Biomedical subjects

T Tezuka

Publications and source records attributed to T Tezuka.

At least 73 records · Page 4Linked to original sources

PubMed1998

Leukemic cells with 11q23 translocations express granulocyte colony-stimulating factor (G-CSF) receptor and their proliferation is stimulated with G-CSF.

We report a 20-month-old boy with acute lymphoblastic leukemia with the 11q23 translocation whose blasts markedly increased in peripheral blood after intravenous granulocyte colony-stimulating factor (G-CSF) administration, but disappeared after stopping G-CSF. The in vitro study showed that the leukemic cells separated from this patient expressed G-CSF receptor (G-CSFR) and an addition of G-CSF stimulated their proliferation by 3H-thymidine incorporation assay (stimulation index, 4.9). To clarify whether or not leukemic cells with 11q23 translocations generally express G-CSFR and show proliferative response to G-CSF, we performed the similar in vitro experiments using eight leukemic cell lines with 11q23 translocations. We found that all cell lines examined expressed G-CSFR (20-98%) and proliferation of seven leukemic cell lines was significantly enhanced in response to G-CSF (stimulation index >1.5 in five cell lines), suggesting a possible participation of the G-CSF/G-CSFR interaction in the process of growth regulation of leukemic cells with 11q23 translocations.

Antineoplastic Combined Chemotherapy Protocols↗

PubMed1998

Expression of CD95 ligand in melanocytic lesions as a diagnostic marker.

CD95 ligand (CD95L) potently induces apoptosis by activating CD95 on target cells. It has recently been reported that melanoma cells in vivo express a significant amount of CD95L, thereby being immediately able to kill CD95-bearing immunocompetent cells specific for cancer antigens, which infiltrate the lesions. In this study, we employed immunohistochemistry using an antibody directed against CD95L to investigate at which stage the melanoma CD95L expression is turned on. Skin biopsies of 49 lesions from 46 patients were assessed. These included benign and dysplastic naevi, melanoma in situ, stage I melanomas (Clark's level 2 or 3), advance-phase melanomas (Clark's level 4 or 5) and lymph node metastases. CD95L was expressed in all of the advance-phase melanomas as well as lymph node metastases of cutaneous origin, whereas neither melanoma in situ, benign naevi nor dysplastic naevi reacted positively with the antibody. To investigate a link between positivity and tumour size, the data were analysed on the basis of Breslow thickness, and indicated that expression was observed only when tumours were thicker than 0.75 mm. We next compared expression of CD95L and HMB-45. CD95L was positive only in melanomas in a more advanced phase than stage I, whereas HMB-45 was not only expressed in melanoma cells but also in benign pigmented naevi. This indicated the advantage of CD95L staining to diagnose melanoma. The present study indicates the significant correlation between tumorigenicity and expression of CD95L, and thereby raises the possibility that CD95L may be a useful diagnostic marker for malignant melanomas.

Antigens, Surface↗

PubMed1998

Localization of sphingomyelinase in lesional skin of atopic dermatitis patients.

Because it has been suggested that the majority of the activity hydrolysing [N-methyl-14C] sphingomyelin is due to sphingomyelin acylase in the lesional skins of atopic dermatitis (AD), in this study we used immunologic techniques to localize and quantitate sphingomyelinase in AD lesional and normal skin. A polyclonal antibody raised against a synthetic polypeptide corresponding to a portion of the amino acid sequence deduced from the cDNA of human acid sphingomyelinase, cross-reacted with a 58 kDa, pI 5.8 human epidermal protein in an immunoblot analysis. The 58 kDa protein-rich fraction, partially purified by immunoprecipitation, converted [N-methyl-14C]-sphingomyelin to 14C-phosphorylcholine and ceramides. The reaction products were immunohistochemically observed in the intercellular domain from the upper spinous cell layer to the upper stratum corneum cell layers in the lesional skin of AD patients. Immunoelectron-microscopically, gold particles appeared to be concentrated in the intercellular domains of the granular-upper stratum corneum cells in the lesional skin of AD patients. The total amount of the 58 kDa protein in a 7 mm2 area of the skin was measured by quantitative immunoblot analysis; and was slightly increased in the lesional skin samples [3.5 +/- 0.3 microg per 7 mm2 (n = 7)], as compared with the nonlesional skin samples of AD patients [2.8 +/- 0.19 microg per 7 mm2 (n = 10)] and with the normal skin samples [2.7 +/- 0.22 microg per 7 mm2 (n = 10)]. This difference (between the lesional skin of AD and the nonlesional skin of AD or the normal control) was significant (nonpaired student's t test, p < 0.05).

Antibodies↗

PubMed1998

Transient increase in CD45RO expression on T lymphocytes in infected newborns.

Although infections are a significant cause of mortality and long-term morbidity of newborns, the early diagnosis of neonatal infections remains difficult. It has been shown recently that up-regulation of CD45RO expression on umbilical cord lymphocytes might be a good indicator for intrauterine viral infections. To evaluate clinical relevance of changes in CD45 isoforms to the diagnosis of neonatal infections, we examined CD45RA and CD45RO expression on total, CD4+, and CD8+ lymphocytes from newborns. Lymphocytes from 55 newborns who were hospitalized in Kiyose Metropolitan Children's Hospital or Yamanashi Medical University Hospital were analyzed. Newborns were classified into three groups according to the possibility of bacterial or viral infection; I ("no" infection, n = 24), II ("possible" infection, n = 17), and III ("definite" infection, n = 14). Results were compared between two of three groups. The CD45RO expression on total or CD4+ lymphocytes in group III (bacterial 12, viral 2) was significantly higher than that in group I or II. Particularly, the CD45RO expression on CD4+ lymphocytes was always greater than 10% in group III, and showed no overlap with the respective value in group I. After termination of infection, the increased CD45RO expression in group III gradually returned to normal levels. These results indicate that not only viral but also bacterial infections trigger transient and reversible changes in CD45 isoform expression by neonatal CD4+ T lymphocytes. This may be a novel marker for the early diagnosis of neonatal infections.

CD3 Complex↗

PubMed1998

[Pharmacokinetic and clinical evaluation of tazobactam/piperacillin in the pediatric field].

Pharmacokinetic and clinical evaluation was conducted on the combination drug, tazobactam/piperacillin (TAZ/PIPC, YP-14), of newly developed beta-lactamase inhibitor, tazobactam (TAZ), and antibiotic agent of penicillin group for injections, piperacillin (PIPC), and the following results were obtained. 1. Absorption and excretion Both serum levels of TAZ and PIPC after the intravenous drip infusion for 30 minutes at the dose of 25 mg/kg or 50 mg/kg of TAZ/PIPC showed peaks at the end of the intravenous drip infusion (in 30 minutes after the commencement of the administration) with the levels of 11.93 or 26.05 micrograms/ml for TAZ and 49.80 or 107.50 micrograms/ml for PIPC. The halflife times were 0.51 or 0.67 hours for TAZ and 0.51 or 0.54 hours for PIPC. The serum level of desethyl-piperacillin (DEt-PIPC), an active metabolite of PIPC, showed a peak one hour after the end of the intravenous drip infusion (in 90 minutes after the commencement of the administration) with the level of 0.79 or 1.47 micrograms/ml. On the other hand, cumulative recovery ratios into urine were 41.3% or 40.4% for TAZ and 38.7% or 37.8% for PIPC. The recovery ratio for DEt-PIPC, an active metabolite of PIPC, was 1.9% or 0.3%. 2. Clinical evaluation TAZ/PIPC was administered to 47 cases (pneumonia: 25 cases, bronchitis: 16 cases, cutaneous soft tissue infection: 3 cases, urinary tract infection: 2 cases and lymphadenitis: 1 case). Clinical efficacy evaluation was conducted in 45 cases, excluding one case with bronchitis complicated by measles and other one case with left cervical cellulitis complicated by mumps. Good or more efficacy was observed in the all cases (excellent efficacy: 28 cases and good efficacy: 17 cases). The eradication rate in 31 cases where the pathogenic bacteria were identified and bacteriological efficacy was revealed was 93.5% (29/31 cases). Out of them, bacterial replacement was observed in 5 cases. Decrease in bacteria (including partial eradication) was observed in the remaining 2 cases. Adverse reactions were not reported in any cases. Abnormal values of clinical laboratory examination were reported in 5 cases (increase in platelet count: 2 cases, decrease in leukocyte count: 2 cases and increase in eosinophil count: 1 case). However, there was no case requiring any treatment.

Bacteria↗

PubMed1997

Pressure-induced changes in the folded structure of lysozyme.

We demonstrate, for the first time in solution, that pressure induces changes in the overall folded structure of a protein (lysozyme). This was made possible by using a home-developed, on-line continuously variable pressure cell on a high resolution NMR spectrometer operating at 750 MHz. We could follow pressure-induced diamagnetic chemical shifts of more than 26 protons of lysozyme at variable pressure in the range of 1 to 2000 bar. The results indicate that the main effect of the pressure is a compaction of the hydrophobic core part of the protein consisting of bulky side-chains. The technique introduced here provides a general method with which one can probe microscopic internal flexibility of a protein in solution.

Animals↗

PubMed1997

Tissue-specific knockout of the mouse Pig-a gene reveals important roles for GPI-anchored proteins in skin development.

Glycosylphosphatidylinositol (GPI)-anchored proteins are widely distributed on plasma membranes of eukaryotes. More than 50 GPI-anchored proteins have been shown to be spatiotemporally expressed in mice with a deficiency of GPI-anchor biosynthesis that causes embryonic lethality. Here, we examine the functional roles of GPI-anchored proteins in mouse skin using the Cre-loxP recombination system. We disrupted the Pig-a gene, an X-linked gene essential for GPI-anchor biosynthesis, in skin. The Cre-mediated Pig-a disruption occurred in skin at almost 100% efficiency in male mice bearing two identically orientated loxP sites within the Pig-a gene. Expression of GPI-anchored proteins was completely absent in the skin of these mice. The skin of such mutants looked wrinkled and more scaly than that of wild-type mice. Furthermore, histological examination of mutant mice showed that the epidermal horny layer was tightly packed and thickened. Electron microscopy showed that the intercellular space was narrow and there were many small vesicles embedded in the intercellular space that were not observed in equivalent wild-type mouse skin preparations. Mutant mice died within a few days after birth, suggesting that Pig-a function is essential for proper skin differentiation and maintenance.

Animals↗

PubMed1997

Quantitative analysis of histidine and cis and trans isomers of urocanic acid by high-performance liquid chromatography: a new assay method and its application.

To elucidate the factors involved in dry skin and the skin damage caused by UV light, it is necessary to analyze small amounts of stratum corneum to determine amino acid contents. A new assay method for this purpose is described. Dabsylated amino acids including histidine and the cis and trans isomers of urocanic acid were analyzed quantitatively by high-performance liquid chromatography (HPLC), using a reversed-phase column. Histidine and the isomers of urocanic acid were separated from 36 other amino acids thought to be present in the extract of stratum corneum. In the presence of the 36 amino acids, standard calibration curves were obtained from 0.25 to 2.5 pmol/microliter, for histidine and for both isomers of urocanic acid. The coefficients of variation for the reproducibility of the analysis at 1.0 pmol/microliter were 3.8%, 2.9% and 2.5% for the cis and trans isomers of urocanic acid and for histidine, respectively. Amounts of 2 to 50 pmol of cis and trans isomers of urocanic acid and histidine in the stratum corneum were detected. The ratio of the cis to the trans isomer of urocanic acid in sunburned stratum corneum was more than three times that in normal stratum corneum. This method appears to be useful for the determination of small amounts of histidine and of the cis and trans isomers of urocanic acid in the stratum corneum.

Chromatography, High Pressure Liquid↗

PubMed1997

Immunohistology of skin and oral biopsies in graft-versus-host disease after bone marrow transplantation and cytokine therapy.

BACKGROUND: Early diagnosis of graft-versus-host-disease (GVHD) after bone marrow transplantation is often difficult, particularly when the patients are immunosuppressed by chemotherapy or irradiation. OBJECTIVE: To investigate the influence of cytokines on skin lesions after bone marrow transplantation. METHODS: Biopsy specimens of skin and oral mucosa were obtained from bone marrow transplant patients with GVHD and were subjected to histologic and immunohistochemical examination. RESULTS: Administration of granulocyte-macrophage colony-stimulating factor caused atopic dermatitis-like lesions in two patients, who had infiltration of neutrophils, eosinophils, and lymphocytes around the hair follicles of the skin and no signs of GVHD in other organs. Only patients who were treated with cytokines developed acute GVHD. Immunohistochemical examination of skin biopsies from 18 patients with acute GVHD and 11 patients with chronic GVHD after cyclophosphamide administration or irradiation showed that the maculopapular skin lesions characteristic of acute GVHD contained infiltrates of CD4+ and CD8+ lymphocytes. There was also an increase in numbers of epidermal keratinocytes expressing intercellular adhesion molecule-I and HLA-DR antigens. CONCLUSION: These findings support the involvement of cytokines in GVHD and suggest that immunostaining of skin biopsies may be useful for the early diagnosis of this condition.

Acute Disease↗

PubMed1997

Decreased level of prosaposin in atopic skin.

In the skin of atopic dermatitis patients, the amount of ceramides in the stratum corneum is decreased. Although the cause of this decrease may be due to the higher activity of acylase, a decrease in the activity of sphingolipid activator proteins may also be the cause. A polyclonal antibody to saposin D, elicited by immunizing rabbits with the synthetic polypeptide from cDNA of saposin D, cross-reacted with a single 65-kDa epidermal protein of pI 5.6 in a 2-dimensional immunoblot study, suggesting that it was prosaposin, the precursor protein of saposin D, from its molecular weight and demonstrating its immunohistochemical localization in the innermost cell layers of the stratum corneum of the skin. The antigenic material was also observed in the epithelium of the esophagus, pneumocytes of the lungs, hepatocytes, and glandular cells of the stomach. Immunoelectron microscopy showed the antigenic material in the cytoplasm of the granular cells and the intercellular spaces, either between the stratum granulosum and the stratum corneum or on the stratum corneum cell envelope. By ELISA, the amount of the 65-kDa protein in the inner surface skin of the upper arm of atopic dermatitis patients (nonlesional skin) [4.1 +/- 2.0 microg per 7 mm2 (mean +/- SD), n = 10] was found to be significantly decreased (p < 0.05) to 66% of that in the normal control (6.2 +/- 1.5 microg per 7 mm2, n = 10). Therefore, the suppression of prosaposin synthesis may be related to the abnormal stratum corneum formation in atopic skin through lower activation of glucosylcerebrosidase or sphingomyelinase.

Amidohydrolases↗

PubMed1997

30-kDa trypsin-like proteases in the plantar stratum corneum.

The casein digestible proteases in human plantar stratum corneum were determined to be about 75-kDa, 30-kDa and 25-kDa in molecular weight by zymography. The enzymatic activity of the 75-kDa and 25-kDa proteases was specifically inhibited by chymostatin, which is an inhibitor of chymotrypsin-like serine proteases, and the proteases around 30-kDa were inhibited by leupeptin, a trypsin-like serine protease inhibitor. The enzymatic activity of all these proteases was inhibited by aprotinin. The 30-kDa trypsin-like proteases were heat-stable; their enzymatic activity still remained even after heating at 100 degrees C for 60 minutes. Their optimal pH was around 9, and the activity was higher in the outer part of the stratum corneum than in the inner part.

Aprotinin↗

PubMed1997

Enhanced production of RANTES, an eosinophil chemoattractant factor, by cytokine-stimulated epidermal keratinocytes.

In allergic skin diseases such as atopic dermatitis (AD), eosinophils migrate from the circulation to the skin. We investigated the mechanisms of eosinophil chemotaxis in atopic dermatitis by examining the effect of stimulation of epidermal keratinocytes (KC) by inflammatory cytokines, interferon-gamma (IFNgamma) and/or tumor necrosis factor-alpha (TNF alpha) on the production of eosinophil chemotactic factors. Simultaneous addition of IFNgamma and TNF alpha to culture KC synergistically increased eosinophil chemotaxis and the expression of RANTES mRNA and protein level on these cells. Anti-RANTES antibody blocked eosinophil chemotaxis by IFNgamma- and TNF alpha-stimulated KC. Our results indicate that the production of RANTES by KC may help to explain eosinophil infiltration into the skin in AD.

Chemokine CCL5↗

PubMed1997

Steroid-induced changes of eosinophils in atopic dermatitis.

We investigated whether apoptosis of eosinophils is specific to atopic dermatitis (AD), or also occurs in other diseases with eosinophilia. We examined the survival of eosinophils cultured with corticosteroids: (1) Clinically, steroid administration significantly decreased high peripheral blood eosinophil cell counts in patients with AD. (2) Treatment with recombinant human (rh) IL-5 prolonged the life span of eosinophils derived from patients with AD and of those derived from non-AD patients with eosinophilia. However, there were differences in the survival rates in the presence of rhIL-5: the eosinophils from non-AD patients showed 1.4-fold higher survival rates than those from AD patients at 24 h. In the presence of steroids, the eosinophils from non-AD patients showed a survival rate double that of those from AD patients at 24 h. (3) In eosinophils from patients with AD, the survival rate decreased significantly in a time- and steroid-concentration-dependent manner. Steroid administration significantly inhibited the survival rate of eosinophils from patients with AD compared to those of monocytes and neutrophils. These findings suggest that apoptosis induced by steroids decreases the eosinophil count in vivo in patients with AD. There may be a difference in the incidence of steroid-induced apoptosis between eosinophil cells from patients with AD and those from patients with eosinophilia due to other underlying diseases.

Apoptosis↗

PubMed1997

Phenylmercury transport mediated by merT-merP genes of Pseudomonas K-62 plasmid pMR26.

The merB-merA-deleted plasmid pMRD141 which contains the intact merT-merP genes of pMRA17 conferred bacterial hypersensitivity not only to Hg2+ but also to C6H5Hg+. The bacterium with pMRD141 took up significantly more C6H5Hg+ than its isogenic strain with the cloning vector Bluescript II. The hypersensitivity to C6H5Hg+ seems to be based on hyperaccumulation of toxic C6H5Hg+ in the absence of detoxifying enzymes encoded by merB and merA. Our results show that bacterial transport of C6H5Hg+ into the cytoplasm is regulated by merT-merP genes.

Bacterial Proteins↗

PubMed1997

Evaluation of rectal mucosal hemodynamics in patients with liver cirrhosis using reflectance spectrophotometry.

OBJECTIVES: Portal hypertensive colonopathy is observed in patients with liver cirrhosis. To determine the correlation between rectal mucosal hemodynamics and portal hypertensive colonopathy, we observed rectal mucosal findings and measured rectal mucosal hemodynamics in patients with liver cirrhosis. METHODS: Thirty-four patients with liver cirrhosis and 16 healthy control subjects were simultaneously examined for rectal mucosal findings by colonoscopy and indices of rectal mucosal Hb concentration (RHb) and rectal mucosal oxygen saturation by reflectance spectrophotometry. Endoscopic findings in the rectal mucosa of patients with liver cirrhosis included vascular ectasias, blue veins, and varices. We investigated the relationship between rectal mucosal hemodynamics and clinical parameters of liver cirrhosis (Child-Pugh classification, the amount of indocyanine green remaining in the blood 15 min after its injection, and ascites). Patients with hepatocellular carcinoma, colonic cancer, multiple colonic polyps, or severe anemia were excluded. RESULTS: Rectal mucosal lesions were observed in 11 patients with liver cirrhosis (32.4%). In the hemodynamic studies, we found significantly increased RHb values in the cirrhosis group as compared with the control group. On the other hand, there was no significant change in rectal mucosal oxygen saturation between the two groups. A significant increase in RHb was observed in patients with rectal mucosal lesions in the cirrhosis group. RHb in the cirrhosis group correlated with grade of Child-Pugh classification. Increased RHb decreased in parallel after portal decompression by creation of a transjugular intravenous portosystemic stent shunt. CONCLUSIONS: The rectal mucosal lesions in liver cirrhosis correlate with an increase in RHb, which correlates with portal hypertension.

Ascites↗

PubMed1996

Filaggrin linker segment peptide and cystatin alpha are parts of a complex of the cornified envelope of epidermis.

We showed that filaggrin linker segment peptide (FLSP) is a glutamine-rich substrate of epidermal transglutaminase (TGase), conjugating enzymatically with a phosphorylated cystatin alpha (P-cystatin alpha) which is a lysine-rich substrate of the enzyme. This finding suggested that FLSP would be a component of cornified envelope of the epidermis as well as P-cystatin alpha. Here, we investigated the in vivo location of FLSP. An antibody against the peptide conjugated with keyhole limpet hemocyanin (FLSP/KLH) reacted specifically with FLSP on immunoblots. Immunofluorescence histochemistry located specific staining with this antibody on keratohyalin granules and the cell membrane region of the stratum corneum. Specific staining was not detected when the antiserum was first absorbed by FLSP. Preembedding immunoelectron microscopic analysis showed that anti-FLSP/KLH antibody labeled with gold particles reacted with cornified envelope prepared from newborn rat stratum corneum. The high molecular weight protein enzymatically synthesized from phosphorylated cystatin alpha and FLSP by TGase reacted with both anti-FLSP/KLH antibody and anti-P-cystatin alpha antibody on Western blotting. These findings suggest that the FLSP-cystatin alpha conjugate is a component of the cornified envelope of the epidermis in rats.

Amino Acid Sequence↗

PubMed1996

Physical and functional association of the cbl protooncogen product with an src-family protein tyrosine kinase, p53/56lyn, in the B cell antigen receptor-mediated signaling.

To identify novel signal transducers involved in signaling mediated by the Src-family protein tyrosine kinases (PTKs), we used a yeast two-hybrid system with a probe corresponding to the regulatory region of p56lyn, a member of Src-family PTKs. One of the isolated clones contained the COOH-terminal 470 amino acid residues of p120c-cbl, the product of the cellular homologue of the v-cbl retroviral oncogene. p120c-cbl is a cytoplasmic protein with nuclear protein-like motifs. Here we show in vivo association of p120c-cbl with p53/56lyn. After stimulation of the B cell antigen receptor (BCR), p120c-cbl was rapidly tyrosine phosphorylated. Studies with lyn- or syk-negative chicken B cells demonstrated that p53/56lyn, but not p72syk, was crucial for tyrosine phosphorylation of p120c-cbl upon stimulation of the BCR. We also show the importance of p59fyn in tyrosine phosphorylation of p120c-cbl in the T-cell receptor-mediated signaling using fyn-overexpressing T cell hybridomas and splenic T cells from fyn-deficient mice. These results suggest that p120c-cbl is an important substrate of Src-family PTKs in the intracellular signaling mediated by the antigen receptors

Cells, Cultured↗

PubMed1996

Decreased expression of filaggrin in atopic skin.

The amounts of the epidermal proteins filaggrin, involucrin, cystatin A and Ted-H-1 antigen produced during the terminal differentiation of keratinocytes were immunohistochemically measured in lesional and nonlesional skin of atopic dermatitis (AD) patients. In addition, the amount of filaggrin in the skin of the inner surface of the upper arm of AD patients (nonlesional skin) and normal controls, obtained by punch biopsy, was measured by an enzyme-linked immunosorbent assay (ELISA) technique. The immunohistochemical study showed that all four proteins were decreased in lesional skin. By contrast, only filaggrin was decreased in nonlesional skin of AD patients. The ELISA showed that the amount of filaggrin in the skin of the inner surface of the upper arm was 2.48 +/- 0.45 microgram/7 mm2 (n = 8) in AD patients, which was 32% of that in the normal controls (7.7 +/- 0.55 microgram/7 mm2; n = 4). This decrease in filaggrin production in atopic skin may be one of the reasons why atopic skin can easily become dry, because filaggrin is thought to be the precursor protein of the emollient factors in the stratum corneum. The evidence that only the expression of filaggrin was suppressed in AD patients, though the genes of filaggrin and involucrin are localized to a very restricted portion of the same gene 1q21, indicates that the filaggrin gene does not share regulatory elements with the involucrin gene.

Adolescent↗