| Schizophrenia is a serious mental disorder with a lifetime prevalence rate 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. While the pathogenesis for schizophrenia remains unknown, several lines of evidence from family, twin and adoption suggest that genetic factors are likely to play an essential role in the developing of schizophrenia and influence susceptibility to schizophrenia. Epidemiological data have demonstrated that schizophrenia is not a simple Mendelian disease but looks like a complex disease involving several genes with each susceptibility gene having only a modest individual effect.The research for 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. So far, the major and specific susceptibility genes leading to schizophrenia remains unidentified.Phospholipid is the important composition of cellular membrane, at the same time, it plays the important role in signal transmission progress in brain. If phospholipid metabolizes disorderly, it will influence the function of nerve cell and lead to psychogenia finally. Phospholipases A2 (PLA2) are key-enzymes in the process of phospholipid metabolism. The abnormality of PLA2 will lead to phospholipid abnormality of nerve cellular, and lead to emotional disturbance accordingly. Several studies suggest increased activity of phospholipase A2 in schizophrenic patients, it thoroughly explains the relationship between PLA2 and schizophrenia. ObjectiveTo investigate a genetic association between the polymophism of cytosolic phospholipase A2(cPLA2) family genes and schizophrenia by detecting SNPs of cPLA2 family genes with the bioinformatics and the molecular genetics. The present study tried to reveal the predisposing genes and molecular genetic mechanism for schizophrenia. MethodsA total of 201 family trios of Chinese Han descent, consisting of fathers, mothers, and affected offspring with schizophrenia, were recruited for the genetic analysis. By using bioinformatics methods 10 SNPs were chosen on cPLA2 family genes including rs7542180 present in the PLA2G4A locus,rs156631,rs2307279,rs2162886 and rs891014 in the PLA2G4C locus, rs1668589 in the PLA2G4D locus,rs4924595 in the PLA2G4E locus,rs1356410 in the PLA2G4F locus,rs3828323 and rs3749117 in the PLA2R1 locus. SNPs were genotyped using PCR-based LDR analysis. Genotyping data were put into the SPSS database. The Hardy-Weinberg (H-W) equilibrium was tested for genotypic frequency distributions of SNPs using the goodness of fit test. The LD between paired SNPs was estimated with UNPHASED programs. The haplotype relative risk (HRR) test and the transmission disequilibrium test (TDT) were applied to detect allelic association between SNPs and schizophrenia with the SPSS13.0 program. To elucidate genetic heterogeneity, in addition, schizophrenic patients were sub-grouped based on their clinical symptoms and the genetic association between SNPs and clinical subgroups was then analyzed. The conditional test was performed with the UNPHASED program. The conditional test was used to test the combined effect of distinct loci on the disease by conditioning on allele(COA)or by conditioning on genotype (COG). Theχ2 test was used to test the association between the SNPs of cPLA2 family genes and clinical symptoms of schizophrenia. Results(1)The analytic results of questionnaire dataThe analytic results showed that first age of onset of schizophrenia was between 14 and 45 years old, the mean of first age of onset was 24.58±6.182 years old. There were no difference between the male and female schizophrenia with first age of onset, course of disease, positive family history and classification diagnosis (t = 0.986,P = 0.325;t = 0.745,P = 0.457;χ2 = 0.872,P = 0.350;χ2 =10.875, P=0.092). There were no difference between the positive family history and negative family history schizophrenia with first age of onset and course of disease(t =0.282,P=0.778;t =1.618,P=0.108). But the incidence rate of the other delusion symptoms was higher in positive family history schizophrenia than in negative family history schizophrenia(χ2 =9.276,P=0.002).There was significant difference between the male and female schizophrenia at the clinical symptoms of experience of being revealed, delusion of love, other delusion, illogic of thinking, bizarre behavior and Apathy ( P < 0.05 ). The incidence rate of the six clinical symptoms were higher in female schizophrenia than in male schizophrenia. Character before illness of most schizophrenia was introvert and there was sex difference(sχ2 =18.717,P<0.001). Introvert was 90.5 percent in all of schizophrenia. The incidence rate of extroversion and intermedial in character before illness were higher in female schizophrenia than in male schizophrenia. Most schizophrenia were paranoid and undifferentiated type and there was no sex differences(χ2 = 0.435,P = 0.510).(2)Hardy-Weinberg equilibrium testThe goodness of fit test showed that genotype frequency distributions of all subjects were not deviated from the H-W equilibrium, thus these samples were suitable for the genetic analysis.(3)LD between paired SNPsThe estimated LD showed that rs2162886 and rs981014 were in the same LD block with UNPHASED programs. (4)HRR analysisIn total samples, HRR analysis showed that there was no significant difference for the frequencies of alleles of these 10 SNPs between the case and the control (P>0.05). This indicated no association between these 10 SNPs and schizophrenia.In male and in female schizophrenia, there was no significant difference for the frequencies of alleles of these 10 SNPs between the case and the control (P>0.05). This indicated no association between these 10 SNPs and the different sex schizophrenia.In paranoid and in undifferentiated schizophrenia, there was no significant difference for the frequencies of alleles of these 10 SNPs between the case and the control (P>0.05). This indicated no association between these 10 SNPs and paranoid schizophrenia,as so as undifferentiated schizophrenia.(5)TDT analysisIn total samples, TDT analysis showed that the probability of the two different alleles from heterozygous parents didn't deviated from 50% at these 10 SNPs(P>0.05). This indicated no association between these 10 SNPs and schizophrenia.In male and in female schizophrenia, the probability of the two different alleles from heterozygous parents didn't deviated from 50% at these 10 SNPs(P>0.05). This indicated no association between these 10 SNPs and male and female schizophrenia.In paranoid and in undifferentiated schizophrenia, the probability of the two different alleles from heterozygous parents didn't deviate from 50% at these 10 SNPs (P>0.05). This indicated no association between these 10 SNPs and paranoid and undifferentiated schizophrenia.(6)Analysis for clinical subgroupsThe results showed that rs156631, rs2307279, rs2162886, rs891014, rs1668589, rs3828323 and rs3749117 were associated with schizophrenia. Since the time at which SNPs occurred differs, each SNP may have its own genetic heritage with a different haplotype and non-random association. They may affect each other in allele frequency distribution, so as to reduce the power to detect their association with the illness. The advantage of clinical subgroup analysis is to limit the interference from different LD signals to reduce the false negative results and validate the hypothesis of genetic heterogeneity.(7)Analysis for the combined effects of multiple locusAnalysis for haplotype transmission showed that the rs2162886-rs981014 haplotype systems did not associated with schizophrenia (P>0.05). The conditional test was used to test the combined effect of distint loci on the disease by conditioning on allele(COA)or by conditioning on genotype(COG). The COA test showed a disease association for the rs2162886-rs1668589 and rs891014-rs1668589 combinations. The COG test also showed a disease association for the rs2162886-rs1668589, rs891014-rs1668589 and rs2307279-rs7542180 combinations.(8)The association between SNPs and psychotic symptoms of schizophreniaIn total samples , seven SNPs, including rs2307279,rs1356410,rs2162886,rs891014,rs1668589,rs3749117 and rs3828323, were associated with some positive symptoms of schizophrenia. The rs2162886 and rs891014 were found to be associated with Character before illness of schizophrenia.In male schizophrenia, five SNPs, including rs1356410,rs2162886,rs891014,rs3749117 and rs1356410, were associated with some positive symptoms of schizophrenia. The rs2162886 and rs891014 were found to be associated with Character before illness of schizophrenia.In female schizophrenia, three SNPs, including rs2307279,rs3749117 and rs1356410, were associated with some positive symptoms of schizophrenia. The rs1668589 was found to be associated with Character before illness of schizophrenia. The rs891014 was found to be associated with some negative symptoms of schizophrenia.ConclusionsAccording to the above mentioned analytic results, we can reach the following conclusions:①PLA2G4C(rs2307279) locus was associated with delusion of sin, one of positive symptoms of schizophrenia.②PLA2G4F(rs1356410) locus was associated with delusion of negation, one of positive symptoms of schizophrenia.③P LA2G4C(rs2162886,rs891014) loci was associated with illogic thought, one of positive symptoms of schizophrenia. ④PLA2G4D(rs1668589) and PLA2R1(rs3749117) loci was associated with bizarre behaviour, one of positive symptoms of schizophrenia.⑤PLA2R1(rs3828323) locus was associated with hallucination-delusion syndrome, one of positive symptoms of schizophrenia.⑥P LA2G4C(rs2162886,rs891014),PLA2G4D(rs1668589),PLA2G4F(rs1356410),PLA2R1(rs3828323) loci was associated with the clinical symptoms of paranoid schizophrenic.⑦PLA2G4F(rs1356410),PLA2R1(rs3749117),PLA2G4C(rs2162886,rs2307279,rs891014) loci was associated with the clinical symptoms of undifferented schizophrenic.⑧The haplotypes, including rs2162886-rs1668589,rs891014-rs1668589 and rs2307279-rs7542180, was associated with schizophrenia.⑨There was no sex difference at the first age of onset, course of disease and positive family history. The female schizophrenia has more clinical symptoms. The frequency of some clinical symptoms, including delusion of being revealed,delusion of love,other delusion,illogic thought,bizarre behaviour and apathy, were greater in the female schizophrenia than that in the male schizophrenia.⑩There was no difference at the first age of onset and course of disease between schizophrenia who had positive family history and ones who had not positive family history. The frequency of other delusion was greater in the schizophrenia who had positive family history than in the ones who had not positive family history.In a word, these findings thoroughly suggested that cPLA2 genes associate with schizophrenia. The genetic association of combined effect of distinct loci with schizophrenia support the polygenic theory of complex disease. At the same time, these findings proved that schizophrenia has the clinical and genetic heterogeneity.This study was very important for elucidating the etiology and genetic mechanisms of schizophrenia at a molecular level, and also for the development of genetic diagnosis, new drugs for the treatment of the illness and prediction of schizophrenia risk. The current study would also provide study strategy and precious experience for other complex genetic disorders.
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