We investigated the intercellular adhesion molecule-1 (ICAM-1) gene polymorphism in 90 patients with young-onset type 1 diabetes, 74 with adult-onset type 1 diabetes, and 171 control subjects. The distribution of C-T genotypes and allele frequencies in exon 6 of the ICAM-1 gene was significantly different between adult-onset type 1 diabetes patients and controls (chi(2) = 9.76, p = 0.0076), and between patients with adult-onset and young-onset type 1 diabetes (chi(2) = 11.28, p = 0.0036). In contrast, we failed to detect any association between patients with young-onset type 1 diabetes and controls. Our data suggest that ICAM-1 exon 6 gene polymorphism affects the age-at-onset of type 1 diabetes and that different pathogenetic mechanisms may exist between young-onset and adult-onset type 1 diabetes.

Adolescent↗

PubMed2000

Tumor necrosis factor, tumor necrosis factor receptors type 1 and 2, lymphotoxin-alpha, and HLA-DRB1 gene polymorphisms in human T-cell lymphotropic virus type I associated myelopathy.

We studied tumor necrosis factor (TNF), lymphotoxin-alpha (LT-alpha), and TNF receptors type 1 (TNFR-1) and type 2 (TNFR-2) gene polymorphisms as well as HLA class II DRB1 alleles in Japanese patients with human T-cell lymphotropic virus type I (HTLV-I) associated myelopathy (HAM) (n = 51), patients with adult T-cell leukemia/lymphoma (ATL) (n = 48), asymptomatic HTLV-I carriers (n = 50), and HTLV-I seronegative, normal controls (n = 112). There were significant differences between HAM patients and normal controls in the distributions of TNF promoter region polymophism at position --857, the LT-alpha gene NcoI polymorphism, and the T-G substitution in exon 6 of the TNFR-2 gene. The distribution of the NcoI polymorphism of the LT-alpha gene was also significantly different between HAM patients and asymptomatic HTLV-I carriers. In contrast, we failed to detect any difference in the frequency of DRB1, TNF promoter at position --1031, --863, or the TNFR-1 promoter --383 polymorphism. The results suggest that the TNF/LT-alpha gene region within the HLA class III of chromosome 6 and the TNFR-2 gene region located on chromosome 1p36 might contribute to susceptibility to HAM, and that aberrant expression or function of these cytokines and the receptor could be involved in the development of HAM.

Alleles↗

PubMed2000

Evidence for stimulation of adenylyl cyclase by an activated G(s) heterotrimer in cell membranes: an experimental method for controlling the G(s) subunit composition of cell membranes.

Heterotrimeric (alphabetagamma) G(s) mediates agonist-induced stimulation of adenylyl cyclase (AC). Cholera toxin (CTx) will ADP-ribosylate the alpha-subunit of G(s) (G(s)alpha). G(s)alpha-deficient cyc(-) membranes were "stripped" of Gbeta. When the stripped cyc(-) were incubated with G(s)alpha and/or Gbetagamma, each was incorporated into the membranes independently of the other. Both G(s)alpha and Gbetagamma had to be present in the membranes, and they had to be able to form a heterotrimer in order for CTx to ADP-ribosylate G(s)alpha, indicating that the membrane bound G(s) heterotrimer is a substrate for CTx, but the G(s)alpha subunit by itself is not. When G(s)alpha was completely and irreversibly activated with GTPgammaS and incorporated into stripped cyc(-), it was a poor substrate for CTx and a weak stimulator of AC unless Gbetagamma was also incorporated. Furthermore, the level of AC stimulation corresponded to the amount of G(s) heterotrimer that was formed in the membranes from GTPgammaS-activated G(s)alpha and Gbetagamma. These data suggest that AC is stimulated by an activated G(s) heterotrimer in cell membranes.

Adenosine Diphosphate Ribose↗

PubMed2000

Genetic analysis of mortality in murine angiostrongyliasis costaricensis using SMXA recombinant inbred mouse strains.

The SMXA recombinant inbred mouse strain set was produced by systematic inbreeding from the F2 generation of a cross between two progenitor inbred strains, A/J and SM/J, which differed markedly with respect to the patterns of infection with Angiostrongylus costaricensis. We have applied this set to genetic analysis of mouse susceptibility to this nematode infection. The mortality was variable among substrains of the SMXA RI strains, indicating the involvement of multiple genes. Linkage analysis showed several chromosomal regions closely linked to mortality; chromosome 6 (D6Rik86, 87; P<==0.001), 10 (D10Rik66-D10Mit12; P=0.0058), 13 (D13Rik79, 80; P=0.0096) and 17 (D17Mit28-D17Rik76; P=0.0088).

Angiostrongylus↗

PubMed2000

Inhibitory effects of carbohydrates on cholesterol esterase biosynthesis in Streptomyces lavendulae H646-SY2.

The inhibitory effects of various carbohydrates on cholesterol esterase (CHE) biosynthesis were investigated in a CHE-producing Streptomyces lavendulae H646-SY2. A marked decrease in CHE production and preferential utilization of glucose were observed when glucose was added to a medium containing palmitic acid as the sole carbon source and an inducer of CHE production. This catabolite repression was also caused by other carbohydrates such as sucrose, glycerol, galactose and arabinose. Sucrose, however, did not inhibit the utilization of palmitic acid by the strain, suggesting that the mechanism of the catabolite repression by sucrose is different from that by glucose.

Journal Article↗

PubMed2000

Transgenic apple (Malus x domestica) shoot showing low browning potential.

Transgenic apple shoots were prepared from leaf disks by using Agrobacterium tumefaciens carrying the kanamycin (KM) resistance gene and antisense polyphenol oxidase (PPO) DNA. Four transgenic apple lines that grew on the medium containing 50 microgram/mL KM were obtained. They contained the KM resistance gene and grew stably on the medium for >3 years. Two transgenic shoot lines containing antisense PPO DNA in which PPO activity was repressed showed a lower browning potential than a control shoot.

Agrobacterium tumefaciens↗

PubMed2000

Alterations of GTP-binding proteins (Gsalpha and Gq/11alpha) in gastric smooth muscle cells from streptozotocin-induced and WBN/Kob diabetic rats.

We investigated possible impairment of the signal transduction system in gastric myocytes of streptozotocin-induced diabetic (STZ) and spontaneous diabetic WBN/Kob (WBN/Kob) rats. Gastric motility 10 weeks after the onset of diabetes mellitus was significantly reduced in both diabetic rats compared with control, and the decreased motility was not recovered by the administration of insulin to maintain normal blood glucose levels. There was no significant difference between both types of diabetic rats and control rats in total number of [3H]quinuclidinyl benzilate ([3H]QNB) binding sites (Bmmax: 545-587 fmol/mg protein) on gastric smooth muscle cell membranes or in the affinity of [3H]QNB for the binding sites (Kd: 0.06-0.07 nM). Immunoblot analysis using polyclonal anti-G-protein antibodies indicated increased expression of Gsalpha in gastric smooth muscle cell membranes, but no significant change in Gialpha or Gq/11alpha expression in STZ rats, and decreased expression of Gq/11alpha with no significant change in Gsalpha and Gialpha in WBN/Kob rats. The cAMP production in gastric smooth muscle cell membranes was augmented in the absence and presence of 100 microM isoproterenol, and 100 microM forskolin in STZ rats, whereas no significant change of cAMP production was observed in WBN/Kob rats irrespective of the presence of the stimulants. These findings suggest that long-standing diabetes may induce alterations in signal transduction at downstream receptors in gastric myocytes, resulting in the impairment of gastric motility, although the mechanism of reduced contractile activity may differ between STZ and WBN/Kob rats.

Adenylyl Cyclases↗

PubMed2000

Bone marrow transplantation for the treatment of X-linked adrenoleukodystrophy.

The effects of bone marrow transplantation (BMT) in X-linked childhood adrenoleukodystrophy (ALD) are described in four Japanese patients. Two older patients, 10-year-old boys with IQ 60 and difficulty in school, had favourable results. IQ levels and MRI findings were generally maintained after BMT. One patient showed improvement of gait disturbance. They have both attended ordinary schools after BMT, although a learning disorder persists. On the other hand, two other younger patients with a rapid course and indeterminate IQ at BMT showed deterioration of neurological functions. Indication for BMT seems to be a maintained IQ level, preferably higher than 80, since it seems to be difficult to normalize IQ level after BMT. Younger patients have higher risk of developing a rapidly progressive form of the disease. Identification of presymptomatic boys, and serial and careful follow-up by neuropsychological and neuroradiological studies, are essential prerequisites to successful BMT in X-ALD.

Adrenoleukodystrophy↗

PubMed2000

Mapping quantitative trait loci for circadian behavioral rhythms in SMXA recombinant inbred strains.

SM/J and A/J inbred strain of mice have different characteristics in circadian behaviors such as free-running period (tau), phase relationship (psi) between light-dark cycles and activity rhythms, and amount of wheel-running activity. To determine the genes which affect these behaviors, a quantitative trait locus (QTL) analysis using SMXA recombinant inbred strains derived from SM/J and A/J mice was performed. Concerning tau, two regions on chromosomes (Chrs) 7 and 18 surpassed the genome-wide suggestive level. As for psi, one suggestive QTL was detected on Chr 7. The QTLs which affect daily activity counts under light-dark cycles and constant darkness were mapped to the same chromosomal regions on Chrs 1 and 17, respectively. The provisional QTLs detected in the present study might be useful for understanding the complex mechanism regulating circadian behaviors.

Animals↗

PubMed2000

Gene polymorphism for microsomal epoxide hydrolase and susceptibility to emphysema in a Japanese population.

Recently, it was reported that gene polymorphism for microsomal epoxide hydrolase (mEPHX), an enzyme involved in the first-pass metabolism of epoxide intermediates, was associated with susceptibility to emphysema. This association was examined in a Japanese population, performing polymerase chain reaction (PCR)-based direct sequencing and restriction fragment length polymorphism assays for variant forms of mEPHX. The subjects consisted of 79 smokers with moderate to severe emphysema diagnosed by lung computed tomography scans, 58 smokers without emphysema, with a comparable smoking history, and 114 consecutive subjects who undertook annual health checkups. The allele frequency of exon 3 Tyr113 to His113, which was reported to confer slow mEPHX activity, was substantially higher in the population control group compared with that of the Caucasian control subjects in a previous study. However, neither the genotype distribution of exon 3, nor that of exon 4 His139 to Arg139, was significantly different between the two groups of smokers. These data indicate that the gene polymorphism for mEPHX is not associated with susceptibility to emphysema in the Japanese population. The discrepancy between the two studies may be explained either by racial difference or by the selection bias of subjects in the previous study, which examined those who had only mild to moderate emphysema with lung cancer or those who were clinically diagnosed as having chronic obstructive pulmonary disease.

Disease Susceptibility↗

PubMed2000

Mutation of conserved aspartates affect maturation of presenilin 1 and presenilin 2 complexes.

Presenilin (PS1 and PS2) holoproteins are transiently incorporated into low molecular weight (MW) complexes. During subsequent incorporation into a higher MW complex, they undergo endoproteolysis to generate stable N- and C-terminal fragments (NTF/CTF). Mutation of either of two conserved aspartate residues in transmembrane domains inhibits both presenilin-endoproteolysis and the proteolytic processing of APP and Notch. We show that aspartate-mutant holoprotein presenilins are not incorporated into the high molecular weight, NTF/CTF-containing complexes. Aspartate-mutant presenilin holoproteins also preclude entry of endogenous wild-type PS1/PS2 into the high molecular weight complexes, but do not affect the incorporation of wild-type holoproteins into lower molecular weight holoprotein complexes. These data suggest that the loss-of-function aspartate-mutants cause altered PS complex maturation, and argue that the functional presenilin moieties are contained in the high molecular weight presenilin NTF/CTF-containing complexes.

Alzheimer Disease↗

PubMed2000

Prognostic value of p53 gene mutations and the product expression in de novo acute myeloid leukemia.

In acute myeloid leukemia (AML), p53 mutations are reportedly infrequent but associated with a poor prognosis. The majority of mutations are missense mutations, which generally lead to accumulation of nuclear p53 protein. However, the prognostic significance of the accumulation remains unknown in AML. In this study, we compared the prognostic value of p53 mutations versus accumulation of the product. p53 mutations were found in 9 (4.5%) of 200 patients with de novo AML. The p53 mutation detectable (mutation+) group had a worse prognosis (p = 0.0009) than the mutation not detectable (mutation-) group. Multivariate analysis showed that the p53 mutation was an independent factor (p = 0.005) for short overall survival as well as 60 yr or older (p = 0.001) and unfavorable karyotypes (p = 0.001). In 79 of the 200 patients, the expression of p53 was studied by immunocytochemistry (ICC) using anti-p53 monoclonal antibody (DO-7). All samples carrying missense mutations (N = 6) were positive for ICC in over 15% of nuclei of each sample, chosen as the optimized cutoff value of p53 accumulation. Accumulation was thus found in 14 of the 79 patients. However, there was no prognostic difference according to the accumulation, because the mutation-/accumulation+ group (N = 8) tended to have a good prognosis. These findings indicate that molecular detection of p53 mutations yields better prognostic information than ICC. In a subset of AML, p53 protein might be accumulated without mutation presumably due to upstream signals of p53.

Acute Disease↗

PubMed2000

Decitabine (5-Aza-2'-deoxycytidine) decreased DNA methylation and expression of MDR-1 gene in K562/ADM cells.

Multidrug resistance (MDR) is a major problem in patients with hematological malignancies. Although drug-resistance is known to be induced by the expression of P-glycoprotein (P-gp) encoded by the MDR-1 gene, little is known about the mechanisms regulating this gene. Herein, we studied the DNA methylation patterns at the enhancer and repressor binding sites of the MDR-1 gene using the human erythroleukemia cell line K562 and its multidrug resistant derivative K562/ADM (adriamycin). Direct DNA sequence analysis demonstrated methylation to be present at the repressor site (minus 110 GC-box) of the MDR-1 gene in K562/ADM cells, but not in parental K562 cells. Methylation-specific PCR (MSP) analysis yielded similar results. Treatment of K562/ADM cells with 5-Aza-2'-deoxycytidine (decitabine; DAC), an inhibitor of DNA methyltransferase, caused demethylation of the repressor binding site of MDR-1 gene, as assessed by MSP, and also decreased P-gp expression, as assessed by flow cytometric and Northern blot analysis. Although it is generally accepted that DAC upregulates gene expression by demethylating the activator binding sites, our present results suggest that DAC induces down-regulation of P-gp expression as a result of demethylation at the repressor binding site in K562/ADM cells. In this regard, methylation-dependent regulation of the MDR-1 gene in K562/ADM cells is unique.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2000

Role of nitric oxide in haemodialysis hypotension.

To investigate the possible involvement of nitric oxide (NO) in haemodialysis hypotension, we measured plasma concentrations of nitrate anion (NO3-), a metabolite of NO, in 114 patients undergoing maintenance haemodialysis. Mean plasma NO3- concentrations before dialysis were greater in subjects with lower blood pressure (155 +/- 16 micromol L-1) than in those with middle (117 +/- 8 micromol L-1) or higher blood pressure (105 +/- 12 micromol L-1) before dialysis. Further, mean plasma NO3- concentrations before dialysis were greater in subjects with lower blood pressure (186 +/- 13 micromol L-1) than in those with middle (112 +/- 7 micromol L-1) or higher blood pressure (64 +/- 11 micromol L-1) after dialysis. Plasma NO3- concentrations before dialysis were inversely correlated with mean blood pressure before dialysis (r=0.318, P=0.0006), and showed a strong inverse correlation with mean blood pressure after dialysis (r=0.608, P=0.0001). In the selected participants who had equal range of mean blood pressure before dialysis, mean plasma NO3- concentrations were greater in subjects with severe hypotension during dialysis (180 +/- 14 micromol L-1) than in those with mild hypotension (99 +/- 11 micromol L-1) or without hypotension (53 +/- 12 micromol L-1); plasma NO3- concentrations before dialysis were inversely correlated with changes in mean blood pressure during dialysis and mean blood pressure after dialysis. Results indicate that enhanced NO production may be involved in acute hypotension during dialysis, and suggest the possible involvement of NO in the pathogenesis of chronic hypotension associated with maintenance haemodialysis.

Adult↗