Association Study Of Single Nucleotide Polymorphisms And Expression Level Of IL12-IFNγ Axis Genes With Spinal Tuberculosis In A Han Population Of Hunan Province

Part1:Association study of single nucleotide polymorphisms in IFNy、IFNyR、IL12and IL12R genes with spinal tuberculosisObjective:To investigate the relationship of single nucleotide polymorphisms in IFNy, IFNyR, IL12and IL12R genes and susceptibility to spinal tuberculosis.Methods:104patients with spinal tuberculosis (sTB),106healthy controls were recruited in our study from December2010to December2012. The polymorphisms at positions:IFNG:rs2430561, IFNGR1: rs2234711、rs7749390、rs1327474, IL12B:rs3212227, IL12RB1: rs436857、rs393548were detected by polymerase chain reaction and direct DNA sequencing. The alleles and genotypes frequency were analyzed by Statistic and the relationship between them and susceptibility to tuberculosis is evaluated.Results:1. In the Spinal tuberculosis group, the Alleles’frequency distributions of A, T at IFNG rs2430561were29.8%,70.2%. The genotypes’frequency distributions of AA, AT, TT at IFNG rs2430561were10.6%,38.5%,53%. The "A" allele frequency was significantly higher at rs2430561in spinal tuberculosis group than normal.(χ2=5.080, p=0.0242). Proportion of spinal tuberculosis AA genotype was significantly higher than normal (χ2=6.090, p=0.048). This indicates that this polymorphism associated with the occurrence of spinal tuberculosis, A allele (OR=1.67, CI=[1.067～2.611]) and AA genotype (OR=4.566,95%CI=[1.211～17.210]) carriers suffer from spinal tuberculosis risk.2. In the Spinal tuberculosis group, the Alleles’frequency distribution of C, T at IFNGR1rs2234711were62.5%,37.5%, the genotypes’frequency distribution of CC, CT, TT at IFNGR1rs2234711were40.4%,44.2%,15.4%. There was no difference between spinal tuberculosis group and normal, no association between this polymorphism and spinal tuberculosis.3. In the Spinal tuberculosis group, the Alleles’frequency distribution of T, C at IFNGR1rs7749390were40.9%,59.1%, the genotypes’frequency distribution of TT, CT, CC at IFNGR1rs7749390were18.3%,45.2%,36.5%. There was no difference between spinal tuberculosis group and normal, no association between this polymorphism and spinal tuberculosis.4. In the spinal tuberculosis group, the Alleles’frequency distribution of A, G at IFNGR1rs1327474were87.5%,12.5%, the genotypes’frequency distribution of AA, AG, GG at IFNGR1rs1327474were75%,25%,0. There was no difference between spinal tuberculosis group and normal, no association between this polymorphism and spinal tuberculosis.5. In the spinal tuberculosis group, the Alleles’frequency distribution of A, C at IL12B rs3212227were60.1%,39.9%, the genotypes’frequency distribution of AA, AC, CC at IL12B rs3212227were34.6%,51%,14.4%. There was no difference between spinal tuberculosis group and normal, no association between this polymorphism and spinal tuberculosis.6. In the spinal tuberculosis group, the Alleles’frequency distribution of A, G at IL12BR1rs436857were15.4%,84.6%, The "A" allele frequency was significantly higher at rs436857in spinal tuberculosis group than normal (χ2=6.371, P=0.012), the genotypes’frequency distribution of GG, AG, AA at IL12BR1rs436857were72.1%、25%、2.9%. Proportion of spinal tuberculosis AA genotype was significantly higher than normal (x2=6.726, P=0.035). This indicates that this polymorphism associated with the occurrence of spinal tuberculosis, A allele carriers suffering from spinal tuberculosis risk (OR=2.2272,95%CI=[1.182～4.197]).7. In the spinal tuberculosis group, the Alleles’frequency distribution of T, A at IL12BR1rs393548were88.7%,11.3%, the genotypes’frequency distributions of TT, AT, AA at IL12BR1rs393548were78%,24%,2%. There was no difference between spinal tuberculosis group and normal, no association between this polymorphism and spinal tuberculosis.8. Polymorphic loci of IFNGR1rs2234711, rs7749390, rs1327474were tested by linkage disequilibrium analysis:rs223711and rs7749390were in the same chain box, haplotype analysis can be carried out. The haplotype of rs223711and rs7749390polymorphisms C-T is the risk factor of spinal tuberculosis (P<0.005, OR=10.838,95%CI=[1.376～85.374]). Polymorphic loci of IL12BR1rs436857and rs393548were tested by linkage disequilibrium analysis, there is no linkage disequilibrium (D’=0.854, r2=0.665), haplotype analysis can not be performed.Conclusions:1. IFNG rs2430561polymorphism associated with the incidence of spinal tuberculosis, A Allele and AA genotype are risk factors for spinal tuberculosis.2. IL12RB1rs436857polymorphism associated with the incidence of spinal tuberculosis, A Allele is a risk factor for spinal tuberculosis.3. IFNGR1rs2234711, rs7749390, rs1327474, IL12B rs3212227, IL12RB1rs393548polymorphic loci have no association with spinal tuberculosis susceptibility.4. IFNGR1polymorphisms rs2234711and rs7749390CT haplotype is a risk factor for tuberculosis of the spine, haplotype CT carrier carry a higher risk of spinal tuberculosisPart2:Association study of serum expression level of IFNy and IL12and related impact factors with spinal tuberculosisObjective:To explore the association between serum IFNy and IL12concentration and spinal tuberculosis. To explore the association between serum IFNy, IL12concentration and IFNG, IL12genes’single nucleotide polymorphisms.Methods:104patients with spinal tuberculosis (sTB),106healthy controls were recruited in our study from December2010to December2012. Their serum IFNy and IL12expression were detected by enzyme linked immunosorbant assay(ELISA). Explore the association between serum IFNy and IL12concentration and development of spinal tuberculosis combined with IFNG and IL12B genotypes which detected before.Results:In spinal tuberculosis patients, the mean serum IFNy concentration was28.19±8.27pg/ml. It was significantly lower than normal controls’(39.54±10.14pg/ml)(t=-8.89, P<0.001). The mean serum IL12concentration was55.11±13.13pg/ml. It was significantly lower than normal controls’(73.16±15.90pg/ml)(t=-8.967,P<0.001). Serum IFNy、 IL12concentrations can be used as serological markers for clinical diagnosis of spinal tuberculosis. In Hunan Han population, IFNG rs2430561AA genotype may cause a decrease in serum IFNy concentration; while IL12B rs3212227polymorphism have no significant correlation with serum concentration of IL12, reduced IL12serum level in spinal tuberculosis group may be caused by other factors.Conclusions:1. Serum IFNy, IL12concentrations of spinal tuberculosis patients are lower than normal.2. IFNG rs2430561AA genotype can cause serum IFNy concentration decreased; IL12B rs3212227polymorphism has no significant correlation with serum concentrations of IL12.