Yutaro Nishida

Severe impairment in adenine metabolism with a partial deficiency of adenine phosphoribosyltransferase

Among three unrelated patients with recurrent 2,8-dihydroxyadenine urolithiasis, two completely lacked adenine phosphoribosyltransferase (APRT) in both erythrocytes and proliferative T cells. The third patient possessed significant enzyme activities in both hemolysates and T-cell extracts at levels comparable to heterozygotes for complete APRT deficiency. Despite significant APRT activities in cell extracts, cultured T cells from the third patient were at least 100-fold more resistant than normal T cells to an adenine analog, 6-methylpurine, whose cytotoxicity is dependent on APRT. These data indicate that APRT activity in T cells from the third patient is positive in cell extracts, but apparently not operating in viable cells. Although the cells from the patients with complete APRT deficiency were as resistant to 6-methylpurine as the cells from the third patient, the cells from the heterozygotes for complete APRT deficiency were almost as sensitive as normal T cells. Therefore, adenine metabolism in the third patient but not in the heterozygotes seems to be as severely impaired as in the patients with complete APRT deficiency, which is quite consistent with the clinical manifestations in these individuals.

Hypouricaemic effect after oral administration in chickens of polyethylene glycol-modified uricase entrapped in liposomes

Liposomal‐entrapped methoxypolyethyleneglycol (PEG) modified uricase was used to study the plasma urate lowering effect after its oral administration to chickens. Plasma uric acid concentrations fell gradually and were accompanied by a rise in plasma uricolytic activity. Serum from chickens given PEG‐modified uricase liposomes did not react with native uricase in immunodiffusion plates. These results suggested the clinical usefulness of liposome‐entrapped PEG modified uricase in the treatment of hyperuricaemia.

Hypoxanthine guanine phosphoribosyltransferase deficiency: nucleotide substitution causing Lesch-Nyhan syndrome identified for the first time among Japanese

SummaryA previously undescribed nucleotide substitution at codon 51 (CGA to TGA) has been identified using the polymerase chain reaction technique in hypoxanthine guanine phosphoribosyltransferase (HPRT) cDNA; this is the first molecular evidence for a point mutation in a Japanese patient with Lesch-Nyhan syndrome. The present mutation is the 19th nucleotide substitution identified as a germ-line mutation at this locus and the second mutation generating a stop codon. The position of the nucelotide substitution is exactly the same as a previously described mutation HPRTToronto, indicating for the first time that nucleotide substitutions at the same position in the sequence of HPRT can generate different mutant alleles, one causing a partial deficiency and the other a complete deficiency. Although the type of nucleotide substitution is different between the two cases, a single base position has twice become the target of a mutation. However, the calculation of the probability of finding substitution mutations at the same base position in the coding region of hprt indicates that there is no evidence for the presence of a hot spot for substitution mutations in the human hprt germ line.

Hyperlipidaemia in patients with down's syndrome

Plasma lipid levels were measured in 20 mongoloid and 16 non-mongoloid mentally retarded subjects. Significant elevations of plasma triglyceride levels were found in patients with Downs syndrome compared with mentally retarded controls. However, no significant difference was found in plasma cholesterol, phospholipid and free fatty acid levels between the mongoloids and control subjects.

Effect of free radicals on lymphocyte response to mitogens and rosette formation

Abstract When normal human lymphocytes were exposed to hypoxanthine plus xanthine oxidase system in vitro , the blastogenic response of lymphocytes to mitogens was markedly suppressed. The lymphocytes which respond to Con A were most sensitive to toxic effects of free radicals. In contrast, lymphocyte blastogenesis induced by PWM was less affected by free radicals. Superoxide dismutase was relatively ineffective in preventing the toxicity of free radicals on lymphocyte blastogenesis induced by PHA. In contrast, catalase showed a preventive effect on the oxidase-induced suppression of lymphocyte response to PHA. Uric acid plus uricase also suppressed blastogenic response of the lymphocytes to PHA. The suppressive effects of uric acid plus uricase on lymphocyte proliferation were reversed by catalase. Furthermore, hypoxanthine plus xanthine oxidase were demonstrated to have effects on the lymphocyte membrane receptors, by markedly inhibiting E rosettes; EAC rosettes were only slightly inhibited. These results suggest that H 2 O 2 suppresses lymphocyte function through damage to the cell membrane, and that T cells are more sensitive than B cells to the toxic effects of free radicals.

Relationship between phosphorylation and cytotoxicity of 2-chloroadenosine and 6-methylmercaptopurine riboside in human cells

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.

Theophylline increases serum uric acid levels

The present study was undertaken to investigate the effect of theophylline on serum uric acid and then to elucidate the mechanisms of action of theophylline as a cause of hyperuricemia. There was a significant increase of serum uric acid levels in male asthmatic patients who received theophylline compared to male control subjects without theophylline (6.3 +/- 0.4 mg/ml, mean +/- SEM, versus 4.3 +/- 0.2 mg/ml, p less than 0.01). A significant correlation of serum levels of uric acid and theophylline was demonstrated in asthmatic patients who received 200 to 400 mg sustained-release theophylline (male group, r = 0.480, p less than 0.001; female group, r = 0.398, p less than 0.01). Intravenous administration of aminophylline in three healthy adult male patients did not inhibit uric acid clearance, suggesting that inhibition of excretion of uric acid by theophylline is unlikely. Theophylline slightly inhibited hypoxanthine guanine phosphoribosyltransferase activity in human erythrocyte lysates at concentrations over 5 mM that is considerably more than therapeutic concentrations of theophylline as determined by the conversion of [14C]hypoxanthine to [14C]inosinic acid. Theophylline caused a moderate inhibition of [14C]hypoxanthine uptake by K-562 cells (approximately 50%) at 10mM that is over 100 times as high as those achieved clinically. Further studies remain to be performed to elucidate the exact mechanisms of theophylline-induced hyperuricemia.

Inhibition of purine nucleoside phosphorylase activity and of T-cell function with allopurinol-riboside

Allopurinol-riboside competitively inhibits the action of purine nucleoside phosphorylase on inosine in vitro with aKi of 277 μmol. After simple incubation of allopurinol-riboside with PNP, allopurinol was not formed. Lymphocyte blastogenesis induced by PHA and Con A was significantly suppressed by allopurinol-riboside in a concentration-dependent manner. When LPS was used as a mitogen, the inhibition of allopurinol-riboside on lymphocyte proliferation was less marked. Humoral immunity was not suppressed by allopurinol-riboside. In contrast, cellular immunity was significantly suppressed by allopurinol-riboside in vivo. These results suggested that allopurinol-riboside is a drug which produces a model of PNP deficiency, and that it may be a useful inhibitor of cellular immunity.

Erythrocyte adenosine deaminase and purine nucleoside phosphorylase activity in gout

1. Erythrocyte adenosine deaminase (EC 3.5.4.4) and purine nucleoside (inosine) phosphorylase (EC 2.4.1.1) were measured in 33 healthy controls and 43 primary gouty subjects. Adenosine deaminase activity in controls and gouty subjects was 0.373 plus or minus 0.108 and 0.457 plus or minus 0.140 A unit per 5-10-3 ml packed red cells per h, respectively. The difference was statistically significant (P less than 0.01). Mean adenosine deaminase: inosine phosphorylase (X10) in primary gout was also significantly higher than in controls (P less than 0.05). Inosine phosphorylase activities in the two groups were not significantly different. 2. When gouty patients were divided into two groups according to weight, normal weight gouty subjects had a higher adenosine deaminase activity and an increased ration of adenosine deaminase to inosine phosphorylase when compared with overweight patients (P less than 0.10). In two control groups divided according to the percentage overweight, such differences were not found. In the case of two gouty groups divided according to the existence of gouty heredity, tophi or renal impairment, adenosine deaminase and inosine phosphorylase activity in the two groups were not significantly different. The possible biochemical role of adenosine deaminase activity in primary gout is discussed.

Establishment and characterization of B cell lines from individuals with various types of adenine phosphoribosyltransferase deficiencies

Patients with 2,8-dihydroxyadenine urolithiasis are either completely or partially deficient in adenine phosphoribosyltransferase activities. Patients with partial enzyme deficiencies, all of whom have been found among Japanese, are homozygotes having a unique mutant adenine phosphoribosyltransferase gene (APRT*J) in double dose (Japanese type deficiency). We have established B-cell lines from heterozygotes and homozygotes of complete and Japanese type adenine phosphoribosyltransferase deficiencies as well as normal individuals. Characterization of the cell lines indicated that all homozygous cells were deficient in adenine phosphoribosyltransferase function while all heterozygous and normal cells had functional adenine phosphoribosyltransferase.