BACKGROUND:Anti-tuberculosis drug induced hepatotoxicity (ATDH) is a major adverse drug reaction associated for anti-tuberculosis therapy. The glutathione S-transferases (GST) plays a crucial role in the detoxification of hepatotoxic metabolites of anti-tuberculosis drugs.An association between GSTM1/GSTT1 null mutations and increased risk of ATDH has been demonstrated in adults. Given the ethnic differences and developmental changes, our study aims to investigate the potential impacts of GSTM1/GSTT1 genotypes on the development of ATDH in Han Chinese children treated with anti-tuberculosis therapy.METHODS:Children receiving anti-tuberculosis therapy with or without evidence of ATDH were considered as the cases or controls, respectively. The GSTM1 and GSTT1 genotyping were performed using the polymerase chain reaction.RESULTS:One hundred sixty-three children (20 cases and 143 controls) with a mean age of 4.7 years (range: 2 months-14.1 years) were included. For the GSTM1, 14 (70.0%) cases and 96 (67.1%) controls had homozygous null mutations. For the GSTT1, 13 (65.0%) cases and 97 (67.8%) controls had homozygous null mutations. Neither the GSTM1, nor the GSTT1 polymorphism was significantly correlated with the occurrence of ATHD.CONCLUSION:Our results did not support the GSTM1 and GSTT1 polymorphisms as the predictors of ADTH in Chinese Han children treated with anti-tuberculosis drugs. An age-related association between pharmacogenetics and ATHD need to be confirmed in the further study.

Host genetic factors play a major role in determining differential susceptibility to human tuberculosis (TB), a re-emerging infectious disease throughout the world. Genetic variations in the IFNG gene coding for interferon gamma (IFN-γ), have been identified in TB patients. To investigate the association of the IFNG polymorphisms with TB susceptibility in Chinese pediatric population. A case–control study of 189 TB patients and 164 controls was performed using single-nucleotide polymorphism (SNP) analysis. Genomic DNA was extracted from leukocytes in peripheral blood. Three SNPs of IFNG, including −1616C/T (rs2069705), +874A/T (rs2430561), and +3234C/T (rs2069718), were selected for genotyping and analysis. The +874A and +3234C alleles were more frequent among TB patients (P = 0.108 and P = 0.088), especially in females (both P = 0.029), although this difference was not significant since Bonferroni corrected significance threshold was 0.025 (two of three SNPs were found to be in linkage disequilibrium). More pronounced differences for the +874 and +3234 polymorphisms were found under the genotype comparison between TB cases and controls in the total population [P = 0.026 (borderline non-significance) and P = 0.020, respectively], and in the female subgroup (P = 0.020 and P = 0.020). The dominant model of inheritance was shown to be significant for +874A and +3234C alleles (both P = 0.019) in the female subgroup. The +874A and +3234C alleles were more frequently found in extrapulmonary TB patients than in controls (P = 0.039). Haplotype analysis carried out on these three SNPs showed the TTT haplotype to be more frequent in controls than in TB cases, and this difference showed a strong significance (P = 0.005). The +874A and +3234C alleles may be related to TB susceptibility in the female subgroup in the Chinese pediatric population of North China. The higher rate of +874A (known to correlate with lower IFN-γ expression) in the extrapulmonary TB subgroup suggests a sufficient IFN-γ expression to be not only an important factor for the onset of TB disease but also for limiting its dissemination to lungs.

OBJECTIVE: Our aim was to describe the patient characteristics, clinical–epidemiological profile, and treatment outcome of childhood tuberculosis (TB). METHODS: A retrospective, descriptive study was undertaken of 1212 children aged 0 to 18 years admitted to Beijing Children’s Hospital for the treatment of TB from January 2002 to December 2010. Statistical significance of category variables was evaluated by using Fisher’s exact test. RESULTS: Fifty-four percent of patients had extrapulmonary tuberculosis (EPTB), 38.8% had tuberculous meningitis, and 31.3% had disseminated TB. The last 2 types were defined as severe TB. Most patients with TB (81.6%) were cured or completed treatment. There were more patients aged <5 years and from rural areas with EPTB than with pulmonary tuberculosis. More severe cases of TB were found in patients aged <1 year than other less severe types of TB. Patients with no bacille Calmette-Guérin vaccination and a contact history at home had a significantly risk of contracting severe TB. Children aged <1 year and those with severe TB were more likely to have poor treatment outcomes (failed to improve or died). Among those with EPTB, only 61.3% and 61.1% had positive results on the purified protein derivative tuberculin skin test and chest radiograph, respectively. CONCLUSIONS: In this referral hospital setting, more pediatric EPTB and severe TB patients were found among children aged <1 year. Age <1 year and having severe TB were risk factors for treatment failure. Thus, prevention and health care in pediatric TB should focus on both EPTB and severe TB.

Objective To analysis risk factors of severe tuberculosis(TB) in children.Method The retrospective study of children under 17 year of age admitted to Beijing Children's Hospital for TB from January 2002 to December 2010 was conducted.The univariate and multivariate logistic regression analysis were used to analyze the relation between risk factors and severe tuberculosis in children.Results The P value of four factors association with severe tuberculosis including age,BCG vaccination,history of exposure to active tuberculosis patient and PPD tuberculin test were less than 0.05,the OR value and 95% CI were 0.716(0.650-0.789),0.606(0.468-0.784),1.483(1.131-1.945) and 0.417(0.317-0.549) respectively.Conclusion The results indicate that the age less than 1 year old,non-BCG vaccinated,close contact with active tuberculosis patient and PPD tuberculin skin test negative are independent risk factors with severe tuberculosis in children.

BACKGROUNDVery few researchers have studied the changes in peripheral lymphocyte patterns in adult tuberculosis (TB) and even less researches have been conducted in pediatric TB. In this study, we obtained blood samples from 114 Chinese pediatric TB patients and 116 matched controls to study the association of phenotypic subsets of peripheral lymphocytes with different clinical phenotypes of TB.METHODSThe subjects were classified as the control group and the TB patients group which were further divided into a pulmonary TB group and an extra-pulmonary TB group (more serious than the former). The distribution of lymphocyte subpopulations, including T lymphocytes, CD4(+) T lymphocytes, CD8(+) T lymphocytes, B lymphocytes, and natural killer (NK) cells, were quantitatively analyzed by flow cytometry.RESULTSCompared to the healthy controls, TB infection was associated with significantly higher B cell (P < 0.0001), and lower T cell (P = 0.029) and NK cell (P < 0.0001) percentages. Compared to pulmonary TB patients, extra-pulmonary TB was associated with relatively higher B cell (P = 0.073), and lower T cell percentages (P = 0.021), higher purified protein derivative (PPD) negative rate (P = 0.061), and poorer PPD response (P = 0.010). Most pulmonary TB cases were primary pulmonary TB (89.1%), and most extra-pulmonary TB cases had TB meningitis (72.1%).CONCLUSIONSThis study demonstrates changes in the lymhocyte distribution in children suffering from different clinical phenotypes of TB; such as primary pulmonary TB, and TB meningitis. These patterns may have significance in understanding the pathogenesis and prognostic markers of the disease, and for developing immunomodulatory modalities of therapy.

OBJECTIVEN-acetyltransferase 2 (NAT2) and cytochrome P450 2EI (CYP2E1) play a crucial role in the drug metabolic process. The aim of this study was to understand the genotype and phenotype polymorphisms of NAT2 and CYP2E1 in the Han Chinese pediatric population in order to provide a theoretical basis for individualized drug treatment.METHODSA total of 341 (211 males and 130 females) randomly sampled Han Chinese children, aged from 2 months to 14 years, were enrolled in this study. Genotyping was carried out by PCR method, and metabolic phenotypes were identified.RESULTSIn this study population, wild genotype was found as a major genotype in seven SNPs of NAT2, rs1801279, rs1041983, rs1801280, rs1799929, rs1799930, rs1208 and rs1799931. The frequency of NAT2 fast metabolism was highest (61.3%), followed by middle to slow metabolism (34.1%). Wild genotype also predominated in the four SNPs of CYP2E1 (rs2031920, rs3813867, rs6413432 and rs72559720) named as CYP2E1*5, *6 and *2, with a frequency of 61.3%, 60.1% and 99.4% respectively. As the relationship between CYP2E1 genotype and phenotype was unknown, phenotyping of CYP2E1 was not done.CONCLUSIONSThe important SNPs of NAT2 and CYP2E1 are predominantly wild genotype in the Han Chinese pediatric population. Fast metabolic phenotype predominates in important SNPs of NAT2.

Objective To analyze the clinical epidemiological characteristics of pediatric tuberculosis (TB) patients.Methods A retrospective,descriptive study was undertaken among pediatric TB patients admitted to Beijing Children's Hospital from January 2002 to December 2010.Clinical data including age,manifestation,type of TB and treatment were collected.Patients were classified into 3 group based on their history of BBC vaccination.Logistic regression analysis was applied to analyze the risk factors to the efficacy of TB treatment.Results ① Overall,1 212 hospitalized TB children were enrolled.Distribution of types of TB in 9 years was similar.Main types of TB were pulmonary TB (45.5%,552/1 212),extra-pulmonary TB (23.5%,285/1 212 and hematogenous disseminated pulmonary TB(15.8%,191/1 212).57.2%(163/285) extra-pulmonary TB was mainly meningitis.② The age of onset for all forms of TB showed a bimodal distribution,under 4 years old and 7-12 years old,accounting for 42.0% (509/1 212) and 30.5% (370/1 212) of the total cases respectively.Severe TB (hematogenous disseminated pulmonary TB or tubercular meningitis) were mainly observed in infants,and the proportion gradually decreased with increasing age.③ The proportion of severe TB in BCG-vaccinated patients was lower than that in non-vaccinated patients,26.9% and 34.6% respectively(P=0.004).④ TB treatment rate of success was related to sex and types of TB in children.Treatment rate of success was lower in females (77.8%) than in males (83.8%),P=0.015.Treatment rate of success was the highest in patients with Pulmonary TB (84.2%) and was the lowest in patients with hematogenous disseminated pulmonary TB (73.8%),P=0.006.Conclusions Pediatric TB was mainly observed in children under 4 years old.Compared with adults TB,children were more susceptible to extra-pulmonary TB and severe TB.Treatment outcome of pediatric TB was poor in severe TB patients and in females.BCG was effective in preventing severe TB especially in infants.BCG vaccination should be strengthened in order to prevent the occurrence of TB in young age children,especially the occurrence of severe TB.

Objective To explore the effect of epitope peptides of Ag85B on in vitro proliferation of human peripheral blood mononuclear cell(PBMC) and to provide theoretical basis for preparation of new tuberculosis vaccines.Methods The epitope of Ag85B was artificially predicted,and polypeptide of Ag85B was then synthesized.After activated by different antigens at the same hours,the proliferation activity of PBMC was analyzed by CCK-8,and compared with those of stimulated by BCG and Ag85B.Results The all five epitope polypeptides of Ag85B activated PBMC proliferation,their OD450 values and proliferation index were higher than that of the control group and traditional BCG antigen.The OD450 value and proliferation index of 28aa epitope peptide was the best in five epitope polypeptides of Ag85B.The OD450 value and proliferation index of peptide of Ag85B were significantly higher than that of BCG(POD4500.01,PPI0.05),but less than that of PHA.The OD450 value and proliferation index of peptide 28aa were significantly lower than that of Ag85B(P0.01).Conclusion The predicted antigens of epitope polypeptides of Ag85B have certain immunogenicity,of which 28aa peptides can be used as an advantage peptide.The further research would be needed to confirm Ag85 as a candidate for epitope polypeptide vaccine.

BACKGROUND & OBJECTIVES:Tuberculosis (TB) bacilli ingested by macrophages evade host immune responses by multiple mechanisms including the inhibition of apoptosis. As the cytochrome-P-450 system (CYP) contributes to apoptosis it has been suggested that genetic variation in CYP may be associated with susceptibility to TB infection. This study was carried out to evaluate cytochrome P-450 polymorphisms in Chinese Han children and to investigate the effect of these polymorphisms in paediatric TB.METHODS:Frequencies for the CYP2C19, CYP3A4, CYP3A5 and CYP2E1 mutated alleles and genotypes were compared between 142 Chinese paediatric TB patients and 150 non-infected controls by real time PCR genotyping on peripheral leukocyte DNA.RESULTS:CYP2C19 (636 G>A, rs4986893) A allele and AG genotype were associated with decreased susceptibility to TB (P = 0.006, OR= 0.33, 95% CI: 0.15-0.76; and P = 0.005, OR =0.31, 95% CI: 0.14-0.72 respectively), as were the CYP3A5 (6986A>G, rs776746) G allele and particularly homozygous GG (recessive mode) genotype (P = 0.004, OR=0.61, 95% CI: 0.43-0.85; and P=0.002, OR=0.47, 95% CI: 0.29-0.76).INTERPRETATION & CONCLUSIONS:The data suggested that CYP2C19 and CYP3A5 polymorphisms affect susceptibility to paediatric TB. Further studies are indicated to confirm and elucidate these observations.

Although interferon gamma release assays (IGRAs) have been widely used for the diagnosis of latent and active tuberculosis in adults, a relative lack of validation studies in children has led to caution in their clinical interpretation. This meta-analysis systematically evaluated two IGRAs (ELISA and ELISPOT) and the tuberculin skin test (TST). We searched databases (PubMed, MEDLINE, Ovid) between January 2000 and January 2011 using search terms of latent tuberculosis infection or tuberculosis and interferon gamma release assay, or T-SPOT.TB test, or QuantiFERON-TB Gold, or ESAT-6, or CFP-10, and child, or childhood, or pediatrics. We also collected data by performing a manual search of references from relevant articles and communicating with selected authors. The meta-analysis was conducted with random effects models to account for heterogeneity between selected studies. The sensitivities of all three tests in active tuberculosis were similar. The pooled sensitivity was 70% for ELISA studies, 62% for ELISPOT studies and 71% for TST. Calculated sensitivities for IGRAs and the TST differ in culture-confirmed tuberculosis [ELISA (85%) vs. ELISPOT (76%) vs. TST (85%)] and clinical diagnosed cases [ELISA (64%) vs. ELISPOT (58%) vs. TST (66%)]. The pooled specificity was 100% for ELISA and 90% for ELISPOT, but was much lower for TST [56% in all included studies and 49% in children with bacillus Calmette-Guerin (BCG) vaccination]. The agreement between the TST and IGRAs in non-BCG-vaccinated children is higher than that in BCG-vaccinated children. In the diagnosis of active tuberculosis in children, the TST and IGRAs have similar sensitivity. By contrast, the specificity of IGRAs is far greater than the TST, particularly in children with previous BCG vaccination.

Objective Human glutathione S-transferases(hGSTs) play a crucial role in the biological detoxification processes of drugs and xenobiotics.In drug metabolic process,the gene polymorphism caused by a homozygous or heterozygous deletion of the GSTM1 and GSTT1 genes was associated with the metabolic speed of glutathione S-transferases.The aim of this study was to investigate the association between gene polymorphisms and the metabolic phenotype of GSTM1 and GSTT1 in normal Chinese Han children,to provide an important theoretical basis for guiding the clinical treatment of drugs which metabolic by liver.Methods Chinese Han healthy children were identified who were randomly sampled from 2005 to 2010 in healthy center of Beijing Children's Hospital.The mean age of them was 5.425 years,ranged from 2 months to 14 years.The participants with positive history of diagnosed cancers,psychiatric disorders,asthma,cataract and cardiovascular disease showing significant association with hGSTs polymorphism were excluded.Demographic and medical information of these children was obtained from the patients' files.Genomic DNA was extracted from peripheral blood by EDTA anticoagulation,using a standard salting-out procedure.The concentration and purity of DNA were estimated spectrophotometrically.Genotyping was carried out by using traditional PCR-RFLP method.Further comparison between our existing data and previously reported frequencies in other ethnic populations,was performed to determine inter-ethnic differences.Results A total of 786 Chinese Han healthy children(486 males and 300 females) were enrolled into the study.In the study population,homozygous deletion(*0/*0) referring to slow metabolism was detected in 59.3%(n=466) for GSTM1 gene and 58.4%(n=459)for GSTT1 gene of the subjects,GSTM1 and GSTT1 homozygous deletions(*0/*0) respectively.The heterozygous deletion(*1/*0) referring to middle metabolism was detected in 34.0%(n=267) for GSTM1 gene and 35.1%(n=276) for GSTT1 gene of the subjects,respectively.The homozygote of non-deletion(*1/*1) referring to fast metabolism was detected in 6.7%(n= 53) for GSTM1 gene and 6.5%(n=51) for GSTT1 gene of the subjects,respectively.There was no co-expression of GSTM1 and GSTT1.And there were no significant statistical differences in gene copy number variation frequency between different genders.The comparisons of GSTM1 and GSTT1 gene genotype frequencies among worldwide populations showed different distribution patterns in Asians,Black and Caucasians.The homozygotes of deletion of the DNA fragments of interest for GSTM1 and GSTT1 gene were more common in Asia population.Conclusions In this study,GSTM1 and GSTT1 slow metabolism(indicated by *0/*0 genotype) was more prone in Chinese Han children.The distributions of GSTM1 and GSTT1 gene polymorphisms varied among different races or regions.It provided an important theoretical basis for formulate treatment plans which suitable for Chinese Han children aiming at the hepatic metabolism drugs.Meanwhile,individualized medical treatment may be fulfilled in Chinese Han children,such as antitubercular treatment.However,observing the frequencies of genetic polymorphism will be insufficient in the absence of any supporting clinical data on catalytic or functional activity concerning the effect of a given genetic variant on the future research,it is planed to present a complete analysis with respect to the role of these variants in influencing the serum levels and therapeutic efficacy of drugs.

Objective To establish the reference ranges of peripheral blood lymphocyte subpopulations(T,B and NK cells)values in healthy Chinese children.Methods Chinese Han children who received admission,preoperative or postoperative physical examination aged of 0-18 years old were enrolled into the study.Included children were divided into infant group(28 d-12 months),toddler age group(1-3 years),preschool age group(3-7 years),school age group(7-12-years)and adolescence age group(12-18 years).Relative counts of lymphocyte subpopulations of T cell(CD3+CD19-),CD4+T cell(CD3+CD4+)、CD8+T cell(CD3+CD8+)、CD4+/CD8+、B cell(CD3-CD19+)and NK cell(CD3-CD16+CD56+)were detected by flow cytometrics dual-color and four-color analysis.Differences in the percentages of lymphocyte subpopulations was compared between gender and among age groups,and the reference ranges of peripheral blood lymphocyte subpopulations were established.Results The adolescence age group was finally deleted becasue of the insufficient sample size.Then the percentages of lymphocyte subpopulations were determined in 592 healthy Chinese Han children aged of 28 d-12 years.①The interior-group comparison between males and females:the interior-group comparison showed significant differences between males and females in T,CD4+T,CD8+T lymphocyte percentages and CD4+/CD8+ value in infant group,toddler age group and school age group(P0.05);②The inter-group comparison between all the lymphocyte subpopulations:the inter-group comparison showed significant differences between all the lymphocyte subpopulations(P0.01)except the T lymphocytes in males(P=0.602),then the result of multiple comparison showed that there were statistical significances between several groups,especially between two younger than 3 years groups and two elder than 3 years of age groups(P0.01),respectively;③The trend of peripheral blood lymphocyte subpopulations with age:the line chart showed T,CD8+T and NK cell percentages were increased with age,CD4+T,B cell percentages and CD4+/CD8+ values were declined with age;the line chart showed a little different trend in increasing or decreasing degree between males and females;④There was a similar trend in the percentages of lymphocyte subpopulations in different ethnic or regional groups such as Chinese,European,American and African,but a little different in numerical value.Conclusions The peripheral blood lymphocyte subpopulations values differ by gender and different age groups.The reference ranges of peripheral blood lymphocyte subpopulations values has been proposed by the current study for healthy Chinese Han children aged 28 d-12 years.