Clinical and molecular features of 38 children with chronic granulomatous disease in mainland china.

PURPOSE: Chronic granulomatous disease (CGD) is an inherited disorder, with phagocytes failing to produce antimicrobial superoxide due to deficient NADPH oxidase activity. Mutations in the gene encoding CYBB are responsible for the majority of the CGD cases. To date, there have been no reports on large samples of children with CGD in China. Therefore, in this study, we described the clinical and molecular features of 38 suspected CGD patients from 36 unrelated Chinese families. METHODS: Clinical diagnosis was performed using dihydrorhodamine assays detected by flow cytometry. Molecular analysis was used to identify underlying CGD-causative genes. RESULTS: The mean age of onset in our 38 patients was 3.4 months, while the mean age at diagnosis was 31.7 months. Apart from recurrent pneumonia and abscesses, tuberculosis (TB) and Bacille Calmette-Guerin (BCG) infections were notable features in our cohort. Overall, 17 cases died and patient 1 did not participate in the follow-up period . In total, we identified 29 different CYBB gene mutations in 31 patients. We found NCF1 and CYBA mutations in 3 and 2 patients, respectively. In addition, we identified 31 carriers and prenatally diagnosed 4 CGD and 4 healthy fetuses. CONCLUSIONS: The results of our study demonstrate that children with BCG infections or recurrent TB infections should have immune function screening tests performed. Moreover, newborns with family histories of primary immunodeficiency diseases should avoid of BCG vaccination. Molecular analysis is an important tool for identifying patients, carriers, and high-risk CGD fetuses.

Molecular diagnosis of severe combined immunodeficiency--identification of IL2RG, JAK3, IL7R, DCLRE1C, RAG1, and RAG2 mutations in a cohort of Chinese and Southeast Asian children.

Severe combined immunodeficiencies (SCID) are a group of rare inherited disorders with profound defects in T cell and B cell immunity. From 2005 to 2010, our unit performed testing for IL2RG, JAK3, IL7R, RAG1, RAG2, DCLRE1C, LIG4, AK2, and ZAP70 mutations in 42 Chinese and Southeast Asian infants with SCID adopting a candidate gene approach, based on patient's gender, immune phenotype, and inheritance pattern. Mutations were identified in 26 patients, including IL2RG (n = 19), IL7R (n = 2), JAK3 (n = 2), RAG1 (n = 1), RAG2 (n = 1), and DCLRE1C (n = 1). Among 12 patients who underwent hematopoietic stem cell transplantation, eight patients survived. Complications and morbidities during transplant period were significant, especially disseminated bacillus Calmette-Guérin disease which was often difficult to control. This is the first cohort study on SCID in the Chinese and Southeast Asian population, based on a multi-centered collaborative research network. The foremost issue is service provision for early detection, diagnosis, management, and definitive treatment for patients with SCID. National management guidelines for SCID should be established, and research into an efficient platform for genetic diagnosis is needed.

Clinical characteristics and molecular analysis of three Chinese children with Omenn syndrome.

Omenn syndrome (OS) is a rare autosomal recessive genetic disorder and presents symptoms of severe combined immunodeficiency characterized by erythrodermia, eosinophilia, hepatosplenomegaly, lymphadenopathy, and elevated serum IgE levels. OS has been found to be caused by mutations in RAG1 or RAG2 gene that result in partial V(D)J recombination activity. No study on OS has been reported in Chinese children so far. In this study, the genotype and phenotypes of three infants with OS from three unrelated Chinese families were investigated. All the three children had most of the characteristics of OS except normal serum IgE level. Compound heterozygosity mutations in RAG1 gene (1983 G>A; 2444 C>T and 2219 C>T; 3127 C>G) were identified in two cases, and a homozygous deletion mutation with a premature stop codon was found at residue 2302 of RAG1 gene (2302delT, I729X) in the remaining case, including three novel mutations (2302delT, I729X; 2219 C>T, R699W; and 3127 C>G, Y1001X). Spectratyping analysis of T-cell receptor ß-chain variable region (TCRVß) gene rearrangement was performed in case 1 and case 2. All the 25 TCRVß subfamilies presented monoclonal or oligoclonal peaks in case 1 and 11 TCRVß subfamilies were very weak or even absent in case 2. This was the first report about OS in Chinese children. Molecular genetic testing represents an important tool for early confirmed diagnosis and may allow accurate carrier detection and prenatal diagnosis.

Clinical characteristics and genotype-phenotype correlation in 62 patients with X-linked agammaglobulinemia.

INTRODUCTION: X-linked agammagobulinemia (XLA) is a primary immunodeficiency disorder caused by Bruton's tyrosine kinase (Btk) gene mutation. Recent studies suggested genotype-phenotype correlation in XLA, but a definitive association remains controversial. PATIENTS AND METHODS: We examined the relationship between specific Btk gene mutations and severity of clinical presentation in 62 patients with XLA. Disease severity was assessed by the age of disease onset and the presence of severe infections, while mutations were classified into severe and mild based on structural and functional consequence by bioinformatics analysis. RESULTS: Fifty-six Btk mutations were identified in 62 patients from 57 kindreds. Variation in phenotypes was observed, and there was a tendency of association between genotype and age of disease onset as well as occurrence of severe infections. CONCLUSION: A critical analysis of the circumstances upon presentation also revealed that under-recognition of recurrent infections and relevant family history are important hurdles to timely diagnosis of XLA.

Genetic variability of respiratory syncytial viruses (RSV) prevalent in Southwestern China from 2006 to 2009: emergence of subgroup B and A RSV as dominant strains.

Respiratory syncytial virus (RSV) is the most commonly identified viral agent in young children with acute respiratory tract infections (ARIs) and often causes repeated infections throughout life. This study investigated the genetic variability of the attachment (G) protein gene among RSV strains prevalent in southwestern China. Reverse transcription-PCR (RT-PCR) for a fragment of the RSV G gene was performed with nasopharyngeal aspirates (NPAs) collected from children with ARIs hospitalized in Chongqing Children's Hospital, Chongqing, China. A total of 1,387 NPA specimens were collected from April 2006 to March 2009, and 439 (31.7%) were positive for RSV. During the study period, subgroup A and B viruses accounted for 79.5% (349/439) and 19.8% (87/439) of the total positive samples, respectively. Both subgroup A and B viruses were identified in three samples. Subgroup A viruses were predominant during two epidemic seasons (2006 to 2008), while subgroup B strains prevailed during the 2008-2009 epidemic season. Phylogenetic analyses showed that all 30 group A strains could be clustered into one genotype, genotype GA2, and 30 group B strains could be clustered into three genotypes, genotypes GB1, GB3, and BA, among which 17 genotype BA strains were detected from 23 group B strains selected during the 2008-2009 epidemic season. The G gene of genotype BA was predicted to encode proteins of five different lengths. These data suggest that group A RSV likely predominated in southwestern China and that a new genotype of subgroup B with a 60-nucleotide insertion, named BA-like virus, became the dominant genotype in the 2008-2009 epidemic season.

Analysis of clinical and molecular characteristics of Wiskott-Aldrich syndrome in 24 patients from 23 unrelated Chinese families.

The clinical data of 24 children with Wiskott-Aldrich syndrome (WAS) from 23 unrelated Chinese families were reviewed in the present study. WAS protein (WASP) expression in peripheral blood mononuclear cells was examined by flow cytometry (FCM); WASP gene was amplified by PCR and directly sequenced to analyze mutations in the WASP gene in patients and their female relatives. FCM analysis of 21 patients showed that 18 cases were WASP-negative, and three had partially WASP expression. WASP gene analysis revealed mutations in 23 patients, including five missense mutations, four nonsense mutations, four deletion mutations, three insertion mutations, six splice site mutations, and one complex mutation, among which, 20 unique mutations were detected, including seven novel mutations (168 C>A, 747-748del T, 793-797del C, 1185 ins C, Dup 1251-1267, 1277 insA and 1266 C>G; 1267-1269del C). Five WAS children underwent stem cell transplantation. After 2 months of transplantation, WASP expression was restored to normal in all five patients whereas one patient died of cytomegalovirus-induced interstitial lung disease. WASP gene analysis can make a definite diagnosis of WAS and identify mutation carriers, beneficial for timely treatment and genetic counseling for children with WAS.

Clinical and molecular characteristics of 35 Chinese children with Wiskott-Aldrich syndrome.

BACKGROUND: Wiskott-Aldrich syndrome (WAS) is a rare primary immunodeficiency disease, with an incidence of 4/1,000,000 live male births. In China, an estimated number of 35 babies with WAS are born each year, but likely many remain undiagnosed. OBJECTIVES: The objectives of study were to review the clinical and molecular characteristics of a cohort of Chinese children with WAS and to describe the long-term outcome of those who underwent hematopoietic stem cell transplant (HSCT). MATERIALS AND METHOD: Records of 35 patients diagnosed with WAS during 1991-2008 were reviewed. Genetic diagnosis was established by direct gene sequencing. RESULTS: All patients had classical WAS phenotype. WASP mutations were identified in 33 patients from 29 families. Nine patients underwent HSCT at a mean age of 22.1 months (match-unrelated donor, n = 5; mismatched related donor, n = 2; matched-sibling donor, n = 2). Post-transplant immune hemolytic anemia and thrombocytopenia occurred in three patients with complete resolution. All patients survived without significant long-term complications and had full platelet, T and B lymphocyte recovery within 2 years post-transplant. CONCLUSION: In the past decade, there has been significant improvement in clinical and genetic diagnosis of WAS in Chinese. We demonstrated excellent long-term survival in patients who underwent HSCT. Early workup for transplant should be advocated for children with classical WAS before they suffer from major disease complications and morbidities.

Lipopolysaccharide induces IL-6 production in respiratory syncytial virus-infected airway epithelial cells through the toll-like receptor 4 signaling pathway.

Respiratory syncytial virus (RSV) is a leading cause of bronchiolitis in young children. Microbial agents such as endotoxin and RSV are implicated in airway inflammation during the development of reactive airway disease (RAD) later in childhood. Toll-like receptors (TLRs) are involved in an inflammation cascade through pathogen-associated molecular pattern recognition including lipopolysaccharide (LPS) and viral components. In this study, we investigated the expression of TLRs and cytokine-chemokine production profiles of RSV-infected epithelial cells. In live-RSV infected human tracheal epithelial cell line (9HTEo), TLRs 1-10 mRNA levels were up-regulated in a time-dependent manner compared with ultraviolet (UV)-inactivated RSV. RSV was shown to alter TLR4 membrane and cytosolic location in epithelial cells. Stimulating RSV-infected epithelial cells with TLR4 agonist LPS increased synthesis of IL-6, IL-8, and reduced regulated on activation, normal T cell expressed and secreted (RANTES) production. TLR4 neutralizing antibody HTA125 and TLR4-targeting RNA interference experiments revealed that TLR4 signaling pathway played a predominant role in mediating LPS-induced-IL-6 production of RSV infected epithelial cells. Altogether, our studies indicated that TLR4 play a critical role in leading LPS mediated-IL-6 response in RSV infected-epithelial cells and might be an important factor influencing the cytokine-chemokine profile of epithelial cells interacting with virus and endotoxin, which is correlated with phenotypes of RSV diseases.

High seroprevalence of human metapneumovirus infection in children in Chongqing, China.

BACKGROUND: The human metapneumovirus (hMPV) is a newly discovered respiratory viral pathogen that was first isolated in 2001 in the Netherlands. Its global distribution and long history of infection in humans have been well documented. In this study, we assessed the seropositivity of hMPV IgG antibodies in children in Chongqing, China. METHODS: The specificity of the enzyme linked immunosorbent assay (ELISA) was first validated by using respiratory syncytial virus (RSV) infected, antigen subtracted reference serum and by performing western blotting using anti-hMPV animal serum. This assay was used to determine the presence of the IgG antibody against hMPV and RSV in 325 serum samples obtained from children aged 0 - 6 years. RESULTS: No crossreaction was detected by ELISA between the antibodies to hMPV and RSV. The seropositivity of the anti-hMPV IgG antibody was 74.5% in children aged 0 to 5 months, 64.0% in 6 to 11 months, 72.7% in 12 to 23 months, 87.1% in 24 to 35 months and 90.3% in 3 to 6 years. CONCLUSIONS: hMPV is a common and significant respiratory pathogen in Chinese children. Almost all individuals are exposed to hMPV by age 6 years.

[Toll-like receptor 4 expression and function of respiratory syncytial virus-infected airway epithelial cells].

OBJECTIVE: To observe the epithelial Toll like receptor (TLR)4 expression changes and the signaling pathway function after respiratory syncytial virus (RSV) infection and to explore the mechanisms of RSV-induced airway inflammation. METHODS: 9HTEo-human tracheal epithelial cell line was infected by RSV (MOI = 10), and TLR1-10 mRNA were detected by RT-PCR assay at 3 h post RSV infection. TLR4 mRNA was detected by real time Q-PCR assay at 3 h, 6 h and 9 h post RSV infection, and TLR4 protein expression and cell apoptosis were determined by flow cytometry at 24 h post RSV infection. IL-8 in supernatant was detected by ELISA after RSV-infected cells exposed to lipopolysaccharide (LPS). A normal control group and a RSV infection group were set up for the RT-PCR and flow cytometry experiments, and the data were analyzed by paired t test using GraphPad 4.0 software. A normal group, a RSV group and a UV-inactivated RSV group were set up for the real time Q-PCR, experiments, and the data were analyzed by Kruskal-Wallis test. The ELISA experiments were divided into 4 groups including a normal control, a RSV, a LPS stimulation, and a RSV plus LPS co-stimulation groups, and the data were analyzed by One-way ANOVA test. RESULTS: (1) TLR2-10 mRNA level was significantly up-regulated (t value of TLR2-10: 3.49 -14.47, P < 0.05), especially TLR-2, 6 enhanced expression, compared with the normal epithelial cells. Real time Q-PCR assay showed that TLR4 mRNA started to increase at 3hr (Kruskal-Wallis test value = 8.82, P < 0.05, n = 6) and significantly elevated at 9 hour (Kruskal-Wallis test value = 6.62, P < 0.05, n = 6). UV inactivated-RSV had no effect on the TLR4 mRNA level. (2) Flow cytometry showed that membrane TLR4 mean fluorescence intensity (MFI) increased (RSV: 1.27 +/- 0.48, normal: 0.97 +/- 0.25; t = 2.39, P > 0.05, n = 10) while cytoplasmic TLR4 MFI simultaneously decreased (RSV: 3.08 +/- 1.38, normal: 3.36 +/- 1.31, t = 2.92, P = 0.225, n = 10). Percentage of membrane TLR4-positive cells was higher in RSV infected population [RSV: (11.99 +/- 7.74)%, normal: (1.16 +/- 0.47)%, Mann-Whitney t value = 0.001, P < 0.01, n = 8], most (93.32 +/- 1.7)% of which were Annexin V positive. IL-8 was significantly induced in the RSV plus LPS costimulation group compared with RSV group (F = 59.29, P < 0.01, n = 3). CONCLUSIONS: RSV induced epithelial TLR4 up-regulation, localization changes from cytoplasm to membrane, IL-8 secretion through TLR4 signaling pathway and epithelial cell apoptosis in membrane TLR4 positive population. These results indicate TLR4 is involved in RSV-induced acute or chronic epithelial-dependent inflammation, which might contribute to acute or chronic airway inflammation.

Inhibition of respiratory syncytial virus of subgroups A and B using deoxyribozyme DZ1133 in mice.

Respiratory syncytial virus (RSV) commonly infects the upper and lower respiratory tracts. Currently, there is no effective treatment available. Deoxyribozymes are a potential therapeutic for RSV and their activity is based on the ability to bind and cleave complementary RNA sequences to inhibit protein expression. DZ1133 is a deoxyribozyme that targets the conserved genomic RNA sequence of the RSV nucleocapsid protein and has been shown to significantly inhibit various strains of RSV including subgroups A and B, standard A2 and CH18537 strains, and CQ381513, CQ381170, BJ01 and BJ04 strains. Treatment with DZ1133 decreased viral plaque formation in lungs of RSV-infected BALB/c mice. In addition, viral mRNA expression was reduced, airway inflammation was alleviated, and leukocyte counts were reduced in bronchoalveolar lavage fluid of RSV-infected mice. The antiviral effect of DZ1133 was dose-dependent (0.2-0.8mg) and more efficient than antisense oligonucleotide inhibition of gene expression. However, levels of cytokines TNF-alpha, IFN-gamma, IL-12, and IL-10 induced by RSV infection were not affected by DZ1133 treatment. Our data demonstrate that DZ1133 is a potential therapeutic agent against both subgroups A and B RSV infection in vivo.

Inhibition of respiratory syncytial virus in cultured cells by nucleocapsid gene targeted deoxyribozyme (DNAzyme).

Respiratory syncytial virus (RSV), which presents the primary cause of bronchiolitis and pneumonia among infants and causes significant morbidity and mortality in immunodeficient patients, remains a health problem worldwide. Unfortunately, an effective vaccine is currently unavailable and pharmacologic treatment needs further optimization for RSV disease. Because RSV is a non-segmented negative-strand RNA virus, it may be sensitive to the genome RNA cleaving by DNAzyme, an artificial nucleic acids molecule with high catalytic capability of cleaving complementary RNA molecules. Thus, RSV-targeted DNAzymes potentially present as a therapeutic candidate of RSV diseases. In this study, DNAzymes targeting the RSV genomic RNA or mRNA were designed and synthesized, one of which (DZn1133) did cleave RSV RNA in vitro, inhibit the transcription and expression of F viral gene, reduce the RSV yield by about 7 logs and protect more than 90% RSV-infected Hep-2 cells from a cytopathic effect at 8 microM. Moreover, 10 wild RSV strains isolated from clinic patients including both subgroups A and B were all suppressed by DZn1133 with greater anti-RSV activity than antisense DNA or ribavirin.

Clinical, molecular, and T cell subset analyses in a small cohort of Chinese patients with hyper-IgM syndrome type 1.

Type 1 hyper-IgM syndrome (HIGM1) is a rare primary immunodeficiency disease caused by mutations in the CD40L gene. Patients often present with recurrent infections and autoimmune manifestations. We investigated the clinical and molecular characteristics of HIGM1 in thirteen patients from the Chinese mainland and examined the proportion of CD4(+)CD25(+)FoxP3(+)Treg, Th17, and Th1 cells in the peripheral blood. We identified ten distinct CD40L mutations in eleven patients: one missense mutation, one nonsense mutation, one insertion mutation (in frame), and seven deletions. Six of these mutations were novel. We observed the percentage of Tregs in the peripheral blood of HIGM1 patients decreased markedly compared with that in healthy controls, but no statistically significant differences was found in the percentages of Th17 and Th1. The identified mutations reflect the heterogeneity of the CD40L gene in HIGM1. Precise genetic diagnosis of HIGM1 will enable appropriate therapeutic interventions, reliable detection of carriers, and genetic counseling. Skewed Treg, Th17/Treg, and Th1/Treg profiles may be associated with immune responses to autoimmunity or infection, which requires replication in larger studies.

Clinical characteristics and mutation analysis of X-linked severe combined immunodeficiency in China.

UNLABELLED: X-linked severe combined immunodeficiency (X-SCID) is a rare, life-threatening immune disorder, caused by mutations of the gene for the γ-chain (γc) of the interleukin-2 receptor, IL2RG. We analyzed the clinical, immunologic, and molecular characteristics of children with X-SCID, attempting to improve the diagnosis and treatment of X-SCID in China. METHODS: X-SCID was suspected in male infants with recurrent or persistent infections. Eleven male infants from ten unrelated Chinese families were included. The IL2RG gene was amplified and sequenced, followed by mutation analysis in these children and their female relatives. X-linked short tandem repeat (X-STR) typing was done to define the maternal lymphocyte engraftment. RESULTS: The 11 children exhibited recurrent infections and 10 of them had lymphopenia. B cells were present in all patients, T cells were markedly reduced in 10, and NK cells were markedly reduced in 9. Nine IL2RG gene mutations were identified in the 11 children, with 5 novel mutations. One patient was found to have the maternal lymphocyte engraftment. CONCLUSION: The clinical presentations and immunologic characteristics of the X-SCID patients were accordingly quite uniform despite the heterogeneity of mutations locating almost in the entire γc gene.

Distribution, clinical features and molecular analysis of primary immunodeficiency diseases in Chinese children: a single-center study from 2005 to 2011.

METHODS: Two hundred three children with genetically proven primary immunodeficiency diseases (PIDs) from 197 unrelated families were enrolled from January 2005 to December 2011. RESULTS: On the basis of criteria developed by the International Union of Immunological Societies, 79 patients were diagnosed as "other well-defined immunodeficiency syndromes" (38.9%), 62 (30.6%) with "predominant antibody deficiencies," 26 (12.8%) with "congenital defects of phagocyte," 25 (12.3%) with "T- and B-cell immunodeficiency" and 11 (5.4%) with "diseases of immune dysregulation." The median time to the diagnosis was 27.9 months and the patients had a wide range of clinical presentations. In addition, a total of 23 pathogenic genes were identified and 213 mutations were detected, including 42 novel mutations. CONCLUSIONS: With the increase in the awareness of PIDs and diagnostic competence, more PID patients will be diagnosed and we will be able to more accurately identify the frequency and the distribution of PIDs in the most populous country in the world.

[Analysis of prenatal diagnosis for seven high-risk fetuses with Wiskott-Aldrich syndrome].

OBJECTIVE: To investigate the value of gene analysis of amniotic fluid exfoliated cells and WASP detection from cord blood in prenatal diagnosis of high-risk fetus with Wiskott-Aldrich syndrome. METHOD: Seven patients with Wiskott-Aldrich syndrome were diagnosed by gene analysis and WASP detected by flow cytometry from 2008 to 2010. After detailed inquiry for medical history and gene analysis of related family members, seven pedigree trees were drawn, including 15 carriers of abnormal genes. From 2008 to 2011, seven samples of amniotic cell gotten by amniocentesis were collected from seven high-risk pregnant women with abnormal gene during 18 to 20 gestational weeks. WASP gene was amplified by polymerase chain reaction (PCR) from DNA of amniotic cell gotten and sequencing was performed directly on the PCR products forward and reversely. Embryo blood sample was collected from one high-risk fetus by needle puncture of umbilical blood vessel and WASP expression was detected by flow cytometry. Karyotyping was performed in amniotic cell gotten cultivated by orthotopic slice and G band staining. Gene analysis of WASP, WASP expression detected by flow cytometry and evaluation of immune function were reexamined in high-risk fetus after delivery. RESULT: Amniocentesis and culture of amniotic cell succeeded in all the seven fetuses. Gene analysis and karyotyping showed that one male fetus and four female fetuses were normal and two female fetuses were carriers. WASP expression detected from embryo blood sample of the patient was normal. After delivery, the result of gene analysis, WASP detection and evaluation of immune function was the same as that of prenatal diagnosis. CONCLUSION: Karyotyping, gene analysis and WASP detection of cord blood can provide reliable service of prenatal diagnosis for high-risk pregnant women with Wiskott-Aldrich syndrome.

Resveratrol Inhibits respiratory syncytial virus-induced IL-6 production, decreases viral replication, and downregulates TRIF expression in airway epithelial cells.

Respiratory syncytial virus (RSV) is the most common pathogen responsible for lower respiratory diseases in children. So far, there is no effective treatment or preventative vaccine available for RSV infection, although ribavirin and dexamethasone are commonly prescribed. Resveratrol has been shown to inhibit the replication of several other viruses, thus the effect of resveratrol on RSV-induced inflammatory mediators in 9HTEo cell cultures was evaluated, and possible mechanisms of action were explored and compared with dexamethasone and ribavirin. Incubation with resveratrol resulted in decreased IL-6 production and partial inhibition of RSV replication. Resveratrol treatment also inhibited virus-induced TIR-domain-containing adapter-inducing interferon-ß (TRIF) and TANK binding kinase 1 (TBK1) protein expression. These data demonstrate the ability of resveratrol to inhibit cytokine production by RSV in airway epithelial cells, indicating that it might be a therapeutic agent with both anti-inflammatory and antiviral potential for the treatment of RSV infection.

[Comparation of inflammation between wild mice and severe combined immunodeficiency mice with endotoxemia induced by lipopolysaccharide].

AIM: To compare the inflammation between wild BALB/c mice and mice with severe combined immunodeficiency (SCID) with endotoxemia induced by lipopolysaccharide (LPS). METHODS: Endotoxemia models of wild and SCID mice were established by injecting LPS intraperitoneally. Serum was taken before and 3 h, 6 h, 12 h after injecting LPS, liver and lung were taken 12 h after injecting LPS. Alanine transarninase (ALT), aspartate aminotransferase (AST) and blood urea nitrogen (BUN) levels were measured by automatic biochemical analyzer. Liver and lung inflammation injury were observed by H.E staining. TNF-α, IFN-γ, IL-6 and MCP-1 levels in serum were detected by Cytometric Bead Array (CBA). RESULTS: (1) All of SCID mice (8/8) were dead at 12-24 h after injecting LPS, and only one BALB/c mouse (1/8) was dead. (2) ALT and AST levels of SCID mice 12 h after injecting LPS were higer than those of BALB/c mice(P<0.05), but there was no difference of BUN levels between them. (3) The blind liver and lung pathology scores of SCID mice were higher than those of BALB/c mice (P<0.05). (4) TNF-α, IFN-γ, IL-6 and MCP-1 levels in serum at 3 h, 6 h, 12 h after injecting LPS increased significantly, and the cytokine levels of SCID mice were higher than those of BALB/c mice(P<0.05). CONCLUSION: SCID mice don't only excessively secrete inflammatory cytokines after injecting LPS, but also lead to more severe endotoxemia and inflammation injury of organs, which are the important cause of mouse death. The above results also show that the adjustment deficiency of adaptive immunoresponse, abnormal augmentation of innate immunoresponse may be an important cause of severe SIRS of endangering life.

[Pulmonary allergic responses were drived by atomization with ovalbumin in BALB/c mice with intestinal microflora disruption].

AIM: The mice in which the intestinal microflora disruption resulted from antibiotic therapy are challenged by atomization with ovalbumin (OVA) to investigate the relation of allergic airway response and intestinal microflora disruption. METHODS: One hundred and twelve female BALB/c mice were divided at random into 6 groups. They were microbiota disruption I group, control I group, microbiota disruption II group, microbiota disruption and challenge group, challenge group and control II group. Cecal contents were collected for quantitative analysis of the intestinal microflora in mice in the former two groups and in mice in the latter four groups on day 6 and day 14, respectively. On day 14, the bronchoalveolar lavage fluid (BALF) was collected for cells counting. OVA-specific IgE in BALF and sera was detected by ELISA. Parts of lungs were collected for histopathology and detection of Th1 and Th2 cell levels by flow of cytometry. RESULTS: The mice which were given antibiotics suffered from intestinal microbiota disruption. In microbiota disruption and challenge group, eosinophil and lymphocyte infiltration was significant and mucus secretion was increased in lung. The number of total cells, eosinophils, lymphocytes, neutrophils and OVA-specific IgE level were increased in BALF in microbiota disruption and challenge group. Th2 cell levels were increased and Th1 cell levels were not significantly different in microbiota disruption and challenge group compared with those in the control II group. CONCLUSION: The allergic (Th2) immune response can be induced by atomization with ovalbumin in the mice in which the intestinal microflora disruption is resulted from antibiotic therapy. The result suggests that the intestinal microflora disruption is a risk factor for allergy and asthma.