Airway inflammation induced by Poly(I:C) stimulation in the late stage of respiratory syncytial virus infection in mice and its mechanism / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To investigate the pathogenic mechanisms of airway inflammation and recurrent wheezing induced by recurrent respiratory virus infection after respiratory syncytial virus (RSV) infection.</p><p><b>METHODS</b>Sixty-four female BALB/c mice (aged 6-8 weeks) were randomly divided into four groups: control, RSV, Poly(I:C), and RSV+Poly(I:C) (n=16 each). The bronchoalveolar lavage fluid (BALF) was collected on the 3rd day after Poly(I:C) administration, and the total cell number and differential counts in BALF were determined. Hematoxylin-eosin staining was used to observe pulmonary pathological changes. The airway responsiveness was detected. ELISA was used to measure the levels of interferon-γ (IFN-γ), interleukin-4 (IL-4), interleukin-13 (IL-13), matrix metallopeptidase-9 (MMP-9), and tissue inhibitor of metalloproteinase-1 (TIMP-1) in BALF.</p><p><b>RESULTS</b>Compared with the other three groups, the RSV+Poly(I:C) group had significant increases in the total number of inflammatory infiltrating cells in the airway, airway responsiveness, and MMP-9 level in BALF (P<0.05). The RSV+Poly(I:C) group showed more severe pulmonary tissue injuries compared with the control and RSV groups (P<0.01). Compared with the RSV group, the RSV+Poly(I:C) group showed significant reductions in the levels of IL-4 and TIMP-1 in BALF (P<0.01).</p><p><b>CONCLUSIONS</b>Viral re-infection in the late stage of RSV infection may cause an imbalance of MMP-9/TIMP-1 expression and thus contribute to aggravated airway inflammation.</p>

Inhibitory effect of chloroquine on airway hyperresponsiveness in asthmatic mice / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the effect of chloroquine on airway hyperresponsiveness in asthmatic mice and explore the possible mechanism.</p><p><b>METHODS</b>Balb/c mouse models of asthma established using OVA received intraperitoneal injections of chloroquine, dexamethasone, or both prior to OVA challenge. Within 24 h after the final challenge, airway hyper- responsiveness (AHR) of the mice was assessed, and the total cell count and the counts of different cell populations in the bronchoalveolar lavage fluid (BALF) were determined under light microscopy. The severity of lung inflammation was evaluated using HE staining, and the concentrations of IL-6 and PGF2α in the BALF were detected by enzyme-linked immunosorbent assay (ELISA).</p><p><b>RESULTS</b>Chloroquine pretreatment significantly decreased AHR (P<0.001) in the asthmatic mice and reduced the total cell count (P<0.01), eosinophils (P<0.001), neutrophils (P<0.01), and PGF2α levels in the BALF. Chloroquine combined with low-dose dexamethasone significantly lessened inflammations around the bronchioles (P<0.05) and blood vessels (P<0.01) in the lung tissue, and obviously lowered IL-6 (P<0.05) and PGF2α (P<0.001) in the BALF in the asthmatic mice.</p><p><b>CONCLUSION</b>Chloroquine can inhibit AHR in asthmatic mice and produce better anti-inflammatory effect when combined with dexamethasone for treatment of neutrophilic asthma.</p>

Difference between Nested-polymerase chain reaction and virus isolation in detection of respiratory syncytial virus and their clinical significances / 中华实用儿科临床杂志

Objective To observe the differences between Nested-polymerase chain reaction(N-PCR) and virus isolation methods used for detection of respiratory syncytial virus(RSV),and to reveal the potential clinical features of them.Methods From Jan.2010 to Aug.2012,nasopharyngeal aspirates (NPAs) were collected from the children with respiratory infection in the Department of Respiratory,the Children's Hospital of Chongqing Medical University.Both N-PCR and virus isolation were applied to detect RSV,and clinical data were collected for statistical analysis.Results A total of 1143 specimens were used for RSV detection by N-PCR and virus isolation.The male-female ratio was 2.16 vs 1.00.The age of patients was ranged from 1 month to 165 months(median:7 months).The most common diagnoses were as follows:bronchopneumonia [478 cases (41.8％)],chronic fibrous pneumonia [223 cases (19.5％)],bron-chiolitis [221 cases (19.3％)],bronchitis [71 cases (6.2％)] and upper respiratory infection [21 cases(1.8％)].For N-PCR,458 cases were RSV positive (total positive rate was 40.1％ ; 31.7％ for RSV-A,7.7％ for RSV-B,0.7％ for both RSV-A and RSV-B).With virus isolation method,204 cases were positive (17.8％).Comparison result of N-PCR and virus isolation showed:165 cases were positive (P+ I+) and 646 cases were negative (P-I-) by both methods (identity was 70.1％),and the most difference was N-PCR positive but virus isolation negative group (P+ I-) (293 cases,25.6％).When compared to P-I-group,the clinical features of P+ I-group were as follows:younger,longer hospital stays,remarkable season distribution (with peak in winter and lowest in summer),lower percentage of fever,higher percentage of cough,wheezing,dyspnea,severe pneumonia and respiratory failure,all these differences were statistically significant(all P ＜ 0.05),the ma-nifestations matched the clinical features of RSV infection.When compared to P + I + group,the symptoms in the P + I-group had longer duration before they were admitted to hospital (P =0.005) and lower percentage of wheezing (P =0.009).Conclusions The differences between N-PCR and virus isolation for the detection of RSV existed in duration of symptoms prior to hospitalization.Both the sensibility and specificity of N-PCR are desirable for RSV detection.

Hypermethylation of testis derived transcript gene promoter significantly correlates with worse outcomes in glioblastoma patients / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Glioblastoma is the most common and lethal cancer of the central nervous system. Global genomic hypomethylation and some CpG island hypermethylation are common hallmarks of these malignancies, but the effects of these methylation abnormalities on glioblastomas are still largely unclear. Methylation of the O6-methylguanine-DNA methyltransferase promoter is currently an only confirmed molecular predictor of better outcome in temozolomide treatment. To better understand the relationship between CpG island methylation status and patient outcome, this study launched DNA methylation profiles for thirty-three primary glioblastomas (pGBMs) and nine secondary glioblastomas (sGBMs) with the expectation to identify valuable prognostic and therapeutic targets.</p><p><b>METHODS</b>We evaluated the methylation status of testis derived transcript (TES) gene promoter by microarray analysis of glioblastomas and the prognostic value for TES methylation in the clinical outcome of pGBM patients. Significance analysis of microarrays was used for genes significantly differently methylated between 33 pGBM and nine sGBM. Survival curves were calculated according to the Kaplan-Meier method, and differences between curves were assessed using the log-rank test. Then, we treated glioblastoma cell lines (U87 and U251) with 5-aza-2-deoxycytidines (5-aza-dC) and detected cell biological behaviors.</p><p><b>RESULTS</b>Microarray data analysis identified TES promoter was hypermethylated in pGBMs compared with sGBMs (P < 0.05). Survival curves from the Kaplan-Meier method analysis revealed that the patients with TES hypermethylation had a short overall survival (P < 0.05). This abnormality is also confirmed in glioblastoma cell lines (U87 and U251). Treating these cells with 5-aza-dC released TES protein expression resulted in significant inhibition of cell growth (P = 0.013).</p><p><b>CONCLUSIONS</b>Hypermethylation of TES gene promoter highly correlated with worse outcome in pGBM patients. TES might represent a valuable prognostic marker for glioblastoma.</p>

Proportion of incidence of etiological agents in children with non-specific chronic cough in Chongqing: a follow-up study / 中华儿科杂志

<p><b>OBJECTIVE</b>To investigate the proportion of incidence of children with non-specific chronic cough in Chongqing and analyze the characteristics of etiology during the follow-up.</p><p><b>METHOD</b>Diagnostic criteria were defined for children with non-specific chronic cough according to the Guidelines of diagnosis and therapy for children with chronic cough that were formulated by the Subspecialty Group, Society of Pediatrics, Chinese Medical Association and Chinese Journal of Pediatrics in 2008. Totally 266 patients in whom cough was the main or the only symptom,lasting > 4 weeks, presenting to Asthma Center of Children's Hospital, Chongqing Medical University between June 2008 and April 2009 were recruited into this study. Based on the Guidelines, diagnosis was made after taking history, physical examination and assistant examination. After etiological treatment, the patients were followed up during the second week, the fourth week and the twelfth week. Etiological diagnosis was confirmed if cough was resolved after specific therapy. If cough was not resolved,the diagnosis was rechecked and a new therapy was applied.</p><p><b>RESULT</b>Totally 125 (47.0%) patients received final diagnoses of cough variant asthma (CVA), 58 (21.8%) was CVA and upper airway cough syndrome (UACS), 44 (16.5%) was diagnosed postinfection cough, 35 (13.2%) of UACS. In different age groups, the proportion of incidence of etiological agents is statistically distinct. In the ≤ 3 years old group, 35 patients (70.0%) were diagnosed CVA, 10 (20.0%) was postinfection cough; in 3 - 6 years group, 71 patients (50.7%) had CVA; the incidence of UACS was significantly higher in ≥ 6 years group.</p><p><b>CONCLUSION</b>It is concluded that CVA, CVA and UACS, post infection cough, and simple UACS were identified as the three top reasons for children with chronic cough in Chongqing. Children with chronic cough of different age groups had different etiology of cough. The characteristic of each etiology need further study.</p>

Determination of the serum mannose binding lectin levels in 738 Han ethnic group children / 中华儿科杂志

<p><b>OBJECTIVE</b>To investigate the distribution of serum mannose binding lectin (MBL) levels in Han ethnic group children.</p><p><b>METHODS</b>The concentrations of MBL in serum were measured by ELISA in 268 umbilical cord blood specimens from Chongqing, Wuhan and Urumqi as well as in serum of 470 normal children aged from 0 to 6 years and 87 adults in Chongqing.</p><p><b>RESULTS</b>The distribution of serum MBL levels in children (28 days to 6 years) was abnormal but there was no significant difference in MBL serum levels in subjects of different ages and genders. The median concentration of MBL in serum was significantly lower in newborns (median: 1597 microg/L, range: 884 - 1825 microg/L), cord blood group (median: 1462 microg/L, range: 0 - 4604 microg/L) than in other groups (children group median: 2536 microg/L, range 0 - 7860 microg/L; adult group median: 2920 microg/L, range 98 - 6495 microg/L). While among the other sub-groups aged from 28 days-6 years (28 day group median 2299 microg/L, range 214 - 4195 microg/L; 6 months-group median 2622 microg/L, range 5 - 4637 microg/L; 2 years-6 years group 2585 microg/L, range 198 - 7860 microg/L), there was no statistically significant difference. The median serum MBL level in normal children aged from 28 days to 6 years was 2563 microg/L and the P(2.5)-P(97.5) was 171 - 5079 microg/L.</p><p><b>CONCLUSIONS</b>The distribution of serum MBL levels in children (28 days to 6 years) was abnormal type but there was no statistically significant difference among different age and sex groups. The reference value of P(2.5)-P(97.5) in children (28 days-6 years) was 171 - 5079 microg/L.</p>

Allergic airway response associated with the intestinal microflora disruption induced by antibiotic therapy / 中华儿科杂志

<p><b>OBJECTIVE</b>Over the past several decades, there has been a significant increase in allergy and asthma in the world, which correlates with alterations in microflora and widespread use of antibiotics. The authors have developed a mouse model of antibiotics-induced microbiota disruption. In that model, mice were challenged by intranasal exposure to Aspergillus fumigatus allergens to explore the relation of allergic airway response and intestinal microflora disruption.</p><p><b>METHODS</b>Sixty female BALB/c mice were divided at random into 6 groups with 10 mice in each. (1) First antibiotic therapy group: the mice were given oral cefoperazone for 7 days, on day 7, mice were inoculated with Candida albicans (10(9)/ml, 50 microl) orally. (2) First control group: the mice were treated as first antibiotic therapy group, but cefoperazone and Candida albicans were replaced by saline. The mice in groups (1) and (2) were sacrificed on day 8, and cecal contents were collected for quantitative analysis of the intestinal bacterial flora. (3) Antibiotic therapy and challenge group: the mice were treated as the first antibiotic therapy group, then challenged (day 9 and 16) by intranasal exposure to Aspergillus fumigatus allergen. (4) Second antibiotic therapy group: the mice were treated as the first antibiotic therapy group, then challenged (day 9 and 16) by intranasal exposure to saline. (5) Challenge group: the mice were treated as the first control group, then challenged (day 9 and 16) by intranasal exposure to Aspergillus fumigatus allergen. (6) Second control group: the mice were treated as the first control group, then challenged (day 9 and 16) by intranasal exposure to saline. The mice in (3) - (6) group were killed for analysis of allergic airway response on day 19.</p><p><b>RESULTS</b>The quantity of Enterobacteriaceae, Enterococcus, Bifidobacterium and Lactobacillus in first antibiotic therapy group was significantly lower than that in the first control group, the quantity of Candida albicans increased in the first antibiotic therapy group as compared with the first control group. Mice intestinal microflora were disrupted with weight reduction and increased moisture in feces. After challenging with Aspergillus fumigatus allergens via intranasal inhalation, the total cell count, eosinophils, lymphocytes and neutrophils increased in BALF, especially in bronchoalveolar lavage fluid (BALF) from the mice in antibiotic therapy and challenge groups. IL-4 level in BALF from antibiotic therapy and challenge group (45.35 +/- 2.36) pg/ml was higher than that in the second control group (35.32 +/- 2.53) pg/ml. The expression of GATA-3 mRNA in the mice lung tissue (0.569 +/- 0.023) was higher than that in the second control group (0.410 +/- 0.020), and the ratios of T-bet/GATA-3 (0.578 +/- 0.021) decreased as compared with that in the second control group (0.804 +/- 0.035). IFN-gamma level in BALF from any group was not significantly different. In the absence of antibiotics, mice exposed to Aspergillus fumigatus allergen did not develop an allergic response in the airways.</p><p><b>CONCLUSIONS</b>The allergic (Th2) immune response can be induced by airway challenge with Aspergillus fumigatus allergen in the mice in which the intestinal microflora disruption resulted from antibiotic therapy, this result suggests that the intestinal microflora disruption resulted from antibiotic therapy is a risk factor for allergy and asthma.</p>

Isolation of human metapneumovirus from children with acute respiratory tract infection in Chongqing, China / 中华儿科杂志

<p><b>OBJECTIVE</b>To isolate and characterize the newly discovered human metapneumovirus (hMPV) in Chongqing, China and elucidate the clinical manifestations of hMPV infection.</p><p><b>METHODS</b>Eighty-six patients hospitalized for acute respiratory tract infection in Children's Hospital, Chongqing University of Medical Sciences from December 2004 to July 2005 were enrolled in the present study. Nasopharyngeal aspirates were collected for screening for common respiratory viruses by direct immunofluorescence assay, including respiratory syncytial virus, influenza virus types A and B, parainfluenza virus types 1, 2, 3 and adenovirus, and for inoculating onto Vero-E6 and LLC-MK2 cells for hMPV isolation. Cultures were maintained in the presence of trypsin and observed for development of cytopathic effect (CPE) for 3 weeks. Presence of hMPV was first indicated by positive CPE and subsequently confirmed by reverse transcription polymerase chain reaction targeting N and F genes. Sequence of amplified F fragments were analyzed and submitted to NCBI GenBank. The clinical findings of hMPV infection were collected and analyzed.</p><p><b>RESULTS</b>Of the collected 86 NPAs, six showed CPE characterized by clustering of infected cells, increased granules and eventuall detachment from cell monolayer and obvious syncytium formation. Successful isolation of hMPVs was confirmed with RT-PCR targeting hMPV F and N genes. Of the six hMPV-positive specimens, two were collected in winter (December, January), two in spring (May) and the other two in autumn (June, July). All the six patients were younger than 2 years of age with disease spectrum of bronchiolitis (2/6), bronchopneumonia (2/6), infantile asthma (1/6) and upper respiratory tract infection (1/6). Clinical findings included fever, cough, wheezing, polypnea, cyanosis and rales. Parainfluenza 3 and adenovirus seemed to beviral pathogens of co-infection.</p><p><b>CONCLUSION</b>Six hMPVs were successfully isolated in the mainland of China for the first time. HMPV appears to be one important viral pathogen for acute lower respiratory tract infections in young children with a detection rate of 7% (6/86) by viral isolation. The virus causes respiratory diseases similar to those caused by respiratory syncytial virus. These findings highlight the need for future investigations to define disease burden of hMPV infection and molecular epidemiology among children in China.</p>

Clinical characteristics of 12 persistently wheezing children with human bocavirus infection / 中华儿科杂志

<p><b>OBJECTIVE</b>The impact of human bocavirus (HBoV), a newly identified human parvovirus, on childhood persistent wheezing has not been identified. In this study, the clinical features of infantile persistent wheezing induced by HBoV was analyzed.</p><p><b>METHODS</b>Tracheal aspirates were collected by bronchofibroscope or nasopharyngeal (NP) aspirates from April, 2006 to January, 2007. HBoV DNA in the tracheal aspirates of 33 children with persistent wheezing and in NP aspirates of 6 children with persistent wheezing, who had at least or more than four weeks wheezing. RSV was identified by virus isolation in Hep-2 cells and antigen detetion by direct immunofluorescence assay (DIFA) which was also used for diagnosis of adenovirus, influenza A and B, parainfluenza 1, 2, 3 infection.</p><p><b>RESULTS</b>Of the 39 children with persistent wheezing, 12 cases (31%) were positive for HBoV DNA. Age of HBoV-positive patients ranged from 2 month to 1 year. The results of sequencing of PCR products proved that sequences of HBoV DNA from these 12 samples were exactly identical to the those of HBoV stored in GeneBank (accession numbers DQ000495 and DQ000496). Two cases with HBoV infection were found to be co-infected with RSV. Ten of the 12 HBoV-positive samples were collected during the period from winter to spring (1 in November, 4 in December, 2 in January and 3 in April), the other two HBoV-positive samples were collected during the period from summer to autumn (1 in May and the other in July). Seven of the 12 HBoV DNA-positive patients had fever, 5 of them had high fever. Significantly more patients with HBoV infection had fever as compared to patients with RSV infection. All the HBoV positive patients showed abnormal findings on chest X ray such as interstitial infiltrates, lung infiltration and hyperinflation. Abnormal findings on chest X ray were found in higher proportion of HBoV positive patients as compared with RSV positive patients. And other manifestations such as wheezing, cough and respiratory distress had no significant difference between HBoV and RSV infected patients.</p><p><b>CONCLUSIONS</b>This study further demonstrated that HBoV probably is a common pathogen of lower respiratory infection in children and might particularly be associated with persistent wheezing.</p>

Role of Foxp3 expression and CD4+CD25+ regulatory T cells on the pathogenesis of childhood asthma / 中华儿科杂志

<p><b>OBJECTIVE</b>To explore the impact of Foxp3 expression and CD(4)(+)CD(25)(+) regulatory T cells on pathogenesis of childhood asthma.</p><p><b>METHODS</b>Totally 15 patients with acute asthma exacerbation, 15 children with asthma remission and 10 children who were hospitalized for skeleton deformity without atopic disorders or history of allergic diseases or respiratory infections within a month as controls were recruited in this study from Sep. 2004 to Mar. 2005. The percentage of CD(4)(+)CD(25)(+) T cells were detected by 2-color flow cytometry. The levels of interleukin (IL)-4, IL-10, interferon (IFN)-gamma, transforming growth factor (TGF)-beta in plasma and supernatant were assayed by ELISA. Both the asthmatic children and the control children were selected to induce sputum by hypertonic saline. Sputum was processed for detecting the expression of Foxp3-mRNA. The expression of Foxp3-mRNA in both sputum and PBMC was detected by semi-quantitative RT-PCR with beta-actin as internal control.</p><p><b>RESULTS</b>The percentage of CD(4)(+)CD(25)(+) regulatory T cells in exacerbation and remission asthmatic children was significantly lower than that of the control children both prestimulation [(10.1 +/- 2.1)% vs. (15.5 +/- 2.7)%, (11.7 +/- 2.5)% vs. (15.5 +/- 2.7)%, P < 0.05] and poststimulation with PHA [(12.4 +/- 2.3)% vs. (26.9 +/- 3.8)%, (17.3 +/- 3.2)% vs. (26.9 +/- 3.8)%, P < 0.05]. The percentage of CD(4)(+)CD(25)(+) regulatory T cells was significantly higher after PHA stimulation in normal children [(15.5 +/- 2.7)% vs. (26.9 +/- 3.8)%, P < 0.01]. The expression of Foxp3-mRNA (Foxp3/beta-actin) in asthmatic children was significantly lower than that in the control children in both PBMC and induced sputum. The expression of Foxp3-mRNA in PBMC was significantly higher after PHA stimulation in the control children (0.77 +/- 0.22 vs. 1.07 +/- 0.21, P < 0.05). However, there was no significant difference in Foxp3-mRNA expression in asthmatic children pre and post PHA stimulation. A significant positive correlation between the Foxp3-mRNA expression and the percentage of CD(4)(+)CD(25)(+) regulatory T cells was detected. The levels of IFN-gamma and TGF-beta were significantly lower in asthmatic children than those in the control children, and the levels of IFN-gamma and TGF-beta correlated positively with Foxp3-mRNA expression and the percentage of CD(4)(+)CD(25)(+) regulatory T cells. The level of IL-4 both in plasma and supernatant was higher in asthmatic children. The levels of IL-10 was higher only in exacerbation than in control children, the levels of IL-4 and IL-10 had no correlation with Foxp3-mRNA expression and the percentage of CD(4)(+)CD(25)(+) regulatory T cells.</p><p><b>CONCLUSION</b>Insufficient secretion of TGF-beta, decreased Foxp3 expression, insufficient number of CD(4)(+)CD(25)(+) regulatory T cells and the defective ability of converting CD(4)(+)CD(25)(-) T cells to CD(4)(+)CD(25)(+) regulatory T cells might play an important role in pathogenesis of asthma.</p>

Combined effects of neonatal Bacillus Calmette-Guerin vaccination and respiratory syncytial infection on experimental asthma in mice / 中华儿科杂志

<p><b>OBJECTIVE</b>Neonatal Bacillus Calmette-Guerin (BCG) vaccination could decrease asthma prevalence in human according to "hygiene hypothesis". The authors proposed a hypothesis that effect of BCG vaccination on inhibiting asthma in human might be reversed by respiratory virus infection. The objective of this study was to observe combined effects of neonatal BCG vaccination and respiratory syncytial virus (RSV) infection on experimental asthma in mice.</p><p><b>METHODS</b>Neonatal BALB/c mice were divided into five groups. Control and ovalbumin (OVA) groups were mock-vaccinated at birth and mock-infected at 3 weeks of age. BCG + OVA group was BCG-vaccinated and mock-infected. RSV + OVA group was mock-vaccinated and RSV-infected. BCG + RSV + OVA group was BCG-vaccinated and RSV-infected. Except for control group, all the other groups underwent ovalbumin (OVA) sensitization and challenge. Airway responsiveness to inhaled methacholine was measured and bronchoalveolar lavage (BAL) was performed after the last challege. Cells in BAL fluid (BALF) were counted. Cytokines in BALF and serum OVA-specific IgE were detected by ELISA and inflammatory characteristics of lungs was scored by staining with hematoxylin and eosin.</p><p><b>RESULTS</b>(1) The numbers of total white cells, lymphocytes, monocytes, neutrophils, and eosinophils in the BALF from all OVA-sensitized/challenged groups were significantly greater than those in control (P < 0.01), and BCG + OVA group had significantly lower total white cells, lymphocytes and eosinophils as compared with other OVA-sensitized/challenged groups (P < 0.05 or 0.01). (2) All OVA-sensitized/challenged groups had significantly lower IFNgamma (P < 0.05) and higher IL-4 (P < 0.05) level in BALF as compared with control, but there was no significant difference among all OVA sensitized/challenged groups. There was no significant difference in IL-10 level between all experimental groups. (3) All OVA-sensitized/challenged groups showed significantly higher serum OVA-specific IgE titers than control (P < 0.05 or 0.01), but no significant difference was found among all OVA sensitized/challenged groups. (4) RSV + OVA and BCG + RSV + OVA groups displayed the highest airway resistance and subsequently in order as follows: OVA group, BCG + OVA group and control group in severity of airway hyperreactivity (AHR), but no significant difference was found between RSV + OVA and BCG + RSV + OVA groups. (5) Histological score of peribronchiolitis, perivasculitis, alveolitis, and peribronchial eosinophilia in all OVA-sensitized/challenged groups was significantly higher than that in control. BCG + OVA group had significantly milder peribronchiolitis and peribronchial eosinophilia than the other OVA-sensitized/challenged groups (P < 0.05) and significantly milder alveolitis than OVA and BCG + RSV + OVA groups (P < 0.05).</p><p><b>CONCLUSION</b>Neonatal BCG vaccination decreased asthmatic inflammation and AHR and RSV infection could reverse anti-asthma effect of neonatal BCG vaccination in OVA-sensitized/challenged mouse model.</p>

Effect of Anti-Respiratory Syncytial Virus Activities by RNA Interference and Ribavirin in vitro / 实用儿科临床杂志

Objective To investigate the effect of RNA interference(RNAi) on inhibiting respiratory syncytial virus(RSV) replication through comparing the anti-RSV activities between pshRNA7816 and ribavirin in cell culture system.Methods The recombinated plasmid pshRNA7816 and ribavirin was added to HEp-2 cells.Methyl thiazolyl tetrazolium(MTT) assay was used to detect cytotoxicity of pshRNA7816 and ribavirin on normal HEp-2 cells and protective effects of them on RSV infected HEp-2 cells.The effects of pshRNA7816 and ribavirin on change of cytopathogenic effect(CPE) of HEp-2 cells induced by RSV infection were observed through microscopically.Results pshRNA7816 had not significant toxicity on the growth of HEp-2 cells,but the ribavirin had significant toxicity when the concentration above 1.0 mmol/L.The pshRNA7816 and ribavirin could alleviate the CPE of HEp-2 cells induced by RSV infection,but the pshRNA7816 showed a more potent inhibition than ribavirin.The inhibition rates of pshRNA7816 were significantly higher than the maximum inhibition rate of ribavirin on RSV infection(P

Construction and effect of the recombinant pshRNA plasmid against respiratory syncystial virus M2-1 gene / 中华儿科杂志

<p><b>OBJECTIVE</b>Respiratory syncystial virus (RSV) is the most common cause of lower respiratory infections in infants worldwide. There is no reliable vaccine or antiviral drug against RSV at present. RNA interference (RNAi) technology is a potent method to degrade expression of the cognate mRNA. In order to inhibit the replication of RSV at gene level, the effects of specific RNAi against M2-1 gene of RSV on inhibition of viral replication in cell culture system was observed in this study.</p><p><b>METHODS</b>RSV M2-1 gene, which plays a key role in RSV transcription, was chosen in this study and was used as target gene and recombinant plasmid pshRNA7816 targeting the mRNA of RSV M2-1 gene coding sequence was constructed. The pshRNA7816 was transfected into Hep2 cells. The effects of the pshRNA7816 on changes of cytopathogenic effect (CPE) of Hep2 cell induced by RSV infection were observed microscopically. Viral plaque forming assay and MTT assay were used to detect the viral titer change and protective function of the pshRNA7816 on RSV infected Hep2 cell.</p><p><b>RESULTS</b>The recombinant RNAi plasmid pshRNA7816 which targets the mRNA of RSV M2-1 gene was successfully constructed. The pshRNA7816 significantly reduced CPE of RSV infected Hep2 cells, reduced the viral titer of RSV in the cells (P < 0.001). The pshRNA7816 raised the survival rate of RSV infected Hep2 cells (P < 0.001). Non-specific pshRNA plasmid did not show anti-RSV effects (P > 0.05).</p><p><b>CONCLUSION</b>The recombinant pshRNA7816 plasmid which targeted the mRNA of RSV M2-1 gene showed a significant and specific anti-RSV effect.</p>

Effects of Bacillus Calmette-Guerin vaccination on immune functional development of splenic T cell in neonatal mice / 中华儿科杂志

<p><b>OBJECTIVE</b>Almost every neonate receives Bacillus Calmette-Guerin (BCG) vaccination in China. The authors' previous study showed that BCG promoted cord blood monocyte-derived dendritic cells maturation and induced high level of interleukin (IL)-10, medium level of interferon (IFN)-gamma, but low level of IL-4 production by cord naive T cells. The experiments in the present study were designed to explore the effects of neonatal BCG vaccination on immune functional development of splenic T cells in mice in vivo.</p><p><b>METHODS</b>Neonatal BALB/c mice were inoculated with BCG intraperiotoneally. Four weeks later, spleen cells of mice were isolated and surface molecular markers of CD4, CD25 and CD44 and intracellular IFN-gamma, IL-10, and IL-4 in CD3(+) T cells were detected by flow cytometry. Furthermore, mRNA expression of transcription factor T-bet, Foxp3 and GATA-3 were analyzed by RT-PCR.</p><p><b>RESULTS</b>The percentage of total CD4(+) T cells decreased [(23.50 +/- 2.59)% vs. (47.38 +/- 10.41)%, P < 0.01] but the percentage of CD25(+) [(24.92 +/- 2.74)% vs. (20.27 +/- 2.85)%, P < 0.05] and CD44(+) [(89.29 +/- 2.56)% vs. (82.98 +/- 5.51)%, P < 0.05] T cells in CD4(+) T cells was higher in BCG-vaccinated mice than that in controls. Meanwhile, the percentage of IFN-gamma positive [(6.52 +/- 2.40)% vs. (3.13 +/- 2.03)%, P < 0.05] and IL-10 positive [(14.81 +/- 3.65)% vs. (10.90 +/- 1.61)%, P < 0.05] but not IL-4 positive [(1.17 +/- 0.46)% vs (1.51 +/- 0.75)%, P > 0.05] cells in CD3(+) T cells of BCG-vaccinated mice was significantly higher than that of non-BCG-vaccinated mice. In comparison with BCG-naive mice, T-bet was significantly high in BCG-vaccinated mice [T-bet/beta-actin 0.44 +/- 0.11 vs. 0.28 +/- 0.06, P < 0.05], but there was no significant difference in GATA-3 [GATA-3/beta-actin 0.46 +/- 0.08 vs. 0.50 +/- 0.10,P > 0.05] and Foxp3 [Foxp3/beta-actin vs. 0.27 +/- 0.11 and 0.30 +/- 0.16, P > 0.05] mRNA expression between the two groups.</p><p><b>CONCLUSION</b>Neonatal BCG vaccination could induce strong Th1 but weak Th2 response as reported previously. Though neonatal BCG vaccination was not capable of inducing CD4(+)CD25(+) regulatory T cell response with Foxp3 expression, it caused increase of IL-10(+) CD3(+) cells which might represent some regulatory T cells producing IL-10.</p>

Identification of two novel WASP gene mutations in 3 boys with Wiskott-Aldrich syndrome / 中华儿科杂志

<p><b>OBJECTIVE</b>The Wiskott-Aldrich syndrome (WAS) is a rare X-linked primary immunodeficiency caused by mutations in the WAS protein (WASP) gene. The disease is characterized by recurrent infections, eczema, and thrombocytopenia with small platelets, and it is known to be associated with extensive clinical variability, and mutation studies indicated that genotypes are also highly variant among WAS patients. The present study was conducted to identify the mutation types of Wiskott-Aldrich syndrome protein (WASP) gene in 3 boys suffering from Wiskott-Aldrich syndrome.</p><p><b>METHODS</b>Based on the typical clinical manifestations of Wiskott-Aldrich syndrome including thrombocytopenia, eczema, and recurrent infections and scanning electron micrographs, 3 patients were suspected of having WAS. The WASP gene of the 3 patients and their mothers were detected by PCR-direct sequencing analysis.</p><p><b>RESULTS</b>By sequence analysis using sense and antisense primer separately, the authors found two novel WASP gene mutations. For the twin brothers, a C deletion at nucleotide 984 was detected in exon 10 of WASP gene (984delC). The consequence of the C deletion involved frameshift mutation after H317 and premature stop at 444 (H317fsX444). Their mother was a carrier of the mutated WASP gene. For another WAS patient, a nonsense mutation with nucleotide substitution of G to T at position 1388 (1388G-->T) in exon 11 of WASP gene, led to premature translational termination at amino acid position 452 (E452X). His mother had not been found to have WASP gene mutation.</p><p><b>CONCLUSION</b>Genetic analysis is useful in definite diagnosis of Wiskott-Aldrich syndrome patients and in carrier detection and prenatal diagnosis, especially of atypical or sporadic WAS patients.</p>

Effect of Specific Immunotherapy on Immune and Pulmonery Function of Children with Asthma / 实用儿科临床杂志

0.05).Concusions There exsits the disequilibrium of Th1/Th2 in asthmatic children,but SIT can recovery the balance of Th1/Th2.We find excllent effects of SIT on immune and pulmonery function of asthmatic infants.

Clinical study on bacille calmette-guerin alone or combined with interferon gamma and vitamin A in preventing asthma children after bronchiolitis / 实用儿科临床杂志

Objective To discuss the effects of bacille calmette guerin(BCG) alone of combined with interferon gamma(IFN-?) and vitamin A in preventing asthma after bronchiolitis.Methods Fourty-four bronchiolitis infants(RBI) were treated with BCG alone or combined with vitamin A/IFN-?.Protein purified derivative(PPD) skin test were observed before and 3 months after interference and all infants enrolled in the study were followed up for average one year to observe the wheezing episodes. Ninteen cases of RBI without BCG treatment were control group.Results The average diameter of PPD skin test in RBI after BCG inoculation was significantly larger than that of before inoculation and control group(P