Biomedical subjects

K Murakami

Publications and source records attributed to K Murakami.

At least 199 records · Page 11Linked to original sources

PubMed1999

Cloning and characterization of a novel membrane-associated antigenic protein of Helicobacter pylori.

Infection by Helicobacter pylori, a noninvasive bacterium, induces chronic leukocyte infiltration in the stomach by still largely unknown molecular mechanisms. We investigated the possibility that a membrane protein of H. pylori induces an inflammatory reaction in the subepithelial tissue of the stomach. By generating an expression library of H. pylori chromosomal DNA and screening with rabbit antiserum raised to a membrane fraction of H. pylori and sera of infected patients, we cloned a 16.0-kDa protein (HP-MP1) which appeared to attach to the inner membrane of the H. pylori in a homodimeric form. Anti-HP-MP1 antibodies were detected in the sera of infected patients but not in those of uninfected controls. Coincubation of monocytes with recombinant HP-MP1 led to cell activation and production of interleukin-1alpha (IL-1alpha), tumor necrosis factor alpha, IL-8, and macrophage inflammatory protein 1alpha. The results indicate that HP-MP1 is an antigenic membrane-associated protein of H. pylori which potentially activates monocytes. This suggests that HP-MP1 may play roles in the pathogenesis of perpetual tissue inflammation associated with H. pylori infection.

Antibodies, Bacterial↗

PubMed1999

Cra-dependent transcriptional activation of the icd gene of Escherichia coli.

The icd gene of Escherichia coli, encoding isocitrate dehydrogenase, was shown to be expressed from two different promoters: the previously identified icd P1 and a newly detected second promoter, icd P2, whose expression is positively regulated by the catabolite repressor-activator protein Cra, formerly called FruR. In each case, we determined the mRNA start site by primer extension analysis of in vivo transcripts and examined the interaction of the icd control region with either RNA polymerase or Cra. We observed that (i) the Cra factor binds to and activates transcription from a site centered at position -76.5 within the icd P2 promoter region and (ii) three particular mutations in the C-terminal end of the alpha subunit of RNA polymerase (L262A, R265A, and N268A) considerably diminish transcription initiating from the icd P2 promoter, as shown by in vitro experiments performed in the presence of mutant RNA polymerases carrying Ala substitutions.

Amino Acid Substitution↗

PubMed1999

Clustering and detection of 5' splice sites of mRNA by k weight-matrices model.

We propose a novel method to detect 5' splice sites of eukaryotic mRNA. We have grouped the 5' splice splice sites into various classes. The clustered sites are represented by a set of PWMs. The clustering algorithm is similar to k-means clustering algorithm but the distance definition and the training score function were arranged. The clustered PWMs were applied to 5' splice site detection. The results showed an improvement in comparison with traditional single PWM. The result of the clusters suggests there are new motifs of 5' splice sites.

Algorithms↗

PubMed1999

Biological roles of angiotensin II via its type 2 receptor during rat follicle atresia.

Type 1 angiotensin II (ANG II) receptors play crucial roles in the regulation of blood pressure and fluid osmolarity, whereas the physiological roles of type 2 ANG II receptors (AT2) remain unclear. Because AT2 is expressed in atretic follicles where granulosa cells undergo apoptosis, we examined the space and time relationship between AT2 expression and follicle atresia in vivo and the effect of AT2 on follicle-stimulating hormone (FSH) actions in vitro. Binding studies, autoradiography, and RT-PCR of AT2 revealed that the AT2 content in granulosa cells was time dependently increased at both protein and mRNA levels in equine chorionic gonadotropin-treated immature female rats. This increase paralleled the progression of atresia. ANG II suppressed FSH-caused prevention of DNA fragmentation, increases in luteinizing hormone receptor content, and estrogen production through AT2 in cultured granulosa cells. Moreover, FSH-induced stimulation of extracellular signal-regulated kinase activity, critical for cell survival, was inhibited by AT2 stimulation. These results suggest that AT2 mediates the progression of follicle atresia through granulosa cell apoptosis by inhibiting FSH actions.

Angiotensin II↗

PubMed1999

Exacerbation of delayed cell injury after transient global ischemia in mutant mice with CuZn superoxide dismutase deficiency.

BACKGROUND AND PURPOSE: We have demonstrated that copper-zinc superoxide dismutase (CuZn-SOD), a cytosolic isoenzyme of SODs, has a protective role in the pathogenesis of superoxide radical-mediated brain injury. Using mice bearing a disruption of the CuZn-SOD gene (Sod1), the present study was designed to clarify the role of superoxide anion in the pathogenesis of selective vulnerability after transient global ischemia. METHODS: Sod1 knockout homozygous mutant mice (Sod1 -/-) with a complete absence of endogenous CuZn-SOD activity, heterozygous mutant mice (Sod1 +/-) with a 50% decrease in the activity, and littermate wild-type mice (male, 35 to 45 g) were subjected to global ischemia. Since the plasticity of the posterior communicating artery (PcomA) has been reported to influence the outcome of hippocampal injury, we assessed the relation between the plasticity of PcomAs and the decrease of regional cerebral blood flow in global ischemia. RESULTS: The fluorescence intensity of hydroethidine oxidation, a measurement of ethidium fluorescence for superoxide radicals, was increased in mutant mice 1 day after both 5 and 10 minutes of global ischemia, compared with wild-type mice. Hippocampal injury in the PcomA hypoplastic brains showed significant exacerbation in mutant mice compared with wild-type littermates 3 days after 5 minutes of global ischemia, although a marked difference was not observed at 1 day. CONCLUSIONS: These data suggest that superoxide radicals play an important role in the pathogenesis of delayed injury in the vulnerable hippocampal CA1 subregion after transient global ischemia.

Animals↗

PubMed1999

Angiotensin II plays a pathogenic role in immune-mediated renal injury in mice.

Several lines of evidence show the importance of angiotensin II (AII) in renal injuries, especially when hemodynamic abnormalities are involved. To elucidate the role of AII in immune-mediated renal injury, we studied anti-glomerular basement membrane (GBM) nephritis in AII type 1a receptor (AT1a)-deficient homozygous (AT1a-/-) and wild-type (AT1a+/+) mice. A transient activation of the renin-angiotensin system (RAS) was observed in both groups of mice at around day 1. A renal expression of monocyte chemoattractant protein-1 (MCP-1) was transiently induced at six hours in both groups, which was then downregulated at day 1. In the AT1a+/+ mice, after RAS activation, the glomerular expression of MCP-1 was exacerbated at days 7 and 14. Thereafter, severe proteinuria developed, and the renal expressions of transforming growth factor-beta1 (TGF-beta1) and collagen type I increased, resulting in severe glomerulosclerosis and interstitial fibrosis. In contrast, glomerular expression of MCP-1, proteinuria, and tissue damage were markedly ameliorated in the AT1a-/- mice. Because this amelioration is likely due to the lack of AT1a, we can conclude that AII action, mediated by AT1a, plays a pathogenic role in anti-GBM nephritis, in which AII may contribute to the exacerbation of glomerular MCP-1 expression. These results suggest the involvement of AII in immune-mediated renal injuries.

Angiotensin II↗

PubMed1999

Retinol esterification activity contributes to retinol transport in stellate cells.

The mechanisms of retinol transport and accumulation in hepatic stellate cells (HSC) remain to be elucidated. Our previous studies suggested that retinol esterification activity, particularly lecithin:retinol acyltransferase (LRAT) activity, in liver retinoid metabolism is important to elucidate the relationship between retinol uptake by HSC and the esterification of retinol. In the present study, using a human HSC-like cell line, LI90, we demonstrated that retinol esterification activity of LI90 cells is similar to that of primary cultures of rat HSC and higher than that of a human hepatoma cell line. Further, since progesterone or diphospho-lauroyl-phosphatidylcholine increased retinol esterification activity of LI90 cells, it is likely that LRAT contributes to retinol esterification in LI90. We examined retinol esterification in LI90 cells and clearance of retinol from culture medium. The percentages of both retinol and esterified retinol in LI90 cells increased in a manner dependent on retinol concentration in medium, whereas that of retinol in medium decreased. The percentages of esterified and unesterified retinol in LI90 cells and of retinol in medium were linearly dependent on the logarithm of the initial concentration of retinol in the medium. These results suggest that retinol esterification activity contributes to retinol uptake by HSC and maintenance of non-toxic retinol levels in plasma.

Actins↗

PubMed1999

Inhibitory mechanisms of glycoprotein fraction derived from Miscanthus sinensis for the immediate phase response of an IgE-mediated cutaneous reaction.

We investigated the inhibitory effect of the glycoprotein fraction (fraction 2) extracted from Miscanthus sinensi ANDERSSON (M. sinensis) on biphasic cutaneous reactions in mice passively sensitized with IgE. Biphasic skin reactions with peak responses at 1 (IPR, immediate phase reaction) and 24 h (LPR, late phase reaction) were caused by passive sensitization with an anti-dinitrophenol IgE monoclonal antibody (anti-DNP IgE mAb) followed by an epicutaneous challenge of 0.1% dinitrofluorobenzene (DNFB) in 100% ethanol. Intraperitoneal injection of fraction 2 before the DNFB challenge significantly inhibited the biphasic ear swelling response in passively sensitized mice in a dose-dependent manner (1-30 mg/kg). We also found that fraction 2 was effective at inhibiting the vascular permeability in mouse ear induced by an injection of compound 48/80, histamine or serotonin. In addition, fraction 2 inhibited scratching behavior as well as ear edema observed within 2 h after DNFB challenge. Marked inhibition was observed in both passively sensitized and non-sensitized mice. The locomotor activity of mice was also reduced by the administration of fraction 2 as well as by diphenhydramine. These results suggest that the inhibitory effect of glycoprotein fraction 2 of M. sinensis on an IgE-mediated allergic inflammatory reaction is due to the protection of mediator-induced vascular permeability and that in addition to the inhibition of an inflammatory reaction, a sedative action is responsible for the inhibition of allergy-induced scratching responses.

Acute-Phase Reaction↗

PubMed1999

Epidemiological analysis of Salmonella enteritidis from human outbreaks by pulsed-field gel electrophoresis.

To determine the extent of genetic diversity among isolates of Salmonella enteritidis obtained from outbreaks in Fukuoka Prefecture, Japan, from 1989 to 1994, we analyzed a total of 55 isolates of S. enteritidis obtained from 13 distinct outbreaks with pulsed-field gel electrophoresis. These isolates showed three different patterns in pulsed-field profile with Bln I digestion. The groups A, B and C consisted of three outbreaks isolates (Dice coefficient, F = 1), of seven outbreaks (F = 0.56-0.94) and of three outbreaks (F = 0.65-0.78), respectively. This result suggests that a few limited clonal lines of S. enteritidis were successively causing outbreaks in this area from 1989 to 1994.

Animals↗

PubMed1999

Vascular remodeling in hypertensive transgenic mice.

We physiologically and histopathologically analyzed vascular damage due to hypertension and vascular remodeling in hypertensive transgenic mice (Tsukuba hypertensive mice; THM). Pubertal (6-week-old) THM already had hypertension similar to blood pressure in adult THM due to an enhanced renin angiotensin system (RAS). They progressively developed remarkable vascular hypertrophy composed of dedifferentiation of vascular smooth muscle cells (VSMCs) and extracellular matrix accumulation in the thoracic aorta, and VSMC hyperplasia was predominant in the abdominal aorta. THM are therefore a useful animal model for studying vascular remodeling mediated by enhanced RAS.

Aldosterone↗

PubMed1999

Effective delivery of a lipophilic 6-chloro-2',3'-dideoxyguanosine (6-Cl-ddG) into rat lymphoid tissues.

Lipophilic 6-chloro-2',3'-dideoxyguanosine (6-Cl-ddG) was evaluated for its improved lymph node delivery by comparison with the parental nucleoside (ddG) in vitro and in vivo. The in vitro studies with rat plasma, lymph node homogenate and stomach content indicated that 6-Cl-ddG converted to ddG more effectively in the lymph node homogenate and that 6-Cl-ddG was more stable than ddG in the stomach content. In an in vivo study, plasma and lymph nodes were collected from rats after a subcutaneous or oral administration of 6-Cl-ddG or ddG. With the subcutaneous administrations of the drugs, the area under the concentration time-curve (AUC) value in the plasma for converted ddG following a 6-Cl-ddG administration was less than half the value for ddG following a ddG administration but the converted ddG AUC values in the lymph nodes due to 6-Cl-ddG administration were 1.4- to 2.0-fold higher than the ddG AUC values due to ddG administration. Moreover, with the oral administrations, the converted ddG AUC value in plasma after a 6-Cl-ddG administration was 3-fold higher than ddG after a ddG administration, and high levels of converted ddG were detected in the lymph nodes, but no ddG was detected in the lymph node following ddG administration. These results suggest that lipophilic 6-Cl-ddG is a useful prodrug for delivering ddG into the lymph nodes by oral administration.

Administration, Oral↗

PubMed1999

Tissue-specific changes of type 1 angiotensin II receptor and angiotensin-converting enzyme mRNA in angiotensinogen gene-knockout mice.

This study examined whether type 1 angiotensin II receptor (AT1) and angiotensin-converting enzyme (ACE) mRNAs are regulated during dietary salt loading in angiotensinogen gene-knockout (Atg-/-) mice which are genetically deficient in endogenous production of angiotensin II. Wild-type (Atg+/+) and Atg-/- mice were fed a normal-salt (0.3% NaCl) or a high-salt (4% NaCl) diet for 2 weeks. The mRNA levels were measured by Northern blot analysis. In Atg+/+ mice, concentrations of plasma angiotensin peptides were decreased by salt loading, whereas the treatment increased the brainstem, cardiac, pulmonary, renal cortex, gastric and intestinal AT1 mRNA levels. Salt loading also enhanced renal cortex ACE mRNA levels in Atg+/+ mice. Although plasma angiotensin peptides and urinary aldosterone excretion were not detected in Atg-/- mice, salt loading increased blood pressure in Atg-/- mice. In Atg-/- mice, pulmonary, renal cortex, gastric and intestinal AT1, and renal cortex and intestinal ACE mRNA levels were higher than those in Atg+/+ mice. However, salt loading upregulated AT1 mRNA expression only in the liver of Atg-/- mice, and the treatment did not affect ACE mRNA levels in Atg-/- mice. Furthermore, although the levels of ACE enzymatic activity showed the same trend with the ACE mRNA levels in the lung, renal cortex and intestine of both Atg-/- and Atg+/+ mice, the results of radioligand binding assay showed that cardiac expression of AT1 protein was regulated differently from AT1 mRNA expression both in Atg-/- and Atg+/+ mice. Thus, expression of AT1 and ACE is regulated by salt loading in a tissue-specific manner that appears to be mediated, at least partly, by a mechanism other than changes in the circulating or tissue levels of angiotensin peptides.

Angiotensin II↗

PubMed1999

Periodontal and computer tomography scanning evaluation of endosseous implants in conjunction with sinus lift procedure. A 6-case series.

The aim of the present study was to evaluate endosseous implants in conjunction with the sinus lift procedure using periodontal parameters and computed tomography (CT) scanning. In 6 patients, 8 sinus lift procedures were performed to the atrophic posterior maxilla. Fourteen endosseous implants were placed. The condition of the peri-implant mucosa was assessed postoperatively at an average of 31 months using periodontal parameters such as plaque index, probing depth, clinical attachment level, and bleeding on probing. All the implants were maintained in a clinically healthy condition throughout the observation period. CT images revealed the difference in the morphology of the sinus floor between the locations of the block-shaped bone and the small pieces of bone which were both grafted. The results indicated that endosseous implants in conjunction with the sinus lift procedure could be maintained by monitoring the periodontal parameters.

Adult↗

PubMed1999

Effects of ascorbic acid on peroxidation of human erythrocyte membranes by lipoxygenase.

The effects of ascorbic acid (AsA) on membrane phospholipids (PLs) and tocopherols (Tocs) of human erythrocyte during peroxidation by soybean lipoxygenase (LOX) were investigated. After extraction of the membrane lipids, alpha-tocopherol (alpha-Toc), gamma-tocopherol (gamma-Toc) and cholesterol were simultaneously measured by high-performance liquid chromatography (HPLC), and the changes of Tocs were expressed on the basis of cholesterol. The phospholipid classes and corresponding hydroperoxides (PL-OOHs) were detected simultaneously by HPLC, and the changes were calculated on the basis of sphingomyelin. These methods are sensitive to the changes of membrane Tocs and PLs by peroxidation. Control incubation without LOX and AsA was done for 45 min at 30 degrees C. After the incubation with LOX, alpha- and gamma-Tocs were exhausted, PLs decreased, and PL-OOHs and malondialdehyde (MDA) increased. Incubation with both AsA and LOX further increased MDA significantly, but it preserved about 30% of alpha-Toc and 45% of gamma-Toc of the control levels and did not decrease PLs or PL-OOHs from the levels after the incubation with LOX. Subsequent incubation with AsA for 45 min after the incubation with LOX (after Tocs were exhausted) showed a 240% increase in MDA, but it decreased PLs by only about 15% of the preincubation values and recovered gamma-Toc to about 13% of the control. The subsequent incubation with AsA after the control incubation increased PLs to higher than that of the control reaction. These results show that AsA protects and regenerates the membrane Tocs against enzymatic peroxidation. The results also indicate that repair of the membrane PLs is promoted in the presence of AsA.

Ascorbic Acid↗

PubMed1999

Expression and characterization of mouse angiotensin II type 1a receptor tagging hemagglutinin epitope in cultured cells.

The octapeptide angiotensin II mediates the physiological actions of the renin-angiotensin system through activation of several angiotensin II receptor (AT) subtypes, in particular AT1 (AT1a and AT1b in the case of rodents). Although we and others have generated mutant mice in which the AT1a gene was disrupted, the function of mouse AT1 remains to be fully elucidated, due to the lack of effective tools involving antibodies against AT1 for detecting biological responses in cellular conditions. To avoid these problems, we constructed the hemagglutinin (HA)-tagged mouse AT1a, and stably introduced this recombinant receptor into human embryonic kidney 293-T cells. Radioligand binding of [(125)I] angiotensin II to AT1a was specific, saturable, and reversible. Scatchard analysis demonstrated that the transfected receptor had a dissociation constant of 1.7 nM with a density of 1.2 x 10(5) sites/cells. Angiotensin II stimulated a rapid increase in cytosolic free calcium, and angiotensin II-induced phosphorylation of extracellular signal-regulated kinases (Erk) was found in a dose-dependent manner. After solubilization, Western blot analysis showed specific interactions between an anti-HA antibody and HA-tagged mouse AT1a. Furthermore, a significant proportion of HA-tagged mouse AT1a was specifically immunoprecipitated with this antibody. In the immunocytochemical and electronmicroscopic studies, treatment of this cell line with angiotensin II resulted in decrease in signals of the surface receptors. Based on these results, the cell line established here provides an excellent tool for studying angiotensin II actions mediated through mouse AT1a, at sub-nanomolar concentrations.

Angiotensin II↗

PubMed1999

[Antibiotic-resistant H. pylori strains in the last ten years in Japan].

An increase in the prevalence of antibiotic-resistant H. pylori strains has been reported. We tested the sensitivity of H. pylori strains to antibiotics before and after the eradication treatment to determine primary and secondary resistance by E-test. A total of 336 H. pylori strains from 1987 to 1998 were tested for sensitivity to clarithromycin (CAM), amoxicillin (AMPC), and metronidazole (MNZ). Primary resistance of H. pylori to CAM, MNZ, and AMPC were found in 9.5%, 26.8%, and 0.3% strains, respectively. The resistant strains to CAM and MNZ were gradually increasing in the last few years. The cure rates of triple therapy with MNZ and CAM for CAM-susceptible and CAM-resistant H. pylori isolates were 83.3% (40/48) and 30.0% (3/10) respectively, and the cure rates of triple therapy with CAM and AMPC for CAM-susceptible and CAM-resistant H. pylori isolates were 79.3% (69/87) and 12.5% (1/8) respectively. After triple therapy, secondary resistant strains to CAM and MNZ were found in 42.1% (8/19) and 31.6% (6/19) of the cases.

Amoxicillin↗

PubMed1999

Inhibition of angiogenesis and intrahepatic growth of colon cancer by TAC-101.

We demonstrated in this study that inhibition of intra-hepatic growth of colon cancer by TAC-101 is mediated by inhibition of angiogenesis. In vitro experiments showed that TAC-101 inhibited the proliferation of murine hepatic sinusoidal endothelial (HSE) cells induced by coculture with murine colon 26-L5 (L5) cells. HSE cell proliferation was also enhanced by conditioned medium of L5 cells (CM-L5), and this enhancement of proliferation was abrogated by anti-vascular endothelial growth factor antibody. CM-L5 also induced in vitro tube formation of HSE cells on Matri-gel, and this activity of CM-L5 was abrogated by TAC-101 in a concentration-dependent manner. On the other hand, p.o. administration of TAC-101 inhibited tumor-induced angiogenesis in vivo and decreased the weights of L5 tumors in the mouse liver. Reverse transcriptase-PCR analysis using in vivo tumor tissue suggested that repression of vascular endothelial growth factor expression by TAC-101 was associated with the antiangiogenic activity. TAC-101 alone and 5-fluorouracil (5-FU)/D,L-leucovorin (LV) significantly inhibited the intrahepatic growth of L5 tumors (P = 0.002 and 0.001, respectively), whereas 5-FU alone did not (P = 0.088). When TAC-101 was administered with 5-FU/LV, marked enhancement of antitumor activity was observed (95% inhibition; P<0.001). This enhanced antitumor effect was also observed in experiments using Co-3 human colon adenocarcinoma. Concurrent treatment with TAC-101 and 5-FU/LV and sequential treatment with 5-FU/LV followed by TAC-101 resulted in significant augmentation of antitumor activity against Co-3 (overall P = 0.007 and 0.015, respectively). These findings indicate that TAC-101 inhibits tumor angiogenesis and suggest that it may be effective against hepatic metastasis of colon cancer.

Adenocarcinoma↗

PubMed1999

[Chemoprophylaxis against Mycobacterium avium complex infection induced in mice].

M. avium complex (MAC) is one of the important causative agents of opportunistic infections among AIDS patients. Recent evidence showed that the entry of infection is through the gastrointestinal tract. In the present study, we compared the prophylactic effect of some antimicrobials against MAC infection induced in mice. Different groups of beige mice were fed with pellets containing 0.0067% (10 mg/kg) of KRM-1648, rifabutin (RFB) and clarithromycin (CAM). Seven days after the administration of drugs, the mice were infected with M. intracellular N-241 (5 x 10(8) CFU) orally, five times, every other day and killed one and 126 days after the last infection. The effect of drug was evaluated using the frequency and severity of gross lung lesions in the mice and by the total CFU recovered from the lungs and spleen. MAC infection was not likely to be established since there was no macroscopic evidence of lesion in organs and the recovery of cultures from lungs and spleen tested was negative, in 3 of 10 mice in the control group, 2 of 9 in the CAM group, 4 of 9 in the RFB group and 4 of 10 in the KRM group. These mice were excluded from the analysis of the study results. Thus, we examined 7 mice in the control group, 7 in the CAM group, 5 in the RFB group, and 6 in the KRM group. Tubercle-like lesions were observed in the lungs of all 7 mice in the control group (severity: 3+ in 5 mice; 4+ in 2 mice), in 5 of 7 mice (71%) in the CAM group (severity: 2+ in 1 mouse; 3+ in 4 mice), and in 4 of 5 mice (80%) in the RFB group (severity: 1+ in 1 mouse; 2+ in 1 mouse; 4+ in 2 mice), while only slight lesions (severity: 1+) were observed in 4 of 6 mice (67%) in the KRM group. There was no macroscopic evidence of lesion in spleen, liver and kidneys. The log CFU was determined at the next day of the completion of the last infection. The log CFU of the lungs was 2.49 and 2.28 in the control group and the CAM group, respectively. The bacteria were not recovered either from the lungs in the RFB and KRM groups, nor from the spleen in all the groups. The order of efficacy of the drugs on the basis of the CFUs recovered from the lungs and spleen in each group determined 126 days after the completion of the last infection was as follows; KRM > CAM > RFB in the lungs and KRM > CAM [symbol: see text] RFB in the spleen, although there was no significant difference among the three drugs (P < 0.05). However, the significantly preferable effect was obtained in the three drug groups as compared with the control group.

Animals↗