Study on the diterpenoids from radix illicii Maji and their antiviral activity against coxsackie B virus / 国际药学研究杂志

Objective To investigate he diterpenoids from he roots of Illicium majus(Radix Illcii Maji) and their antiviral activity against the Coxsackie B virus. Methods The compounds were isolated by column chromatography over silica gel, octadecylsi-ane chemically bonded silica gel(ODS), and Sephadex HL-20 coupled with preparative HPLC. Their stuctures were elucidated by spectroscopic analysis and the in situ dimolybdenum circular dichroism(CD) method, and their antiviral activities against the Coxsackie B3 virus were evaluated by cytopathic effect(CPE) method. Results Twelve diterpenoids were isolated from the roots of Illicium ma-jus, which were identified as 4-epi-dehydroabietic acid(l), 8,11,13,15-abietatraen-19-oic acid(2), jiadifenoic acids B(3), C(4), G(5) and 1(6), majusanic acids B(7) and D(8), lambertic acid(9), angustanoic acids F(10) and G(ll), and 13-hydroxy-8,11, 13-podocarpatrien-19-oic acid(12). These diterpenoids displayed antiviral activity against the Coxsackie B3 virus, with IC50 values of 3. 3-66. 7 μmol/ml. Conclusion The antiviral activity and cytotoxicity of the diterpenoids relate o he substituent species and position. Compounds 3-6 and 9 were obtained from his plant for the first time.

Development of a new method for diagnosis of Group B Coxsackie genome by reverse transcription loop-mediated isothermal amplification.

Background: Coxsackie B viruses (genus, Enterovirus; family, Picornaviridae) can cause aseptic meningitis, encephalitis, pleurodynia, and fatal myocarditis, and are implicated in the pathogenesis of dilated cardiomyopathy. The differentiation of the group B Coxsackieviruses into their subtypes has potential clinical and epidemiological implications. Objective: In this study, we developed a one-step, single-tube genogroup-specific reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for the detection of group B Coxsackie genomes targeting 5′ UTR region. Materials and Methods: The amplification can be obtained in less than 1 hour by incubating all the reagents in a single tube with reverse transcriptase and Bst DNA polymerase at 63°C. Detection of gene amplification could be accomplished by agarose gel electrophoresis and the monitoring of gene amplification can also be visualised with the naked eye by using SYBR green I fluorescent dye. Results: A total of 40 samples comprising 31 positive samples and 9 negative samples were used in this study for comparative evaluation. The results were compared with those from Real-Time Polymerase Chain Reaction (RT-PCR). None of the RT-PCR-positive samples were missed by RT-LAMP, thereby indicating a higher sensitivity of the RT-LAMP assay. Conclusion: Thus, due to easy operation without a requirement of sophisticated equipment and skilled personnel, the RT-LAMP assay reported here is extremely rapid, cost-effective, highly sensitive, and specific and has potential usefulness for rapid detection of non-polio enterovirus (NPEV) not only by well-equipped laboratories but also by peripheral diagnostic laboratories with limited financial resources in developing countries.

Advancement of the relationship between coxsackie virus infection and type 1 diabetes / 国际儿科学杂志

In recent years,the studies have shown that the incidence of type 1 diabetes(T1D)is closely related to the infection of human enteroviruses,particularly Coxsackie B Viruses(CVB).On one hand.the researches show that vires infection can induce T1D,and it is associated with the viral amount.the viral strain,and the host microenvironment.On the other hand,the studies confirm that viral infection also can inhibit the incidence of T1D.In terms of the factors,which is related to the vires infection-inducing.T1D.and the possible mechanism of the virus infection-inhibiting-T1D,the article aims to explore the relationship between CVB infection and the incidence of T1D based on referring to foreign literatures in recent years.

Circulating Adhesion Molecules in Patients with Keshan Disease and Their Relationship with Coxsackie B Virus Infection / 华中科技大学学报（医学）（英德文版）

This study determined the levels of serum soluble intercellular adhesion molecule-1 (sI-CAM-1) and soluble vascular cell adhesion molecular-1 (sVCAM-1) in patients with different types of Keshan disease (KD),examined the relationship between Coxsackie B virus-specific lgM antibody (CBV-IgM) and sICAM-1 or sVCAM-1 in KD patients,and investigated the role of these adhesion molecules in the pathogenesis of KD and their clinical implications.The levels of serum slCAM-1,sVCAM-1 and CBV-IgM were measured by using enzyme-linked immunosorbent assay in 22 patients with chronic Keshan disease (CKD),27 with latent Keshan disease (LKD) and 28 healthy controis.The subjects in different groups were adjusted for sex and age.Echocardiography was adopted to determine left ventrieular ejection fraction (LVEF) in 22 patients with CKD.The results showed that CKD patients had significantly higher levels of sICAM-1 and sVCAM-1 than LKD patients and healthy controls (P<0.01 for all).And there was significant difference in the levels of the 2 adhesion molecules between LKD patients and healthy controls (P<0.05).A negative correlation was found between LVEF and sICAM-1 or sVCAM-1 in CKD patients.The percentage of CBV-specific IgM positive individuals in KD patients was significantly higher than that of healthy controls.In CVB-specific IgM positive patients,the levels of serum sICAM-1 and sVCAM-1 were significantly greater than those in CBV-specific IgM negative counterpart.It was concluded that the increase in the levels of sICAM-1 and sVCAM-1 suggests the progression of inflammation in KD.sICAM-1 and sVCAM-1 can promote the development of myocardial pathology and lead to poor myocardial function.The increased serum sICAM-1 and sVCAM-1 in KD patients may be related to CBV infection.

Construction of double siRNA expressing vector of Coxsackie virus B4 / 吉林大学学报(医学版)

Objective To construct the double hairpin siRNA and green fluorescent protein(GFP) expressing vector pU6/double-siRNA/Neo/GFP/1A/2A to interfere 1A and 2A gene of Coxsackie virus B4.Methods 21 bp fragments of the Coxsackie virus B4 2A and 1A gene were chosen as the targets and 65 bp complimentary fragments were synthesized,then the target gene fragments were cloned into pSilencer2.1U6 Neo and pGCsi-U6/Neo/GFP/siNeGative,respectively,then the double siRNA expressing vector pU6/double-siRNA/Neo/GFP/1A /2A was constructed by restrict endonuclease digestion,elctrophoresis isolation and reclaimer,ligatied by T4 DNA ligase;then the double siRNA expressing plasmid was transfected into Hela cells,and the GFP was observed under fluorescent microscope.Results The correct results showed that the recombinant plasmid had the correct special fragments and DNA sequence detected by restrict endonuclease digestion, electrophoresis and DNA sequencing;and GFP was also observed in Hela cells tansfected with pU6/double-siRNA/Neo/GFP/1A /2A under fluorescent microscope more than 15 d after transfection.Conclusion The double siRNA expressing vector pU6/double-siRNA/Neo/GFP/1A/2A is constructed successfully;it has the correct target viral gene sequences and can express GFP gene in Hela cells more than 15 d after transfection.

Immunological Manifestation of Children with Recurrent Respiratory Tract Inflammation Caused by Superinfection / 实用儿科临床杂志

Objective To investigate the influence of coxsackie B virus (CVB) and mycoplasma pneumoniae (MP) superinfection on immunological manifestation of recurrent respiratory tract inflammations(RRI) in children.Methods One hundred and thirty-two hospitalized children with RRI between Jan.to Dec.2005 were divided into negative control group,MP infection group,CVB infection group and superintection group by determining anti-MP IgM and anti-CVB IgM.Blood sedimentation,C-reactive protein,IgG,IgA,IgM and T lymphocyte subpopulation etc.were determined in four groups.The anti-MP IgM was determined by specificity immune agglutination test.Enzyme linked immunosorbent assay was used to detect anti-CVB IgM.The IgG,IgA,IgM were determined by simple agar diffusion method.T lymphocyte subpopulation was tested by flow cytometry.Results The percentage of CVB infection was 32.1%,mainly expressed in the 1-3 years old children;the percentage of MP infection was 22.7%,mainly expressed in children over 3 years old.In both CVB and MP infection group,the ten-dency of IgG increased and that of IgA decreased.IgM in the CVB and MP superinfection group was obviously higher than that in negative control group and MP infection group (Pa

Construction of recombinant nonstructural P_2C plasmid in coxsackie B4 virus / 中国免疫学杂志

Objective:To construct the recombinant nonstructural P_2C plasmid in coxsackie B4 virus.Methods:Using the RT-PCR,we fishing the nonstructural protein P_2C cDNA fraction from Coxsackie B4 virus(CVB4) and cloning into the PUCm-T vector, then transfer it into E.coli to construct the recombinant plasmid.Results:Using the total RNA of Coxsackie B4 virus as model, we amplied 987 bp fraction and cloned into PUCm-T vector identifying with BamH Ⅰ and Hind Ⅲ double enzyme.Its products were consistent with nonstructural protein P_2C PCR productions.Conclusion:The recombinant nonstructural P_2C plasmid was constructed succesfully in coxsackie B4 virus.

Comparison Study between Immunohistochemical Staining and Enzyme Linked Immunosorbent Assay in Metalloproteinases and Tissue inhibitor of Metalloproteinases in Virus induced Myocarditis

PURPOSE: Most tissue disruption of extracellular matrix is mediated by extracellular proteinases. Matrix metalloproteinases(MMP) and tissue inhibitor of matrix metalloproteinases(TIMP) are associated with wound healing and repair. There has been no study done on MMP and TIMP in myocarditis. METHODS: Coxsackie B virus(4,000 plaque forming unit) was injected into Balb/c mice by intraperitoneal injection. Histopathological finding was observed by H-E staining. Immunohistochemical staining was performed using the Labelled Streptavidin Biotin(LSAB) kit for MMP-2, TIMP-2, and Interleukin(IL) 6. The results were compared to the serum levels by ELISA method. RESULTS: MMP-2 was strongly expressed in complicated myocarditis such as calcification, severe fibrosis, thrombosis, or dilated cardiomyopathy compared to normal or uncomplicated myocarditis. TIMP-2 expression was decreased in severe myocarditis. Serum MMP-2 levels were significantly higher in complicated myocarditis, but TIMP-2 levels were significantly lower. There was significant correlation between the grade of immunohistochemical staining and serum MMP-2 or TIMP-2 level by ELISA method. IL-6 was strongly expressed in immunohistochemical staining according to the severity of inflammation. CONCLUSION: There was significant correlation between grade of immunohistochemical staining and serum levels of MMP or TIMP by ELISA method. Accurate estimations of serum MMP and TIMP levels would be useful for the diagnosis and follow up of myocarditis.

Virus Coxsackie B em crianças com miocardite aguda / Coxsackie B virus in children with acute myocarditis

Estudar aspectos etiopatológicos e de evoluçäo em portadores de miocardite. Dentre 44 crianças com miocardite aguda estudadas clínica e virologicamente, foram selecionados 16 casos positivos para Coxasackie B. O protocolo clínico incluiu dosagens enzímicas, radiografia de tórax, eletro e ecocardiograma. A investigaçäo virológica para Coxsacke B1, B3, B4, B5 e B6 se baseou em cultura, teste de neutralizaçäo e pesquisa de IgM por imunofluorescência indireta. Obtivemos vírus B4 em 9 (57%), B5 em 4 (25%), B1 em 2 (12%) e B3 em 1 (6%). Nenhum paciente foi submetido à terapêutica imunossupressora. A evoluçäo de pelo menos 1 ano foi; 7 (43%) permaneceram com miocardiopatia dilatada, 1 (6%) faleceu e 4 (25%) tiveram alta hospitalar, mas näo seguiram o acompanhamento. Uma das pacientes que teve evoluçäo crônica (extra-sístoles ventriculares, BAV do 2§ grau) está agora assintomática. Näo observamos diferença significativa entre os vários tipos de Coxsackie B em relaçäo à evoluçäo clínica

The Protective Effect of Astragalus saponins on Coxsackie B, Virus Infected New Born Rat Myocardiac Cells in Culture / 第二军医大学学报

The protective effect of Astragalus saponins or Astragalus mongholicus bunge on the micro -model of cultured new born rat heart cells infected with Coxsackie B, virus was observed. After inoculation with 1000 TCID Coxsackie B, virus, the cardiac enzyme lactic acid dehydrogenase (LDH) was much lower in the Astragalus saponins or Astragalus mongholiais bunge treated groups than in those untreated groups, but the synthesis rate of DNA was higher. Virus titer in the supernatant of cultures was detected 48 h after virus challenge. The virus titer of the Astragalus saponins (Lg 3.78 TCID) or the Astragalus mongholiais bunge (Lg 4.33 TCID50) treated groups was lower than in those untreated groups (Lg 5.78 TCID50). These results show that the Astragalus saponins may be a useful drug in treatment of acute viral myocarditis.

The Measurement of NK Cells Activity and Interferon Titers in Murine Coxsackievirus B_3 Induced Myocarditis / 第二军医大学学报

The level of interferon (IFN) in serum and homogenized cardiac tissue, and the activity of natural killer cells in Coxsackievirus B3 induced myocarditis in BALB/c mice were determined. NK cell cytolytic activity and IFN titers peaked on day 3~ 7 postinoculation (p.i.) and then declined. Virus titers in heart tissue reached a maximum on day 7 p.i. and then declined. These results suggest that the increase of NK cell activity and IFN titers provides some protection against Cox B, induced myocarditis by limiting virus replication in heart tissue.

ESTABLISHMENT OF COXSACKIE B_5 VIRUS INFECTED RAT HEART CELLS IN CULTURES / 第二军医大学学报

A micro-model of Coxsackie B5 virus infected rat beating heart cells was introduced.In the infected group, the cardiac enzymes-lactic acid dehydrogenase (LDH) and glutamic oxaloacetic transaminase(SGOT)werc significantly increased and the synthesis rate of DNA was decreased 48 h after inoculation with 400 TCID50 Coxsakie B6 virus.Virus titer(Lg 6.95 TCID50)in the supernatant of the infected cultures was detected 96 h after virus challenge.In addition, the specific fluorescence in cytoplasma of infected cells could be observed.These results suggest that Coxsackie B5 virus might be replicated, assembled and released in cultured rat myocardial cells.