Biomedical subjects

Shinya Kato

Publications and source records attributed to Shinya Kato.

5 recordsLinked to original sources

PubMed2007

Hydroxyapatite formed on/in agarose gel induces activation of blood coagulation and platelets aggregation.

We reported earlier that hydroxyapatite (HA) formed on/in agarose gels (HA/agarose) produced by alternate soaking process is a bone-filling material possessing osteoconductive and hemostatic effects. This process could allow us to make bone-like apatite that was formed on/in organic polymer hydrogel matrices. Here, we investigated the mechanism of hemostasis induced by HA/agarose and found that HA/agarose, but not agarose or HA powder, significantly shortened activated partial thromboplastin time (APTT). While HA/agarose did not show significant platelet aggregation, it markedly enhanced adenosine diphosphate (ADP)-induced platelet aggregation. Moreover, Western blot analysis revealed selective adsorption of vitronectin onto HA/agarose. We also observed marked differences between HA powder and HA/agarose in their XRD patterns. The crystallinity of HA powder was much higher compared to that of HA/agarose. Furthermore, 50-100 nm of tube-form aggregations was observed in HA powder on the other hand 100-200 nm of particles was observed in HA/agarose by SEM observation. Thus 100-200 nm of low crystallized particles on the surface structure of HA/agarose may play an important role in hemostasis. Our results demonstrated a crucial role of HA/agarose in the mechanism of hemostasis and suggested a potential role for HA/agarose as a bone-grafting material.

Adenosine Diphosphate↗

PubMed2005

Rho/Rho-kinase pathway contributes to C-reactive protein-induced plasminogen activator inhibitor-1 expression in endothelial cells.

OBJECTIVE: Rho/Rho-kinase pathway plays pivotal roles in cardiovascular diseases including arteriosclerosis and hypertension. Recently it has become evident that C-reactive protein (CRP), a powerful marker for cardiovascular events, has direct proatherothrombotic effects on vascular cells. However, its molecular mechanism has not been fully investigated. We examined the involvement of Rho/Rho-kinase signaling in CRP-induced plasminogen activator inhibitor-1 (PAI-1) expression in bovine aortic endothelial cells (BAECs). METHODS AND RESULTS: PAI-1 expression was determined by Western blotting. RhoA activation was determined by an affinity pull-down assay using Rho-binding fragment of rhotekin. NF-kappaB activity was determined using the luciferase reporter gene. Incubation of BAECs with human recombinant CRP (> or =25 microg/mL) induced a significant increase in PAI-1 expression. Stimulation of BAECs with CRP significantly increased RhoA activation. Pretreatment with TAT-C3 (a membrane-permeable RhoA inhibitor) and Y-27632 (Rho-kinase inhibitor) significantly inhibited CRP-induced PAI-1 expression. NF-kappaB activity was markedly enhanced by CRP and pretreatment with Y-27632 inhibited its activation. Parthenolide, SN50, and BAY 11-7082 (NF-kappaB inhibitors) significantly blocked CRP-mediated PAI-1 expression. CONCLUSIONS: These data suggested that CRP activates Rho/Rho-kinase signaling, which in turn activates NF-kappaB activity, resulting in PAI-1 expression in BAEC. These observations provide evidence for the possible involvement of Rho/Rho-kinase signaling in CRP-induced atherothrombogenesis.

Animals↗

PubMed2005

The "spanning protocol": a new DNA extraction method for efficient single-cell genetic diagnosis.

PURPOSE: We evaluated methods of preparation of DNA from single cells for amplification and preimplantation genetic diagnosis (PGD), including our "spanning protocol." METHODS: Dystrophin gene exons 45 and 51 were amplified by nested polymerase chain reaction (PCR) from a single lymphocyte or blastomere. Amplification efficiencies were compared between DNA extraction by (A) lysis in distilled water with freeze-thawing and boiling; (B) two-step lysis involving potassium hydroxide and dithiothreitol; and (C) the spanning protocol, using N-lauroylsarcosine. RESULTS: With method A, amplification efficiency was 66/120 (55%) and false-positive such as amplification failure or allele drop out was 42/120 (35%); with B, 96/120 (80%) and 21/120 (17.5%); and with C, 111/120 (92%) and 5/120 (4.2%), using single blastomeres and unaffected lymphocytes from male. Occurrence of false-negative such as contamination of another DNA with method A was 4/120 (3.3%); with B, 10/120 (8.3%); and with C, 2/120 (1.7%) from using single lymphocytes from affected males. CONCLUSION: The spanning protocol was most efficient for extracting DNA from a single cell and should be particularly useful for preimplantation genetic diagnosis.

Blastomeres↗

PubMed2005

Phosphatidylserine- and integrin-mediated phagocytosis of apoptotic luteal cells by macrophages of the rat.

Corpora lutea disappear from ovaries in the absence of conception. The present study was undertaken to examine the hypothesis that disappearance of corpora lutea is accomplished through apoptosis-dependent phagocytosis of luteal cells. When bone marrow cells expressing green fluorescence protein were transplanted into X-ray-irradiated mice, macrophages derived from donor mice were detected within corpora lutea, suggesting macrophage infiltration into the tissue. Dispersed rat luteal cells underwent spontaneous apoptosis during culture and were phagocytosed by luteal macrophages. Treatment with doxorubicin increased the extent of apoptosis in luteal cells, and those cells were more efficiently phagocytosed than cells left untreated. The phagocytosis was inhibited by liposomes containing phosphatidylserine or a peptide containing the integrin-targeted sequence, and was stimulated by milk fat globule epidermal growth factor 8. These results collectively indicate that apoptotic luteal cells are phagocytosed by macrophages in a manner mediated by phosphatidylserine and integrin.

Animals↗

PubMed2004

Substrate-dependent cellular behavior of Swiss 3T3 fibroblasts and activation of Rho family during adhesion and spreading processes.

Recent biochemical studies revealed that intracellular Rho guanosine triphosphatases (Rho, Rac1, and Cdc42) are key regulatory molecules that link surface receptors to cytoskeletal organization and regulation of cell shape/morphology/motility. In this study, Swiss 3T3 fibroblasts were cultured on three representative substrates [tissue culture polystyrene dishes, nontreated polystyrene, and poly(ethylene terephthalate)] for 24 h after plating. Time-dependent changes in cell shape, morphology, cytoskeletal dynamics, and motility as well as Rho family activities were determined on each substrate. The cells on tissue culture polystyrene and on poly(ethylene terephthalate), which induced rapid and relatively rapid cell spreading, respectively, expressed Rac1 and Cdc42 activities continuously during the observation period. In contrast, such activities were suppressed in cells on polystyrene, which induced slow spreading but the highest cell motility compared with the other two substrates. Although a clear-cut relationship between cellular behavior and Rho family activation was not obtained, substrate-dependent coordinated control of cellular activities by Rho family is discussed.

Animals↗