Title page
ACKNOWLEDGEMENT
Contents
CHAPTER 1. INTRODUCTION 10
1. Introduction 11
1-1. Infertility 11
1-2. Female Infertility 12
1-3. Male Infertility 13
1-3-1. Azoospermia 14
2. Cause of Male Infertility 15
2-1. Pre-Testicular Infertility 15
2-2. Testicular Infertility 15
2-3. Post-Testicular Infertility 16
3. Unexplained Male Infertility 17
4. Single Nucleotide Polymorphism 18
CHAPTER 2. ASSOCIATION STUDY OF FOUR POLYMORPHISMS IN THREE FOLATERELATED ENZYME GENES WITH NONOBSTRUCTIVE 20
ABSTRACT 21
INTRODUCTION 23
MATERIALS AND METHODS 28
RESULTS 33
DISCUSSION 50
CHAPTER 3. ASSOCIATION OF POLYMORPHISM IN THE H2BFWT GENE WITH MALE INFERTILITY IN KOREAN 54
ABSTRACT 55
INTRODUCTION 57
MATERIALS AND METHODS 59
RESULTS 63
DISCUSSION 70
CHAPTER 4. CONCLUSIONS 72
REFERENCES 74
ABSTRACT IN KOREAN 87
Table 1-I. Distribution of the MTHFR 677C]T, 1298A]C, MTR 2756A]G and MTRR 66A]G genotypes in the infertile and fertile males 43
Table 1-II. Genotype and allelic frequencies for MTHFR 677C]T, 1298A]C, MTR 2756A]G and MTRR 66A]G in the azoospermia group (n=174) 44
Table 1-III. Genotype and allele frequencies for MTHFR 677C]T, 1298A]C, MTR 2756A]G and MTRR 66A]G in the OAT group (n=186). 45
Table 1-IV. Gene to Gene interaction analyses for MTHFR 677C]T, MTR 2756A]G and MTRR 66A]G in the azoospermia group and the OAT group 46
Table 1-V. Sperm count, motility, vitality, morphology according to genotype of each folate metabolism relate genes in the OAT group (N=186). 47
Table 1-VI. The serum homocysteine and folate levels of selected 46 patients depending on the genotypes of folate metabolism enzymes. 48
Table 1-VII. Sperm count, motility, vitality, morphology according to genotype of each folate metabolism relate genes, smoking and alcohol drinking in the OAT group (N=114). 49
Table 2-I. Distribution of H2BFWT -14C/T, -9C/T and +368A/G genotypes in infertile and fertile males 67
Figure 1-1. Folate metabolism with three enzymes. Three enzymes of folate methabolism play an essential role in both DNA synthesis and methylation processes. MTR catalyzes 24
Figure 1-2. Representative diagrams of pyrosequencing data to analyze the four SNP. At each diagram, the X axis represents the sequence of near target SNP site of 34
Figure 1-3. The RFLP results from the male controls and the infertile patients. PCR-RFLP analysis was carried out to confirm the pyrosequencing method. The RFLP 36
Figure 1-4. The frequency of the MTR 2756A]G genotypes according to the MTHFR 677C]T genotypes for the control and the azoospermic group. When the MTHFR 41
Figure 2-1. DNA and protein sequence of H2BFWT. Open quadrangles were shown the location of SNPs and mutation. Two SNPs, -14C/T and -9C/T locate in 5 untranslation 65
Figure 2-2. DNA sequencing results of the new single nucleotide polymorphisms identified in this study. Square indicated the positions (-14 region) of mucleotide changes. 66
Figure 2-3. Reporter assay of human H2BFWT gene. A. Sequences of H2BFWT reporters including 5UTR. The bold letters were at -9 position. B. 293T cells were transfected with 69