정신분열병과 양극성 정동장애 환자에서 세로토닌 운반체 유전자 다형성과 질환의 감수성간의 연관성 연구 = Association studies between the polymorphism within the promoter of the serotonin transporter gene and disease susceptibility of schizophrenia and bipolar affective disorder

국문 초록 (Abstract)

정신분열병의 발병에 있어서 세로토닌의 연관성은 대뇌 세로토닌 수용체를 길항하는 물질인 LSD가 정신분열병과 유사한 증상을 유발한다는 것, 정신분열병 환자에서 혈소판 또는 혈중 세로토닌 농도가 증가되어 있다는 것, 세로토닌에 의해 이차적으로 세로토닌 수용체가 하향조절(down-regulation)되는 것이 정신분열병과 관련이 있다는 것, 세로토닌-도파민 가설, 정신분열병환자의 대뇌에서 세로토닌 재흡수 부위가 감소된다는 보고, 정신분열병 환자의 전두엽과 측두엽에서 세로토닌 운반체(5-HTT)의 mRNA가 의미있게 증가한다는 보고 등에서 확인되고 있다.
양극성 정동장애환자에서 낮은 활성도의 5-HTT변이가 더 많다는 보고와 우울증 환자와 자살시도 환자에서 5-HTT의 기능이 감소되어 있다는 보고는 양극성 정동장애와 세로토닌의 연관성을 강하게 시사한다. 이와 같은 세로토닌계와 주요 정신과 질환 간의 연관관계에 대한 연구 결과들은 5-HT계가 주요 정신질환의 병태생리와 생물학적 치료에 관여한다는 점을 지지해주고 있는데 5-HTT 유전자의 대립유전자가 한 장애에 결정적 유전자로서 작용하기 보다는 세로토닌계와 관련이 되었을 것이라고 생각되는 여러 장애군에서 조절유전자로 작용한다고 가정해 볼 수 있다.
따라서, 본 연구에서는 중합효소 연쇄반응을 이용하여 정신분열병 및 양극성 정동장애 환자와 정상대조군을 대상으로 각 군간에 백혈구내 5-HTTLPR의 대립유전자의 유전자형을 알아보고 그 빈도와 발현율을 비교하여 유의한 차이가 있는지를 살펴보고 5-HTTLPR의 특정변이와 위의 두 가지 질환간의 연관성을 밝혀보고자 하였다.
대상 환자군은 정신분열병과 양극성 정동장애로 진단된 20세 이상 70세이하의 환자 100명씩을 대상으로 하였다. 환자들 및 정상 대조군 대상자로부터 말초 혈액을 채취하여 백혈구를 분리한 뒤 genomic DNA를 추출하였다. 중합효소연쇄반응을 이용하여 Serotonin transporter 유전자에서 다형성 (Polymorphism)을 보이는 부위에 대한 대립유전자형을 확인하였다. 대조군, 양극성 정동장애 및 정신분열병 환자군들 간의 대립유전자형들의 분포의 차이를 보기 위해서 χ^(2) 검증과 logistic 회귀분석을 시행하였다. 결과는 다음과 같았다.
본 연구에서는 대조군과 양극성 정동장애 환자들 사이에서 5-HTTLPR 유전자형과 대립 유전자 분포에 있어서 유의한 차이를 발견하지 못하였다. 이는 5-HTTLPR 유전자 다형성과 양극성 정동장애가 서로 관련될 수 있다는 가설을 지지하지 않는다는 것을 의미한다.
정신분열증의 경우에는 정신분열증 환자들이 정상 대조군에 비해서 ls 유전자 형을 더 많이 보였으며, 남자 정신분열증 환자들이 남자 정상 대조군에 비해서 ls 유전자 형을 더 많이 보였다. 또한 남자 정신분열증 환자들이 남자 정상 대조군에 비해서 대립 유전자 l의 빈도가 더 높았으며, 발병위험도는 l 대립 유전자가 s 대립 유전자에 비해서 높은 위험도를 보였다.

다국어 초록 (Multilingual Abstract)

Recently, the development of molecular biology has provided lots of evidence of genetic and environmental factors contributing to a better understanding of psychiatric illness. In particular, many studies are underway to find the genes for the causes of different diseases these days. Much research is being done on biological factors of psychiatric diseases, and among these studies, the work on serotonin has been valuable.
The associations between genetic factors and development of major psychiatric illness such as schizophrenia and bipolar affective disorder were being suggested for a long time. Previous studies involved family, adoption and twins. Nowadays, molecular genetic approaches contribute to the understanding of the underlying genetic mechanism for major psychiatric illness. However, the results are contradictory and somewhat confusing due to genetic heterogeneity, incomplete penetrance, misspecification of genetic model. The association study is the proper method for evaluation of diseases associated with many genetic loci and with inapparent types of transmission such as schizophrenia and bipolar affective disorder.
The schizophrenia starts from a very early age and makes disruptions of human thought, affect, cognition and behavior with various features of symptoms, progress, treatment response, and prognosis. There was much research to find out the causes of this disease with biological, psychological and social viewpoints, but little evidence has been found until now. For a long time, many people thought that serotonin was related to schizophrenia. Guddum and Hameed(1954), Wooley and Shaw(1954) noticed that serotonin antagonist, LSD, provoked schizophrenia-like symptoms. Garelis et al(1975), Freedman et al(1981), Jackman et al(1983), and King et al(1985) found that the concentrations of serotonin in the platelet of schizophrenic patient were elevated. Meltzer(1989) suggested the serotonin-dopamine hypothesis that serotonin regulated the activity of dopaminergic system. Several postmortem studies with radioligand reported that serotonin uptake sites were reduced in brains of schizophrenic patients. Hernandez and Sokolov(1997) reported that mRNA of 5-HTT were increased significantly.
The bipolar affective disorder, which has a lifetime prevalence of about 1% and is characterized by fluctuations in mood, ranging from extreme elation or mania to severe depression usually accompanied by disturbances in thinking and behavior. Although the family, twin and adoption studies provide strong evidence of important genetic factors, the mode of inheritance is complex and non-mendelian.
Serotonin(5-HT) is a key neurotrasmitter in the central and peripheral nervous system which is implicated in the control of mood, sleep, appetite and a variety of traits and behaviors. A possible dysregulation of serotonergic neurotransmission has been implicated in the aetiology of schizophrenic psychoses, depression, bipolar affective disorder, obsessive-compulsive disorder. By directing the magnitude and duration of postsynaptic responses, carrier-facilitated 5-HT trasport into and release from the presynaptic neuron are essential for the fine tuning of serotonergic neurotrasmission. So, the gene of 5-HTT is thought to be related to the pathogenesis of affective disorders. Many pharmacological studies, neurobehavioral and therapeutic evidence have suggested the important roles of serotonin in pathogenesis of affective disorders. 5-HTT is thought to have an important role in affective disorders based on the fact that blocking agents of 5-HTT activity have antidepressant effects and can provoke manic symptoms in susceptible patients. The human serotonin transporter gene has been clonde and maps to chromosome 17q11.1-1q12, and two common polymorphism have been described: a Variable-Number-Tandem-Repeat(VNTR) located in intron 2(5-HTT-VNTR) and a deletion/insertion in the transcriptional control region approximately 1kb upstream of the transcription initiation site (5-HTTLPR). The promoter polymorphism has been shown to influence transcription activity and 5-HTT function. There are two alleles in 5-HTTLPR polymorphism and they have different activity of promoters. The long(l) variant has 3 times more transcription activity than short(s) variant. So, the s variant has less expression of 5-HTT and lower uptake of 5-HT. Collier(1996) reported that the affective disorder has more frequent number of low activity variants of 5-HTT. Routleedge and Middlemiss(1996) found that the 5-HTT activity in depressive patients and suicide patients were low.
The purpose of this study is to compare the frequency of allelic combinations of 5-HTTLPR polymorphism in the leukocytes in schizophrenia, bipolar affective disorders and normal controls by PCR and to determine the associations between specific variants and these two diseases.
The control subjects are 100 normal healthy adults, and 100 bipolar disorder pateints and 100 schizophrenic patients were diagnosed according to DSM IV criteria. The peripheral blood from all subjects were taken and the genomic DNAs were extracted. After PCR amplification of 5-HTTLPR region, the PCR products were separated by polyacrylamide gel electrophoresis. The gene products were visualized by ethidium bromide staining and photographed under UV transilluminator. After statistical analysis, following results were taken.
1) The patients with schizophrenia were more likely to have the l/s genotype than controls.
2) The male patients with schizophrenia were more likely to have the l/s genotype than male controls
3) The male patients with schizophrenia were more likely to have the l allele than male controls.
4) In male subjects, a significant association between l allele and schizophrenia.
This results means that there is no relationship between 5-HTTLPR polymorphism and bipolar disorder. And in male subjects the development of schizphrenia is related l allele.

목차 (Table of Contents)

• 목차
• 국문요약 = 1
• Ⅰ. 서론 = 3
• Ⅱ. 재료 및 방법 = 8
• 1. 대상 = 8
• 2. 방법 = 8
• 가. Genomic DNA의 정제 = 8
• 나. 중합효소연쇄반응(Polymerase Chain Reaction : PCR)을 이용한 유전자형의 판별 = 9
• 다. 증폭된 생성물의 분석 = 9
• 라. PCR 생성물의 결정 = 9
• 3. 통계학적 분석 = 10
• Ⅲ. 결과 = 11
• 1. 연구 대상군의 특성 = 11
• 2. 대조군에 대한 양극성 정동장애 환자들의 5-HTTLPR의 유전자형 및 대립 유전자 분포의 비교 = 12
• 가) 전체 환자 분석 = 12
• 나) 성별에 따른 분석 = 13
• 3. 대조군에 대한 정신분열증 환자들의 5-HTTLPR의 유전자형 및 대립 유전자 분포의 비교 = 15
• 가) 전체 환자 분석 = 15
• 나) 성별에 따른 분석 = 16
• Ⅳ. 고찰 = 20
• Ⅴ. 결론 = 22
• 참고문헌 = 23
• Abstract = 26