A case of factitious skin disease.
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Adjustment Disorders↗
Biomedical subjects
H Miyoshi
Publications and source records attributed to H Miyoshi.
Retroviral vector targeting to human immunodeficiency virus type 1-infected cells by receptor pseudotyping.
We report the generation of retroviral vectors based on Moloney murine leukemia virus that specifically transduce cells infected with T-cell-tropic human immunodeficiency virus type 1 (HIV-1). This vector was pseudotyped with T-cell-tropic HIV-1 receptors CD4 and CXCR4. We demonstrate that transduction is contingent upon HIV-1 gp120 and gp41 expression.
Animals↗
Bile salts mediate hepatocyte apoptosis by increasing cell surface trafficking of Fas.
Toxic bile salts induce hepatocyte apoptosis by a Fas-dependent, Fas ligand-independent mechanism. To account for this observation, we formulated the hypothesis that toxic bile salts induce apoptosis by effecting translocation of cytoplasmic Fas to the cell surface, resulting in transduction of Fas death signals. In McNtcp.24 cells the majority of Fas was cytoplasmic, as assessed by cell fractionation and immunofluorescence studies. However, cell surface Fas increased sixfold after treatment with the toxic bile salt glycochenodeoxycholate (GCDC) in the absence of increased Fas protein expression. Moreover, in cells transfected with Fas-green fluorescence protein, cell surface fluorescence also increased in GCDC-treated cells, directly demonstrating Fas translocation to the plasma membrane. Both brefeldin A, a Golgi-disrupting agent, and nocodazole, a microtubule inhibitor, prevented the GCDC-induced increase in cell surface Fas and apoptosis. In conclusion, toxic bile salts appear to induce apoptosis by promoting cytoplasmic transport of Fas to the cell surface by a Golgi- and microtubule-dependent pathway.
Apoptosis↗
Cathepsin B contributes to TNF-alpha-mediated hepatocyte apoptosis by promoting mitochondrial release of cytochrome c.
TNF-alpha-induced apoptosis is thought to involve mediators from acidic vesicles. Cathepsin B (cat B), a lysosomal cysteine protease, has recently been implicated in apoptosis. To determine whether cat B contributes to TNF-alpha-induced apoptosis, we exposed mouse hepatocytes to the cytokine in vitro and in vivo. Isolated hepatocytes treated with TNF-alpha in the presence of the transcription inhibitor actinomycin D (AcD) accumulated cat B in their cytosol. Further experiments using cell-free systems indicated that caspase-8 caused release of active cat B from purified lysosomes and that cat B, in turn, increased cytosol-induced release of cytochrome c from mitochondria. Consistent with these observations, the ability of TNF-alpha/AcD to induce mitochondrial release of cytochrome c, caspase activation, and apoptosis of isolated hepatocytes was markedly diminished in cells from CatB(-/-) mice. Deletion of the CatB gene resulted in diminished liver injury and enhanced survival after treatment in vivo with TNF-alpha and an adenovirus construct expressing the IkappaB superrepressor. Collectively, these observations suggest that caspase-mediated release of cat B from lysosomes enhances mitochondrial release of cytochrome c and subsequent caspase activation in TNF-alpha-treated hepatocytes.
Animals↗
Accuracy of detection of colorectal neoplasia using an immunochemical occult blood test in symptomatic referred patients: comparison of retrospective and prospective studies.
OBJECTIVE: In this study the sensitivity and specificity of immunochemical tests for colorectal neoplasia were evaluated in retrospective and prospective studies. METHODS: Four types of fecal blood tests--a chemical test (Hemoccult II) and three different immunochemical tests including a test which detects hemoglobin and transferrin- were performed in the retrospective study. In the prospective study the test for hemoglobin and transferrin was used for all patients that underwent total colonoscopy. PATIENTS: One hundred seven patients with colorectal neoplasia, 57 with gastroduodenal bleeding, and 62 with normal digestive tracts were examined retrospectively. One thousand two hundred and ninety-eight nonspecifically symptomatic patients whose endoscopic examination was negative for hemorrhagic lesions in the upper digestive tract were examined prospectively. RESULTS: In the retrospective study, sensitivities for the detection of colorectal cancers and adenomas with diameters > or =10 mm using the tests which detect hemoglobin and transferrin were 98% and 89%, respectively. These were the highest sensitivity among the four tests. The specificity of this test was 97%, which was higher than that of the Hemoccult II test. In the prospective study, the sensitivities of the tests for hemoglobin and transferrin for the detection of colorectal cancers and adenomas with diameters > or =10 mm were 79% and 33%, respectively. The specificity was 95%. CONCLUSIONS: The test for hemoglobin and transferrin showed the highest sensitivity and specificity for colorectal neoplasia in the retrospective study. The sensitivity and specificity of this test were not so high in the prospective study, but they may be clinically applicable in the evaluation of patients with various nonspecific symptoms.
Adenoma↗
[Diabetic nephropathy and low molecular weight advanced glycation end-product in urine].
Glucose reacts non-enzymatically with amino groups of proteins to produce diverse groups of protein-bound moieties with cross-linking properties called advanced glycation end-products (AGEs). A large number of studies suggest that tissue and circulating AGEs, especially those bound to small peptides, play an important role in the pathogenesis and development of diabetic complications. The aim of the present study was to evaluate AGE-peptide levels in the urine in various stages of diabetic nephropathy and to analyze structures of AGEs generated in vivo. Urine samples were obtained from 100 type-2 diabetic patients with various stages of nephropathy, 39 non-diabetic patients with renal dysfunction, and 20 control subjects with neither diabetes nor renal disease. Low molecular weight peptides (< 10 kDa) isolated from each sample were used. With the use of size-exclusion HPLC monitored by AGE-specific fluorescence, three main fluorescence peaks were detected at molecular weights of 6,000 Da, 2,500 Da, and 300 Da. The fluorescence intensity of total AGE-peptide and 6,000 Da peak AGE-peptide is enhanced together with the development of renal dysfunction in diabetic patients, which strongly correlated with the level of serum creatinine. There was no difference in total urinary AGE between diabetic and non-diabetic hemodialysis patients, suggesting that insufficient clearance during dialysis is more important than AGE production. A competitive ELISA using a polyclonal anti-AGE antibody, anion-exchange HPLC, and reversed-phase HPLC were performed on the peaks appearing in size-exclusive HPLC to further characterize the AGE composition. The 300 Da product was specific to diabetic patients with chronic renal failure and hemodialysis patients, which may be a good marker to distinguish diabetic from non-diabetic renal failures. We also speculated that because of its low molecular weight, the 300 Da product might represent the final degradation product of AGE-peptides in the kidney.
Biomarkers↗
Allergic contact dermatitis due to beclometasone dipropionate in an inhalant for asthma.
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Administration, Inhalation↗
Gastric and duodenal polyps in Smad4 (Dpc4) knockout mice.
The SMAD4 (DPC4) gene was initially isolated as a candidate tumor suppressor from the convergent site of homozygous deletions on 18q in a panel of pancreatic carcinoma cell lines. It encodes a common cytoplasmic signaling molecule shared by the transforming growth factor-beta, activin, and bone morphogenic pathways. We recently inactivated its mouse homologue Smad4 and demonstrated its role in the malignant progression of benign adenomas to invasive adenocarcinomas by analyzing mice with Apc and Smad4 compound mutations. Although simple Smad4 homozygotes were embryonically lethal, the heterozygotes were fertile and appeared normal up to the age of 1 year. Upon further investigation, however, they have developed inflammatory polyps in the glandular stomach and duodenum. By PCR genotyping and immunohistochemical staining, the wild-type Smad4 allele has been lost in the polyp epithelial cells, ie., loss of heterozygosity. On the other hand, we have not found any mutations in such genes as K-Ras, H-Ras, N-Ras, p53, or PTEN. Histologically, the polyps are similar to human juvenile polyps showing moderate stromal cell proliferation and infiltrations by eosinophils and plasma cells. In addition, foci of adenocarcinoma with signet ring cells are also found. These results are consistent with a recent report that germ-line SMAD4 mutations are found in a subset of familial juvenile polyposis.
Animals↗
Vibrational modes of ubiquinone in cytochrome bo(3) from Escherichia coli identified by Fourier transform infrared difference spectroscopy and specific (13)C labeling.
In this study we present the infrared spectroscopic characterization of the bound ubiquinone in cytochrome bo(3) from Escherichia coli. Electrochemically induced Fourier transform infrared (FTIR) difference spectra of DeltaUbiA (an oxidase devoid of bound ubiquinone) and DeltaUbiA reconstituted with ubiquinone 2 and with isotopically labeled ubiquinone 2, where (13)C was introduced either at the 1- or at the 4-position of the ring (C=O groups), have been obtained. The vibrational modes of the quinone bound to the discussed high-affinity binding site (Q(H)) are compared to those from the synthetic quinones in solution, leading to the assignment of the C=O modes to a split signal at 1658/1668 cm(-)(1), with both carbonyls similarly contributing. The FTIR spectra of DeltaUbiA reconstituted with the labeled quinones indicate an essentially symmetrical and weak hydrogen bonding of the two C=O groups from the neutral quinone with the protein and distinct conformations of the 2- and 3-methoxy groups. Perturbations of the vibrational modes of the 5-methyl side groups are discussed for a signal at 1452 cm(-)(1). Only negligible shifts of the aromatic ring modes can be reported for the reduced and the protonated form of the quinone. Alterations of the protein upon quinone binding are reflected in the electrochemically induced FTIR difference spectra. In particular, difference signals at 1640-1633 cm(-)(1) and 1700-1670 cm(-)(1) indicate variations of beta-sheet secondary structure elements and loops, bands at 1706 and 1678 cm(-)(1) are tentatively attributed to individual amino acids, and a difference signal a 1540 cm(-)(1) is discussed to reflect an influence on C=C modes of the porphyrin ring or on deprotonated propionate groups of the hemes. Further tentative assignments are presented and discussed. The (13)C labeling experiments allow the assignment of the vibrational modes of a bound ubiquinone 8 in the electrochemically induced FTIR difference spectra of wild-type bo(3).
Binding Sites↗
IkappaB kinases phosphorylate NF-kappaB p65 subunit on serine 536 in the transactivation domain.
Recent investigations have elucidated the cytokine-induced NF-kappaB activation pathway. IkappaB kinase (IKK) phosphorylates inhibitors of NF-kappaB (IkappaBs). The phosphorylation targets them for rapid degradation through a ubiquitin-proteasome pathway, allowing the nuclear translocation of NF-kappaB. We have examined the possibility that IKK can phosphorylate the p65 NF-kappaB subunit as well as IkappaB in the cytokine-induced NF-kappaB activation. In the cytoplasm of HeLa cells, the p65 subunit was rapidly phosphorylated in response to TNF-alpha in a time dependent manner similar to IkappaB phosphorylation. In vitro phosphorylation with GST-fused p65 showed that a p65 phosphorylating activity was present in the cytoplasmic fraction and the target residue was Ser-536 in the carboxyl-terminal transactivation domain. The endogenous IKK complex, overexpressed IKKs, and recombinant IKKbeta efficiently phosphorylated the same Ser residue of p65 in vitro. The major phosphorylation site in vivo was also Ser-536. Furthermore, activation of IKKs by NF-kappaB-inducing kinase induced phosphorylation of p65 in vivo. Our finding, together with previous observations, suggests dual roles for IKK complex in the regulation of NF-kappaB.IkappaB complex.
Amino Acid Sequence↗
Origin of selective inhibition of mitochondrial complex I by pyridinium-type inhibitor MP-24.
Positively charged pyridiniums are unique inhibitors to probe the structural and functional properties of the ubiquinone reduction site of bovine heart mitochondrial complex I. In this study, we synthesized a series of neutral as well as pyridinium analogues of MP-24 (N-methyl-4-[2-methyl-2-(p-tert-butylbenzyl)propyl]pyridinium), a selective inhibitor of one of the two proposed binding sites of these pyridinium-type inhibitors of complex I (H. Miyoshi et al., J. Biol. Chem. 273 (1998) 17368-17374), to elucidate the origin of its selectivity. Inhibitory potencies of all neutral and pyridinium analogues with tetraphenylboron (TPB(-)), which forms an ion-pair with pyridiniums, were comparable, although the degrees of selective inhibition by pyridiniums without TPB(-) were entirely different. In contrast to MP-24, the dose-response curves of nonselective pyridiniums and all neutral analogues were not affected by incubation conditions. These results strongly suggested that the process of the inhibitor passage to the binding sites is responsible for the selective inhibition.
Animals↗
Role of a bound ubiquinone on reactions of the Escherichia coli cytochrome bo with ubiquinol and dioxygen.
To probe the functional role of a bound ubiquinone-8 in cytochrome bo-type ubiquinol oxidase from Escherichia coli, we examined reactions with ubiquinol-1 and dioxygen. Stopped-flow studies showed that anaerobic reduction of the wild-type and the bound ubiquinone-free (DeltaUbiA) enzymes with ubiquinol-1 immediately takes place with four kinetic phases. Replacement of the bound ubiquinone with 2,6-dibromo-4-cyanophenol (PC32) suppressed the anaerobic reduction of the hemes with ubiquinol-1 by eliminating the fast phase. Flow-flash studies in the reaction of the fully reduced enzyme with dioxygen showed that the heme b-to-heme o electron transfer occurs with a rate constant of approximately 1x10(4) s(-1) in all three preparations. These results support our previous proposal that the bound ubiquinone is involved in facile oxidation of substrates in subunit II and subsequent intramolecular electron transfer to low-spin heme b in subunit I.
Cytochrome b Group↗
Role of a bound ubiquinone on reactions of the Escherichia coli cytochrome bo with ubiquinol and dioxygen.
To probe the functional role of a bound ubiquinone-8 in cytochrome bo-type ubiquinol oxidase from Escherichia coli, we examined reactions with ubiquinol-1 and dioxygen. Stopped-flow studies showed that anaerobic reduction of the wild-type and the bound ubiquinone-free (delta UbiA) enzymes with ubiquinol-1 immediately takes place with four kinetic phases. Replacement of the bound ubiquinone with 2,6-dibromo-4-cyanophenol (PC32) suppressed the anaerobic reduction of the hemes with ubiquinol-1 by eliminating the fast phase. Flow-flash studies in the reaction of the fully reduced enzyme with dioxygen showed that the heme b to heme o electron transfer occurs with a rate constant of approximately 10(4) s-1 in all three preparations. These results support our previous proposal that the bound ubiquinone is involved in facile oxidation of substrates in subunit II and subsequent intramolecular electron transfer to low-spin heme b in subunit I.
Catalysis↗
Topographical characterization of the ubiquinone reduction site of glucose dehydrogenase in Escherichia coli using depth-dependent fluorescent inhibitors.
Membrane-bound glucose dehydrogenase in Escherichia coli possesses a binding site for ubiquinone as well as glucose, metal ion and pyrroloquinoline quinone. To probe the depth of the ubiquinone binding site in the membrane environment, we synthesized two types of fluorenyl fatty acids which bear an inhibitor mimic moiety (i.e., specific inhibitor capsaicin) close to the fluorene located at different positions in the alkyl tail chain; one close to the polar carbonyl head group (alpha-(3, 4-dimethoxyphenyl)acetyloxy-7-nonyl-2-fluoreneacetic acid, alpha-DFA), and the other in the middle of the chain (theta-(3, 4-dimethoxyphenyl)acetyloxy-7-ethyl-2-fluorenenonanoic acid, theta-DFA). Mixed lipid vesicles consisting of phosphatidylcholine (PC) and alpha-DFA or theta-DFA were prepared by sonication method, and fluorescent quenching against a hydrophilic quencher, iodide anion, was examined. The vesicles containing alpha-DFA were more susceptible to quenching than those containing theta-DFA, indicating that the fluorene and consequently capsaicin mimic moiety are located at different depths in the lipid bilayer depending upon the position of attachment to the alkyl tail chain. The purified glucose dehydrogenase was reconstituted into PC vesicles which consisted of PC and alpha-DFA or theta-DFA with various molar ratios. For both types of reconstituted vesicles, the extent of inhibition of short-chain ubiquinone reduction activity increased with increases in the molar ratio of fluorenyl fatty acid to PC. The ubiquinone reduction activity was more significantly inhibited in the reconstituted vesicles containing alpha-DFA compared to those containing theta-DFA. Our findings strongly suggested that the ubiquinone reduction site in glucose dehydrogenase is located close to the membrane surface rather than in the hydrophobic membrane interior.
Binding Sites↗
Functional interactions of transforming growth factor beta-activated kinase 1 with IkappaB kinases to stimulate NF-kappaB activation.
Several mitogen-activated protein kinase kinase kinases play critical roles in nuclear factor-kappaB (NF-kappaB) activation. We recently reported that the overexpression of transforming growth factor-beta-activated kinase 1 (TAK1), a member of the mitogen-activated protein kinase kinase kinase family, together with its activator TAK1-binding protein 1 (TAB1) stimulates NF-kappaB activation. Here we investigated the molecular mechanism of TAK1-induced NF-kappaB activation. Dominant negative mutants of IkappaB kinase (IKK) alpha and IKKbeta inhibited TAK1-induced NF-kappaB activation. TAK1 activated IKKalpha and IKKbeta in the presence of TAB1. IKKalpha and IKKbeta were coimmunoprecipitated with TAK1 in the absence of TAB1. TAB1-induced TAK1 activation promoted the dissociation of active forms of IKKalpha and IKKbeta from active TAK1, whereas the IKK mutants remained to interact with active TAK1. Furthermore, tumor necrosis factor-alpha activated endogenous TAK1, and the kinase-negative TAK1 acted as a dominant negative inhibitor against tumor necrosis factor-alpha-induced NF-kappaB activation. These results demonstrated a novel signaling pathway to NF-kappaB activation through TAK1 in which TAK1 may act as a regulatory kinase of IKKs.
Adaptor Proteins, Signal Transducing↗
Transduction of human CD34+ cells that mediate long-term engraftment of NOD/SCID mice by HIV vectors.
Efficient gene transfer into human hematopoietic stem cells (HSCs) is an important goal in the study of the hematopoietic system as well as for gene therapy of hematopoietic disorders. A lentiviral vector based on the human immunodeficiency virus (HIV) was able to transduce human CD34+ cells capable of stable, long-term reconstitution of nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice. High-efficiency transduction occurred in the absence of cytokine stimulation and resulted in transgene expression in multiple lineages of human hematopoietic cells for up to 22 weeks after transplantation.
Animals↗
The quinohemoprotein alcohol dehydrogenase of Gluconobacter suboxydans has ubiquinol oxidation activity at a site different from the ubiquinone reduction site.
Alcohol dehydrogenase (ADH) of acetic acid bacteria functions as the primary dehydrogenase of the ethanol oxidase respiratory chain, where it donates electrons to ubiquinone. In addition to the reduction of ubiquinone, ADHs of Gluconobacter suboxydans and Acetobacter aceti were shown to have a novel function in the oxidation of ubiquinol. The oxidation activity of ubiquinol was detected as an ubiquinol:ferricyanide oxidoreductase activity, which can be monitored by selected wavelength pairs at 273 and 298 nm with a dual-wavelength spectrophotometer. The ubiquinol oxidation activity of G. suboxydans ADH was shown to be two times higher in 'inactive ADH', whose ubiquinone reductase activity is 10 times lower, than with normal 'active' ADH. No activity could be detected in the isolated subunit II or subunit I/III complex, but activity was detectable in the reconstituted ADH complex. Inactive and active ADHs exhibited a 2-3-fold difference in their affinity to ubiquinol despite having the same affinity to ubiquinone. Furthermore, the ubiquinol oxidation site in ADH could be distinguished from the ubiquinone reduction site by differences in their sensitivity to ubiquinone-related inhibitors and by their substrate specificity with several ubiquinone analogues. Thus, the results strongly suggest that the reactions occur at different sites. Furthermore, in situ reconstitution experiments showed that ADH is able to accept electrons from ubiquinol present in Escherichia coli membranes, suggesting the ubiquinol oxidation activity of ADH has a physiological function. Thus, ADH of acetic acid bacteria, which has ubiquinone reduction activity, was shown to have a novel ubiquinol oxidation activity, of which the physiological function in the respiratory chain of the organism is also discussed.
Acetobacter↗
Neuronal impairment of adult moyamoya disease detected by quantified proton MRS and comparison with cerebral perfusion by SPECT with tc-99m HM-PAO: a trial of clinical quantification of metabolites.
We evaluated neuronal impairment of adult moyamoya patients by proton magnetic resonance spectroscopy (MRS) and cerebral perfusion scintigraphy. After evaluating two methods for compensating relaxation times, we selected a long TR, short TE sequence to achieve better reproducibility. The cerebral blood flow (CBF) value was measured by scintigraphy following the method of quantification reported in the previous literature. N-acetyl aspartate (NAA) concentrations and CBFs value were decreased statistically (P < 0. 05) compared with those of the age-matched normal controls. However, the decreased rate of NAA concentration was changed more than the CBF value, and no linear correlation was found between the two values. We considered that the NAA concentration was not always correlated with that of the present cerebral perfusion. The NAA concentration showed a more dispersive distribution in patients than in controls, indicating a large individual variation in neuronal impairment. We concluded that proton MRS may provide useful information about neuronal impairment in individual patients. J. Magn. Reson Imaging 1999;10:124-129.
Adult↗