ABSTRACT
<p><b>OBJECTIVE</b>To study the roles of rhoassociated coiled coil forming protein kinase 2 (Rock2) and transforming growth factor-β1 (TGF-β1) mRNA in acute asthma and the effect of glucocorticoid intervention on the Rock2 and TGF-β1 mRNA expression in rats.</p><p><b>METHODS</b>Forty-eight male rats were randomly divided into 4 groups (n=12 each): asthma, control, dexamethasone treated (DXM) and budesonide treated (BUD). Rat model of asthma was prepared by the ovalbumin (OVA) challenge. The animals were sacrificed 24 hrs after the last challenge. The total cell number and differentiation cell number were counted in bronchoalveolar lavage fluid (BALF). The protein expression of Rock2 was ascertained by immunohistochemistry and the mRNA expression of TGF-β1 was ascertained by hybridization in situ.</p><p><b>RESULTS</b>The pathological changes in the BUD and the DXM groups were alleviated when compared with the asthma group. The total cell number and the percentage of eosinophil (EOS), polymorphonuclear leukocytes (PMN) and lymphocytes (Lym) in BALF in the asthma group were significantly higher than those in the control group (P<0.01). The percentage of macrophage (Mф) in the asthma group was significantly lower than that in the control group (P<0.01). The total cell number and the percentage of EOS and Lym in BALF in the DXM and the BUD groups decreased, while the percentage of Mф increased significantly compared with those in the asthma group (P<0.01). The Rock2 and TGF-β1 mRNA expression in lung tissues in the asthma group increased significantly compared with those in the control, BUD and DXM groups, while there were no significant differences in the Rock2 expression and TGF-β1 mRNA expression between the DXM or BUD group and the control group.</p><p><b>CONCLUSIONS</b>The expression of Rock2 and TGF-β1 mRNA in lung tissues is increased in rats with acute asthma. Glucocorticoids can significantly decrease the expression of Rock2 and TGF-β1 in lung tissues, thus alleviates airway inflammation.</p>
Subject(s)
Animals , Male , Rats , Asthma , Drug Therapy , Metabolism , Budesonide , Therapeutic Uses , Dexamethasone , Therapeutic Uses , Glucocorticoids , Therapeutic Uses , Lung , Metabolism , Pathology , RNA, Messenger , Rats, Sprague-Dawley , Transforming Growth Factor beta1 , Genetics , rho-Associated Kinases , GeneticsABSTRACT
<p><b>OBJECTIVE</b>To clone and express VP, gene from HBoV, and the expressed VP, protein was as the antigen in order to detect serum from children in Wenling area with lower respiratory tract infections.</p><p><b>METHODS</b>The VP, gene was recombined with the genome of Baculovirus, which infected the insect cell. The fusion protein with HA tag was applied to confirm the specificity of expressed protein. Furthermore, the recombinant protein was observed using electron microscopy. The 176 serum from children in Wenling area with lower respiratory tract infections was screened using Western blot.</p><p><b>RESULTS</b>The expressed VP2 protein was more than 60% in total proteins from insect cell, and MWt about 60 x 10(3). The virus-like particle (VLP) was observed using electron microscopy, and size about 20 nm. The 176 serum from children in wenling area with lower respiratory tract infections was screened using Western blot. The HBoV positive rate was 2.28% (4/176).</p><p><b>CONCLUSION</b>The VP2 protein from human bocavirus was expressed in insect cell successfully. Through HA tag the VP2 protein was specific, and then the assay using SDS-PAGE with Western blot could detect and screen the antibody in serum from children with lower respiratory tract infections rapidly and accurately.</p>
Subject(s)
Animals , Child, Preschool , Female , Humans , Infant , Male , Antibodies, Viral , Blood , Bocavirus , Genetics , Allergy and Immunology , Capsid Proteins , Genetics , Allergy and Immunology , Gene Expression , Parvoviridae Infections , Blood , Diagnosis , Allergy and Immunology , Virology , Recombinant Proteins , Genetics , Allergy and Immunology , SpodopteraABSTRACT
<p><b>OBJECTIVE</b>To investigate pave a way for studying pathogenicty of HBoV.</p><p><b>METHODS</b>Isolation and cell culture of HBoV by human bronchial epithelial cell line, which was founded in our laboratory. The morphology of the virus were primarily studied with a transmission electron microscope. In addition, transcript mRNA was detected in human bronchial epithelial cells, which was passaged and infected within HBoV, using the reverse-transcription polymerase chain reaction (RT-PCR). The amplified products nucleotide sequence of HBoV were sequencing and sequence analysis.</p><p><b>RESULTS</b>Cytopathic effect (CPE) was observed after the aseptic residue of filtration of 2 case sputum specimens with HBoV, which was inoculated to the human bronchial epithelial cell line. The virus particles were observed in the cytoplasm, which were hexagonal or spherical in shape and 18-26 nm in diameter,bulk was 20 nm. cDNA amplicon obtained 295 bp fragment results of electrophoresis bands as same as NS1 region of the conserved matrix gene of publish sequence of HboV. PCR products nucleotide sequence of HboV were compared with corresponding HboV GeneBank sequences. The comparison/alignment and construction of phylogenetic trees also point to an affiliation of the parvovirus to the species HBoV.</p><p><b>CONCLUSION</b>Isolation and identification of HBoV could be done in the human bronchial epithelial cell, and we found some characterizing CPE in the human bronchial epithelial cell after HBoV infection. The above studies pave a way for studying pathogenicty of human bocavirus.</p>
Subject(s)
Child , Child, Preschool , Humans , Infant , Male , Bronchi , Cell Biology , Virology , Cell Culture Techniques , Cell Line , Epithelial Cells , Virology , Human bocavirus , Classification , Genetics , Molecular Sequence Data , Parvoviridae Infections , Virology , Phylogeny , Respiratory Tract Infections , Virology , Virus CultivationABSTRACT
WU polyomavirus, which was firstly discovered in 2007, is a new human polyomavirus belonging to Polyomaviridae and containing circular double-stranded genomic DNA. In this study, the 278 clinical sputum specimens from children under 5 years old were collected from Wenzhou Medical College affiliated Wenling First Hospital, Zhejiang Province. Based on identification assay of WU polyomavirus previously reported, a WU polyomavirus was identified from clinical samples successfully, the positive rate was 0.4%. The sequences of PCR products were identical to that of VP2 gene and large T antigen gene derived from WU polyomavirus reported. The above results strongly suggested that the WU polyomavirus isolated was firstly found in Chinese children with acute lower respiratory tract infections. This study provides a firm basis for further research of WU polyomavirus.
Subject(s)
Child, Preschool , Humans , Infant , Infant, Newborn , Base Sequence , Molecular Sequence Data , Polymerase Chain Reaction , Polyomavirus , Genetics , Sputum , VirologyABSTRACT
KI polyomavirus, which was firstly discovered in 2007, is a new human polyomavirus belonging to Polyomaviridae and containing circular double-strand genomic DNA. This study was based on identification assay of KI polyomavirus reported. Total 2293 clinical sputum specimens from children under 3-years-old were collected and screened from Wenzhou Medical College affiliated Wenling Hospital, Zhejiang Province. A KI polyomavirus was detected and identified, the positive rate was 0.04%. The sequences of PCR products was identical to that of the viral capsid protein (VP1) gene derived from KI polyomavirus. The results strongly suggested that the KI polyomavirus was found firstly in Chinese children with acute lower respiratory tract infections from Zhejiang region. This study provided new information for further investigation of etiopathogenisis and diagnosis in children with lower respiratory tract infections.
Subject(s)
Child, Preschool , Humans , Infant , China , Polymerase Chain Reaction , Polyomavirus , Respiratory Tract Infections , VirologyABSTRACT
<p><b>OBJECTIVE</b>In this study, human bronchial epithelial cells were inoculated with positive sputum specimens of HBoV. After four days' infection, cytopathic effects (CPE) were observed by inverted microscopy. These viruses all cause typical cell damages such as rounded and shrivelled, fusion and fallout. These damages got quick following increased future degenerations. The other assay result of CPE within the infected cells were observed by inverted microscopy, have typical "owl's eye" plaque and above 90 percent hemadsorption within the infected cells by erythrocytes for hemadsorption technique. The typical fluorescence lump of nucleus within the infected cells was found by indirect immunofluorescence technique.</p><p><b>CONCLUSION</b>Isolation and identification of HBoV could be done in the human bronchial epithelial cell, and we found some characterizing CPE in the human bronchial epithelial cell after HBoV infection. The above studies pave a way for studying pathogenicity of human bocavirus.</p>
Subject(s)
Humans , Bocavirus , Physiology , Bronchi , Cell Biology , Cell Death , Physiology , Cell Survival , Physiology , Cells, Cultured , Epithelial Cells , Cell Biology , Virology , Fluorescent Antibody Technique, Indirect , Host-Pathogen Interactions , Microscopy, FluorescenceABSTRACT
Human bocavirus, which was firstly discovered in 2005, is a new human parvovirus associated with lower respiratory tract infection in children. In this study, a human bocavirus, named WLL-1 isolate, was identified in Wenlin County, Zhejiang Province. The genome of bocavirus WLL-1 has been sequenced and analyzed. Phylogenetic analyses showed that WLL-1 shares 99% homology with other bocaviruses recently reported, but also has some special variations.