[Overview: practical psychotherapy and technical psychotherapy].
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Biofeedback, Psychology↗
Biomedical subjects
Kenji Yamamoto
Publications and source records attributed to Kenji Yamamoto.
Involvement of cathepsin E in exogenous antigen processing in primary cultured murine microglia.
We have attempted to elucidate an involvement of cathepsin E (CE) in major histocompatibility complex class II-mediated antigen presentation by microglia. In primary cultured murine microglia, CE was localized mainly in early endosomes and its expression level was markedly increased upon stimulation with interferon-gamma. Pepstatin A, a specific inhibitor of aspartic proteases, significantly inhibited interleukin-2 production from an OVA-(266-281)-specific T helper cell hybridomas upon stimulation with native OVA presented by interferon-gamma-treated microglia. However, pepstatin A failed to inhibit the presentation of OVA-(266-281) peptide. The possible involvement of CE in the processing of native OVA into antigenic peptide was further substantiated by that digested fragments of native OVA by CE could be recognized by OVA-specific Th cells. Cathepsin D also degraded native OVA into antigenic peptide, whereas microglia prepared from cathepsin D-deficient mice retained an ability for antigen presentation. On the other hand, the requirement for cysteine proteases such as cathepsins S and B in the processing of invariant chain (Ii) was confirmed by immunoblot analyses in the presence of their specific inhibitors. In conclusion, CE is required for the generation of an antigenic epitope from OVA but not for the processing of Ii in microglia.
Animals↗
Selection-by-function: efficient enrichment of cathepsin E inhibitors from a DNA library.
A method for efficient enrichment of protease inhibitors out of a DNA library was developed by introducing SF-link technology. A two-step selection strategy was designed consisting of the initial enrichment of aptamers based on binding function while the second enrichment step was based on the inhibitory activity to a protease, cathepsin E (CE). The latter was constructed by covalently linking of a biotinylated peptide substrate to each of the ssDNA molecule contained in the preliminarily selected DNA library, generating 'SF-link'. Gradual enrichment of inhibitory DNAs was attained in the course of selection. One molecule, SFR-6-3, showed an IC(50) of around 30 nM, a K(d) of around 15 nM and high selectivity for CE. Sequence and structure analysis revealed a C-rich sequence without any guanine and possibly an i-motif structure, which must be novel to be found in in vitro-selected aptamers. SF-link technology, which is novel as the screening technology, provided a remarkable enrichment of specific protease inhibitors and has a potential to be further developed.
Amino Acid Sequence↗
Roles for Arg- and Lys-gingipains in the disruption of cytokine responses and loss of viability of human endothelial cells by Porphyromonas gingivalis infection.
Accumulating evidence indicates that periodontal disease is associated with human cardiovascular diseases. The periodontal pathogen Porphyromonas gingivalis was shown to be present in atherosclerotic plaques in addition to periodontal pockets. This bacterium is known to produce two individual cysteine proteinases, Arg-gingipain (Rgp) and Lys-gingipain (Kgp). Here we show that these two enzymes are responsible for either the disruption of cytokine responses in human umbilical vein endothelial cells (HUVEC) to the bacterium infection or the loss of cell viability. The expression of interleukin-8 and monocyte chemoattractant protein-1 mRNA in HUVEC was greatly induced when infected with the wild-type strain, nevertheless, their protein levels in the culture medium were markedly decreased. This decrease was completely abolished in the cells infected with the Rgp/Kgp-null mutant, but not in either the Rgp- or Kgp-null mutants. Loss of the adhesion activity and viability of HUVEC were greatly induced by the culture supernatant of the wild-type strain and strongly inhibited by either a combination of the Rgp- and the Kgp-specific inhibitors or the deficiency of the Rgp- and Kgp-encoding genes. These findings indicate that P. gingivalis modulates the cytokine response in the cells and disrupts the adhesion activity and the viability through the cooperative action of Rgp and Kgp and thereby may contribute to pathogenesis of cardiovascular diseases as well as periodontal disease.
Adhesins, Bacterial↗
Phase I and pharmacological study of single paclitaxel administered weekly for heavily pre-treated patients with epithelial ovarian cancer.
We have reported that paclitaxel results in cisplatin sensitization in cisplatin-resistant ovarian cancer cell lines in vitro and in nude mice. The purpose of this trial was to determine the maximum tolerated dose and recommend phase II dose of weekly single agent paclitaxel for outpatients with recurrent or persistent epithelial ovarian carcinoma (REOC), with standard chemotherapy containing platinum in the initial setting. Patients with REOC were eligible for this protocol regardless of the number and kind of previous chemotherapy regimens. The starting dose was paclitaxel 70 mg/m2/week in 1-hour infusion, 3 weeks on, 1 off and repeated at least twice. This dose was increased by 10 mg per step to 100 mg/m2/week. Three patients were accrued to each dose cohort. Three new patients were to be entered at escalation doses unless dose-limiting toxicities (DLT) occurred, defined as grade 4 hematological or grade 3/4 non-hematological toxicities. If 1 out of 3 patients developed DLT, 3 additional patients were entered at the same dose level. Sixteen patients were accrued. All the patients had received at least one prior platinum-containing regimen (1 regimen 14 cases, 2 regimens 1 case, 3 regimens 1 case). At the level I dose of 70 mg/m2/week no hematological or non-hematological toxicity more than grade 2 was observed. At the level II dose of 80 mg/m2/week, 1 patient had grade 4 non-hematological toxicity, showing difficulty-walking. Three new additional patients were treated with the same dose. Except for this patient, 1 had grade 3 leukopenia and grade 4 neutropenia, but these toxicities were overcome within 3 days without support of granulocyte-colony stimulating factor (G-CSF). At the level III dose, 90 mg/m2/week, 1 of 3 patients showed grade 4 leukopenia and 2 had grade 4 neutropenia, requiring support by G-CSF. Similarly, when using 100 mg/m2/week of paclitaxel, 2 out of 4 patients had more than grade 3 hematological toxicity. However, at levels II or IV, no non-hematological toxicity exceeding grade 2 was observed. Even if the weekly single paclitaxel was repeated, the toxicity did not seem to accumulate. According to dose-escalation, use of G-CSF and treatment delay were increased. The use of G-CSF was significantly (p<0.05) increased between levels I, II, III and IV. Although treatment with 90 or 100 mg/m2/week, at 3 weeks on, 1 off was tolerable and safe with support of G-CSF, these doses cannot be recommended for out-patients because of treatment delay. In this phase I trial, 80 mg/m2/week of paclitaxel was recommended as the phase II dose for outpatients.
Adult↗