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Takayasu arteritis (TAK) is an autoimmune systemic vasculitis of unknown etiology. Although previous studies have revealed that HLA-B*52:01 has an effect on TAK susceptibility, no other genetic determinants have been established so far. Here, we performed genome scanning of 167 TAK cases and 663 healthy controls via Illumina Infinium Human Exome BeadChip arrays, followed by a replication study consisting of 212 TAK cases and 1,322 controls. As a result, we found that the IL12B region on chromosome 5 (rs6871626, overall p = 1.7 × 10(-13), OR = 1.75, 95% CI 1.42-2.16) and the MLX region on chromosome 17 (rs665268, overall p = 5.2 × 10(-7), OR = 1.50, 95% CI 1.28-1.76) as well as the HLA-B region (rs9263739, a proxy of HLA-B*52:01, overall p = 2.8 × 10(-21), OR = 2.44, 95% CI 2.03-2.93) exhibited significant associations. A significant synergistic effect of rs6871626 and rs9263739 was found with a relative excess risk of 3.45, attributable proportion of 0.58, and synergy index of 3.24 (p ≤ 0.00028) in addition to a suggestive synergistic effect between rs665268 and rs926379 (p ≤ 0.027). We also found that rs6871626 showed a significant association with clinical manifestations of TAK, including increased risk and severity of aortic regurgitation, a representative severe complication of TAK. Detection of these susceptibility loci will provide new insights to the basic mechanisms of TAK pathogenesis. Our findings indicate that IL12B plays a fundamental role on the pathophysiology of TAK in combination with HLA-B(∗)52:01 and that common autoimmune mechanisms underlie the pathology of TAK and other autoimmune disorders such as psoriasis and inflammatory bowel diseases in which IL12B is involved as a genetic predisposing factor.

The maximum dosage of methotrexate (MTX) for treatment of rheumatoid arthritis (RA) formally approved in Japan is 8 mg/week. We intended to examine the efficacy and safety of MTX at dosages over 8 mg/week in Japanese rheumatoid arthritis patients using the large Institute of Rheumatology, Rheumatoid Arthritis (IORRA) cohort database. Among 9,122 patients registered in the IORRA database from the October 2000 survey to the October 2007 survey, 5,201 patients who had been treated with MTX were selected. We attempted to overcome the drawbacks innate to nonrandomized studies by using longitudinal analyses and multifactorial logistic regression analyses. Cross-sectional analysis of data obtained from the October 2007 survey indicated that dosages of MTX higher than 8 mg/week were used in 27.5% of patients treated with MTX. Longitudinal analyses based on data from three consecutive phases showed that final Disease Activity Score-28 (DAS28) values were significantly lower [n = 260, mean difference 0.563, 95% confidence interval (CI) 0.438-0.688, P < 2.2 × 10(-22), two-sided paired t test] than initial values when MTX was increased from 8 mg/week or lower to over 8 mg/week. In addition, longitudinal analyses based on data from two consecutive phases indicated decreases in DAS28 values of 0.26 ± 1.04 (n = 690, P = 6.78 × 10(-11), two-sided paired t test) when MTX dosages were increased from 8 mg/week or lower to over 8 mg/week, compared with decreases of 0.07 ± 0.89 (n = 2,125, P = 0.000307) when the dosage was maintained at 8 mg/week. The decreases in DAS28 values were significantly larger in the former than the latter (P = 2.27 × 10(-6), two-sided unpaired t test). Concerning safety of MTX at dosages over 8 mg/week, we performed logistic regression analysis in which the objective variable was the existence or nonexistence of self-reported side-effects and the explanatory variable was the MTX dosage in the former phase, with adjustments made for age, sex, body mass index (BMI), steroid administration, folic acid administration, concomitant pulmonary diseases, and renal dysfunction. The results indicated that MTX dosages over 8 mg/week did not have any association with either severe or severe + moderate side-effects. These data regarding both efficacy and safety of MTX at dosages over 8 mg/week in Japanese RA patients would provide the basis for use of the drug at dosages currently not formally approved by the Japanese government.

2,8-Dihydroxyadenine urolithiasis associated with partial deficiencies of adenine phosphoribosyltransferase (APRT) has been found only among Japanese families. All Caucasian patients with the same lithiasis are completely deficient in this enzyme. Partially purified APRT from one of the Japanese families with the lithiasis associated with a partial deficiency of APRT had a reduced affinity for 5-phosphoribosyl-1-pyrophosphate (PRPP). In the present investigations, we have shown that this characteristic is common in mutant enzymes from all the four separate Japanese urolithiasis families associated with partial APRT deficiencies so far tested. The mutant enzymes also had several other characteristics in common including increased resistance to heat in the absence of PRPP and reduced sensitivity to the stabilizing effect of PRPP. These data suggest that these families have a common mutant allele (APRT * J) at the APRT gene locus.

若齢ラットに自然発生した横紋筋肉腫を病理組織学的に検索した. 腫瘍は筋芽細胞に類似した長紡錘形の細胞, 横紋を示す多核で大型の細胞および細胞質の乏しい小型の細胞から構成されていた. 電顕的に小型の細胞には細線維が, 長錘形および多核で大型の細胞にはアクチンおよびミオシン線維の規則的配列とZ膜様の構造が認められた. これら腫瘍細胞の構造は, それぞれ分化程度の異なる筋芽細胞の特徴を示すものと考えられた.

A screening method using high-performance liquid chromatography (HPLC) for the simultaneous detection of deficiencies of adenine phosphoribosyltransferase (APRT) and hypoxanthine phosphoribosyltransferase (HPRT) activities in human erythrocytes is described. Both enzyme reactions of APRT and HPRT in lysates treated with a charcoal-dextran were simultaneously carried out in the same reaction tube and the enzyme activities were determined by measuring the increases in absorbance at 260 nm of adenosine and inosine converted from adenosine-5'-monophosphate and inosine-5'-monophosphate with alkaline phosphatase. Adenosine and inosine were separated from adenine and hypoxanthine by a reversed-phase column. The method could detect 1% of normal APRT activity and 0.3% of normal HPRT activity. The within-run coefficients of variation for APRT and HPRT activities were 3.2 and 3.4%, respectively.

痛風,痛風腎を合併したBartter症候群の1例を経験した,症例は17才の女性で,手指関節の変形腫脹にて入院,高尿酸血症・低K性アルカローシス,アンジオテンシン-IIに対する昇圧反応の低下が認められた,腎生検では,傍糸球体装置の過形成および遠位尿細管内尿酸結晶が確認された,本症候群では高尿酸血症,痛風の合併頻度が高いとされているが,本邦での報告例は少なく,また組織学的に痛風腎を証明しえた症例の報告はない.

2-Chloroadenosine (CIAdo) has been reported not to be a substrate for either adenosine deaminase(1) or adenosine kinase(2) and for this reason, has been widely used as a ligand for the adenosine receptor assay(3,4). However, since it is quite toxic to human growing cells, we suspected it might be, as most other toxic nucleosides, phosphorylated by some mammalian enzymes. We cultured a human B cell line with this adenosine analog and found that human cells are able to phosphorylate CIAdo, and, in the course of its phosphorylation, this compound induces the overproduction of hypoxanthine in the culture medium.

2-Chloroadenosine but not 6-methylmercaptopurine riboside was phosphorylated by adenosine kinase negative human B cells, and the adenosine negative cells were resistant to 6-methylmercaptopurine riboside but not to 2-chloroadenosine. Phosphorylation of 6-methylmercaptopurine was totally dependent on adenosine kinase, but 2-chloroadenosine seemed to be phosphorylated by other enzyme(s) as well. The cytotoxicity of both of these analogs depends on the phosphorylation.

Since a number of patients with severe combined immunodeficiency syndrome have been found to be deficient in adenosine deaminase (ADA) or purine nucleotide phosphorylase (PNP)(1, 2), purine metabolism has been postulated to be deeply involved in human immune system. A series of evidences has shown that abnormally accumulated purine deoxynucleosides in these patients have selective toxicity on T or B lymphocytes. Another experiments demonstrated critical roles of purine enzymes on lymphocyte maturation or differentiation. (3, 4, 5).

Histopathological observations were made on pleomorphic liposarcoma found in the omentum and mesentery of a 68-week-old rat. The tumor consisted of two types of tumor cells; round cells that had abundant cytoplasm with a clear large nucleus and spindle-shaped, slender cells with dark oval nuclei. Some round cells contained features suggesting ongoing lipid synthesis in the dilatated cistern of the rough endoplasmic reticulum. Both types of cells had various sizes of lipid droplets with no limited membrane.

Xanthinuria is a rare hereditary disease characterized by a deficiency of xanthine oxidase (E C 1,2,3,2), which metabolized hypoxanthine and xanthine to uric acid. In Japan, two cases of xanthinuria have been reported. However, during the past two years, we have investigated four patients with hypouricemia, hypouricosuria and xanthinuria. In almost all of these patients, hypouricemia (less than 1.0 mg/dl) was the most important index information to make a definite diagnosis of this disorder. After clinical profiles of these four patients were described, an easy method of detection of xanthine oxidase activity from the duodenal mucosa was developed by the authors. Urinary oxypurine was also analyzed by high-pressure liquid chromatography, and the results are reported here.

Urolithiasis composed of 2,8-dihydroxyadenine is not only formed in a complete defect of adenine phosphoribosyltransferase, but also in a partial deficiency of this enzyme. To our knowledge these cases reported in this paper are the third and fourth cases of dihydroxyadenine urinary calculi in patients with partial deficiency of adenine phosphoribosyltransferase.

Peripheral T cells from 3 Lesch-Nyhan patients, 3 normal subjects, and 3 brothers with hypoxanthine-guanine phosphoribosyltransferase (HGPRT) deficiency but without Lesch-Nyhan syndrome (so-called partial deficiency) have been analyzed. Although these brothers contained HGPRT activities neither in the hemolysates nor in the T cell extracts at levels detectable by the regular radioenzyme assay, the enzyme deficiency had not caused any typical neurological symptoms of the Lesch-Nyhan syndrome. Although the T cells from these brothers were at least 10-fold more resistant to 6-thioguanine than normal T cells, they were more than 30-fold less resistant than the T cells from 3 Lesch-Nyhan patients indicating that there is a clear difference in the severity of the enzyme deficiency between the brothers and the Lesch-Nyhan patients. These data indicate that the long-term T cell culture in the medium containing a purine analog whose toxicity depends on a salvaging enzyme is useful for evaluating the severity of the enzyme deficiency in viable cells.

Background Tofacitinib is an oral Janus kinase inhibitor for the treatment of rheumatoid arthritis. Here, tofacitinib safety and efficacy data from a long-term extension study in Japanese patients are presented. Methods Study A3921041 was a multi-centre, open-label, long-term extension study that included Japanese patients who had participated in a prior Phase 2 or Phase 3 study of tofacitinib as monotherapy or with background methotrexate. Patients received tofacitinib 5 mg twice daily (BID) or tofacitinib 10 mg BID. Dose adjustment of tofacitinib during treatment period, and concomitant usage of disease-modifying antirheumatic drugs including methotrexate after week 12 were permitted. Primary endpoints were adverse events, laboratory parameters and vital signs. Secondary efficacy endpoints included American College of Rheumatology (ACR)20/50/70 response rates, Disease Activity Score (DAS)28-4(erythrocyte sedimentation rate (ESR))

Purine nucleotide degradation during ethanol catabolism, inhibition of renal excretion of urate by lactic acid, and high purine content of certain kinds of beverage are responsible for the elevation of serum uric acid level following alcohol drinking. It is well documented that rapid consumption of ATP produces uric acid via purine nucleotide degradation. Since individuals with ALDH2*1 can catalyze ethanol readily, they consume large amount of ATP, and thus produce more hypoxanthine than those with ALDH2*2. Thus, daily drinker and heavy drinker tend to readily induce hyperuricemia after alcohol ingestion. A high purine content in beer may contribute to the increasing frequency of hyperuricemia. Restriction of any kind of alcohol beverages, especially beer, is necessary in the medical control of patients with gout or hyperuricemia.

Guideline for the management of hyperuricemia and gout, first edition was published in 2002 from Japanese Society of Gout and Nucleic Acid Metabolism, and has contributed for the improvement in the management of patients by primary physicians. During five years after the publication of this first edition, large amount of evidences has been accumulated, thus it is time to revise the guideline. Japanese Society of Gout and Nucleic Acid Metabolism has organized the committee for the revision of guideline, and the operations have begun. In this revision, we pay attention to the quality in the process of guideline edition, application of AGREE checklist and patients perspective is considered.

Human IgM that express the variable region genes of the VH3 family bind staphylococcal protein A (SPA). We previously reported that the SPA-binding IgM can be divided into two groups based on the differential binding avidities for solid-phase SPA. To study the molecular basis for these differences, we cloned B cells from human blood by EBV transformation. The nucleotide sequences of the expressed Ig heavy chain genes were determined on 20 B cell clones that produce SPA-binding IgM. The germ-line VH3 gene usage in IgM with high avidities for SPA were distinct from the germ-line VH3 genes used in IgM with low avidities for SPA. There was no correlation in the usage of D or JH genes or in the usage of light chains in IgM according to the SPA binding avidity. These results suggest that the differential binding avidities for SPA are at least partly due to specific germ-line VH3 gene usage. An investigation of direct binding of SPA to the synthetic peptides corresponding to the portions of the variable regions of SPA-binding and non-SPA-binding IgM showed that the peptides corresponding to the VH3 family specific framework region 3 sequences had significant SPA binding capacities, while the peptides corresponding to the other subdomains and those corresponding to framework region 3 of the reported VH3 sequences from non-SPA-binding IgM showed little or no binding. It is of interest that the Ig-framework region 3 subdomain corresponds to the fourth hypervariable region, which in the TCR-beta chain has been implicated as a critical site for T cell superantigen binding.

Rheumatoid arthritis (RA) is a complex polygenic disease characterized by progressive joint destruction. Anti-citrullinated peptide antibody (ACPA) is the most specific autoantibody for RA. Genetic polymorphisms in the PADI4 gene, encoding citrullinating enzyme peptidylarginine deiminase 4 (PADI4), have been associated with susceptibility to RA [1,2]. They have also been reported to be associated with radiographic joint destruction in patients with RA [3,4]. We focused on ACPA-negative RA patients to investigate whether a PADI4 polymorphism is associated with joint damage in ACPA-negative patients. DNA samples from 122 Japanese ACPA-negative RA patients were used for the study; 81.1% were female, 51.6% were rheumatoid factor (RF)-positive, and the mean age was 55 years. Sharp/van der Heijde score of the hands at a 5-year disease duration, which represents joint damage, was scored and log-transformed as described elsewhere [4]. Single-nucleotide polymorphism (SNP) rs2240340 was selected and genotyped by using a TaqMan method as described elsewhere [4]. The genetic risk of joint damage associated with rs2240340 was assessed by multiple regression analysis adjusted for HLA-DRB1 shared-epitope alleles and RF that are thought to be associated with joint damage in patients with RA [4]. The PADI4 SNP was significantly associated with radiographic joint destruction in the ACPA-negative RA patients in a recessive model (P = 0.0287) (Table 1 and Figure 1). The overall genotyping success rate was 99.2% and the genotype concordance rate was 100% as assessed by duplicate samples. Although the sample size is one of the major limitations to the study of ACPA-negative patients because of the high positivity of ACPA (up to 90%) in RA populations, we were able to collect DNA samples from 122 ACPA-negative RA patients with radiographic data. Table 1 Association of PADI4 SNP (recessive model) with Sharp/van der Heijde score of hands at 5-year disease duration in ACPA-negative RA patients Figure 1 Boxplots representing the distribution of Sharp/van der Heijde score of hands at 5-year disease duration according to the number of the susceptibility allele of rs2240340 (minor allele, A) in the PADI4 locus. Each box represents the interquartile range ... PADI4 encodes citrullinating enzymes that may play an important role in ACPA formation. It has been shown that PADI4 alleles were associated with the presence of ACPA in patients with RA [1]. In the present study, we have shown that PADI4 polymorphism contributes to joint destruction in ACPA-negative RA patients. A recent study has suggested that PADI4 gene contributes to the development of RA, regardless of ACPA status [3]. We have also reported that the PADI4 risk allele has an impact on joint damage after adjustment for ACPA status [4]. The PADI4 gene is likely to play a role in the disease progression of RA in addition to its role in ACPA formation. The results of this study provide important knowledge of the risks on progressive joint damage in patients with RA.

Although all Caucacian patients with 2,8-dihydroxyadenine urolithiasis completely lack adenine phosphoribosyltransferase (APRT), more than half of Japanese patients with the same lithiasis only partially lack APRT. The degrees of APRT deficiencies in these Japanese patients are broadly the same as healthy heterozygotes for complete APRT deficiencies. We found that cultured T-cells from the Japanese patients with partial APRT deficiencies were, like those from the patients with complete APRT deficiencies but unlike the heterozygotes, more than 100-fold less sensitive to adenine analogs, 6-methylpurine and 2,6-diaminopurine, suggesting that APRT of these Japanese patients is non-functional in viable cells, although it functions normally in test tubes. Therefore, it is not APRT activity in test tubes but whether APRT is functional in viable cells that is important for clinical symptoms (lithiasis). The mutant enzymes from four separate families were found to have reduced affinities to PRPP but not to adenine, and at physiological concentrations of PRPP, normal but not mutant enzymes showed significant activities. The mutant enzymes contained several other altered characteristics in common possibly suggesting that they are coded by a common mutant allele.

Poly(ADP-ribose) polymerase (ADPRP) is highly activated by DNA with strand breaks and transfer poly (ADP-ribose)chains outo nuclear proteins. Poly (ADP-ribosyl)action rapidly consumes cellular NAD and leads cells to die. This reaction has been implied to be involved in the deoxyadenosine toxicity toward resting human lymphocytes, and could be an important mechanism in the toxicity of all DNA-damaging agents. To analyze the characteristics of ADPRP, we have utilized human autoantibodies to ADPRP which we have identified in the sera of rheumatic patients. cDNA clones encoding ADPRP were isolated from human libraries and used in these studies. The results of these studies indicate that ADPRP is an abundant nuclear enzyme. Eukaryotic cells synthesize ADPRP throughout S-phase, and keep the cellular ADPRP level constantly in proportion to the amount of DNA. Interestingly more than 99% of ADPRP is present in cells as inactive form. These data may suggest the importance of ADPRP as a structural protein in chromatin conformation. In addition, these results are consistent with the hypothesis of “suicide mechanism” of ADPRP. Eukaryotic cells have approximately 100-fold more enzyme than that is required for the regular cell growth, and keep it for an emergency event. When cellular DNA is severely damaged, ADPRP is activated and eliminates the cell. This mechanism could be applicable to the toxicity of any DNA damaging agents, including many purine and pyrimidine analogs.

Exogenous addition of methylthioadenosine (MeSAdo) inhibits proliferation of cells which are deficient in MeSAdo phosphorylase. A series of evidences has shown that the target of toxic mechanism of MeSAdo is not totally dependent on polyamine synthesis. Eukaryotic cells contain unique modified amino acid in elongation factor 2 (EF-2), designated as diphthamide. This residue is the specific target of mono(ADP-ribosyl)ation catalyzed by diphtheria toxin (DT) or Pseudomonous toxin. The structure of diphthamide is 2- [ 3-carboxyamido-3-(trimethylammonio)propyl]histidine, and the first reaction to modify histidine involves the transfer of an aminocarboxypropyl group from S-adenosylmethionine. MeSAdo should be the nucleoside product of this reaction. By these reasons, we have analyzed the effect of MeSAdo on the biosynthesis of diphthamide. A mutant cell line H3 which is deficient in MeSAdo phosphorylase and resistant to MeSAdo, has been isolated from murine lymphoma cell line R1.1. for the study. As measured by susceptibility to DT induced ADP-ribosylation, MeSAdo inhibits the formation of diphthamide in a dose dependent manner. In addition, MeSAdo substantially protected H3 cells from the lethal effect of DT. These results suggest that the modulation of diphthamide synthesis can be, at least a part of, the mechanism of MeSAdo toxicity toward eukaryotic cells.

Peripheral T-lymphocyte populations from two Lesch-Nyhan heterozygotes were found to consist of 95% hypoxanthineguanine phosphoribosyltransferase (HGPRT)-positive and 5% negative cells by the culture of the T-cells with 6-thioguanine. The T-cells frome each heterozygote were cultured in the conditions in which either enzyme-positive (azaserine-hypoxanthine medium) or negative (6-thioguanine medium) cells selectively proliferate. After two weeks, each separated T-cell subpopulation was considered to be pure with regard to the enzyme expression. Bewteen the enzyme-positive and negative subpopulations, percentages of OKT3, OKT4 or OKT8-positive T-cells were compared. The fact that only small percentage of peripheral T-cells are HGPRT-negative in the heterozygotes suggests that in vivo differentiation of HGPRT-negative cells is disturbed in the heterozygotes. Our data have shown that both enzyme-positive and negative T-cell subpopulations from each heterozygote possessed equivalent percentages of OKT4, OKT8 and OKT3-positive cells. These results suggest that, in the heterozygotes, growth or differentiation of HGPRT-nnegative lympoid cells of T-cell linkage was disturbed before OKT4 and OKT8-bearing thymocyte subpopulations separate from each other.

2-COROADENOSINE IS PHOSPHORYLATED AND INCREASES THE PRODUCTION OF HYPOXANTHINE IN HUMAN CELLLS: 234

The patients with purine metabolic disorders have been registered at our institute from about 1000 various clinical institutes in overall Japan. This survey was done from 1982 to 1984. Results were summarized in Table 1. One hundred six patients were registered and the patients with disorders of purine salvage enzymes were frequent.

Although carriers of deficient genes for Lesch-Nyhan syndrome are detectable using cultured fibroblasts or hair roots, peripheral blood has been insufficient specimen for the diagnosis. We have established a method using peripheral T-cells for the diagnosis of the heterozygotes. Mononuclear cells from five mothers of Lesch-Nyhan boys were stimulated with phytohemagglutinin and then maintained in the medium containing T-cell growth factor. The growing viable T-cells were then exposed to 6-thioguanine. Most of the cells were killed by the treatment, but after 2-4 days, rapid growth of 6-thioguanine resistant cells was observed. T-cells from Lesch-Nyhan boys were totally resistant to 6-thioguanine and normal T-cells were sensitive. We estimated the percentages of HGPRT negative cells among T-cell growth factor-responsive T-cells from each heterozygote by comparing the growth curves for these cell populations with those for mixed cell populations containing enzyme-negative (Lesch-Nyhan's cells) cells and positive (normal) cells in various ratios. The Lesch-Nyhan mothers were estimated to contain 5-40% enzyme negative T-cells, indicating that mosaicism is present in T-cell populations in heterozygotes and the carrier state for Lesch-Nyhan syndrome is detectable using peripheral blood.

The transition of adult patients with childhood-onset rheumatic disorders from pediatric to non-pediatric healthcare systems has received attention in Japan. However, the clinical course of patients transferred to non-pediatric rheumatologists has not been adequately communicated to pediatric rheumatologists.