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Biomedical subjects

K Kume

Publications and source records attributed to K Kume.

At least 91 records · Page 5Linked to original sources
PubMed1996

Predominant expression of platelet-activating factor receptor in the rat brain microglia.

Cellular localization of platelet-activating factor (PAF) receptor in the rat brain was determined by (1) in situ hybridization, (2) Northern blot analysis in primary cell cultures of neurons, microglia, astrocytes, and fibroblasts, and (3) Ca2+ imaging in hippocampal culture. In situ hybridization revealed that the PAF receptor mRNA is expressed intensely in microglia and moderately in neurons. Northern blot analysis revealed that PAF receptor mRNA is the most abundant in microglia. In primary hippocampal cultures, PAF elevated intracellular Ca2+ concentration in microglia and also in neurons, but to a lesser extent. These results suggest predominant presence of PAF receptor in microglia. Cultured microglia also expressed cPLA2 mRNA the most intensely. PAF-activated microglia released arachidonic acid in a Ca(2+)-dependent manner and without conversion to its derivatives. We propose that microglia as well as neurons contribute to PAF-related events in the CNS by releasing arachidonic acid.

Animals↗
PubMed1996

Wortmannin inhibits mitogen-activated protein kinase activation by platelet-activating factor through a mechanism independent of p85/p110-type phosphatidylinositol 3-kinase.

We have shown previously that wortmannin partially inhibits mitogen-activated protein kinase (MAPK) activated by platelet-activating factor (PAF) in guinea pig neutrophils (Ferby, M. I., Waga, I., Sakanaka, C., Kume, K., and Shimizu, T. (1994) J. Biol. Chem. 269, 30485-30488). To identify whether p85-dependent phosphatidylinositol 3-kinase is a target molecule of wortmannin in this inhibitory process, we established a murine macrophage cell line (P388D1), inducibly expressing a dominant-negative p85, delta p85. Upon induction of delta p85 by isopropyl-beta-D-thiogalactopyranoside, PAF still induced unaltered activation of MAPK, which was inhibited completely by wortmannin and 1,2-bis-(O-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid acetoxymethyl ester in an additive manner. Thus, PAF activates MAPK in P388D1 cells via two distinct pathways, one calcium-dependent and another calcium-independent, but wortmannin-sensitive. The inhibition of calcium-independent activation of MAPK by wortmannin does not involve p85-dependent phosphatidylinositol 3-kinase.

Androstadienes↗
PubMed1996

Micro-trap phosphorylation assay of mitogen-activated protein (MAP) kinases to detect their activation by lipopolysaccharides.

We designed a microplate-based assay method for mitogen-activated protein (MAP) kinase. Using anion-exchanger resin, MAP kinases from murine macrophages were partially purified in 96-well plates. The activities of these purified enzymes correlated well with those detected in heretofore used assays. The micro-trap phosphorylation assay has advantages over conventional methods (immunoprecipitation, Western blotting for the detection of mobility shift, or kinase detection assay in myelin basic protein (MBP)-containing gel), in terms of sensitivity, economy and rapid execution for hundreds of samples. Using micro-trap phosphorylation assay, it was demonstrated that MAP kinase activities in macrophages were persistently increased by lipopolysaccharide (LPS) stimulation, and this activation was inhibited by polymyxin B or tyrosine kinase inhibitors. This method is expected to give a wide range of application, such as determining effects of drug inhibitors or antisense oligonucleotides on MAP kinases, or measuring the various protein kinases after specificity controls were done.

Animals↗
PubMed1996

A murine platelet-activating factor receptor gene: cloning, chromosomal localization and up-regulation of expression by lipopolysaccharide in peritoneal resident macrophages.

A murine gene encoding a platelet-activating factor receptor (PAFR) was cloned. The gene was mapped to a region of the D2.2 band of chromosome 4 both by fluorescence in situ hybridization and by molecular linkage analysis. Northern blot analysis showed a high expression of the PAFR message in peritoneal macrophages. When C3H/HeN macrophages were treated with bacterial lipopolysaccharide (LPS) or synthetic lipid A, the PAFR gene expression was induced. Bacterial LPS, but not lipid A, induced the level of PAFR mRNA in LPS unresponsive C3H/HeJ macrophages. These induction patterns were parallel to those of tumor necrosis factor-alpha mRNA. Thus the PAFR in macrophages is important in LPS-induced pathologies.

Animals↗
PubMed1996

cDNA cloning, expression, and mutagenesis study of leukotriene B4 12-hydroxydehydrogenase.

Leukotriene B4 12-hydroxydehydrogenase catalyzes the conversion of leukotriene B4 into its biologically less active metabolite, 12-oxo-leukotriene B4. This is an initial and key step of metabolic inactivation of leukotriene B4 in various tissues other than leukocytes. Here we report the cDNA cloning for porcine and human enzymes from kidney cDNA libraries. A full-length cDNA of the porcine enzyme contains an open reading frame consisting of 987 base pairs, corresponding to 329 amino acids. The human enzyme showed a 97.1% homology with the porcine enzyme. Northern blotting of human tissues revealed its high expression in the kidney, liver, and intestine but not in leukocytes. The porcine enzyme was expressed as a glutathione S-transferase fusion protein in Escherichia coli, which exhibited similar characteristics with the native enzyme. Because the enzymes have a homology, in part, with NAD(P)(+)-dependent alcohol dehydrogenases, a site-directed mutagenesis study was carried out. We found that three glycines at 152, 155, and 166 have crucial roles in the enzyme activity, possibly by producing an NADP+ binding pocket.

Alcohol Oxidoreductases↗
PubMed1996

Up-regulation of the intracellular Ca2+ signaling and mRNA expression of platelet-activating factor receptor by estradiol in human uterine endometrial cells.

Platelet-activating factor (PAF, 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine), a potent chemical mediator in inflammation, plays a role in reproduction. Using primary culture of human uterine endometrial cells, we investigated the effect of sex steroid hormones on the PAF-induced signal and its receptor mRNA expression. After a 24 hr treatment with estradiol, PAF increased the intracellular calcium ion ([Ca2+]i) in the glandular cells, but not in the stromal cells. This response was not observed in the non-treated cells, and was blocked by a PAF antagonist, WEB2086. Two types of mRNA (transcript 1 and transcript 2) occurred for PAF receptor by alternative splicing, which are under control of two distinct promoters. Using RT-PCR analysis, it was shown that both transcripts existed in endometrial cells and that estradiol alone or a combination of estradiol and progesterone induced the accumulation of transcript 2, the promoter of which responded to estrogen in our previous studies. The regulation of PAF receptor by sex steroid hormones in human uterine endometrial cells suggests that PAF is involved in the physiological process of reproduction.

Alternative Splicing↗
PubMed1996

PAF-induced MAPK activation is inhibited by wortmannin in neutrophils and macrophages.

In the present study we examined the mechanism by which PAF activates MAPK in native cells such as guinea-pig neutrophils and P388D1 macrophage-like cells. We found that PAF activates MAPK through two distinct pathways. One calcium-dependent pathway that likely involves cPKC, and another calcium-independent but wortmannin-sensitive pathway. Using molecular biological methods we are presently examining whether hetrodimeric (p85/p110) type PI 3-kinase is the actual target of wortmannin involved in PAF mediated activation of MAPK.

Androstadienes↗
PubMed1996

Platelet-activating factor and somatostatin activate mitogen-activated protein kinase (MAP kinase) and arachidonate release.

Platelet-activating factor (PAF) receptor and somatostatin receptor (SSTR4) were cloned, and their primary structures were identified. They are both highly expressed in the rat hippocampus. When expressed in Chinese hamster ovary cells, these receptors activated mitogen-activated protein (MAP) kinase cascade and phospholipase A2. Arachidonic acid or its derivatives, thus produced by the activation of these receptors may play some roles in synaptic transmission and synaptic plasticity.

Animals↗
PubMed1996

Sjögren's syndrome with membranous glomerulonephritis detected by urine screening of schoolchildren.

A case of Sjögren's syndrome with glomerulonephritis is presented. The patient was a 13 year old male with hematuria and proteinuria discovered by urine screening of school children. Evaluation showed no evidence of any associated connective tissue disease. Kidney biopsy was consistent with membranous glomerulonephritis. Sjögren's syndrome with membranous glomerulonephritis is rare and the patient was the youngest case in the literature.

Adolescent↗
PubMed1996

Long-term results and prognostic factors after repair of abdominal aortic aneurysm with concomitant malignancy.

OBJECTIVE: Long-term results after repair of abdominal aortic aneurysm (AAA) with concomitant malignancy were reviewed, and factors which may affect survival were analyzed. DESIGN: Retrospective series with follow-up of three to 125 months. Setting. Department of Surgery, Matsuyama Red Cross Hospital, Matsuyama, Japan. PATIENTS: Among 112 consecutive repairs of AAA, 16 cases had concomitant malignancy. The malignant lesions included eight gastric cancers and eight other malignant tumours. The malignancies were divided using TNM Classification into an early stage (stage O or I) group (n=9) and an advanced stage (stage II, III, or IV) group (n=7). INTERVENTIONS: All aneurysms were successfully repaired, and simultaneous resection of the concomitant malignancy was performed in five cases. While 13 malignant lesions were resected completely, three could not be resected completely, but were treated by other surgical procedures. MEASURES: Survival rates were predicated using the Kaplan-Meier method. The log-rank test was used to compare survival rates. RESULTS: The one-, two-, and five-year survival rates after repair of AAA were 80%, 72% and 63%, respectively. The survival rates for the early stage group were significantly higher than those for the advanced stage group (p<0.05). Patients with concomitant gastric cancer or who underwent complete resection of the malignant lesion survived longer. CONCLUSION: In patients with concomitant AAA and malignancy, factors influencing survival for those with malignant lesions also affected survival after aneurysmectomy. Detection of early-stage concomitant malignancy and more aggressive treatment for the malignancy may improve the outcome.

Age Factors↗
PubMed1996

[Clinicopathological study of Henoch-Schönlein purpura nephritis with special reference to C3c deposits].

The aim of this study was to evaluate glomerular C3c deposits of Henoch-Schönlein purpura nephritis (HSPN) in children. Fifty-one patients aged 7-15 years (20 males and 31 females) were studied. On histological investigation, crescent formation seen under light microscopy and subepithelial electron dense deposits (EDD) under electron microscopy were found to be related to the degree of proteinuria and the duration of proteinuria and/or hematuria. A comparative clinicopathological study was performed on C3c-positive patients (n = 22) and C3c-negative patients (n = 25). Histological findings, such as crescent formation and subepithelial EDD, had no relation to glomerular C3c-deposits. At renal biopsy, C3c deposits were positive in 65 % of patients with heavy proteinuria ( > 100mg/kg/day), and in 30% of mild proteinuria patients ( < 50mg/kg/day). The difference between the two groups was statistically significant (p < 0.05). The duration of proteinuria and/or hematuria in C3c-positive patients had a tendency to persist in comparison with that in C3c-negative cases. Renal biopsies on many cases of C3c-negarive patients were performed following the lapse of three months, while the biopsies on patients showing global (+) C3c deposits (n = 15) were conducted within the three-month period. These results suggest that glomerular C3c deposits influence the clinical conditions of patients with HSPN, and complement activation is generated in the early stage of HSPN.

Adolescent↗
PubMed1996

Gene differentiation at the AB0, MN and Rhesus loci among Albanians and their relation with other Balkanic populations.

The population of 26 Albanian districts were analysed for AB0, MN and Rhesus blood group systems. The dendrogram based on genetic distances showed a closer relationship between districts with a higher population density and a higher gene flow. It was shown that, of all Balkan populations, the most closely related were those from Slavic areas and that the Albanian population was the most distinct.

ABO Blood-Group System↗