| Systemic lupus erythematosus(SLE) is a chronic noninfectious autoimmune disease, both genetic factors and environmental factors play important roles in the development of this disease. In order to explore the association of the genetic variations of several genetic factors (including cytotoxic T lymphocyte-associated antigen 4, programmed cell death-1, and Methyl-CpG-binding protein 2) with SLE, as well as interactions between genetic factors (including cytotoxic T lymphocyte-associated antigen 4 and programmed cell death-1) and environmental factors for SLE in Han nationality females in Southern regions of Yangtze River in China, study designs as case-control, case-only and family-based association were adopted, with the aid of some molecular biologic techniques, such as polymerase chain reaction(PCR) and restriction fragement length polymorphism(RFLP). The results are as follows:Part I Study on the main effect and interaction effect of genetic polymorphisms of the CTLA-4 and PDCD1 and environmental factors for SLEThe study on association of genetic polymorphisms of the CTLA-4 and PDCD1 with SLE1. For locus CTLA-4 -318, both genotypic and allelic frequencies were not significantly different between SLE patients and controls (x2=2.248, P=0.325; x2=0.08, P=0.370).2. For locus CTLA-4-1722, the results showed that genotypic frequency in case group was significantly different from that in control group (x2=9.300, P=0.010) Individuals with genotype TC or TT had a higher onset risk of SLE as compared with those having genotype CC, with OR=2.004 for TC (95%CI: 1.013-3.968) and OR=2.953 for TT (95%CI:1.451-6.010). For the allelic frequencies of locus-1722, there was also significant difference between patients and controls (x2=9.110, P=0.003). There was a higher proportion of allele-1722T in SLE patients than that in controls, which may imply that the T allele of -1722 may increase the risk for SLE (OR=1.664,95%CI:1.194-2.318)3. For locus PDCD1-PD1.2, the results showed that genotypic frequency in case group was significantly different from that in control group (x2=20.596, P＜0.001). Individuals with genotype AG or GG had a higher onset risk of SLE as compared with those having genotype AA(OR=2.031 for AG,95%CI:1.445-2.856; OR=3.111 for GG, 95%CI:1.284- 7.537). For the allelic frequency of PD1.2, there was significant difference between patients and in controls (x2=17.000, P＜0.001), and there was a higher proportion of allele G in SLE patients than that in controls, which indicates that the G allele of PD1.2 may increase the risk for SLE(OR=2.121,95%CI: 1.479-3.042)4. For locus PDCD1-PD1.5, the results showed that genotypic frequency in case group was significantly different from that in control group (x2=7.235, P=0.027) Individuals with genotype TC had a higher onset risk of SLE as compared with those having genotype CC(OR=1.681,95%CI:1.039-2.718). For the allelic frequency of PD1.5, there was significant difference between patients and controls (x2=6.020, P=0.014), and there was a higher proportion of allele G in SLE patients than that in controls, which indicates that the T allele of PD1.5 may increase the risk for SLE(OR= 1.661,95%CI:1.105-2.496)5. For locus PDCD1-PD1.6, the results showed that genotypic frequency in case group was significantly different from that in control group (x2=7.658,P=0.022) Individuals with genotype AG had a higher onset risk of SLE as compared with genotype AA(OR=1.756,95%CI:1.085-2.841). For the allelic frequencies of PD16, there was also significant difference between patients and controls (x2=7.070, P=0.008), and there was a higher proportion of allele G in SLE patients than that in controls, which indicates that the G allele of PD1.6 may increase the risk for SLE(OR=1.698,95%CI:1.147-2.514)6. It was found that there was significant linkage disequilibrium between alleles of the locus PD1.2 and PD1.5, as well as between the alleles of the locus PD1.2 and PD1.6 of PDCD1 gene (D'=0.195, R0.05; D'=0.035, R0.05). However, no significant difference was found between alleles of two polymorphic sites of CTLA-4 gene (D'=0.363,P＞0.05)7. It was found that the frequencies of haplotypes in PDCD1 gene were significantly different between SLE patients and controls. The haplotypes of A-C-A, G-T-A and G-C-G that were composed of the alleles of PD1.2, PD1.5 and PD1.6 were found significantly associated with SLE, while other haplotypes were not shown any significant association with SLE. When using individuals with no certain haplotype as reference, the haplotype of G-T-A and G-C-G in PDCD1 gene had a higher onset risk of SLE under the additive model (for G-T-A,β=1.6619, Z=3.4976, P=0.0005, OR=5.2693;for G-C-G,β=1.5567, Z=2.8338, P=0.0046,OR=4.7431); and the same effect was shown under the dominant model for haplotype G-T-A and G-C-G (for G-T-A,β=1.5799, Z=3.9499, P=0.0001, OR=4.8545; for G-C-G,β=1.5722, Z=3.5024, P=0.0005,OR=4.8172); whereas the haplotype of A-C-A in PDCD1 showed a protective effect on SLE under the recessive model(β=-0.8062, Z=-3.0525, P=0.0023, OR=0.4466). Moreover, the additive model was selected as the optimal model according to the value of Akaike's information criterion(AIC).Multiple logistic regression analysis of genes and environmental factors for SLE1. The multiple logistic regression model was fitted to by the variables of the relevant genotypes of CTLA-4, PDCD1 and environmental factors. It was shown that both genotypes of TT on -1722 site and AG on PD1.6 site were associated with SLE as compared with genotypes of CC and AA, respectively, under the additive model; and genotype of AG on PD1.6 site was associated with SLE as compared with genotype of AA under the dominant model; while genotype of TT on -1722 site was associated with SLE as compared with genotype of CC under the recessive model. However, the history of chilblain, damp of inhabited environment, photosensitivity, ultraviolet exposure, measles and hazardous substances exposure increased the onset risk of SLE under any of the above models. Moreover, the additive model was selected as the optimal model according to the value of AIC.2. The multiple logistic regression model was fitted to by the variables of the relevant haplotypes of PDCD1 and environmental factors. When using individuals with no certain haplotype as reference, it was shown that both the haplotypes of G-T-A and G-C-G in PDCD1 gene had higher onset risk of SLE under the additive model and dominant model, respectively; while the haplotype of A-C-A in PDCD1 showed a protective effect on SLE under the recessive model; and the haplotype of A-T-G was not responsible for the susceptibility of SLE in any of the above models. However, the history of chilblain, damp of inhabited environment, photosensitivity, ultraviolet exposure, measles and hazardous substances exposure increased the onset risk of SLE under any of the above models. Moreover, the additive model was selected as the optimal model according to the value of AIC.Interactions of gene polymorphisms of CTLA-4 and PDCD1 with environment risk factors for SLE1. In case-only study, we performed log-linear model analysis, the results showed that there were interactions between the genetic polymorphisms of CTLA-4 and PDCD1 and environmental factor. However, no interactions were found between genetic polymorphisms of CTLA-4 and PDCD1, and environmental factors. For the model composed of ultraviolet exposure history, locus-1722 of CTLA-4 gene and locus PD1.6 of PDCD1, according to the optimal model, it was found that interaction existed between the genotype of TT on -1722 site and ultraviolet exposure (OR=4.744,95% CI:1.037-21.737), as well as between the genotype of TC on -1722 site and ultraviolet exposure (OR=4.973,95% CI:1.110-22.287) under the additive model; and interaction was also found existed between the genotype of GG on PD1.6 site and ultraviolet exposure (OR=3.199,95% CI:1.023-10.004),while no statistical siginificance was found for interaction between the genotype of GG genotype on PD1.6 site and ultraviolet exposure under the additive model. For CTLA-4 locus, there existed interactions between the genotype of TT or TC on -1722 site and ultraviolet exposure under the dominant model (OR=4.874,95% CI:1.119-21.242); while no evidence was found that there were statistical significance for interaction between the genotypes on PD1.6 site and ultraviolet exposure under the dominant model. Moreover, interaction between the genotype of GG on PD1.6 site and ultraviolet exposure was found existed under the recessive model (OR=3.714,95% CI:1.235-11.179)2. In case-control study, we performed logistic regression analysis. The results showed that except for the interaction existed between the genotype of TT on -1722 site and UV exposure history (β=3.250, P=0.041) under the additive model, no statistical significance was found under other genetic models between the former two factors and between other genetic polymorphisms and risk environmental factors under different genetic models both from the view of genotype and haplotype.PartⅡStudy on the association of genetic polymorphisms of MECP2 with SLEPopulation-based genetic association study for SLE 1. For locus MECP2-rs2239464, the results showed that genotypic frequency in case group was significantly different from that in control group (x2=6.902, P=0.009) Individuals with genotype AA had a higher onset risk of SLE as compared with those having genotype AG and GG(OR=2.378,95%CI:1.246-4.537). For the allelic frequencies of rs2239464, there was significant difference between patients and controls (x2=6.73, P=0.009), There was a higher proportion of allele A in SLE patients than that in controls, which indicates that the A allele of rs2239464 may increase the risk for SLE(OR=2.170,95%CI:1.196-3.937)2. For locus MECP2-rs2075596, the results showed that genotypic frequency in case group was significantly different from that in control group (x2=14.432, P＜0.001). Individuals with genotype AA had a higher onset risk of SLE as compared with those having genotype AG and GG(OR=3.259,95%CI: 1.772-5.995). For the allelic frequencies of rs2075596, there was significant difference between patients and controls (x2=14.16, P＜0.001). There was a higher proportion of allele A in SLE patients than that in controls, which indicates that the A allele of rs2075596 may increase the risk for SLE (OR=2.807,95%CI:1.613-4.884)3. It was found that there was significant linkage disequilibrium between the two sites of MECP2 gene (D'=0.19,P＜0.05)4. The frequencies of haplotypes in MECP2 gene were significantly different between SLE patients and controls. The haplotypes of A-A and G-G that composed of the alleles of rs2239464A/G and rs2075596A/G were found significantly associated with SLE, while other haplotypes did not show any statistical significance. When using individuals with no certain haplotype as reference, the haplotype of A-A in MECP2 gene had a higher onset risk of SLE under the additive model (β=1.0038, Z=2.7300, P=0.0063, OR=2.7286); and the haplotype of G-G in MECP2 gene had a lower onset risk of SLE under the dominant model (β=-0.9080, Z=-2.4191,P=0.0156, OR=0.4033); while the haplotype of A-A in MECP2 gene had a higher onset risk of SLE under the recessive model (β=1.0076, Z=3.5349, P=0.0004, OR=2.739) Moreover, the additive model was selected as the optimal model according to the value of AIC.The family-based association test on SLE1. The transmission disequilibrium test showed that rs2239464 may not increase the transmission of the A allele from heterozygous parents to affected offspring (x2=0.2, P>0.05); and univariate (single-marker) family-based association tests demonstrated that alleles at SNP rs2239464 of the MECP2 gene was not associated with genetic susceptibility to SLE under either of the additive and recessive model, independently (for additive model, Z=0.447, P=0.655; for recessive model, Z=0.447, P=0.655)2. The transmission disequilibrium test showed that rs2075596 have an excess of transmission of the A allele from heterozygous parents to affected offspring (x2=6, P<0.05); and univariate (single-marker) family-based association tests demonstrated that A allele at rs2075596 site of the MECP2 gene was significantly associated with genetic susceptibility to SLE under additive and recessive mode, independently (for additive model, Z=2.646, P=0.008; for recessive model, Z=2.646, P=0.008), which indicates that A allele at rs2075596 site of the MECP2 gene may be a conservative susceptible allele.
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