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Biomedical subjects

A Abe

Publications and source records attributed to A Abe.

At least 55 records · Page 3Linked to original sources
PubMed2000

Glucosylceramide synthase: assay and properties.

Glucosylceramide synthesis is a key step in the formation of most mammalian glycosphingolipids. The expanding number of cellular functions that may be glycosphinolipid dependent and the identification of this glucosylceramide synthase as a potential therapeutic target for several sphingolipid storage disorders necessitate the availability of a reliable assay for glucosylceramide synthase. Coupled with the recent sequencing of this enzyme, the liposome-based assay utilizing a single extraction step should aid in the understanding of this critical early pathway in glycosphingolipid formation.

Cations, Divalent↗
PubMed2000

In vivo interactions of rabbit enteropathogenic Escherichia coli O103 with its host: an electron microscopic and histopathologic study.

A family of human and animal pathogens, including enteropathogenic and enterohemorrhagic Escherichia coli (EPEC and EHEC), trigger formation of 'attaching and effacing' lesions on cultured and intestinal epithelial surfaces. However, our understanding of these events in vivo is incomplete. To further study these interactions in a natural infection model, weaned rabbits were infected with rabbit enteropathogenic E. coli O103 (REPEC O103), followed clinically, and infected tissues were evaluated by electron and light microscopy. Of the 36 rabbits challenged, morbidity and mortality were 65 and 23%, respectively. Twenty-four hours after infection, expression of fimbriae-like organelles was observed on the bacterial surface. Microvilli of ileal Peyer's patches (PP) became disorganized, and intestinal mucus secretion increased which coincided with intraluminal binding of the pathogen in the proximal colon. Forty-eight hours after infection, there was conspicuous lack of fimbriae-like organelle expression, while bacterial adherence preferentially occurred at the domed villi of PP. Seventy-two hours after infection, broad morphological heterogeneity was noted within pedestals beneath attached bacteria, including extended pseudopods. We conclude that REPEC O103 express surface organelles during initial exposure to the host, that the initial target sites of adherence are the domed villi of ileal PP, and that increased mucus secretion occurs during REPEC O103 infection. As well, extended pseudopod formation was demonstrated in vivo.

Animals↗
PubMed2000

Microbial demetallization of crude oil: nickel protoporphyrin disodium as a model organo-metallic substrate.

A soil isolate designated as YA-1 strain was selected for its ability to degrade nickel protoporphyrin disodium (NiPPDS). The strain was capable of utilizing NiPPDS as the sole source of carbon. This strain, a gram-negative aerobic rod, was identified as Pseudomonas azelaica YA-1 based on the result of its 16S rRNA analysis. Product analyses by HPLC showed that this strain can decompose the porphyrin ring to which a metal ion is bound. However, the use of whole bacterial cells cannot result in extensive NiPPDS degradation; therefore, the YA-1 enzyme was extracted and purified. This NiPPDS-degrading enzyme named as protoporphyrinase was purified from P. azelaica YA-1 by ammonium sulfate fractionation and sequential chromatographies using DEAE Toyopearl 650 M, CM Toyopearl 650 M and Biogel P-60 columns, with a yield of 11.3% based on the enzyme activity and an overall purification of 498-fold. The molecular weight of this enzyme is estimated to be 39,000 Da by SDS-PAGE and 34,000 Da by gel filtration. The optimum pH and temperature for the enzyme were 7.0 and 30 degrees C, respectively. The activity was stable at pH 2.0-11.0 and at temperatures below 50 degrees C. The enzyme activity was inactivated by ferric chloride, potassium ferricyanide, ZnCl2 and CdCl2.

Journal Article↗
PubMed2000

Glycosphingolipid depletion in fabry disease lymphoblasts with potent inhibitors of glucosylceramide synthase.

BACKGROUND: Fabry disease is an inherited X-linked disorder resulting in the loss of activity of the lysosomal hydrolase alpha-galactosidase A and causing the clinical manifestations of renal failure, cerebral vascular disease, and myocardial infarction. The phenotypic expression of this disorder is manifest by the accumulation of glycosphingolipids containing alpha-galactosyl linkages, most prominently globotriaosylceramide. METHODS: Based on quantitative structure activity studies, we recently reported two newly designed glucosylceramide synthase inhibitors based on 1-phenyl-2-palmitoylamino-3-pyrrolidino-1-propanol (P4). These inhibitors, 4'-hydroxy-P4 and ethylenedioxy-P4, were evaluated for their ability to deplete globotriaosylceramide and other glucosylceramide-based lipids in Fabry lymphocytes and were compared with N-butyldeoxynojirimycin, another reported glucosylceramide synthase inhibitor. RESULTS: Concentrations as low as 10 nmol/L of 4'-hydroxy-P4 and ethylenedioxy-P4 resulted in 70 and 80% depletion, respectively, of globotriaosylceramide, with maximal depletion occurring at three days of treatment. There was no impairment of cell growth. In contrast, N-butyldeoxynojirimycin only minimally lowered globotriaosylceramide levels, even at concentrations as high as 10 micromol/L. Globotriaosylceramide depletion was confirmed by the loss of binding of FITC-conjugated verotoxin B subunit to the lymphoblasts. CONCLUSIONS: These findings suggest that selective glucosylceramide synthase inhibitors are highly effective in the depletion of globotriaosylceramide from Fabry cell lines. We suggest that these compounds have potential therapeutic utility in the treatment of Fabry disease.

1-Deoxynojirimycin↗
PubMed2000

Segmented filamentous bacteria prevent colonization of enteropathogenic Escherichia coli O103 in rabbits.

Despite their distribution in the intestines of many mammals, including man, segmented filamentous bacteria (SFB) have not been found in rabbits, nor has any function been identified for these uncultivable microbes. New Zealand White rabbits were infected with rabbit enteropathogenic Escherichia coli O103 (REPEC O103) derivatives, followed up clinically, and randomly killed 1-4 days after inoculation. Intestinal tissue samples were examined by electron and light microscopy to search for SFB and to evaluate REPEC O103 colonization. Twelve of 21 rabbits showed SFB colonization on ileal absorptive villi. The presence of SFB was correlated with lack of REPEC 0103 ileal colonization (P<.01) and disease. Rabbits without SFB were always colonized by this pathogen. SFB appear to inhibit intestinal colonization by REPEC O103 and thus protect against REPEC 0103 disease. SFB colonization in rabbits is also described for the first time.

Animals↗
PubMed2000

omega-hydroxylation activity toward leukotriene B(4) and polyunsaturated fatty acids in the human hepatoblastoma cell line, HepG2, and human lung adenocarcinoma cell line, A549.

The addition of glucose to the culture medium of HepG2 or A549 cells for 22 h caused a dose-dependent increase in leukotriene B(4) omega-hydroxylation activity in the homogenate. The addition of genistein to the culture medium of HepG2 or A549 cells for 22 h caused a dose-dependent decrease in the activity, although the number of living cells was not influenced by the addition of genistein. The inhibition by genistein was reversed by removal of genistein from the culture medium in 22 h. The specific leukotriene B(4) omega-hydroxylation activity was high in the nuclear envelope fraction of HepG2 or A549 cells, and a large portion of the activity was concentrated in the nuclear envelope fraction. In the nuclear envelope fraction, leukotriene B(4) omega-hydroxylation activity was accompanied by high polyunsaturated fatty acid omega-hydroxylation activity. The apparent K(m) values for arachidonic acid and leukotriene B(4) in the fractions of HepG2 or A549 cells were 25 and 50 microM, or 22 and 66 microM, respectively. The V(max) values were 222 and 104 pmol/min/mg protein, or 175 and 370 pmol/min/mg protein, respectively. NADPH-dependent omega-hydroxylation of LTB(4) in the nuclear envelope fraction of HepG2 or A549 cells was strongly inhibited by metyrapone and CO. The expression of cytochrome P450 4F2 mRNAs was detected in HepG2 and A549 cells, and thus the arachidonic acid and leukotriene B(4) omega-hydroxylation activities in the nuclear envelope fractions of HepG2 and A549 cells are likely due to cytochrome P450 4F2.

Adenocarcinoma↗
PubMed2000

Functional analysis of the type III secretion system in enteropathogenic Escherichia coli O157:H45.

A mass outbreak of Escherichia coli O157:H45 was first reported in Japan in 1998. This pathogen was classified as an enteropathogenic E. coli (EPEC) O157 because it was characterized by the Shiga toxin gene (stx)-negative and bundle-forming pilus (bfp) gene-positive genotypes. In this study, we investigated the type III secretion system in EPEC O157. Although no type III secreted proteins, Esps (E. colisecreted proteins), in EPEC O157:H45 were detectable in culture supernatant, secreted proteins were induced by the introduction of an EPEC plasmid-encoded regulator, per. In further contrast to EHEC O157:H7, EPEC O157:H45 triggered the accumulation of tyrosine phosphorylated proteins beneath the adherent bacteria. These results suggest that regulation of the type III secretion apparatus and host signal transduction events between E. coli O157:H45 and O157:H7 are completely different.

Adhesins, Bacterial↗
PubMed2000

Role of EspB in experimental human enteropathogenic Escherichia coli infection.

Enteropathogenic Escherichia coli (EPEC), a leading cause of diarrhea among infants in developing countries, induces dramatic alterations in host cell architecture that depend on a type III secretion system. EspB, one of the proteins secreted and translocated to the host cytoplasm via this system, is required for numerous alterations in host cell structure and function. To determine the role of EspB in virulence, we conducted a randomized, double-blind trial comparing the ability of wild-type EPEC and an isogenic DeltaespB mutant strain to cause diarrhea in adult volunteers. Diarrhea developed in 9 of 10 volunteers who ingested the wild-type strain but in only 1 of 10 volunteers who ingested the DeltaespB mutant strain. Marked destruction of the microvillous brush border adjacent to adherent organisms was observed in a jejunal biopsy from a volunteer who ingested the wild-type strain but not from two volunteers who ingested the DeltaespB mutant strain. Humoral and cell-mediated immune responses to EPEC antigens were stronger among recipients of the wild-type strain. In addition, four of the volunteers who ingested the wild-type strain had lymphoproliferative responses to EspB. These results demonstrate that EspB is a critical virulence determinant of EPEC infections and suggest that EspB contributes to an immune response.

Adolescent↗
PubMed2000

Reduction of globotriaosylceramide in Fabry disease mice by substrate deprivation.

We used a potent inhibitor of glucosylceramide synthase to test whether substrate deprivation could lower globotriaosylceramide levels in alpha-galactosidase A (alpha-gal A) knockout mice, a model of Fabry disease. C57BL/6 mice treated twice daily for 3 days with D-threo-1-ethylendioxyphenyl-2-palmitoylamino-3-pyrrolidi no-propanol (D-t-EtDO-P4) showed a concentration-dependent decrement in glucosylceramide levels in kidney, liver, and spleen. A single intraperitoneal injection of D-t-EtDO-P4 resulted in a 55% reduction in renal glucosylceramide, consistent with rapid renal glucosylceramide metabolism. A concentration-dependent decrement in renal and hepatic globotriaosylceramide levels was observed in alpha-Gal A(-) males treated for 4 weeks with D-t-EtDO-P4. When 8-week-old alpha-Gal A(-) males were treated for 8 weeks with 10 mg/kg twice daily, renal globotriaosylceramide fell to below starting levels, consistent with an alpha-galactosidase A-independent salvage pathway for globotriaosylceramide degradation. Complications observed with another glucosylceramide synthase inhibitor, N-butyldeoxynojirimycin, including weight loss and acellularity of lymphatic organs, were not observed with D-t-EtDO-P4. These data suggest that Fabry disease may be amenable to substrate deprivation therapy.

1-Deoxynojirimycin↗
PubMed2000

Antitachycardia burst pacing for pleomorphic reentrant ventricular tachycardias associated with non-coronary artery diseases: a morphology specific programming for ventricular tachycardias.

To study the role of antitachycardia burst pacing in patients with reentrant pleomorphic ventricular tachycardia (VT) associated with non-coronary artery diseases, the efficacy of antitachycardia pacing and appropriate antitachycardia pacing cycle length were evaluated in each pleomorphic VT morphology of seven patients. Seven patients were included in this study. Clinically documented pleomorphic VTs were reproduced in an electrophysiologic study. For each VT, rapid ventricular pacing was attempted from the apex of the right ventricle at a cycle length which was 20 ms shorter than that of VT and repeated after a decrement of the cycle length in steps of 10 ms until the VT was terminated or accelerated. All 16 VTs could be entrained by the rapid pacing, and 13 of the 16 VTs (81%) were terminated, whereas pacing-induced acceleration was observed in the other 3 VTs of the 3 patients. VT cycle length (VTCL), block cycle length (BCL) which was defined as the longest VT interrupting paced cycle length, %BCL/VTCL and entrainment zone which was defined as VTCL minus BCL, varied in each VT morphology of each patient. In two patients, antitachycardia pacing was effective in all VT morphologies and the maximum difference of the %BCL/VTCL among the pleomorphic VTs was less than 10%. Thus, antitachycardia pacing seemed to be beneficial for these patients. In the other 5 patients, a difference of more than 10% in %BCL/VTCL was observed among the pleomorphic VT morphologies and/or at least one VT morphology showed pacing-induced acceleration. Compared to the 13 terminated VTs, three accelerated VTs had a wide entrainment zone [160 +/- 44 vs 90 +/- 48 ms, p < 0.04] and small %BCL/VTCL [61 +/- 6 vs 77 +/- 11%,p<0.03]. In pleomorphic VTs associated with non-coronary artery diseases, responses to rapid pacing was not uniform; VT might be terminable or accelerated even in the same patient. We need to pay close attention when programming antitachycardia pacing in patients with pleomorphic VT.

Accelerated Idioventricular Rhythm↗
PubMed2000

[The current status and future applications of gene therapy and immunogene therapy for malignant lymphoma].

The present therapy for malignant lymphoma including stem cell transplantation, has been greatly developed. However, treatments still remain ineffective for many patients. Gene therapy is providing new strategies for the treatments of malignant lymphoma. There are three major approaches; 1) killing the tumor cell itself by introducing anti-sense genes against oncogene, tumor suppressor genes or drug-sensitive genes. 2) modifying the immune response by introducing genes that will trigger anti-tumor response or tumor specific genes to antigen presenting cell. 3) decreasing the sensitivity of hemopoietic cells by introducing drug resistance genes. We describe here the current and future applications of gene therapy for malignant lymphoma.

Antibodies, Anti-Idiotypic↗
PubMed2000

[Ambulatory blood pressure monitoring in hypertensive CAPD patients].

The periodic structure of 24-hour blood pressure variation(circadian rhythm of blood pressure by ambulatory blood pressure monitoring(ABPM) in hypertensive CAPD patients was investigated by a new method of analysis based upon the maximum entropy method(MEM). In addition, this method allows the adequacy of antihypertensive therapies to be evaluated in such patients. The results were as follows; 1) The frequency of non-dipper type hypertension was 88%(36/41 cases), and the remaining 12% (5/41) were dipper type hypertension patients. The rise in morning blood pressure(morning surge: MS) was noted in 64% of the former. 2) Night time systolic blood pressure(182 +/- 22 mmHg, n = 36) was higher in patients with non-dipper type hypertension than in those with the dipper type(151 +/- 17 mmHg, n = 5, p < 0.01). 3) The standardized level of systolic blood pressure(SLSBP) calculated by MEM analysis in patients with non-dipper type hypertension(177 +/- 7 mmHg) was comparable with that in those with dipper type hypertension(168 +/- 13 mmHg, ns). 4) Treatment with long-acting Ca antagonist alone significantly reduced both SLSBP and the area over the SLSBP from 188 +/- 18 mmHg to 160 +/- 7 mmHg(p < 0.01, n = 8), and area over the SLSBP from 2,735 +/- 340 mmHg.hr to 1,945 +/- 298 mmHg.hr(p < 0.01, n = 8). 5) In addition to long-acting Ca antagonist, administration of alpha 1-blocker given at bed time was significantly efficacious in reducing the rise in morning blood pressure, MS. The present study using MEM analysis of ABPM suggests that the blood pressure profile of hypertensive CAPD patients is characterized by a non-dipper type dominance and a frequent morning surge. Furthermore, the combined therapy with long-acting Ca antagonist and alpha 1-blocker was substantially effective both in reducing the overall blood pressure level, and in inhibiting the MS. This combined antihypertensive therapy may be potentially useful to prevent CAPD patients from the future development of cardiovascular complications.

Adrenergic alpha-Antagonists↗
PubMed1999

Improved inhibitors of glucosylceramide synthase.

Previous work has led to the identification of inhibitors of glucosylceramide synthase, the enzyme catalyzing the first glycosylation step in the synthesis of glucosylceramide-based glycosphingolipids. These inhibitors have two identified sites of action: the inhibition of glucosylceramide synthase, resulting in the depletion of cellular glycosphingolipids, and the inhibition of 1-O-acylceramide synthase, resulting in the elevation of cell ceramide levels. A new series of glucosylceramide synthase inhibitors based on substitutions in the phenyl ring of a parent compound, 1-phenyl-2-palmitoylamino-3-pyrrolidino-1-propanol (P4), was made. For substitutions of single functional groups, the potency of these inhibitors in blocking glucosylceramide synthase was primarily dependent upon the hydrophobic and electronic properties of the substituents. An exponential relationship was found between the IC50 of each inhibitor and the sum of derived hydrophobic (pi) and electronic (sigma) parameters. This relationship demonstrated that substitutions that increased the electron-donating characteristics and decreased the lipophilic characteristics of the homologues enhanced the potency of these compounds in blocking glucosylceramide formation. A novel compound was subsequently designed and observed to be even more active in blocking glucosylceramide formation. This compound, D-threo-4'-hydroxy-P4, inhibited glucosylceramide synthase at an IC50 of 90 nM. In addition, a series of dioxane substitutions was designed and tested. These included 3',4'-methylenedioxyphenyl-, 3',4'-ethylenedioxyphenyl-, and 3'4'-trimethylenedioxyphenyl-substituted homologues. D-threo-3', 4'-Ethylenedioxy-P4-inhibited glucosylceramide synthase was comparably active to the p-hydroxy homologue. 4'-Hydroxy-P4 and ethylenedioxy-P4 blocked glucosylceramide synthase activity at concentrations that had little effect on 1-O-acylceramide synthase activity. These novel inhibitors resulted in the inhibition of glycosphingolipid synthesis in cultured cells at concentrations that did not significantly raise intracellular ceramide levels or inhibit cell growth.

Cells, Cultured↗
PubMed1999

Successful culture and sustainability in vivo of gene-modified human oral mucosal epithelium.

Human oral mucosal cells are an attractive site for tissue engineering because they are the most accessible cells in the body and easy to manipulate in vitro. They thus have possibilities for targeting by somatic gene therapy. We examined the efficiency of retrovirus-mediated gene transfer and the construction of mucosal epithelium in vivo. Human oral mucosal cells were transduced with a retroviral vector carrying the lacZ gene at high efficiency and constructed epithelium after G418 selection with 3T3 cells in vitro. The cultured oral mucosal epithelium membrane was then grafted onto immunodeficient mice. Beta-Gal expression was detected histochemically in vivo 5 weeks after grafting. Furthermore, we transduced factor IX cDNA into the mucosal epithelium membrane, and it was then transplanted into nude mice. Between 0.6 and 1.8 ng of human factor IX per milliliter was found in mouse plasma, and the production was continued for 23 days in vivo. These results confirmed that the oral mucosal epithelium is an ideal target tissue for gene therapy or tissue engineering.

Animals↗
PubMed1999

Distribution of 1,4-dioxane in relation to possible sources in the water environment.

The distribution of 1,4-dioxane was investigated in surface and groundwater in relation to possible sources. Water samples from river, coastal sea, and groundwater were collected and examined in Kanagawa Prefecture, Japan, from 1995 to 1998. Almost all samples were contaminated by 1,4-dioxane. On annual changes, the concentrations did not vary significantly in river water. However, some samples were still heavily polluted in groundwater. The highest correlation was between the concentration of 1,4-dioxane and 1,1,1-trichloroethane in the polluted groundwater samples. Soil and groundwater that was polluted by 1,1,1-trichloroethane in the past is still a possible source of 1,4-dioxane. In regard to a survey of other sources, effluents from the combined collection treatment from apartment houses, river basin sewage systems, and chemical plants were examined. The load unit of 1,4-dioxane from the treated effluent of domestic wastewater was assumed to be approximately 0.25 mg/person/day. This was derived from commonly used household chemicals, which contained 1,4-dioxane as a by-product. In river basin sewage systems, the concentrations in effluents varied significantly. High concentration inflows were probably not degraded and released. Although 50-60% of removal was recognized in acclimatized activated sludge from a chemical plant, 1,4-dioxane was easily desorbed with water from sludge.

Dioxanes↗