The Study On Relationship Between Gene Polymorphism Of Surfactant Protein B (SPB) And Premature Lung Diseases

Part1Relationship between Polymorphisms of Surfactant Protein B Gene and Neonatal Respiratory Distress Syndrome SusceptibilityObjective To study the relationship between gene polymorphism of Surfactant Protein B (SPB) and neonatal respiratory distress symdrome susceptibilityMethods Genomic DNA was extraced from peripheral blood by EDTA anticoagulation,using a standard Phenol/chloroform extraction procedure, The genetic polymorphism of SPB-18A/C and SPB1580C/T were analyzed with polymerase chain reaction restriction fragment lenth polymorphism(PCR-RFLP) using the restriction enzyme Apapl and Dde I in 91 children diagnosed neonatal respiratory distress syndrome (NRDS)and 182 children born at the same gesation age as control group. Statistical analyses were carred out using SPSS software,version 13.0.The Hardy-weinberg equilibrium was determined for both NRDS and control group. x2test was used to detect the different between two groups.Results:φThe SPB1580TT, CT, and TT genotype frequencies in NRDS patients are 5.5%,30.8%,63.7% versus 6.6%,45.1%, and 48.4% in control group, with no significant difference. The frequencies of C allele were higher in NRDS group than thant of control group (79.1%vs70.9%),P<0.05.(?)The SPB-18AA, AC, and CC genotype frequencies in NRDS patients are 10.9%,40.6%,48.4% versus 6.6%,31.1%, and 62.1% in control group, with no significant difference The frequencies of A allele were higher in NRDS group than thant of control group (31.3%vs22.3%, P<0.05.(?) The Genotype frequency of 1580C/T among Han and American, Brazilian,Dane is 74%VS26%;35%VS65%; 41%VS57%; 46%VS54%。P< 0.05, Japanese is 72%VS28%, P>0.05. The Genotype frequency of -18A/C among Han and American, Brazilian,Dane is 25%VS75%;58%VS42%;57%VS43%;61%VS39%.P< 0.05.Conclusions The genetic polymorphism of SPB-18A/C and SPB1580C/T are independent risk factor of neonatal respirtory distress symdrome. The polymorphism of SPB-18A/C and 1580C/T are different among different races.Part 2The relationship between gene polymorphism of SPB and bronchopulmonary dysplasia susceptibility.Objective To study the relationship of gene polymorphism of SPB and bronchopulmonary dysplasia susceptibility.Methods The genetic polymorphism of SPB-18A/C and 1580C/T were analyzed with polymerase chain reaction restriction fragment lenth polymorphism(PCR-RFLP) and gene sequence in 57 children with bronchopulmonary dysplasia(BPD) and 103 children born at the same gesation age as control group. Statistical analyses were carred out using SPSS software,version 13.0.Results The SPB-18AA, AC, and CC genotype frequencies in BPD patients are 14%,45.6%,40.4%VS4.9%,31.1%.64.1%; in control group, with significant difference (P<0.05),Compared with the SFPB-18CC genotype, the AA genotypes were significantly associated with BPD patients (adjusted OR= 4.69,95%CI 1.4-16.2). SPB1580 polymorphism show no different between BPD group and control grop(P>0.05).Conclusions SPB-18A/C serve as a marker for BPD patients.AA genotype may be a risk factor for BPD. There is no relationship was found between SPB1580 and BPD.Part3Detecting Polymorphisms of Surfactant Protein B-18 Gene using high resolution melting curve analysisObjective:To evaluate the feasibility and sensitivity of detecting the Polymorphisms of Surfactant Protein B Gene-18 infants using high resolution melting(HRM) curve analysis.Methods:HRM was applied to detect the Polymorphisms of Surfactant Protein B-18 in 80 DNA samples, the results were confirmed by DNA Sequence Using 3730XLDNA Sequencer.Result:Three melting curves were obtained by HRM anlysis for Polymorphisms of Surfactant Protein B-18, Type A(42/80), B(33/80), C(5/80) melting curves represent SPB-18CC,CA,and AA respectively. Results of HRM were consistent with DNA Sequence.Conclusion:HRM is an accurate technique to detect Polymorphisms of Surfactant Protein B-18. It is also a low_cost test which is suitable for screening. Part 4Relationship between gentic polymorphism of Surfactant Protein B (SPB)-18 and Neonatal Respiratory Distress SyndromeObjective:To study the relationship between gene polymorphism of Surfactant Protein B (SPB)-18 and neonatal respiratory distress symdrome susceptibility.Methods:100 neonates diagnosed as NRDS and 186 gestational age,gender and birth weight matched neonates were enrolled from May 2009 to Dec 2010. The genetic polymorphism of SPB-18A/C were analyzed with High Resolution Melting(HRM)and gene sequence. Non brochoscopic bronchoalveolar lavage(BALF)was performed. The levels of SPB in BALF were measured using enzyme linked immunosorbent assay(ELISA).Statistical analyses were carred out using SPSS software,version 13.0.Results:The SPB-18AA, AC, and CC genotype frequencies in NRDS patients are 11%,40%,49% versus 6.5%,31.7%, and 61.8% in control group,P>0.05, with no significant difference.The frequencies of A allel is higher in NRDS group than that of in control group, (31%VS22.3%), P<0.05. There was difference in amounts of SP-B in BALF from SPB-18 CC and CA individuals, 2824±486ng/mlVS5167±927ng/ml, P<0.05.Conclusions Our study suggested that SPB-18 polymorphism may confer susceptibility to DRDS in Chinese. Differences in amounts of SP-B resulting from the promoter SNP could affect the clinical presentation of NRDS.