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Single-cell expression analysis is an effective tool for studying the dynamics of cell population profiles. However, the majority of statistical methods are applied to individual profiles and the methods for comparing multiple profiles simultaneously are limited. In this study, we propose a nonparametric statistical method, called Decomposition into Extended Exponential Family (DEEF), that embeds a set of single-cell expression profiles of several markers into a low-dimensional space and identifies the principal distributions that describe their heterogeneity. We demonstrate that DEEF can appropriately decompose and embed sets of theoretical probability distributions. We then apply DEEF to a cytometry dataset to examine the effects of epidermal growth factor stimulation on an adult human mammary gland. It is shown that DEEF can describe the complex dynamics of cell population profiles using two parameters and visualize them as a trajectory. The two parameters identified the principal patterns of the cell population profile without prior biological assumptions. As a further application, we perform a dimensionality reduction and a time series reconstruction. DEEF can reconstruct the distributions based on the top coordinates, which enables the creation of an artificial dataset based on an actual single-cell expression dataset. Using the coordinate system assigned by DEEF, it is possible to analyze the relationship between the attributes of the distribution sample and the features or shape of the distribution using conventional data mining methods.

The genetic factors affecting the natural history of nonalcoholic fatty liver disease (NAFLD), including the development of nonalcoholic steatohepatitis (NASH) and NASH-derived hepatocellular carcinoma (NASH-HCC), are still unknown. In the current study, we sought to identify genetic factors related to the development of NAFLD, NASH, and NASH-HCC, and to establish risk-estimation models for them. For these purposes, 936 histologically proven NAFLD patients were recruited, and genome-wide association (GWA) studies were conducted for 902, including 476 NASH and 58 NASH-HCC patients, against 7,672 general-population controls. Risk estimations for NAFLD and NASH were then performed using the SNPs identified as having significant associations in the GWA studies. We found that rs2896019 in PNPLA3 [p = 2.3x10-31, OR (95%CI) = 1.85 (1.67-2.05)], rs1260326 in GCKR [p = 9.6x10-10, OR (95%CI) = 1.38(1.25-1.53)], and rs4808199 in GATAD2A [p = 2.3x10-8, OR (95%CI) = 1.37 (1.23-1.53)] were significantly associated with NAFLD. Notably, the number of risk alleles in PNPLA3 and GATAD2A was much higher in Matteoni type 4 (NASH) patients than in type 1, type 2, and type 3 NAFLD patients. In addition, we newly identified rs17007417 in DYSF [p = 5.2x10-7, OR (95%CI) = 2.74 (1.84-4.06)] as a SNP associated with NASH-HCC. Rs641738 in TMC4, which showed association with NAFLD in patients of European descent, was not replicated in our study (p = 0.73), although the complicated LD pattern in the region suggests the necessity for further investigation. The genetic variants of PNPLA3, GCKR, and GATAD2A were then used to estimate the risk for NAFLD. The obtained Polygenic Risk Scores showed that the risk for NAFLD increased with the accumulation of risk alleles [AUC (95%CI) = 0.65 (0.63-0.67)].We demonstrated that NASH is genetically and clinically different from the other NAFLD subgroups. We also established risk-estimation models for NAFLD and NASH using multiple genetic markers. These models can be used to improve the accuracy of NAFLD diagnosis and to guide treatment decisions for patients.

Non-restorative sleep (NRS) was suggested to be associated with cardiovascular outcomes. However, causative factors for NRS have not been fully elucidated. This study aimed to clarify factors and their relationships with NRS to better understand the clinical and epidemiological implications of NRS and to develop a score that can objectively evaluate NRS status.Study subjects consisted of 9,788 community residents (age 53.6 ± 13.4 y). Subjective NRS as well as possible clinical and lifestyle factors for NRS were investigated by questionnaires. Other clinical parameters were obtained from personal records of information obtained at the baseline examination.A total of 3,261 participants complained of NRS. Factors independently associated with subjective NRS were younger age (odds ratio = 1.43), use of a hypnotic drug (2.04), irregular sleep schedule (2.02), short sleep duration (

We developed a new approach for pairwise kinship analysis in forensic genetics based on chromosomal sharing between two individuals. Here, we defined "index of chromosome sharing" (ICS) calculated using 174,254 single nucleotide polymorphism (SNP) loci typed by SNP microarray and genetic length of the shared segments from the genotypes of two individuals. To investigate the expected ICS distributions from first- to fifth-degree relatives and unrelated pairs, we used computationally generated genotypes to consider the effect of linkage disequilibrium and recombination. The distributions were used for probabilistic evaluation of the pairwise kinship analysis, such as likelihood ratio (LR) or posterior probability, without allele frequencies and haplotype frequencies. Using our method, all actual sample pairs from volunteers showed significantly high LR values (i.e., ≥ 108); therefore, we can distinguish distant relationships (up to the fifth-degree) from unrelated pairs based on LR. Moreover, we can determine accurate degrees of kinship in up to third-degree relationships with a probability of > 80% using the criterion of posterior probability ≥ 0.90, even if the kinship of the pair is totally unpredictable. This approach greatly improves pairwise kinship analysis of distant relationships, specifically in cases involving identification of disaster victims or missing persons.

Association of knee and low back pain with sleep disturbance is poorly understood. We aimed to clarify the independent and combined effects of these orthopedic symptoms on sleep in a large-scale general population.Cross-sectional data about sleep and knee/low back pain were collected for 9,611 community residents (53±14 years old) by a structured questionnaire. Sleep duration less than 6 h/d was defined as short sleep. Sleep quality and the presence of knee and low back pain were evaluated by dichotomous questions. Subjects who complained about knee or low back pains were graded by tertiles of a numerical response scale (NRS) score and a Roland-Morris disability questionnaire (RDQ) score respectively. Multivariate regression analyses were performed to determine the correlates of short sleep duration and poor sleep quality.Frequency of participants who complained of the orthopedic symptoms was as follows; knee pain, 29.0%; low back pain, 42.0% and both knee and low back pain 17.6%. Both knee and low back pain were significantly and independently associated with short sleep duration (knee pain: odds ratio (OR) = 1.19, p

Profiles of sequence variants that influence gene transcription are very important for understanding mechanisms that affect phenotypic variation and disease susceptibility. Using genotypes at 1.4 million SNPs and a comprehensive transcriptional profile of 15,454 coding genes and 6,113 lincRNA genes obtained from peripheral blood cells of 298 Japanese individuals, we mapped expression quantitative trait loci (eQTLs). We identified 3,804 cis-eQTLs (within 500 kb from target genes) and 165 trans-eQTLs (>500 kb away or on different chromosomes). Cis-eQTLs were often located in transcribed or adjacent regions of genes; among these regions, 5' untranslated regions and 5' flanking regions had the largest effects. Epigenetic evidence for regulatory potential accumulated in public databases explained the magnitude of the effects of our eQTLs. Cis-eQTLs were often located near the respective target genes, if not within genes. Large effect sizes were observed with eQTLs near target genes, and effect sizes were obviously attenuated as the eQTL distance from the gene increased. Using a very stringent significance threshold, we identified 165 large-effect trans-eQTLs. We used our eQTL map to assess 8,069 disease-associated SNPs identified in 1,436 genome-wide association studies (GWAS). We identified genes that might be truly causative, but GWAS might have failed to identify for 148 out of the GWAS-identified SNPs; for example, TUFM (P = 3.3E-48) was identified for inflammatory bowel disease (early onset); ZFP90 (P = 4.4E-34) for ulcerative colitis; and IDUA (P = 2.2E-11) for Parkinson's disease. We identified four genes (P

Podocyte injury is the first step in the progression of glomerulosclerosis. Previous studies have demonstrated the beneficial effect of bone morphogenetic protein 7 (Bmp7) in podocyte injury and the existence of native Bmp signaling in podocytes. Local activity of Bmp7 is controlled by cell-type specific Bmp antagonists, which inhibit the binding of Bmp7 to its receptors. Here we show that the product of Twisted gastrulation (Twsg1), a Bmp antagonist, is the central negative regulator of Bmp function in podocytes and that Twsg1 null mice are resistant to podocyte injury. Twsg1 was the most abundant Bmp antagonist in murine cultured podocytes. The administration of Bmp induced podocyte differentiation through Smad signaling, whereas the simultaneous administration of Twsg1 antagonized the effect. The administration of Bmp also inhibited podocyte proliferation, whereas simultaneous administration of Twsg1 antagonized the effect. Twsg1 was expressed in the glomerular parietal cells (PECs) and distal nephron of the healthy kidney, and additionally in damaged glomerular cells in a murine model of podocyte injury. Twsg1 null mice exhibited milder hypoalbuminemia and hyperlipidemia, and milder histological changes while maintaining the expression of podocyte markers during podocyte injury model. Taken together, our results show that Twsg1 plays a critical role in the modulation of protective action of Bmp7 on podocytes, and that inhibition of Twsg1 is a promising means of development of novel treatment for podocyte injury.

OBJECTIVE: To identify similarities and differences in the clinical features of adult Japanese patients with individual anti-aminoacyl-tRNA synthetase antibodies (anti-ARS Abs). METHODS: This was a retrospective analysis of 166 adult Japanese patients with anti-ARS Abs detected by immunoprecipitation assays. These patients had visited Kanazawa University Hospital or collaborating medical centers from 2003 to 2009. RESULTS: Anti-ARS Ab specificity included anti-Jo-1 (36%), anti-EJ (23%), anti-PL-7 (18%), anti-PL-12 (11%), anti-KS (8%), and anti-OJ (5%). These anti-ARS Abs were mutually exclusive, except for one serum Ab that had both anti-PL-7 and PL-12 reactivity. Myositis was closely associated with anti-Jo-1, anti-EJ, and anti-PL-7, while interstitial lung disease (ILD) was correlated with all 6 anti-ARS Abs. Dermatomyositis (DM)-specific skin manifestations (heliotrope rash and Gottron's sign) were frequently observed in patients with anti-Jo-1, anti-EJ, anti-PL-7, and anti-PL-12. Therefore, most clinical diagnoses were polymyositis or DM for anti-Jo-1, anti-EJ, and anti-PL-7; clinically amyopathic DM or ILD for anti-PL-12; and ILD for anti-KS and anti-OJ. Patients with anti-Jo-1, anti-EJ, and anti-PL-7 developed myositis later if they had ILD alone at the time of disease onset, and most patients with anti-ARS Abs eventually developed ILD if they did not have ILD at disease onset. CONCLUSION: Patients with anti-ARS Abs are relatively homogeneous. However, the distribution and timing of myositis, ILD, and rashes differ among patients with individual anti-ARS Abs. Thus, identification of individual anti-ARS Abs is beneficial to define this rather homogeneous subset and to predict clinical outcomes within the "anti-synthetase syndrome."

Rheumatoid arthritis (RA) is a joint-destructive autoimmune disease. Three composite indices evaluating the same 28 joints are commonly used for the evaluation of RA activity. However, the relationship between, and the frequency of, the joint involvements are still not fully understood. Here, we obtained and analyzed 17,311 assessments for 28 joints in 1,314 patients with RA from 2005 to 2011 from electronic clinical chart templates stored in the KURAMA (Kyoto University Rheumatoid Arthritis Management Alliance) database. Affected rates for swelling and tenderness were assessed for each of the 28 joints and compared between two different sets of RA patients. Correlations of joint symptoms were analyzed for swellings and tenderness using kappa coefficient and eigen vectors by principal component analysis. As a result, we found that joint affected rates greatly varied from joint to joint both for tenderness and swelling for the two sets. Right wrist joint is the most affected joint of the 28 joints. Tenderness and swellings are well correlated in the same joints except for the shoulder joints. Patients with RA tended to demonstrate right-dominant joint involvement and joint destruction. We also found that RA synovitis could be classified into three categories of joints in the correlation analyses: large joints with wrist joints, PIP joints, and MCP joints. Clustering analysis based on distribution of synovitis revealed that patients with RA could be classified into six subgroups. We confirmed the symmetric joint involvement in RA. Our results suggested that RA synovitis can be classified into subgroups and that several different mechanisms may underlie the pathophysiology in RA synovitis.

Rheumatoid arthritis is an autoimmune disease with a complex etiology, leading to inflammation of synovial tissue and joint destruction. Through a genome-wide association study (GWAS) and two replication studies in the Japanese population (7,907 cases and 35,362 controls), we identified two gene loci associated with rheumatoid arthritis susceptibility (NFKBIE at 6p21.1, rs2233434, odds ratio (OR) = 1.20, P = 1.3 × 10(-15); RTKN2 at 10q21.2, rs3125734, OR = 1.20, P = 4.6 × 10(-9)). In addition to two functional non-synonymous SNPs in NFKBIE, we identified candidate causal SNPs with regulatory potential in NFKBIE and RTKN2 gene regions by integrating in silico analysis using public genome databases and subsequent in vitro analysis. Both of these genes are known to regulate the NF-κB pathway, and the risk alleles of the genes were implicated in the enhancement of NF-κB activity in our analyses. These results suggest that the NF-κB pathway plays a role in pathogenesis and would be a rational target for treatment of rheumatoid arthritis.

Systemic lupus erythematosus (SLE) is an autoimmune disease that causes multiple organ damage. Although recent genome-wide association studies (GWAS) have contributed to discovery of SLE susceptibility genes, few studies has been performed in Asian populations. Here, we report a GWAS for SLE examining 891 SLE cases and 3,384 controls and multi-stage replication studies examining 1,387 SLE cases and 28,564 controls in Japanese subjects. Considering that expression quantitative trait loci (eQTLs) have been implicated in genetic risks for autoimmune diseases, we integrated an eQTL study into the results of the GWAS. We observed enrichments of cis-eQTL positive loci among the known SLE susceptibility loci (30.8%) compared to the genome-wide SNPs (6.9%). In addition, we identified a novel association of a variant in the AF4/FMR2 family, member 1 (AFF1) gene at 4q21 with SLE susceptibility (rs340630; P = 8.3×10(-9), odds ratio = 1.21). The risk A allele of rs340630 demonstrated a cis-eQTL effect on the AFF1 transcript with enhanced expression levels (P

Nonalcoholic fatty liver disease (NAFLD) includes a broad range of liver pathologies from simple steatosis to cirrhosis and fibrosis, in which a subtype accompanying hepatocyte degeneration and fibrosis is classified as nonalcoholic steatohepatitis (NASH). NASH accounts for approximately 10-30% of NAFLD and causes a higher frequency of liver-related death, and its progression of NASH has been considered to be complex involving multiple genetic factors interacting with the environment and lifestyle.To identify genetic factors related to NAFLD in the Japanese, we performed a genome-wide association study recruiting 529 histologically diagnosed NAFLD patients and 932 population controls. A significant association was observed for a cluster of SNPs in PNPLA3 on chromosome 22q13 with the strongest p-value of 1.4 × 10(-10) (OR = 1.66, 95%CI: 1.43-1.94) for rs738409. Rs738409 also showed the strongest association (p = 3.6 × 10(-6)) with the histological classifications proposed by Matteoni and colleagues based on the degree of inflammation, ballooning degeneration, fibrosis and Mallory-Denk body. In addition, there were marked differences in rs738409 genotype distributions between type4 subgroup corresponding to NASH and the other three subgroups (p = 4.8 × 10(-6), OR = 1.96, 95%CI: 1.47-2.62). Moreover, a subgroup analysis of NAFLD patients against controls showed a significant association of rs738409 with type4 (p = 1.7 × 10(-16), OR = 2.18, 95%CI: 1.81-2.63) whereas no association was obtained for type1 to type3 (p = 0.41). Rs738409 also showed strong associations with three clinical traits related to the prognosis of NAFLD, namely, levels of hyaluronic acid (p = 4.6 × 10(-4)), HbA1c (p = 0.0011) and iron deposition in the liver (p = 5.6 × 10(-4)).With these results we clearly demonstrated that Matteoni type4 NAFLD is both a genetically and clinically different subset from the other spectrums of the disease and that the PNPLA3 gene is strongly associated with the progression of NASH in Japanese population.

HLA-DRB1, especially the shared epitope (SE), is strongly associated with rheumatoid arthritis (RA). However, recent studies have shown that SE is at most weakly associated with RA without anti-citrullinated peptide/protein antibody (ACPA). We have recently reported that ACPA-negative RA is associated with specific HLA-DRB1 alleles and diplotypes. Here, we attempted to detect genetically different subsets of ACPA-negative RA by classifying ACPA-negative RA patients into two groups based on their positivity for rheumatoid factor (RF). HLA-DRB1 genotyping data for totally 954 ACPA-negative RA patients and 2,008 healthy individuals in two independent sets were used. HLA-DRB1 allele and diplotype frequencies were compared among the ACPA-negative RF-positive RA patients, ACPA-negative RF-negative RA patients, and controls in each set. Combined results were also analyzed. A similar analysis was performed in 685 ACPA-positive RA patients classified according to their RF positivity. As a result, HLA-DRB1*04:05 and *09:01 showed strong associations with ACPA-negative RF-positive RA in the combined analysis (p = 8.8×10(-6) and 0.0011, OR: 1.57 (1.28-1.91) and 1.37 (1.13-1.65), respectively). We also found that HLA-DR14 and the HLA-DR8 homozygote were associated with ACPA-negative RF-negative RA (p = 0.00022 and 0.00013, OR: 1.52 (1.21-1.89) and 3.08 (1.68-5.64), respectively). These association tendencies were found in each set. On the contrary, we could not detect any significant differences between ACPA-positive RA subsets. As a conclusion, ACPA-negative RA includes two genetically distinct subsets according to RF positivity in Japan, which display different associations with HLA-DRB1. ACPA-negative RF-positive RA is strongly associated with HLA-DRB1*04:05 and *09:01. ACPA-negative RF-negative RA is associated with DR14 and the HLA-DR8 homozygote.

PURPOSE: Recently, a complement component 1 inhibitor (SERPING1) gene polymorphism was identified as a novel risk factor for age-related macular degeneration (AMD) in Caucasians. We aimed to investigate whether variations in SERPING1 are associated with typical AMD or with polypoidal choroidal vasculopathy (PCV) in a Japanese population. METHODS: We performed a case-control study in a group of Japanese patients with typical AMD (n = 401) or PCV (n = 510) and in 2 independent control groups--336 cataract patients without age-related maculopathy and 1,194 healthy Japanese individuals. Differences in the observed genotypic distribution between the case and control groups were tested using chi-square test for trend. Age and gender were adjusted using logistic regression analysis. RESULTS: We targeted rs2511989 as the haplotype-tagging single nucleotide polymorphism (SNP) for the SERPING1 gene, which was reported to be associated with the risk of AMD in Caucasians. Although we compared the genotypic distributions of rs2511989 in typical AMD and PCV patients against 2 independent control groups (cataract patients and healthy Japanese individuals), SERPING1 rs2511989 was not significantly associated with typical AMD (P = 0.932 and 0.513, respectively) or PCV (P = 0.505 and 0.141, respectively). After correction for age and gender differences based on a logistic regression model, the difference in genotypic distributions remained insignificant (P>0.05). Our sample size had a statistical power of more than 90% to detect an association of a risk allele with an odds ratio reported in the original studies for rs2511989 for developing AMD. CONCLUSIONS: In the present study, we could not replicate the reported association between SERPING1 and either neovascular AMD or PCV in a Japanese population; thus, the results suggest that SERPING1 does not play a significant role in the risk of developing AMD or PCV in Japanese.

Rheumatoid arthritis (RA) is a major cause of adult chronic inflammatory arthritis and a typical complex trait. Although several genetic determinants have been identified, they account for only a part of the genetic susceptibility. We conducted a genome-wide association study of RA in Japanese using 225,079 SNPs genotyped in 990 cases and 1,236 controls from two independent collections (658 cases and 934 controls in collection1; 332 cases and 302 controls in collection2), followed by replication studies in two additional collections (874 cases and 855 controls in collection3; 1,264 cases and 948 controls in collection4). SNPs showing p

Myopia is one of the most common ocular disorders worldwide. Pathological myopia, also called high myopia, comprises 1% to 5% of the general population and is one of the leading causes of legal blindness in developed countries. To identify genetic determinants associated with pathological myopia in Japanese, we conducted a genome-wide association study, analyzing 411,777 SNPs with 830 cases and 1,911 general population controls in a two-stage design (297 cases and 934 controls in the first stage and 533 cases and 977 controls in the second stage). We selected 22 SNPs that showed P-values smaller than 10(-4) in the first stage and tested them for association in the second stage. The meta-analysis combining the first and second stages identified an SNP, rs577948, at chromosome 11q24.1, which was associated with the disease (P = 2.22x10(-7) and OR of 1.37 with 95% confidence interval: 1.21-1.54). Two genes, BLID and LOC399959, were identified within a 200-kb DNA encompassing rs577948. RT-PCR analysis demonstrated that both genes were expressed in human retinal tissue. Our results strongly suggest that the region at 11q24.1 is a novel susceptibility locus for pathological myopia in Japanese.