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Biomedical subjects

J Honda

Publications and source records attributed to J Honda.

At least 19 recordsLinked to original sources
PubMed2003

IGF-I regulates pro-opiomelanocortin and GH gene expression in the mouse pituitary gland.

IGF-I is expressed in somatotrophs, and IGF-I receptors are expressed in most somatotrophs and some corticotrophs in the mouse pituitary gland. Our recent study demonstrated that IGF-I stimulates the proliferation of corticotrophs in the mouse pituitary. These results suggested that somatotrophs regulate corticotrophic functions as well as somatotrophic functions by the mediation of IGF-I molecules. The present study aimed to clarify factors regulating pituitary IGF-I expression and also the roles exerted by IGF-I within the mouse anterior pituitary gland. Mouse anterior pituitary cells were isolated and cultured under serum-free conditions. GH (0.5 or 1 microg/ml), ACTH (10(-8) or 10(-7) M), GH-releasing hormone (GHRH; 10(-8) or 10(-7) M), dexamethasone (DEX; 10(-8) or 10(-7) M) and estradiol-17beta (e2; 10(-11) or 10(-9) M) were given for 24 h. IGF-I mRNA levels were measured using competitive RT-PCR, and GH and pro-opiomelanocortin (POMC) mRNA levels were measured using Northern blotting analysis. GH treatment significantly increased IGF-I mRNA levels (1.5- or 2.1-fold). ACTH treatment did not alter GH and IGF-I mRNA levels. IGF-I treatment decreased GH mRNA levels (0.7- or 0.5-fold), but increased POMC mRNA levels (1.8-fold). GH treatment (4 or 8 microg/ml) for 4 days increased POMC mRNA levels. GHRH treatment increased GH mRNA levels (1.3-fold), but not IGF-I mRNA levels. DEX treatment significantly decreased IGF-I mRNA levels (0.8-fold). e2 treatment did not affect IGF-I mRNA levels. GH receptor mRNA, probably with GH-binding protein mRNA, was detected in somatotrophs, and some mammotrophs and gonadotrophs by in situ hybridization using GH receptor cDNA as a probe. These results suggested that IGF-I expression in somatotrophs is regulated by pituitary GH, and that IGF-I suppresses GH expression and stimulates POMC expression at the transcription level. Pituitary IGF-I produced in somatotrophs is probably involved in the regulation of somatotroph and corticotroph functions.

Adrenocorticotropic Hormone↗
PubMed2002

Intrapituitary regulatory system of mammotrophs in the mouse.

Estrogen stimulates the proliferation of pituitary cells, in particular mammotrophs. The present study was designed to clarify involvement of transforming growth factor alpha (TGF-alpha) in the estrogen-induced growth of mouse pituitary cells in vitro. Anterior pituitary cells obtained from ICR male mice were cultured in a primary, serum-free culture system. Proliferation of pituitary cells was detected by monitoring the cellular uptake of a thymidine analogue, bromodeoxyuridine. Secretory cell types were immunocytochemically determined. Treatment with TGF-alpha (0.1 and 1 ng/ml) for 5 days stimulated cell proliferation. Since TGF-alpha binds to the epidermal growth factor (EGF)-receptor, this action may be exerted through this receptor. Estradiol-17beta (E2, 10(-9) M) stimulated proliferation of mammotrophs. RG-13022, an EGF receptor inhibitor, reduced the cell proliferation induced by EGF or E2, showing that the EGF receptor was involved in this induction of mammotroph growth. Treatment with TGF-alpha antisense oligodeoxynucleotide (ODN) inhibited the cell proliferation induced by E2, but treatment with EGF antisense ODN did not. Dual detection of TGF-alpha mRNA and growth hormone by in situ hybridization and fluorescence-immunocytochemistry demonstrated that TGF-alpha mRNA was detected in most somatotrophs. Our recent RT-PCR analysis revealed that E2 stimulated TGF-alpha-mRNA and EGF-receptor mRNA expression. These results indicate that TGF-alpha produced in somatotrophs mediates the stimulatory effect of estrogen on pituitary cell proliferation in a paracrine manner, and that EGF-receptor expression is stimulated by estrogen. These findings indicate that intrapituitary cell-to-cell interaction plays an important role in the control of pituitary secretory cells.

Animals↗
PubMed2002

Elevation of serum KL-6 levels in patients with hematological malignancies associated with cytomegalovirus or Pneumocystis carinii pneumonia.

The level of serum KL-6 antigen has been reported to be a sensitive indicator of various interstitial pneumonitis, but in patients with hematological malignancies who were accompanied by infective interstitial pneumonitis like Pneumocystis carinii or cytomegalovirus (CMV) pneumonia, it is still unknown whether serum KL-6 level is useful as a good marker for the diagnosis or disease activity. In this study, the serum levels of KL-6 and soluble intercellular adhesion molecule 1 (sICAM-1) were evaluated in five patients with malignant lymphoma or adult T-cell leukemia. Serum KL-6 and sICAM-1 levels at the time of diagnosis of P. carinii or CMV pneumonia were 1220+/-323 U/ml (mean+/-SD) and 1246+/-485 ng/ml, respectively. These levels were apparently high, when compared with standard value of serum KL-6 (<520 U/ml) and that of sICAM-1 (115-306 ng/ml). In patients without P. carinii or CMV pneumonia, who had hematological malignancies or AIDS, serum level of KL-6 was not high (299+/-122 U/ml), but sICAM-1 was high (651+/-495 ng/ml) because of the elevation of sICAM-1 in four of five cases. These findings suggest that, in patients with hematological malignancies, serum level of KL-6 antigen rather than sICAM-1 may be useful in the evaluation of CMV or P. carinii pneumonia.

Aged↗
PubMed2001

Observation of charge state and conformational change in immobilized protein using surface plasmon resonance sensor.

Behaviors of proteins immobilized on a solid surface were investigated using BIACORE, a biosensor utilizing surface plasmon resonance. This sensor is usually used for analyzing binding events during biomolecular interactions. Here we propose a novel use of this sensor to monitor two kinds of intramolecular changes in immobilized proteins. Several proteins were covalently attached to dextran chains on the sensor surface in the flow cell and were then exposed to a series of buffers with varying pH. Signal changes derived from changes of refractive index around the sensor surface were detected during and after the exposure to each of these buffers, which we denoted as in situ values and postvalues, respectively. The in situ value reflects the behavior of immobilized proteins in these buffers and was revealed to have a correlation with total charge state of the proteins, while the postvalue reflects how immobilized proteins react after the exposure and was suggested to represent the degree of conformational changes of the proteins. This method is expected to be applicable to various analyses and can provide us with new information about the behavior of proteins on solid phase.

Apoproteins↗
PubMed2001

Clinical utility of capillary polymerase chain reaction for diagnosis of Cytomegalovirus pneumonia.

The purpose of this retrospective study was to assess the diagnostic efficacy of CMV DNA detection by capillary PCR in patients with interstitial pneumonia. Of 882 samples taken from 363 patients, 317 were obtained from sputum, 94 from BAL fluid, 291 from blood and 180 from urine. PCR for CMV was positive in 58 samples (6.6%), with positive detection for 6.9% of sputum, 10.6% of BAL fluid, 4.1% of blood and 7.8% of urine samples. CMV pneumonia was diagnosed retrospectively in 34 (9.4%) of the 363 patients by demonstration of CMV antigen-positive cytomegalic inclusion bodies in lung tissue sections. The positive and negative predictive values were 100% (10/10) and 98.8% (83/84) for the BAL fluid samples and 95.5% (21/22) and 99.7% (294/295) for the sputum samples, respectively. Clinical sensitivity and specificity were 90.9% (10/11) and 100% (83/83) for the BAL fluid samples and 95.5% (21/22) and 99.7% (294/295) for the sputum samples, respectively. However, the blood and urine samples showed poor clinical sensitivity and low positive predictive values. We suggest that the use of capillary PCR for BAL fluid and sputum samples is very useful for diagnosing CMV pneumonia in patients with interstitial pneumonia in whom CMV pneumonia is suspected.

Adolescent↗
PubMed2001

Identification of the HLA-A24 peptide epitope within cytomegalovirus protein pp65 recognized by CMV-specific cytotoxic T lymphocytes.

Among cytomegalovirus (CMV) tegument proteins, phosphoprotein 65 (pp65) has been identified as the important target antigen of the cytotoxic T lymphocyte (CTL) response against the virus. We synthesized seven CMV-pp65-derived peptides carrying an HLA-A24-binding motif, and investigated the ability of these peptides to induce CMV-specific CTL. We identified one nonamer peptide (pp65113-121; VYALPLKML) able to bind HLA-A24 and induce CTL responses in vitro in peripheral blood mononuclear cells (PBMC) from CMV-seropositive individuals. The peptide-specific CTLs generated were capable of recognizing pp65 expressed on CMV-infected fibroblasts as well as pp65113-121 peptide bound to the surface of C1R-A*2402 cells in an HLA-A24-restricted manner. The pp65113-121 peptide thus might be considered a synthetic peptide vaccine in HLA-A24-positive individuals.

Antigens, Viral↗
PubMed2001

Off-pump Coronary Artery Bypass Grafting in patients on chronic hemodialysis.

BACKGROUND: Off-pump Coronary Artery Bypass Grafting(CABG) has gained much support but has been scarcely reported in patients on chronic hemodialysis (HD). The details of CABG for such patients on chronic HD are presented. METHODS: Between January 1998 and December 1999, off-pump CABG was performed in 5 patients on chronic HD. All patients presented with unstable angina pectoris. The indication for off-pump CABG was suitable coronary anatomy (N = 5) and expected high risks associated with cardiopulmonary bypass due to reoperation (N = 1) and impaired cerebral blood flow (N = 2). The approach was median sternotomy (N = 4) and left anterior short thoracotomy (N = 1). The mean graft number was 2.0 +/- 1.0 (range, 1-3). In situ arterial grafts were mainly used: the in situ left internal thoracic artery was used in four patients, the in situ right gastroepiploic artery in two, the in situ right internal thoracic artery in two, and the saphenous vein in two patients. RESULTS: There were no deaths or complications. Nine of 10 grafts were confirmed fully patent by postoperative angiography before hospital discharge. In a mean follow-up of 17.2 months, no patients had developed recurrence of angina or any cardiac symptom. CONCLUSION: Off-pump CABG using in situ arterial grafts can be performed on patients on chronic hemodialysis subject to suitable anatomy of the target coronary arteries.

Adult↗
PubMed2001

[Clinical significance of eosinophilic cationic protein in serum and bronchoalveolar lavage fluid of adult patients with mycoplasmal pneumonia].

Pathogenic mechanisms of mycoplasmal pneumonia is not fully understood at present though some kind of cell-mediated hypersensitivity is closely related to its mechanisms. Though eosinophilia in peripheral blood are sometimes revealed in patient with mycoplasmal pneumonia, it is not unclear whether eosinophils related to its pathogenesis, or not. We evaluated the clinical significance of ECP in serum and BAL fluid in patients with mycoplasmal pneumonia. The diagnosis of mycoplasmal pneumonia was confirmed both by serological diagnosis from paired serum and by the polymerase chain reaction (PCR) methods using specific primers of the Mycoplasma pneumoniae for detecting specific DNA from bronchial washing fluids. ECP level in serum were measured in 27 patients (11 male, 16 female, average age 31.7 yo) with mycoplasmal pneumonia by ELISA methods. ECP level in BALF were also measured in ten of all patients. The level of ECP in serum was high in 17 cases (63%) of the total cases. In addition the level of ECP in BALF was also high in all tested patients (10 cases). There was a correlation between serum ECP level and days from onset. There was also a correlation between serum ECP level and WBC counts, the degree of PaO2. These results suggested that ECP derived from activated eosinophils in the lung might in part play a role in the pathogenesis of mycoplasmal pneumonia.

Adult↗
PubMed2001

Interleukin-4 is needed for the infection of monocytes by human cytomegalovirus.

OBJECTIVE: We studied the ability of human cytomegalovirus (HCMV) to infect peripheral blood mononuclear cells (PBMC) pretreated with or without Th2-cytokine interleukin-4 (IL-4) in vitro. METHODS: Adherent cells and nonadherent cells were obtained from PBMC. We inoculated these cells with HCMV at concentrations ranging from 0 to 10 ng/ml of IL-4. Immediate-early antigen-1 (IE-1) and glycoprotein H (gH) mRNAs were detected using the reverse-transcription polymerase chain reaction. RESULTS: IE-1 and gH mRNAs could be detected in monocytes pretreated with IL-4. In contrast, no IE-1 mRNA was detected in monocytes pretreated without IL-4. We tested whether higher infectious titers could result in the infection of monocytes whether or not they were pretreated with IL-4. However, no IE-1 mRNA was detected in the monocytes not pretreated with IL-4. To elucidate how HCMV-infected monocytes affect lung tissue, human embryonic lung fibroblasts MRC-5 were cocultured with HCMV-infected monocytes. The cytopathic effects of HCMV were observed microscopically and was confirmed by direct immunoperoxidase staining with a human monoclonal antibody against the HCMV IE-1. CONCLUSION: Our data strongly suggest that the ability of HCMV to infect monocytes may correlate with the presence of IL-4.

Cytomegalovirus↗
PubMed2001

Expanded-bed protein refolding using a solid-phase artificial chaperone.

An efficient solid-phase protein refolding method based on artificial chaperone-assisted refolding is proposed. The method employs insoluble cyclodextrin polymer beads and the expanded-bed technique. Alpha-glucosidase, whose spontaneous refolding yield from a urea-denatured state is up to 30% at a protein concentration of up to 10 microg/ml, could be refolded with a yield that was improved more than two-fold at a protein concentration more than five-fold higher when protein solution was circulated through an expanded bed under optimized conditions. Unlike the conventional liquid-phase artificial system, further steps to purify the refolded product, which are generally needed to remove detergent-cyclodextrin complex and excess cyclodextrin, were unnecessary. In addition, the polymer beads were reusable after simple washing with water, and the continuous system is suitable for easy-scale-up using commercially available devices. This new method is considered to be a powerful means of achieving large-scale protein refolding for industrial protein production.

Journal Article↗
PubMed2001

In vitro transmission of HCMV between fibroblasts and peripheral blood leukocytes in the presence of IL-4.

We demonstrated transmission of human cytomegalovirus (HCMV) from the human lung fibroblast MRC-5 to peripheral blood leukocytes (PBLs). mRNA of the HCMV immediately-early (IE) antigen was detected in PBLs cultured with IL-2 or IL-2 + IL-4 that made direct contact with HCMV-infected MRC-5, whereas it was not detected in PBLs prevented from making cell-to-cell contact. However, mRNA of HCMV IE was not detected in PBLs cultured with IL-2 and IFN-gamma that made direct contact with HCMV-infected MRC-5. Transmission of the pp65 antigen was increased in culture medium containing IL-4. At a higher viral infection titer, cell-free HCMV infected adherent PBLs cells. The subset, which did not adhere, did not infect cell-free viruses even at a very high multiplicity of infection. Moreover, the adhered subset of PBLs infected with HCMV was able to transmit HCMV to non-infected fibroblasts. Our results suggest that cell-to-cell contact (when PBLs make direct contact with HCMV-infected cells) is important in the mechanism of HCMV transmission and that the adherent cells of PBLs are one of the most important vehicles for HCMV infection. Moreover, we suggest that type 2 cytokines such as IL-4 enhance the transmission of HCMV to PBLs.

Antigens, Viral↗
PubMed2001

Immediate and long-term results of coronary artery bypass operation in hemodialysis patients.

This study evaluated the early and late results of coronary artery bypass grafting (CABG) in patients on long-term maintenance hemodialysis (chronic HD) at Teikyo University Ichihara Hospital between January 1996 and June 2000. Thirty-six patients on chronic HD underwent CABG. There were 26 males (72%) and 10 females (28%) ranging from 41 to 81 years (mean +/- SD, 61.8 +/- 9.2 years) of age. Twenty-one patients (58%) had unstable angina, 14 (39%) stable angina, and 1 acute myocardial infarction. Eleven patients (31%) had urgent or emergency CABG. The average graft number was 2.5 +/- 0.8 (arterial graft 1.3 +/- 0.7/patient). Six patients had concomitant cardiac operations. Three patients underwent re- or a second re-CABG. Five patients underwent off-pump CABG. Principally, HD was performed during cardiopulmonary bypass and was followed by continuous hemodiafiltration in the early postoperative period. The early mortality was 11%; 25% in emergency and urgent CABG and 4% in elective CABG. In the follow-up period between 1 and 53 months (mean +/- SD 21.9 +/- 15.1 months), 4 patients died, and 9 patients developed recurrence of angina pectoris (6, occlusion of saphenous vein graft and 3, native coronary progression). Six patients had coronary intervention. The postoperative angiogram showed that all arterial grafts were patent, but the patency of the vein grafts was only 61.5%. The early results of CABG in patients on chronic HD was satisfactory. The late recurrence of angina pectoris mostly was caused by occlusion of the saphenous vein graft. In conclusion, the aggressive use of arterial grafts is crucial in CABG for patients on chronic HD.

Adult↗
PubMed2000

Direct refolding of recombinant human growth differentiation factor 5 for large-scale production process.

An efficient downstream process for the production of recombinant human growth differentiation factor 5 (rhGDF5) has been developed for industrial application utilizing a novel "direct refolding" method. In this method, the starting material is an inclusion body produced in Escherichia coli, and the critical step is the direct refolding step that follows directly after solubilization of the inclusion body. rhGDF5 can be refolded at a markedly high concentration of 2.4 mg.ml(-1), which is 24 times that hitherto achieved by the proteins of the TGF-beta superfamily. The refolding yield is 63%, and after purification by diafiltration, isoelectric precipitation and reverse-phase chromatography, the final purification yield is 20% with purity higher than 99%. The yield is more than twice that of a conventionally established process having three chromatography steps and the purity is equivalent. The first pilot-scale trial shows a refolding yield of 51% and a final yield of 11%. The final yield is 1.4 times that of the conventional process, and further optimization at pilot-scale is expected to bring this figure up to or above that of laboratory-scale. As a result, the calculated production cost of rhGDF5 has been reduced dramatically. This type of efficient and simple process is beneficial particularly in the large-scale production of recombinant proteins in which high yield and quality are required.

Journal Article↗
PubMed2000

[Clinical study of infection associated hemophagocytic syndrome].

Hemophagocytic syndrome (HPS) causes pancytopenia, increased blood LDH level, liver dysfunction, DIC, etc. with macrophages appearing in the bone marrow, spleen, lymph nodes, etc. Adult HPS is mostly secondary to various infections, malignant tumors, etc. and sometimes has a serious outcome. Particularly infection associated HPS (IAHS) is triggered by viral, bacterial and fungal infections. The cases of unknown primary disease and suspected IAHS of unidentified pathogenic microorganism are often encountered in the clinical setting. The authors compared IAHS and malignant associated HPS (MAHS) and classified IAHS into viral associated HPS (VAHS), bacterial associated HPS (BAHS) and fungal types to compare the test values based on the test findings at the onset in the HPS cases treated at our Department. The patients consisted of 21 HPS cases, 11 IAHS cases (VAHS 4, BAHS 5, fungal 2) and 10 MAHS cases. Based on the test findings (WBC, Hb, Plt, LDH, ferritin, myelogram, cytokines, [IFN alpha, TNF gamma, IL-6, sIL-2R, M-CSF], adhesion molecules [sICAM-1, sVCAM-1, sELAM-1, sL-selectin]) at the onset, a comparison between IAHS and MAHS and among the IAHS cases classified by pathogenic microorganism was made. In the comparison between IAHS and MAHS, the Hb value was significantly decreased and sIL-2R tended to be increased at the onset in MAHS. When comparing the IAHS cases by pathogenic microorganism, Plt was significantly decreased and sICAM-1 and sVCAM-1 were increased at the onset in the BAHS, The BAHS cases had serious underlying diseases and poor prognosis with high incidence of DIC complications. We are going to accumulate more cases for early diagnosis and treatment of IAHS.

Adolescent↗
PubMed2000

Clinical use of capillary PCR to diagnose Mycoplasma pneumonia.

In the present study, serologic data were compared with data obtained by capillary PCR to establish the efficacy of capillary PCR for the determination of Mycoplasma infection in samples obtained from throat swabs, bronchoalveolar lavage fluids (BALF), and sputum of patients with Mycoplasma pneumonia. We performed PCR analysis for Mycoplasma DNA on a total of 325 samples from 197 patients with community-acquired pneumonia and in whom Mycoplasma pneumonia was suspected. There were 68 PCR-positive specimens. Review of the differences in PCR positivity rates based on the site of specimen collection showed the highest rate of detection (28.6%) from throat swabs. From among the 31 patients with significantly elevated titers of serum Mycoplasma antibodies, the PCR results were positive for 25 patients. Thus, capillary PCR had a sensitivity of 80.6% (25 of 31). Five of the six false-negative results were from throat swab specimens. Moreover, testing (PCR) had been performed only once for these five patients with false-negative results. From among the PCR-positive findings from BALF specimens, there were no false-positive results. BALF specimens were very useful, except for the technical procedures and increased patient burden required to obtain these specimens. We suggest that the use of throat swab specimens in capillary PCR is much more suitable for diagnosing Mycoplasma pneumonia in routine clinical practice; however, careful throat swab specimen collection and an increase in the number of times that the PCR is performed are necessary to reduce the rate of false-negative results.

Bronchoalveolar Lavage Fluid↗
PubMed2000

Identification of epidermal growth factor mRNA-expressing cells in the mouse anterior pituitary.

Epidermal growth factor (EGF) produced within the pituitary gland is associated with the growth of pituitary cells in rats. The aim of the present study was to localize EGF- and EGF receptor-expressing cells, and to clarify the involvement of EGF in DNA replication in 2-month-old male mouse pituitary cells. In situ hybridization of the pituitaries of these mice demonstrated that EGF mRNA was expressed in the anterior and intermediate lobes. Within the anterior pituitary, EGF mRNA-expressing cells were medium-sized and round, and made up 40% of the total number of secretory cells. EGF receptor mRNA was only detected in anterior pituitary cells. Forty-seven percent anterior pituitary cells expressed EGF receptor mRNA. An immunocytochemical study showed that most somatotropes and some mammotropes expressed EGF mRNA. When anterior pituitary cells were enzymatically dissociated and cultured in serum-free medium, RT-PCR demonstrated both EGF mRNA and EGF receptor mRNA expression. Treatment with EGF (1 and 10 ng/ml) for 5 days stimulated DNA replication in mammotropes and corticotropes. These results indicate that the DNA replication in mammotropes and corticotropes is regulated by the paracrine and/or autocrine activity of EGF produced at least in part by these cell types themselves.

Animals↗
PubMed2000

Synthesis and secretion of A-type natriuretic peptide in the auricular cardiocytes during pregnancy and lactation in mouse.

A-type (atrial) natriuretic peptide (ANP) levels in the auricular cardiocytes and plasma were examined by immunohistochemistry, electron microscopy, and radioimmunoassay in pregnant and lactating mice. Additionally, the cardiocyte ANP mRNA expression was measured by the polymerase chain reaction method. ANP-immunoreactivity (IR) and the number of ANP-granules in the cardiocytes on the 18th day of gestation were greater than those in virgin controls, but the plasma ANP concentration decreased on the 18th day of gestation. On the day of delivery, ANP-IR and the number of ANP-granules in the cardiocytes were decreased compared to those during the pregnancy and to those in virgin controls, and then began to increase continually until the 15th day of lactation. Plasma ANP concentration after delivery was significantly higher than that during pregnancy, and than that in virgin controls, and continued to increase until the 15th day of lactation. Cardiocyte ANP mRNA expression was highest on the day of delivery compared to that in all the other times. In conclusion, these results suggested that the circulating systems of ANP during pregnancy and lactation were regulated differentially.

Animals↗