複数遺伝マーカーを用いた日本人における非アルコール性脂肪性肝疾患のリスク予測モデル

京都大学新制・論文博士博士(医学)乙第13398号論医博第2222号新制||医||1051(附属図書館)(主査)教授 妹尾 浩, 教授 中山 健夫, 教授 西浦 博学位規則第4条第2項該当Doctor of Medical ScienceKyoto UniversityDFA

Genome-wide association study of individual differences of human lymphocyte profiles using large-scale cytometry data

Human immune systems are very complex, and the basis for individual differences in immune phenotypes is largely unclear. One reason is that the phenotype of the immune system is so complex that it is very difficult to describe its features and quantify differences between samples. To identify the genetic factors that cause individual differences in whole lymphocyte profiles and their changes after vaccination without having to rely on biological assumptions, we performed a genome-wide association study (GWAS), using cytometry data. Here, we applied computational analysis to the cytometry data of 301 people before receiving an influenza vaccine, and 1, 7, and 90 days after the vaccination to extract the feature statistics of the lymphocyte profiles in a nonparametric and data-driven manner. We analyzed two types of cytometry data: measurements of six markers for B cell classification and seven markers for T cell classification. The coordinate values calculated by this method can be treated as feature statistics of the lymphocyte profile. Next, we examined the genetic basis of individual differences in human immune phenotypes with a GWAS for the feature statistics, and we newly identified seven significant and 36 suggestive single-nucleotide polymorphisms associated with the individual differences in lymphocyte profiles and their change after vaccination. This study provides a new workflow for performing combined analyses of cytometry data and other types of genomics data

Diabetic condition induces hypertrophy and vacuolization in glomerular parietal epithelial cells

Diabetic nephropathy (DN) is accompanied by characteristic changes in the glomerulus, but little is known about the effect of diabetes on parietal epithelial cells (PECs). In this study, a descriptive analysis of PECs was undertaken in diabetic db/db mice and in diabetic patients. PEC hypertrophy was significantly more prominent in diabetic mice than in nondiabetic mice, and this was evident even at the early stage. Additionally, the number of vacuoles in PECs was markedly increased in diabetic mice, suggesting the presence of cellular injury in PECs in DN. Although rare, binuclear cells were observed in mice with early diabetes. In cultured PECs, a high glucose condition, compared with normal glucose condition, induced cellular hypertrophy and apoptosis. Flow cytometry showed that some PECs in the G0 phase reentered the cell cycle but got arrested in the S phase. Finally, in human diabetic subjects, hypertrophy and vacuolization were observed in the PECs. Our data showed that PECs undergo substantial changes in DN and may participate in rearrangement for differentiation into podocytes

Tetraspanin CD63 Promotes Targetion and Lysosomal Proteolysis of Membrance-Type 1 Matrix Metalloproteinase

金沢大学がん研究所Membrane-type 1 matrix metalloproteinase (MT1-MMP) is known to be internalized from cell surface, however, the fate of internalized MT1-MMP is still unknown. Here we demonstrate that at least a part of internalized MT1-MMP is targeted for lysosomal proteolysis. Treatment with an inhibitor of lysosomal proteinases chloroquine suppressed degradation of internalized MT1-MMP and induced accumulation of MT1-MMP in CD63-positive lysosomes. Ectopic expression of CD63 accelerated degradation of MT1-MMP, which was blocked by chloroquine. MT1-MMP, and CD63 were shown to form a complex through hemopexin-like domain of MT1-MMP and N-terminal region of CD63, and thus accelerated degradation of MT1-MMP was not observed with mutants lacking these domains. CD63 mutant lacking lysosomal targeting motif was unable to promote MT1-MMP degradation. These results suggest that CD63 regulates MT1-MMP by targeting to lysosomes

Nonimmunoglobulin target loci of activation-induced cytidine deaminase (AID) share unique features with immunoglobulin genes.

Activation-induced cytidine deaminase (AID) is required for both somatic hypermutation and class-switch recombination in activated B cells. AID is also known to target nonimmunoglobulin genes and introduce mutations or chromosomal translocations, eventually causing tumors. To identify as-yet-unknown AID targets, we screened early AID-induced DNA breaks by using two independent genome-wide approaches. Along with known AID targets, this screen identified a set of unique genes (SNHG3, MALAT1, BCL7A, and CUX1) and confirmed that these loci accumulated mutations as frequently as Ig locus after AID activation. Moreover, these genes share three important characteristics with the Ig gene: translocations in tumors, repetitive sequences, and the epigenetic modification of chromatin by H3K4 trimethylation in the vicinity of cleavage sites

Population-genetic nature of copy number variations in the human genome

Copy number variations (CNVs) are universal genetic variations, and their association with disease has been increasingly recognized. We designed high-density microarrays for CNVs, and detected 3000–4000 CNVs (4–6% of the genomic sequence) per population that included CNVs previously missed because of smaller sizes and residing in segmental duplications. The patterns of CNVs across individuals were surprisingly simple at the kilo-base scale, suggesting the applicability of a simple genetic analysis for these genetic loci. We utilized the probabilistic theory to determine integer copy numbers of CNVs and employed a recently developed phasing tool to estimate the population frequencies of integer copy number alleles and CNV–SNP haplotypes. The results showed a tendency toward a lower frequency of CNV alleles and that most of our CNVs were explained only by zero-, one- and two-copy alleles. Using the estimated population frequencies, we found several CNV regions with exceptionally high population differentiation. Investigation of CNV–SNP linkage disequilibrium (LD) for 500–900 bi- and multi-allelic CNVs per population revealed that previous conflicting reports on bi-allelic LD were unexpectedly consistent and explained by an LD increase correlated with deletion-allele frequencies. Typically, the bi-allelic LD was lower than SNP–SNP LD, whereas the multi-allelic LD was somewhat stronger than the bi-allelic LD. After further investigation of tag SNPs for CNVs, we conclude that the customary tagging strategy for disease association studies can be applicable for common deletion CNVs, but direct interrogation is needed for other types of CNVs

Increased aortic wave reflection and smaller pulse pressure amplification in smokers and passive smokers confirmed by urinary cotinine levels: The Nagahama Study.

[Background]Central blood pressure (cSBP) is suggested to be a better predictor of cardiovascular risk than brachial BP. Although brachial BP levels among smokers have been reported to be the same or somewhat lower than those in nonsmokers, it is suggested that smoking might have a substantial impact on cSBP. [Methods]We conducted a cross-sectional study to clarify the association of smoking habit with arterial tone and cSBP in a general population of 8557 participants using urinary cotinine levels as an objective marker of smoking intensity. Absolute pressure of the late systolic peak (SBP2) was obtained by calibrating the radial waveform with brachial systolic BP (bSBP) and considered to be the cSBP. [Results]Confounding factor-adjusted mean pulse pressure amplification (PPa = bSBP − cSBP) was significantly smaller in habitual smokers (current, 9.3 ± 0.15; past, 10.2 ± 0.13; never, 10.6 ± 0.10 mm Hg; p < 0.001). Further, among smokers, PPa was linearly decreased with increasing urinary cotinine quartile (Q1, 10.9 ± 0.38; Q2, 10.9 ± 0.39; Q3, 10.4 ± 0.39; Q4, 9.7 ± 0.41 mm Hg; p = 0.020). Multiple linear regression analysis identified both smoking habit (p = 0.003) and urinary cotinine levels (p = 0.008) as independent determinants of PPa. Urinary cotinine was also detected in a small fraction of never smokers (1.8%). These passive smokers showed a smaller PPa (passive smoker, 9.4 ± 0.4; never smoker, 10.4 ± 0.12 mm Hg, p = 0.020) but not bSBP (122.7 ± 0.6, 123.1 ± 0.2 mm Hg, p = 0.474). [Conclusions]Not only habitual smoking but also passive smoking had harmful effects on AIx and central BP. Our results strongly emphasize the importance of avoiding passive smoking to the prevention of cardiovascular risks of which the subject is likely unaware

IL-6, IL-8, and IL-10 Are Associated with Hyperferritinemia in Rapidly Progressive Interstitial Lung Disease with Polymyositis/Dermatomyositis

Objective. Hyperferritinemia is frequently accompanied by rapidly progressive (RP) interstitial lung disease (ILD) with polymyositis (PM)/dermatomyositis (DM). To clarify the mechanism of RP-ILD with hyperferritinemia, we investigated the associations between serum ferritin levels and various cytokines in patients with PM/DM. Methods. This retrospective study included 38 patients admitted to our hospital with PM/DM. Levels of serum ferritin and cytokines (IL-1 , IL-2, IL-4, IL-6, IL-8, IL-10, IL-12, IL-13, IL-17, IL-18, TNF-, IFN-, IFN-, and IP-10) were measured. Disease activity was evaluated using the tool proposed by the International Myositis Assessment and Clinical Studies Group. We analyzed the associations between disease activity and levels of serum ferritin and cytokines. Results. The levels of serum ferritin, IL-8, IL-10, IL-18, and TNF-, were significantly correlated with disease activity. In a multivariate analysis, IL-6 ( = 3.6, = 0.0010), IL-8 ( = 4.8, &lt; 0.0001), and IL-10 ( = 5.7, &lt; 0.0001) significantly contributed to serum ferritin levels. The levels of serum ferritin, IL-6, IL-8, and IL-10, were higher in the RP-ILD subset than in the non-ILD subset or the chronic ILD subset. Conclusion. IL-6, IL-8, and IL-10 are significant contributors to hyperferritinemia in PM/DM. The regulation of these cytokines might offer a possible treatment strategy for RP-ILD with PM/DM