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Biomedical subjects

M Kawakami

Publications and source records attributed to M Kawakami.

At least 19 recordsLinked to original sources
PubMed1992

Assignment of the magnetic resonances of the imino protons and methyl protons of Bombyx mori tRNA(GlyGCC) and the effect of ion binding on its structure.

The magnetic resonances in the low-field H-NMR spectra of Bombyx mori tRNA(GlyGCC), corresponding to the hydrogen-bonded imino protons of the helical stems and tertiary base pairs, could be tentatively assigned by means of the sequential nuclear Overhauser effects. While B. mori tRNA(GlyGCC) does not contain the G19C56 tertiary base pair, the D20G57 base pair exists between the D and T loops, which was not found in the X-ray crystal structure of yeast tRNA(Phe). The effects of Mg2+, spermine and temperature on the conformation of this tRNA have also been examined based on the behavior of the assigned resonance signals. Mg2+ stabilize the D and T stems and the tertiary structure between the D and T loops. Spermine affects the resonances of the D and anticodon stems, and A23G9, but does not stabilize them. While the acceptor stem melts sequentially from both ends (G7C66 and G1C72) with increasing temperature, the anticodon stem melts from only one end (G39C31) and the G26C44 base pair is the most stable. In the tertiary structure between the variable loop and D stem, G10G45 melts first and G22G46 last. Yeast tRNA(Phe) has also been examined, and the results were compared with those for B. mori tRNA(Gly).

Animals↗
PubMed1992

An ultrasensitive system to detect IL-4: enzyme-linked immunosorbent assay (ELISA) combined with an avidin-biotin and enzyme amplification system.

We established an ultrasensitive interleukin-4 enzyme-linked immunosorbent assay by combining ELISA with an avidin-biotin and enzyme amplification system. The resultant system (AB-EA ELISA) was 250 times more sensitive than conventional ELISA and 2.5 times more sensitive than enhanced ELISA using an enzyme amplification system alone. The ultrasensitive assay was specific to IL-4 alone; there was no cross reaction with other cytokines. Using the ultrasensitive assay, we measured IL-4 synthesis in vitro by unstimulated and stimulated peripheral blood mononuclear cells (PBMC) from patients with allergic rhinitis. PBMC from patients spontaneously produced measurable amounts of IL-4, whereas IL-4 production from PBMC of normal controls, if any, was below detectable levels. Stimulation of the cultures with LPS significantly increased IL-4 production in two of six patient PBMC cultures but in none of the control cultures; stimulation with Con A markedly increased IL-4 production in all patient PBMC cultures but in only two of seven control cultures. These results suggest that the AB-EA ELISA is a useful method to study the mechanism of IL-4 synthesis in type-I allergic diseases.

Animals↗
PubMed1992

cDNAs and deduced amino acid sequences of subunits in the binding component of mouse bactericidal factor, Ra-reactive factor: similarity to mannose-binding proteins.

The complement-dependent bactericidal factor, Ra-reactive factor, binds specifically to Ra polysaccharide, which is common to some strains of Gram-negative enterobacteria, and its is a complex of proteins composed of a polysaccharide-binding component and a component that is presumably responsible for the complement activation. The former component consists of two different 28-kDa polypeptides, P28a and P28b. We determined the partial amino acid sequences of P28a and P28b, and the results indicated that these polypeptides were similar to two species of mannose-binding protein, MBP-C and MBP-A (alternative names, liver and serum mannan-binding proteins, respectively), which have been isolated from rat liver and/or serum [Drickamer, K., Dordal, M. S., & Reynolds, L. (1986) J. Biol. Chem. 261, 6878-6887; Oka, S., Itoh, N., Kawasaki, T., & Yamashina, I. (1987) J. Biochem. 101, 135-144]. Thus, we cloned the respective cDNAs, using as probes synthetic oligonucleotides for which the sequences had been deduced from the amino acid sequences of P28a and P28b and of rat MBP cDNAs. The primary structures of P28a and P28b deduced from the cloned cDNAs are homologous to one another. They have three domains, a short NH2-terminal domain, a collagen-like domain, and a domain homologous to regions of some carbohydrate-binding proteins, as has been reported for rat MBPs. Southern and Northern blotting analyses using these cDNAs indicated that the P28a and P28b polypeptides are the products of two unique mouse genes which are expressed in hepatic cells.

Amino Acid Sequence↗
PubMed1992

Increased interleukin 1 receptor, type I, at messenger RNA and protein level in skin fibroblasts from patients with systemic sclerosis.

Summary. To elucidate the mechanisms of the fibrosis in systemic sclerosis (SSc) through the action of a cytokine, interleukin 1 (IL-1), we studied the specific biologic and biochemical features of interleukin 1 receptor (IL-1R) as expressed on the surfaces of fibroblast cells in cultures from 3 SSc patients and 3 normal donors. 125I-IL-1 beta binding assays revealed a high density of IL-1R on the cell surfaces of SSc fibroblasts as compared to those of normal subjects. We also found an enhanced expression of IL-1R messenger RNA (mRNA) in SSc fibroblasts, using Northern blot or slot blot analysis. These findings indicate that the expression of IL-1R on SSc fibroblasts were spontaneously induced at the transcriptional level. It is suggested that SSc fibroblasts are more sensitive to IL-1, and that the signal transduction of IL-1 through IL-1R may be eventually involved in the fibrosis of SSc.

Cells, Cultured↗
PubMed1992

Human mannose-binding protein is identical to a component of Ra-reactive factor.

Human Ra-reactive factor (RaRF) and mannose-binding protein (MBP) were isolated from sera by utilizing their affinity to Ra chemotype Salmonella typhimurium and yeast mannan, respectively. A predominant polypeptide of human RaRF with an Mr of 32 kDa (P32) has the same mobility as human MBP on SDS-PAGE gels. A monoclonal antibody against P32 of human RaRF, 3E7, was found to react not only with P32 but also with human MBP, as assessed by immunoblotting. In addition, 3E7 blocked the complement-activating capacity of human MBP. The NH2-terminal amino acid sequence of P32 of human RaRF was determined and found to coincide with that of human MBP. Based on these results, it is clear that human MBP is identical to a component of RaRF.

Antibodies, Monoclonal↗
PubMed1992

Serum granulocyte colony-stimulating factor in patients with repeated infections.

We have already reported significant elevation of serum granulocyte colony-stimulating factor (G-CSF) in the acute phase of infection. In this study, we compared the responses to infection between patients with frequently repeated infection (repeaters) and others (non-repeaters). We examined the clinical data and serum G-CSF levels in 48 patients with acute infections. Serum G-CSF levels were significantly lower in repeaters than in non-repeaters (197.7 +/- 370.0 vs. 1014.1 +/- 924.4 pg/ml. P less than 0.001). There were no significant differences in age, serum total protein, or cholinesterase between the groups, but serum albumin was significantly lower in repeaters than in non-repeaters (2.87 +/- 0.5 vs. 3.31 +/- 0.4 g/dl. P less than 0.005). It is suggested that administration of recombinant G-CSF may be useful for patients with repeated infections.

Aged↗
PubMed1992

On the variability of the 3' terminal sequence of the turnip mosaic virus genome.

The sequence of the 3'-terminal 1223 nucleotides (nts) of a Japanese isolate of turnip mosaic virus (TuMV-Jap) RNA has been determined. The sequence reveals a single open reading frame (ORF) which terminates at a position 212 nts upstream of the 3' poly(A)-tract. Determination of the N-terminal amino acids of TuMV-Jap coat protein (CP) mapped the CP cistron within this ORF and revealed a Glu-Ala dipeptide sequence as the putative cleavage site by which the CP is released from the viral polyprotein. The predicted amino acid sequence of the TuMV-Jap CP shows 97.2% identity with that of a Canadian isolate of TuMV (TuMV-Can) and 99% with a second, Chinese, isolate (TuMV-Chi). However, the 3'-terminal non-translated region (NTR) of TuMV-Jap RNA is significantly shorter (212 nts) than the 3'-NTR of TuMV-Can RNA (668 nts), but of equal length as the 3'-NTR of the TuMV-Chi isolate which also measures 212 nts. The 3'-NTRs of both the TuMV-Jap and TuMV-Chi RNAs show homology with the first 201 nucleotides of the TuMV-Can RNA 3'-NTR. A search in the EMBL nucleotide sequence database revealed that the 467 nt-long unique extension of the 3'-NTR of TuMV-Can RNA has 89.8% homology to a part of the chloroplast ribosomal protein 12 gene (rsp12-gene). Irrespective of the origin of this extra sequence in the reported TuMV-Can sequence, which may have been introduced by a genuine RNA recombination event, it is concluded that the standard TuMV genome has a CP gene of 864 nts and an conserved 3'-NTR of approximately 212 nucleotides in length.

Amino Acid Sequence↗
PubMed1992

Presence of autoantibodies to peptidyl-prolyl cis-trans isomerase (cyclosporin A-binding protein) in systemic lupus erythematosus.

Several autoantibodies against cytoplasmic or nuclear components of cells have been reported in autoimmune diseases. We report here a previously unrecognized autoantibody to peptidyl-prolyl cis-trans isomerase (PPIase) in patients with systemic lupus erythematosus (SLE). PPIase, which catalyzes the cis-trans isomerization of proline imidic peptide bonds in oligopeptides, has recently been found to be identical to cyclophilin, a specific binding protein of a potent immunosuppressant, cyclosporin A. IgG and IgM anti-PPIase antibodies were detected in 40 and 20% of unselected patients with SLE, respectively, by ELISA. The reactivity of these sera was confirmed by immunoblotting experiments. Sera from rheumatoid arthritis patients showed no reactivity and 1 of 8 sera from systemic sclerosis patients and 1 of 25 sera from normal controls showed only weak reactivity. Unexpectedly, the anti-PPIase antibody was unable to inhibit PPIase activity, indicating that the autoantibody recognizes an epitope of PPIase which is different from the active site of PPIase. The levels of the anti-PPIase antibody in SLE patients correlated with remissions and flares of the disease. The anti-PPIase antibody was higher in patients with active SLE than those with inactive disease. The prevalence of the active stage of the disease was significantly higher in IgG anti-PPIase antibody-positive SLE patients as compared to antibody-negative SLE patients. These data define the presence of a new autoantibody against PPIase and its association with the activity and certain clinical manifestations in SLE.

Adult↗
PubMed1992

Influence of the electrical stimulation of the hypothalamus on adrenocortical steroidogenesis in hypophysectomized rats.

The electrical stimulation of the ventromedial hypothalamus (VMH), lateral hypothalamic area (LHA), periventricular arcuate nucleus (ARC), and posterior hypothalamus (PHY), on 14C transfer rates from 14C-1-acetate into adrenocortical steroids in adrenal slices of hypophysectomized rats were investigated. The 14C transfer rates into corticosterone and cortisol were increased by the stimulation of the VMH, ARC, and PHY, but decreased by the stimulation of the LHA. From these results, it might be suggested that these hypothalamic structures were involved in the regulation of adrenocortical steroidogenesis without participation of the pituitary.

Acetates↗
PubMed1992

The role of limbic-hypothalamic systems in metabolic responses of phenylalanine to repeated cold exposures in rabbits.

In this study we examine the effects of lesions to limbic or hypothalamic structures on the metabolic responses of phenylalanine in liver slices prepared from rabbits exposed to cold stress (-20 degrees C) for 12 hours at a fixed time once a day. The results were as follows: 1. The 1st cold exposure (cold exposure on the 1st day) had marked and various effects on phenylalanine metabolism, and the metabolic responses of phenylalanine to cold exposure gradually decreased and then completely disappeared with repetitions of exposure in intact rabbits. 2. The metabolic patterns and responses of phenylalanine to the 1st cold exposure were altered by lesioning the periventricular arcuate nucleus (ARC), ventromedial hypothalamus (VMH), stria terminalis (ST) or dorsal fornix (FX). 3. The complete abolishment of the metabolic responses to cold exposure by its repetition was observed in rabbits with lesions in the ARC or VMH as well. 4. In contrast, metabolic responses of phenylalanine to cold exposure in rabbits with lesions in the ST or FX remained even after six exposures. These results suggest that the ARC, VMH, amygdala (AMYG)-ST system and dorsal hippocampus (HPC)-FX system played a role in the metabolic regulation of phenylalanine and in the metabolic responses of phenylalanine to the 1st cold exposure and that the AMYG-ST and HPC-FX systems, but not the ARC or VMH, participated in the process of the metabolic adaptation of phenylalanine to cold exposure.

Amygdala↗
PubMed1992

Influence of the electrical stimulation of the medial amygdala on adrenocortical sensitivity to adrenocorticotrophin in hypophysectomized rats.

The effects of electrical stimulation of medial amygdala on the adrenocortical sensitivity to adrenocorticotrophin (ACTH) were investigated in hypophysectomized rats. The intravenous injection of ACTH increased the rates of 14C transfer from 14C-l-acetate into corticosterone and cortisol in the adrenal slices of hypophysectomized rats. The electrical stimulation of the medial amygdala produced a further increase in the rates of 14C transfer from 14C-l-acetate into corticosterone and cortisol. From these results, it might be suggested that the medial amygdala was capable of enhancing the adrenocortical steroidogenesis in response to ACTH.

Acetates↗
PubMed1992

Morphologic and quantitative changes in neurotransmitters in the lumbar spinal cord after acute or chronic mechanical compression of the cauda equina.

Changes in the neurotransmitters associated with pain transmission and regulation in the lumbar spinal cord were studied after acute or chronic mechanical compression of the cauda equina in rats. Using glyoxylic acid histofluorescence and immunohistochemical methods, it was morphologically apparent that substance P-containing nerve ending were decreased after chronic compression of the cauda equina. Somatostatin nerve terminals were reduced, and aminergic fibers and serotonin were enhanced after both acute and chronic mechanical compressions. In addition, quantitative analysis revealed that the levels of norepinephrine and serotonin remained elevated after mechanical compression of the cauda equina. It is suggested that pain in the lower back and extremities after mechanical compression of the cauda equina is controlled by these complicated changes of neurotransmitters in the lumbar spinal cord.

Animals↗
PubMed1992

Autostimulatory effects of IL-6 on excessive B cell differentiation in patients with systemic lupus erythematosus: analysis of IL-6 production and IL-6R expression.

Introducing avidin-biotin complex ELISA for anti-DNA antibody, the mechanism of in vitro production of anti-ssDNA antibody as well as of polyclonal immunoglobulin mediated by an IL-6-IL-6R loop was studied in patients with systemic lupus erythematosus (SLE). Regardless of the presence or absence of T cells, B cells from SLE patients could produce IgG anti-ssDNA antibody as well as total IgG without any stimulation. Low density B cells obtained by Percoll gradient density centrifugation responded to rIL-6 to produce IgG and IgG anti-ssDNA antibody. rIL-2 and rIL-4 had lesser effects on the differentiation of low density B cells. In fact, IL-6R was preferentially expressed on low density B cells from active SLE patients, as detected by anti-IL-6R MoAb, MT18, which did not inhibit IL-6 binding. SLE B cells, especially high density B cells, produced greater amounts of IL-6 in culture supernatants than did T cells, regardless of whether disease was active or inactive. Normal T cells and B cells did not produce significant amounts of IL-6. Thus, endogenous IL-6 produced by high density B cells bound to the IL-6R preferentially expressed on the low density B cells, and drove them into terminal differentiation, especially in active SLE patients. Further, addition of polyclonal anti-IL-6 or anti-IL-6R MoAb (PM1), which inhibited IL-6 binding, both inhibited IgG anti-ssDNA antibody as well as total IgG production by SLE B cells in a dose-dependent manner. These results suggest that interruption of the autocrine IL-6 loop would be of therapeutic value in SLE.

Antibodies, Antinuclear↗
PubMed1992

Siblings with normal LDL receptor activity and severe hypercholesterolemia.

We report about a brother and sister having clinical symptoms similar to those of homozygous familial hypercholesterolemia (FH) but surprisingly who have normal low density lipoprotein (LDL) receptor activities (M. Harada-Shiba et al, J Jpn Atheroscler Soc 1991; 19:227-242). The LDL receptor activities in the cultured fibroblasts of the patients were compared with those of FH heterozygotes and homozygotes for the LDL receptor mutation. The LDL receptor activities in the cultured fibroblasts of the patients were in the normal range, but their plasma cholesterol concentrations were similar to patients with homozygous FH. After the plasma LDL was removed by plasmapheresis in both patients, plasma cholesterol levels started to increase. The "rebound" of plasma cholesterol was compared with those for heterozygous and homozygous FH. The plasma cholesterol levels of the patients, which were greater than 410 mg/dl 2 weeks after plasmapheresis, were much higher than those of FH heterozygotes (232-311 mg/dl) but similar to those of FH homozygotes (345-464 mg/dl). The urinary mevalonate excretion rate, which reflects the rate of whole-body cholesterol synthesis, was higher for the brother (patient 1, 32.6 nmol/kg.day-1) than for the normal subjects (17.7 +/- 4.1 nmol/kg.day-1) but was similar to those of FH homozygotes (31.2 +/- 4.3 nmol/kg.day-1) and heterozygotes (29.8 +/- 10.9 nmol/kg.day-1). To estimate the catabolic and production rates of cholesterol in the brother, the time course for the increment in the total cholesterol level after plasmapheresis was analyzed by the two-compartment model.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1992

The effect of local application of 1,25-dihydroxycholecalciferol on osteoclast numbers in orthodontically treated rats.

Orthodontic tooth movement requires remodeling of periodontal tissues, especially alveolar bone. 1,25-(OH)2D3, the active form of vitamin D3, is known to be a potent stimulator of osteoclastic bone resorption. The purpose of this study was to investigate the effect of local application of 1,25-(OH)2D3 on osteoclast numbers induced by experimental tooth movement. A piece of orthodontic elastic band was inserted between the first and second upper molars of male Wistar rats weighing about 200 g each. Twenty microL of 1,25-(OH)2D3 (10(-12)-10(-7) mol/L) was injected locally into the submucosal palatal area of the root bifurcation of the right first molar. The left side was injected with phosphate-buffered saline (PBS). The number of osteoclasts was counted in a 700 x 1050 microns 2 area of the interradicular septum. The local injection of 1,25-(OH)2D3 caused a dose-dependent increase in osteoclast number. The effect of 1,25-(OH)2D3 reached a response plateau at 10(-10) mol/L when greater than a three-fold rise in osteoclast number was attained compared with the PBS-treated controls. While the insertion of a piece of elastic band for three days induced a significant increase in osteoclasts in the alveolar bone, daily injections of 20 microL of 10(-10) mol/M 1,25-(OH)2D3 for three days markedly stimulated the numbers of osteoclasts induced by the insertion of an elastic band. 1,25-(OH)2D3 was apparently synergistic with mechanical stimuli, resulting in enhancement of the numbers of osteoclasts induced by mechanical stimuli alone.(ABSTRACT TRUNCATED AT 250 WORDS)

Alveolar Process↗
PubMed1992

Effect of age on the rate of tooth movement in combination with local use of 1,25(OH)2D3 and mechanical force in the rat.

The purpose of this study was to compare the amount and rate of tooth movement in young and mature rats administered 1,25(OH)2D3 simultaneous with application of mechanical force. In 30 seven-week-old and 30 28-week-old male Wistar rats, the right maxillary first molar was moved buccally with a fixed appliance. The appliances delivered forces ranging from 5 to 20 g. Twenty microL of 1,25(OH)2D3 (10(-10) and 10(-8) mol/L) was injected locally into the submucosal palatal area of the root bifurcation of the right first molar. The left side was injected with phosphate-buffered saline (PBS). In young rats receiving 10(-10) mol/L 1,25(OH)2D3 every three days, tooth movement significantly increased to 126% of that in PBS-injected control rats on day 20. In 1,25(OH)2D3-injected mature rats, tooth movement was stimulated markedly and increased with 10(-10) mol/L to 245% and with 10(-8) mol/L to 154% of the amount of tooth movement seen in the PBS-injected controls by the end of the experiment. PBS-injected rats had a plateau stage where tooth movement did not occur at all, while there was no such lag-time in the 1,25(OH)2D3-injected group which showed continuous tooth movement. The local injection of 1,25(OH)2D3 did not change serum calcium, phosphate, and alkaline phosphatase activity, and there were no apparent clinical or microscopic side-effects.

Activator Appliances↗