Biomedical subjects

Yukako Yamazaki

Publications and source records attributed to Yukako Yamazaki.

4 recordsLinked to original sources

PubMed2003

Polycation liposome-mediated gene transfer in vivo.

The polycation liposome (PCL), a recently developed gene transfer system, is simply prepared by a modification of liposomes with cetylated polyethylenimine (PEI), and shows remarkable transgene efficiency with low cytotoxicity. In the present study, we investigated the applicability of PCLs for in vivo gene transfer, since the PCL-mediated transgene efficiency was found to be maintained in the presence of serum. PCLs composed of dioleoylphosphatidylethanolamine (DOPE) with 5 mol% cetyl PEI (PEI average mr. wt. 1800), were superior for transfection to those of dipalmitoylphosphatidylcholine (DPPC) and cholesterol (2:1 as molar ratio) with 5 mol% cetyl PEI in vitro, although the latter PCLs were more efficient for gene transfer in vivo. PCL-DNA complexes were injected into mice via a tail or the portal vein, with the DNA being a plasmid encoding green fluorescent protein (GFP) or luciferase; and the expression was monitored qualitatively or quantitatively, respectively. Tail vein injection resulted in high expression of both GFP and luciferase genes in lung, and portal vein injection resulted in high expression of both genes in the liver. Concerning the gene delivery efficiency, the PCL was found to be superior to PEI or cetyl PEI alone. The optimal conditions for in vivo transfection with PCLs were also examined.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed2003

Super-parallel MR microscope.

A super-parallel MR microscope in which multiple (up to 100) samples can be imaged simultaneously at high spatial resolution is described. The system consists of a multichannel transmitter-receiver system and a gradient probe array housed in a large-bore magnet. An eight-channel MR microscope was constructed for verification of the system concept, and a four-channel MR microscope was constructed for a practical application. Eight chemically fixed mouse fetuses were simultaneously imaged at the 200 micro m(3) voxel resolution in a 1.5 T superconducting magnet of a whole-body MRI, and four chemically fixed human embryos were simultaneously imaged at 120 micro m(3) voxel resolution in a 2.35 T superconducting magnet. Although the spatial resolutions achieved were not strictly those of MR microscopy, the system design proposed here can be used to attain a much higher spatial resolution imaging of multiple samples, because higher magnetic field gradients can be generated at multiple positions in a homogeneous magnetic field.

Anatomy, Cross-Sectional↗

PubMed2002

New and better protocols for a short-term Caco-2 cell culture system.

The aim of the present study was to develop new and better protocols for a short-term Caco-2 cell culture system for use in rapid screening of intestinal drug absorption. Caco-2 cells were cultured according to several protocols for short-term cell culture to obtain monolayers. The effects of serum (fetal bovine serum, FBS) in the culture medium and of the period of cell culture on the barrier function and transporter activities of the monolayers were examined. The barrier function was estimated both from the transepithelial electrical resistance (TEER) and the permeability of [(14)C]mannitol. Transporter activities were monitored by measuring the permeability of [(14)C]glycylsarcosine for oligopeptide transporter (PepT1) and of rhodamine 123 for P-glycoprotein (P-gp). Caco-2 monolayers obtained by 3-day culture in the BIOCOAT HTS Caco-2 Assay System, developed by Becton Dickinson Bioscience, showed much higher permeability to hydrophilic compounds, such as mannitol, compared with those obtained by the standard 21-day culture system, due to the leaky structure of cell junctions. The newly developed 3-day protocol, which includes 10% FBS in the culture medium during the first day of culture, markedly enhanced TEER and lowered mannitol permeability of the monolayers. This protocol allowed us to better determine the rank order of permeability of compounds, giving results equivalent to those in the 21-day culture system. The longer culture period gave tighter monolayers, and the maximum value of TEER was obtained with 5 days in culture. However, after 5 days in culture, the integrity of monolayers decreased gradually. The highest activities of transporters, PepT1 and P-gp, in monolayers were obtained at days 5 or 6 of culture by the new protocol with FBS-containing medium. These results indicate that by a simple modification of the short-term culture protocol, it is possible to obtain Caco-2 monolayers with better barrier properties and higher activity of transporters that are equivalent to those found in the 21-day Caco-2 culture system.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2002

Role of integrin alpha(v)beta3 in the early phase of liver metastasis: PET and IVM analyses.

To clarify the function of integrin alpha(v)beta3 in the early stage of liver metastasis, we investigated the interactions of metastatic cells with their target organ under the actual blood flow by using positron emission tomography (PET). The cells used were CHO-K1 cells and their transfectants bearing human integrin alpha(v)beta3 cDNA (alpha(v)beta3-CHO-K1 cells). The liver accumulation of alpha(v)beta3-CHO-K1 cells was significantly higher than that of CHO-K1 cells after injection via the portal vein, whereas no significant difference was observed in the lung accumulation after tail vein injection, suggesting a specific interaction of alpha(v)beta3-CHO-K1 cells with the hepatic sinusoids. Furthermore, to clarify the precise location of each cell in the liver, i.e., to determine whether individual cells were intravascularly localized or had extravasated, we performed intravital fluorescence microscopy (IVM) on the liver by using stable transfectants bearing the green fluorescent protein (GFP) gene, namely, GFP-CHO-K1 and GFP-alpha(v)beta3-CHO-K1 cells. Both types of cells remained in the hepatic blood vessels 1 h after injection via the portal vein. On the other hand, expression of integrin alpha(v)beta3 promoted the cells to reach the extravascular region after 24 h. These results suggest the possibility that the specific accumulation of alpha(v)beta3-CHO-K1 cells in the liver is followed by migration of the cells into the extravascular region. Interestingly, the adhesion of the two types of cells to hepatic sinusoidal endothelial cells in vitro did not correspond to in vivo accumulation of these cells. Therefore, integrin alpha(v)beta3 may function to promote extravasation of integrin alpha(v)beta3-expressing tumor cells in liver through a process possibly mediated by vitronectin produced by this organ.

Animals↗