrochloride, and their therapeutic effects were observed.

RESULTSThe heroin dependence (acute stage) were effectively withdrawn in the two groups. The treatment group in change of total scores for withdrawal symptoms before and after treatment, the total scores for withdrawal symptoms at the 4th and 5th days, treatment of insomnia and the score for self-Hamilton Anxiety Scale and the score after at the 4th day was superior to the control group (P < 0.05, P < 0.01, P < 0.001).

CONCLUSIONAcupuncture has a satisfactory, rapid, safe and reliable clinical therapeutic effect.

Acupuncture Therapy ; Anxiety ; Heroin ; Heroin Dependence ; Humans ; Substance Withdrawal Syndrome

OBJECTIVETo detect the duplication or deletion of peripheral myelin protein 22(PMP22) gene in Chinese patients with Charcot-Marie-Tooth disease(CMT) or hereditary neuropathy with liability to pressure palsies(HNPP) using real-time quantitative polymerase chain reaction.

METHODSDuplications or deletions of PMP22 gene were detected in 113 CMT cases, 4 HNPP cases and 50 normal controls by using real-time quantitative PCR.

RESULTSThirty-six of 113 CMT cases had the PMP22 duplication, 4 HNPP cases had the PMP22 deletion. No duplication or deletion was found in 50 normal controls.

CONCLUSIONThe PMP22 duplication rate in Chinese patients with CMT is 31.9%(36/113). PMP22 deletion is the common cause of HNPP.

Adult ; Charcot-Marie-Tooth Disease ; genetics ; Female ; Gene Duplication ; Humans ; Male ; Myelin Proteins ; genetics ; Polymerase Chain Reaction ; methods ; Sequence Deletion ; Young Adult

OBJECTIVETo investigate the features of small heat shock protein 27 (HSP27) gene mutation in Chinese patients with Charcot-Marie-Tooth disease (CMT).

METHODSDNA samples from 114 CMT probands were screened for mutations in HSP27 gene by polymerase chain reaction and direct sequencing, and haplotype analysis was further carried out on the mutation detected families.

RESULTSOne missense mutation C379T was detected in 4 autosomal dominant CMT2 families. Haplotype analysis indicated that the 4 families probably had a common ancestor.

CONCLUSIONTo the authors' knowledge, this is the first report of HSP27 gene mutation in Chinese patients with CMT, but it may be not common(0.90%). The C379T mutation in HSP27 gene also causes CMT2 except for distal hereditary motor neuropathy, thus providing further evidence that even the same mutation in the same gene may lead to distinct phenotypes.

Asian Continental Ancestry Group ; genetics ; Base Sequence ; Charcot-Marie-Tooth Disease ; ethnology ; genetics ; DNA Mutational Analysis ; methods ; Female ; HSP27 Heat-Shock Proteins ; genetics ; Haplotypes ; Humans ; Male ; Mutation ; Mutation, Missense ; Pedigree

OBJECTIVETo investigate the children with hearing loss from the age 0 to 6, and discuss the found age, found way and audiological characteristics.

METHODSGeneral information of found age and found way of 265 children, were investigated with self-made questionnaire and routine audiological evaluations, and then made statistical analysis.

RESULTSThe average (x +/- s) found age for the children with hearing loss was (23.21 +/- 10.02) months, and the first average coming age was (28.01 +/- 13.41) months. The found age of girls [(27.11 +/- 13.13) months] was 6.1 months later than the boys' [(21.03 +/- 12.32) months] and the countryside children [(28.27 +/- 11.09) months] later than the city's [(19.52 +/- 13.05) months] 8.65 months in the average found age. The found age of children who were found with speech disability was later than others. As the hearing loss degree of children went milder, the found age might later.

CONCLUSIONSThe popularization of knowledge in preventing from hearing loss must be strengthened. It is also necessary to popularize newborn hearing screening and early intervention while to enhance the parents' consciousness.

Child ; Child, Preschool ; Female ; Hearing Loss ; diagnosis ; prevention & control ; Hearing Tests ; Humans ; Infant ; Infant, Newborn ; Male ; Mass Screening ; Surveys and Questionnaires