Genetic Research Into Schizophrenia Associated Genes In A Chinese Han Population

Schizophrenia is a serious mental disorder with a lifetime prevalence of 1% in the general population worldwide. It was characterized by the abnormal mental functions and disturbed behaviors, which characteristically appear as a series of clinical features, such as positive and negative symptoms, and disturbances in basic cognitive functions. Both the incidence and the prevalence of psychiatric diseases are gradually increasing year by year with the development of social economy and the change of a life style and the prevalence of schizophrenia is the highest one among them. The investigation of the cause for schizophrenia has always drawn great attention from the public.In the past decades, people have studied all the risk factors related to schizophrenia, especially the genetic factor to which a high attention was paid. Work on genetic epidemiology showed that schizophrenia was not a simple Mendelian disease but it was very likely to be a complex disease with a polygenic mechanism.The research into schizophrenia susceptibility genes has been a hot spot of mental disease. The completion of the Human Genome Project (HGP) has provided a good opportunity for mapping all the genes involved in human diseases, including schizophrenia. Basically, there are two steps for mapping a disease-related gene in the human genome, the linkage-based genome-wide scan and the regional mapping with linkage disequilibrium (LD) analysis. Genome-wide scanning shows the consistent linkage findings on some chromosome regions, such as 1q21-22, 5q22-23, 6p24-21, 8p22-21, 13q14-33 and 22q11-12.A total of 296 family trios of Chinese Han descent, consisting of healthy fathers, healthy mothers and affected offspring with schizophrenia, and 350 unrelated patients with schizophrenia and 351 unrelated healthy individuals were recruited. Ten SNPs were chosen on each known gene using bioinformatics. SNPs were genotyped using PCR-based RFLP analysis. Genotyping data were put into the SPSS database and pedigree database. The Hardy-Weinberg (H-W) equilibrium was tested for genotypic distributions of SNPs using the chi-square (X2) goodness-of-fit test. The LD between paired SNPs was estimated with UNPHASED programs (Version 2.404). The haplotype-based haplotype relative risk (HHRR) test and the transmission/disequil- ibrium test (TDT) were applied to detect allelic association between an SNP and schizophrenia. Haplotypes consisting of two or more SNPs were tested by the UNPHASED program.The patients were divided into two groups according to the clinical psychotic symptoms. We analyzed psychotic symptoms versus allelic and genotypic frequency of each SNP.To investigate the association between the all genes and schizophrenia, the case-control study was designed. The following are the details of methodology and major results obtained in this study.1. DNA markersSix SNPs present in five genes on the 22q11-12 region were chosen by accessing the databases at http://www.ncbi.nlm.nih.gov/, http://www.ncbi.nlm.nih.gov/SNP and http://snp.cshl.org/ web sites. The candidate SNPs included SNP1 present in the FAAH locus, SNP2 and SNP3 in the PPARD locus, SNP4 in the CLDN5 locus, SNP5 and SNP6 in the IL3 locus, SNP7 and SNP8 in the IL3R locus, SNP9 in the IL6 locus, SNP10 in the IL10 locus respectively.2. The H-W equilibriumThe X2 goodness-of-fit test showed that the genotypic distributions of all 10 SNPs were not deviated from the H-W equilibrium, and thus this sample pool was suitable for the genetic analysis.3. LD between paired SNPsThe estimated LD showed that SNP6–SNP5 and SNP7–SNP8 were in the same LD block.4. Association between SNPs and schizophrenia4.1 The TDT analysisThe TDT analysis showed that both SNP3 on the PPARD and SNP7 on the IL3R were associated with schizophrenia (P=0.002 and P=0.034, respectively).4.2 The HHRR analysis The HHRR analysis revealed allelic association between two SNPs and schizophrenia, one the SNP1 and the other one the SNP3. These findings were consistent with TDT results.5. Analysis for case-controlThe SNP3, SNP5 and SNP7 were associated with schizophrenia. There was a significant difference in frequency of allele and genotype between the patient group and the control group.The association could be also showed between SNP9 and schizophrenia. There was only a significant difference in frequency of allele between the patient group and the control group.6. Analysis for multi-SNP hapoltypeThe analysis for multi-SNP hapoltype transmission is essential to look for a specific haplotype or chromosome possibly carrying a gene for schizophrenia.6.1 Analysis for haplotype transmission.6.1.1 A number of haplotype systems were constructed with these 10 SNPs studied, including 3 two-SNP haplotype systems. The two–SNP haplotype systems included SNP2-SNP3, SNP6-SNP5 and SNP7-SNP8. The globalχ2 test showed that the SNP2-SNP3, SNP6-SNP5 and SNP7-SNP8 haplotype system were associated with schizophrenia.6.1.2 The 1-df test for individual haplotypesIn the study of trios we found that the SNP2(C)-SNP3(C) and SNP7(T)-SNP8(G) haplotypes were excessively transmitted (P<0.05). They may contain a susceptibility allele for schizophrenia respectively.In the study of case-control we found that SNP6(C)-SNP5(T) and SNP7(T)-SNP8(A) haplotypes were excessively transmitted (P<0.05). They may contain a susceptibility allele for schizophrenia respectively. But the SNP7(G)-SNP8(A) haplotype was excessively non-transmitted (P<0.05), this haplotype may carry a disease-resistant allele providing a protective mechanism from the attack of schizophrenia.6.2 Analysis for conditional testThe conditional test was used to test the combined effect of 4 loci on the IL3 and IL3R gene. In the study of trios, we found that the SNP7(G)-SNP5(C), SNP8(G)-SNP5(C) and SNP7(G)-SNP8(A)- SNP5(C) combinations as the protective haplotypes were associated with schizophrenia(P<0.05). In the study of case-control, we found that the SNP6(T)-SNP7(G), SNP6(T)-SNP5(T)–SNP7(G) and SNP6(T)-SNP5(C)–SNP7(G)–SNP8(A) combinations as the protective haplotypes were associated with schizophrenia (P<0.05). But the SNP5(T)-SNP7(T), SNP5(T)-SNP7(T)–SNP8(A), SNP5(C)-SNP7( T)–SNP8(A), SNP6(T)-SNP5(T)–SNP7(T), SNP6(T)-SNP5(T)–SNP7(T)–SNP8(A) and SNP6(C)-SNP5(C)–SNP7(T)–SNP8(A) combinations as the risk haplotypes were associated with schizophrenia(P<0.05).7. The association between SNPs and psychotic symptoms of schizophreniaBecause the genotypic distributions of all 10 SNPs were in the H-W equilibrium, we were able to analyze psychotic symptoms versus allelic and genotypic frequency of each SNP using a case-control design. To investigate the association between SNPs and positive and negative symptoms of schizophrenia, the SPSS program and the UNPHASED program were applied for the statistical analysis. The results were shown as follows:7.1 The allele of SNP1 was associated with hallucination-delusion syndrome.7.2 The allele of SNP2 was associated with genuine auditory hallucination.7.3 The allele of SNP7 was associated with delusion of love.7.4 The allele of SNP9 was associated with illogic thought.7.5 The genotype of SNP2 was associated with genuine auditory hallucination and delusion of jealousy. The genotype of SNP7 was associated with delusion of love and other delusion. The genotype of SNP8 was associated with delusion of negation. The genotype of SNP9 was associated with delusion of persecution and illogic thouhgt. 7.6 The quantitative trait analysis showed that the SNP5 was associated with barrier of emotion exchang, lack of spontaneity and mechanical thought (P=0.029, P=0.047, P=0.014 ). The SNP7 was associated with exaggerationg (P=0.045). The SNP9 was associated with concept disorder (P=0.025 ).8. The association between SNPs and sex of schizophreniaIn the study of case-control, we found that there was a significant difference in frequency of allele and genotype of SNP5 and SNP7 between the female patient group and the male patient group. The T allele of SNP5 and T allele of SNP7 were significant more in the male patient group than in the female patient group.These results suggested that SNP5 was associated with male schizophrenia and SNP7 was also associated with male schizophrenia.