Preliminary Study On The Genetic Effects And Genetic Mode Of First-episode Depression, As Well As Association And Expression Study Of CREB1, BDNF Gene In Major Depression

(Part I)A preliminary study on the genetic effects and genetic mode of first-episode major depressionObjective The pilot study was to probe into the genetic effects and genetic mode of first-episode major depression.Methods According to strict inclusion and exclusion criteria, 107 casesdiagnosed of first-episode major depression were studied using family history method. We investigated the illness history of 4 439 relatives, drew the pedigree chart and calculate the prevalence rate of mental disorders, especially major depression. A mathematic method in medical genetics, including chi square test and offspring correction of segregation analysis and the Falconer's multifactorial threshold theory, was applied for the research into the inheritance mode.Results (1) In all the pedigrees investigated, 28 probands had family history of major depression, accounting for 26.17%, higher than that of other psychiatric disorders. (2) The prevalence of first-episode major depression in the proband's relatives was 0.87%, higher than 0.02% in general population from the epidemic survey in the seven areas of China in 1993 (P<0.01). The prevalence in the first-, second- and third-degree relatives was 7.46%, 0.37%, and 0.05%, respectively. The prevalence of female is higher than that of male. (3) The adjusted segregative rate was 0.20, and it was not significantly different from the rate of 0.25 in autosome recessive inheritance. The weighted mean of the heritability and standard error was (109.73±5.26)%. The expected prevalence (7.20%, 0.62% and 0.13%) was not significantly different from the real prevalence (7.46%, 0.37% and 0.05%) in the first-, second- and third-degree relatives, respectively.Conclusions First-episode depression has evident genetic effect, its mode ofinheritance may conform to polygenic inheritance with a major autosome recessivegene.(Part II)Association and expression study of the cyclic AMP response element-binding protein gene in major depressionObjective The genetic pathogenesis of major depression has not been elucidated, although family, twin and adopt studies suggested genetic factor was very important. The aim of the present study was to investigate relationship between cyclic AMP response element-binding protein (CREB1) gene and major depression.Methods (1) We recruited 105 parent-offspring trios of Chinese descent, drew the peripheral whole blood and extracted genomic DNA. By means of restriction fragment length polymorphism (RFLP)/polymerase chain reaction (PCR), we genotyped the two single nucleotide polymorphisms (SNPs), including rsl0932201 and rs6740584, of CREB1 gene. Then we performed the single-marker transmission disequilibrium test (TDT), linkage disequilibrium (LD) between the two SNPs, and haplotype-based TDT to search for the association between CREB1 gene and major depression, applying the ETDT software, EMLD program and TRANSMIT version 2.5.2 software. (2) 36 first-episode drug-naive depressive patients, 15 depressive patients administrated with new antidepressants for 12 weeks, and 44 healthy controls were included. We performed relative TaqMan real-time quantitative reverse transcription (RT)-PCR of CREB1 gene in the leukocytes using the standard curve method on ABI PRISM(?) 7900 Sequence Detection System. Amplification of an endogenous control, house-keeping GAPDH gene, was performed to standardize the amount of unknown samples. Furthermore, we analysed the relationship between CREB1 gene expression levels and gender, onset age, severity of depressive symptoms, anxiety symptoms, scores of HAMD-24 and HAMA rating scale, as well as therapeutic efficacy.Results (1) Single-marker TDT analysis showed that both the two SNPs including rs 10932201 and rs6740584 were not significantly associated with major depression in all parent-offspring trios (x~2=2.700 and 0.458, respectively. P=0.1004 and 0.4986, respectively.). (2) Pairwise LD analysis by EMLD program indicated that there was significant linkage disequilibrium between SNPs rs 10932201 and rs6740584, with D' value 0.884. (3) Haplotype-based TDT analysis demonstrated that there was strongpositive association between the rsl0932201-rs6740584 haplotype and major depression (global x~2 (3df)=23.458. P=0.00003241), and haplotypes A-C and A-T both were associated with major depression (x~2=5.405 and 13.623, respectively. P=0.020 and 0.00022, respectively.). (4) CREB1 gene expression level (amole/0.3μl cDNA) in peripheral blood of 36 pre-treatment patients, 15 pre-treatment* patients, 15 post-treatment* patients and 44 healthy controls was 0.197±0.103, 0.232±0.140, 0.186±0.065 and 0.168±0.055, respectively. There was no significant difference of expression levels between pre-treatment group and healthy control group (P=0.139), and between pre-treatment* group and post-treatment* group (P=0.321). (5) There were no significant differences between male patients-controls, female patients-controls and male-female patients (P=0.157, 0.669 and 0.133, respectively). (6) After dividing 36 patients into early-onset and late-onset group, we found that CREB1 gene expression levels were not associated with first-episode age (P all>0.05). However, there seemed to be a trend of difference between early-onset group and healthy control group (P=0.054). (7) After dividing 36 patients into severe depression and mild-moderate depression group, as well as severe anxiety and mild-moderate anxiety group, we found no significant differences of CREB1 gene expression levels in different subgroup (P all>0.05). Moreover, CREB1 gene expression levels were not correlated with total scores and factor scores of HAMD-24 and HAMA rating scales (P all>0.05). (8) There was no significant correlation between CREB1 gene expression level change and reduce rate of HAMD-24 rating scale (r=-0.172, P=0.540).Conclusion (1) Single-marker TDT analysis indicated that both the SNPs rsl0932201 and rs6740584 were not significantly associated with major depression in 105 parent-offspring trios. However, there was strong positive association between the rs10932201-rs6740584 haplotype and major depression. The result showed that this haplotype may play an important role in the pathogenesis of major depression, and CREB1 gene may be a candidate gene conferring the susceptibility to major depression. (2) CREB1 gene expression level in peripheral blood of depressive patients was not different significantly from that of healthy controls, also was not related to gender, onset age and symptom clusters. Moreover, antidepressant therapy might not exert significant influence on CREB1 gene expression level in peripheral blood.(Part III)Association study between the brain-derived neurotrophic factorgene and major depressionObjective The genetic pathogenesis of major depression has not been elucidated, although family, twin and adopt studies suggested genetic factor was very important. The aim of the present study was to ascertain for the association between brain-derived neurotrophic factor (BDNF) gene and major depression.Methods We recruited 105 parent-offspring trios of Chinese descent, drew the peripheral whole blood and extracted genomic DNA. By means of restriction fragment length polymorphism (RFLP)/polymerase chain reaction (PCR), we genotyped the three single nucleotide polymorphisms (SNPs), including rs6265, rs 10835210 and rs2030324 of BDNF gene. Then we performed the single-marker transmission disequilibrium test (TDT), pairwise-SNP linkage disequilibrium (LD), and haplotype-based TDT to search for the association between BDNF gene and major depression, applying the ETDT software, EMLD program and TRANSMIT version 2.5.2 software.Results (1) Single-marker TDT analysis showed that the three SNPs including rs6265, rsl0835210 and rs2030324 were not significantly associated with major depression in all parent-offspring trios (x~2=0.374, 0.474 and 0.000, respectively. P=0.5408, 0.4913 and 1.0000, respectively.). (2) Pairwise LD analysis by EMLD program indicated that there was significant linkage disequilibrium between SNPs rs6265 and rs10835210, rs10835210 and rs2030324, and rs6265 and rs2030324 (D' value was 0.447, 0.922 and 0.669, respectively). (3) Multiple-marker TDT demonstrated that there was no association between the BDNF haplotypes from rs6265-rs10835210-rs2030324 and major depression (global x~2 (6df)=9.216. P=0.162). Every haplotypes showed no association with major depression (P all >0.05), but it seemed that there was a trend of association between A-A-T haplotype and major depression (x~2=3.516, df=1, P=0.061).Conclusion single-marker TDT and haplotype-based TDT showed that three SNPs (rs6265, rsl0835210 and rs2030324) and haplotype of BDNF gene were not associated with major depression. It seemed to be reasonable that the three SNPs ofBDNF gene are unlikely to play a critical role in the genetic susceptibility to major depression.