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Background— Apoptosis is an important cause of early graft loss after heart transplantation. Bcl-xL was reported to protect the heart against normothermic ischemia and reperfusion injury. In this study, we determined whether overexpression of Bcl-xL could inhibit tissue injury resulting from prolonged cold preservation followed by warm reperfusion of heart transplants. Methods and Results— Lewis rat hearts were transduced with an adenovirus vector harboring Bcl-xL cDNA (AxCAhBclxL) 4 days before collection of tissue. After preservation in University of Wisconsin solution at 4°C for 24 hours, the heart was either perfused with a Langendorff device ex vivo or used for heterotopic heart transplantation in vivo. Bcl-xL gene transfer significantly reduced the infarct size (23.0±2.6% versus 47.7±7.0% in saline control and 48.6±6.1% in vector control, P P c release from the mitochondria; it also significantly decreased cardiac cell apoptosis and improved graft survival rate after long cold preservation, followed by warm reperfusion. Conclusions— Bcl-xL gene transfer inhibited the translocation of Bax and prolonged the cold preservation time of cardiac transplants. This may be a potential therapeutic method in clinical practice.

Cracks in concrete structure cause deterioration of durability due to corrosion of reinforcement. Although crack width is controlled in less than the allowable value in the present design method, concrete structure with cracks deteriorate more easily than that without cracks. The objective of this study is to develop the reinforced concrete member that a crack doesn't occur in serviceability limit state. This study gave attention to a super high strength of Reactive Powder Composite (RPC), especially a high tensile strength on crack development. RPC consists of premixed powder in selected combination of cement, silica particles, siliceous sand, special water reducing agent, and steel fibers.A part of tension zone in reinforced concrete beam was fortified by RPC. The cross-sectional area of RPC part was varied in three ways. Flexural load tests of beam were carried out. The crack development moment of the beams fortified by RPC increased with the increase of reinforcing area of RPC. When the bending moment that the cracks with permissible crack width for corrosion of reinforcement develop in the usual reinforced concrete beam acted, cracks didn't occur in the beams reinforced with RPC suitably. The flexural behaviours of the beam reinforced with RPC could be analysed by elastic theory in the range of service state. When the bending moment on the occurrence of the crack is estimated, the restraint stress due to autogeneous shrinkage of RPC must be taken into consideration.

Mesenchymal stem cells (MSC) were reported to ameliorate functional deficits after stroke in rats, with some of this improvement possibly resulting from the action of cytokines secreted by these cells. To enhance such cytokine effects, we previously transfected the telomerized human MSC with the BDNF gene using a fiber-mutant adenovirus vector and reported that such treatment contributed to improved ischemic recovery in a rat transient middle cerebral artery occlusion (MCAO) model. In the present study, we investigated whether other cytokines in addition to BDNF, i.e., GDNF, CNTF, or NT3, might have a similar or greater effect in this model. Rats that received MSC-BDNF (P < 0.05) or MSC-GDNF (P < 0.05) showed significantly more functional recovery as demonstrated by improved behavioral test results and reduced ischemic damage on MRI than did control rats 7 and 14 days following MCAO. On the other hand, rats that received MSC-CNTF or MSC-NT3 showed neither functional recovery nor ischemic damage reduction compared to control rats. Thus, MSC transfected with the BDNF or GDNF gene resulted in improved function and reduced ischemic damage in a rat model of MCAO. These data suggest that gene-modified cell therapy may be a useful approach for the treatment of stroke.

Gene targeting using short interfering RNA (siRNA) has become a common strategy to explore gene function because of its prominent efficacy and specificity. For the application of siRNA technology to gene therapy, however, still more efficient transduction of siRNA into target cells is needed. In this study, we developed an adenoviral vector harboring a tandem-type siRNA expression unit, in which sense and antisense strands composing the siRNA duplex were separately transcribed by two human U6 promoters. Targeting survivin, an antiapoptotic molecule widely overexpressed in malignancies but not detected in terminally differentiated adult tissues, this type of adenoviral vector (Adv-siSurv) successfully exerted a gene knockdown effect and induced apoptosis in HeLa, U251, and MCF-7 cells. These cancer cells, once infected with Adv-siSurv, displayed remarkably attenuated growth potential, both in vitro and in vivo. Moreover, intratumoral injection of Adv-siSurv significantly suppressed tumor growth in a xenograft model using U251 glioma cells. This novel modality may be a promising tool for cancer therapy.

Examination of the clinical therapeutic efficacy of using bone marrow stromal cells, including mesenchymal stem cells (MSC), has recently been the focus of much investigation. MSC were reported to ameliorate functional deficits after stroke in rats, with some of this improvement possibly resulting from the action of cytokines secreted by these cells. To enhance such cytokine effects, we transfected telomerized human MSC with the BDNF gene using a fiber-mutant F/RGD adenovirus vector and investigated whether these cells contributed to improved functional recovery in a rat transient middle cerebral artery occlusion (MCAO) model. BDNF production by MSC-BDNF cells was 23-fold greater than that seen in uninfected MSC. Rats that received MSC-BDNF showed significantly more functional recovery than did control rats following MCAO. Specifically, MRI analysis revealed that the rats in the MSC-BDNF group exhibited more significant recovery from ischemia after 7 and 14 days. The number of TUNEL-positive cells in the ischemic boundary zone was significantly smaller in animals treated with MSC-BDNF compared to animals in the control group. These data suggest that MSC transfected with the BDNF gene may be useful in the treatment of cerebral ischemia and may represent a new strategy for the treatment of stroke.

The expression of CD154 (CD40 ligand) on activated CD4+ T cells is known to be transient and tightly regulated for antigen-specific immune responses, and is increased and prolonged among patients with systemic lupus erythematosus (SLE). We investigated the regulation of CD154 expression by determining the protein and mRNA expression with PMA and ionomycin stimulation in CD4+ T cells, and confirmed their increase and prolongation in SLE T cells. Treatment with actinomycin D, a transcription inhibitor, after PMA and ionomycin stimulation was performed, and the findings revealed that the stability of CD154 mRNA increased significantly in activated SLE T cells compared with that of controls. However, alternations or abnormal sequences were not identified in the 3″ untranslated region, including AU-rich elements and CU-rich sequences, while their partial involvement in the posttranscriptional regulation of CD154 mRNA stability has been reported. With 96 h culture in vitro, the destabilization of CD154 mRNA was demonstrated, resulting in a corresponding decrease and normalization of surface expression on activated SLE T cells. We speculate that the CD154 expression on T cells from SLE patients may be increased and prolonged, with mRNA stabilization being related to a continuous stimulation in vivo.

Background Although naked plasmid injection is the safest and most convenient method for gene delivery, a major limitation of this approach is currently poor transgene expression. The CA promoter (chicken β-actin promoter with cytomegalovirus, CMV, enhancer) is one of the strongest transcriptional control modules found; however, it is uncertain whether a CA promoter-based vector is efficient enough for naked gene therapy in a cardiovascular context. Methods The β-galactosidase (LacZ) expression provided by CA promoter plasmid (pCAZ2) injection into the skeletal muscle or the heart of Lewis rats was compared with CMV promoter plasmid or adenoviral vector (AxCAZ3). The effect of Simian virus 40 of the replication origin (SV40ori) deletion from pCAZ2 on transgene expression was also evaluated. Results pCAZ2 showed the highest LacZ expression in both skeletal muscle and heart in comparison with the CMV promoter-based vector 5 days after naked plasmid injection. LacZ expression in the heart obtained using 20 µg of pCAZ2 was almost equivalent to that shown with AxCAZ3 at 6.0 × 109 optical particle units. The time course of transgene expression driven by CMV and CA promoters in the heart were similar, with the CA promoter providing significantly higher gene expression than the CMV promoter across all time points examined. SV40ori deletion from pCAZ2 did not affect transgene expression in either skeletal muscle or heart. Conclusions Transgene expression mediated by naked CA promoter-based plasmid injection was shown to be quite efficient in the heart. We propose that the CA promoter vector is suitable for myocardial gene therapy. Copyright © 2003 John Wiley & Sons, Ltd.

In the present study, unpurified peripheral blood mononuclear cells (PBMCs) from various sources, including steady-state blood (normal donors) and granulocyte colony-stimulating factor (G-CSF)-mobilized blood (cancer patients and normal donors) (G-PBSC), were cultured in RPMI-1640 in the presence of IL-2 and alpha-galactosylceramide (alpha-GalCer) to expand V alpha 24(+) T cells, and their expansion kinetics were compared. G-CSF-mobilized cells showed markedly higher expansion potential (350-fold expansion of V alpha 24(+) T cells, regardless of whether the cells were from cancer patients or normal donors) than steady-state cells (15-fold expansion, compared to the initial inoculums) (n = 5, P < 0.01). We also confirmed that the CD14(-) fraction of G-PBSCs contained a large number of precursors of V alpha 24(+) T cells, compared to PBSCs, as well as a large number of CD14(+) cells, which assist V alpha 24(+) T cell proliferation. Our simple and practical procedure, which eliminates complicated cell manipulation (including cell purification), produces efficient expansion of V alpha 24(+) T cells when G-CSF-mobilized blood cells are cultured with alpha-GalCer.

Vapor grown carbon fiber (VGCF) is a kind of carbon nanotube (CNT), which has outstanding properties such as high mechanical strength and high electrical conductivity. In this study, injection molding properties of polystryrene (PS) filled with VGCF and evaluation of mechanical and electrical properties are discussed in comparison with composites in which conventional carbon fillers were filled. As a result, volume resistivity of VGCF/PS composites dropped significantly between VGCF concentration of 3 and 4 vol.%. Resistivity of the composites filled with VGCF was 1.2 × 10 2 Ω.cm when CGCF concentration was 11.6 vol.%. The resistivity was significantly lower than that of composites which were filled with conventional carbon fillers. The elastic modulus slightly increases with increasing VGCF concentration. whereas the tensile strength slightly decreases in the VGCF concentration in the range from 0 to 12 vol.%.

Sequence variability of the control region of mitochondrial DNA in five Sebastes species (S. thompsoni, S. joyneri, S. inermis, S. schlegeli and S. owstoni) was investigated. Of 324 nucleotide sites in the control region of S. thompsoni, 56 sites (17.3%) varied, and all 20 individuals had different haplotypes. The other four species had between five and 17 sites (1.5–5.9%), which was fewer than that observed in S. thompsoni. The nucleotide diversity of S. thompsoni was highest (3.45%) among the five species, and that of S. schlegeli was the lowest (0.19%). The results demonstrated that sequence variability exists in the control region of the five Sebastes fishes investigated, and that sequence data in the control region may be suitable for stock structure analysis. Also, the extent of genetic variablitiy in the control region differed among these species. In particular, the mitochondrial DNA control region in S. thompsoni was characterized by high sequence variability, which may indicate a large effective population size.

U1B3.3 monoclonal antibody (mAb) was established by immunizing a rat with the tMK-2U lymphoma cell line, derived from athymic nude mice. U1B3.3 mAb recognizes some T cell populations, B cells and natural killer (NK) cells. U1B3.3+ T cells express CD3/T cell receptor (TCR) complex at a unique intensity: between that of TCR-intermediate cells (TCRint cells) and TCRbright cells. TCRint cells strongly express the interleukin-2 receptor β chain (IL-2Rβ), but almost all U1B3.3+ T cells express high or low levels of IL-2Rβ. When purified U1B3.3+ NK1.1-T cells were cultured with IL-2, approximately 14% of the cells acquired expression of NK1.1 molecules, which are expressed on NK cells and natural killer T (NKT) cells. Furthermore, U1B3.3+ NK1.1-T cells killed tumor cells after culture with IL-2. These results indicate that U1B3.3 mAb recognizes a precursor of NK1.1+ cytotoxic T cells generated by IL-2.

The ratio of the surface area to the volume of materials increases in inverse proportion to their size and therefore the surface area of nanostructures and nanomaterials is extremely large compared to that of macroscopic materials of the same volume, thanks to which it is supposed that chemical and biochemical reactions may be greatly enhanced and target molecules and cells may be efficiently trapped on the surface of nanomaterials. It is well known that C60 molecules are stable both physically and chemically and the affinity of C60 molecules with biomolecules is rather high. Here, we synthesise fibres composed of C60 and sulphur and immobilise the surface of the fibres with the primary antibody; i.e., epithelial cell adhesion molecules (anti-EpCAM), to trap target cells. The primary antibody is evenly immobilised on the fibres confirmed by a fluorescent secondary antibody attached to the primary one and then TE2 esophageal and DLD-1 colon cancer cells are successfully trapped by the primary antibody immobilised on the fibres thanks to its high affinity with TE2 and DLD-1 cells, whereas few IM9 B lymphoblast cells are captured on the fibres since the affinity of the primary antibody with IM9 cells is extremely low. Furthermore, those cells trapped by the primary antibody immobilised on the fibres proliferate faster than native cells thanks to the primary antibody acting as a growth factor. The present result suggests that different types of cells can be trapped and grown on nano fibres by immobilising appropriate antibody molecules on the surface of the fibres. Even an extremely small number of cells in sample fluids may be analysed and characterised for the detection of diseases such as cancer in the early stage by trapping and proliferating target cells on the fibres.

We present evidence that T-cell-conditioned media (TCCM) can efficiently induce human immature dendritic cells (DC) to express high levels of immune accessory molecules commonly found on mature DC. TCCM prepared from cell-free supernatants of anti-CD3-activated T cells contained several soluble factors including CD40-ligand (sCD40L), TNF-α, and IFN-γ. In contrast to moderate up-regulation of costimulatory molecules by the addition of individual cytokines or monocyte-conditioned medium, treatment of immature DC with TCCM induced a marked increase in the expression of costimulatory molecules in a dose-dependent manner. The ability of TCCM to induce such phenotypic changes could be abrogated by neutralizing antibodies specific for CD40L, TNF-α, and IFN-γ, indicating that these factors present in TCCM are mainly implicated in the maturation of DC. Importantly, TCCM-treated DC can produce significantly higher levels of IL-12 and are highly effective stimulators in allogenenic and autologous mixed-lymphocyte reactions. Overall, these findings show that cultivation with TCCM is an efficient approach for the induction of mature DC that should be useful in eliciting antigen-specific immune responses against cancer and viruses.

The authors previously proposed an energy method using finite-element division and applied it to the analysis of the flow of material. In this work, the authors develop this method further to obtain the stress distribution in caliber rolling. Recently, Tomita obtained the stress distribution using the grid method so that the equilibrium condition may be satisfied approximately, whereas Mori evaluated the accuracy of the result obtained by the rigid-plastic finite-element method, by considering the equivalent nodal force which is assumed to be in equilibrium. Hence, the authors combined these methods and applied them to determine the stress distribution. The following results were obtained. First, in the flat rolling of bars, the roll force agrees well with the theoretical equation proposed by Saito. Second, in square-diamond rolling and round-oval rolling, the roll force depends on the ratio of the indirectly reduced area in the cross-section to the area of the cross-section. Finally, in square-oval rolling, the roll force agrees with the theoretical equation proposed by Saito when the projected material-roll contact length is replaced by the mean projected material-roll contact length in the width direction.

Melt front roughness was examined firstly, and it was found that an injected part is separated into two regions: a rough surface region and a fine surface region. The rough surface region results from melt front asperities generated by the blowout of voids at the front surface which are mixed into the material by jetting flow in the sprue. It was also found that voids in the sprue can be eliminated by applying counter-pressure. Secondly, the generation property of welding defects, weld line and voids, has been examined by counter-flow joining and their elimination criteria are obtained. It was shown that both criteria approximately coincide with each other. The welding-defect elimination criterion obtained is applied to the evaluation of welding-defect generation under various injection conditions and it was found that the criterion does not depend greatly on the asperities of the flow front. Furthermore, the criterion is also effective for estimating welding defects in injection molding using a mold cavity with inserts. Finally, it was confirmed that the above criterion approximately coincides with that in a previous paper which was obtained by the use of simple void-reducing conditions in the joining of green ceramic blocks with artificially grooved surfaces.

A new method of joining metal sheets using a short pipe rivet is proposed. A pipe rivet is set between two sheets and compressive force is applied, then both ends of the pipe are impressed into the sheets, respectively, and joining is made. Here, the inner corners of both ends of the pipe rivet are cut off conically so that the diameter of ends may be enlarged during impression and a tight joining is obtained. Consequently, it was found that joining strength against in-plane and out-of-plane forces is quite high. Furthermore, a modified method by using a wavy ribbon rivet is proposed and it was found that this method can be applied to the joining of a long joining line successfully.

In injection molding of an optical disk, mis-transcription of pits sometimes occurs, especially when pit density is low and smooth surface part is dominant. Deformation of a mold and injection molding machine may possibly make influence on transcription property in such situation. In this paper, direct measurement of mold cavity gap has been carried out, together with microscopic observation of transcription state, and the relationship between them was discussed. Finally, the following conclusions were obtained. (1) Low stiffness of mold closing system is suitable for precise transcription of a fine pattern, because cavity pressure decrease in cooling state is retarded. (2) Mold closing force should be applied to the mold itself, settled on the fixed platen, both in usual injection and injection compression molding, for preventing detachment of the mold from the platen due to nozzle contacting force.

In the injection molding of an optical disk, a precise minute pit pattern is transcribed from a nickel sheet stamper in 0.3mm thickness. Various defects or mis-transcriptions are sometimes generated, but the causes of these defects have not yet been completely clarified. Thus ideal molding conditions and optimum mold designs can not be precisely determined.In this paper, the injection molding of an optical disk for a sample-servo-type application is performed by using an injection-compression molding system and the generation mechanism of transcription defects was studied. The following conclusions were obtained.: (1) Mis-transcriptions are classified into three types; multiple transcription errors, scratches in radial directions and scratches in other direction. Their occurrence is affected by the compression pressure holding time and mold temperature. (2) In the case of rather low mold temperatures, multiple transcription errors occur when the compression time is short, but scratches occur when the compression holding time is long. (3) A relative displacement of the polymer molding from the stamper cccurs when a cooling time is longer than 10s, which results in multiple transcription error.

In injection molding of short fiber-polymer melt mixture, influence of the interaction between a fiber and a mold wall on fiber distribution and flow property cannot be neglected, when fiber length is not very short comparatively to the thickness of a mold cavity. In the flow through a cooled mold cavity, solidifying layer grows and it affects that relation further. In this paper, effiects of these causes on fiber orientation and velocity distribution have been made clear by using numerical analysis, in which each fiber motion is analyzed from equilibrium conditions and fiber orientation distribution is obtained by integrating frequency of orientation angles of each fiber over the whole angles, and the following results have been obtained. (i) Near a mold wall, mixture is cooled and no flow layer is generated. On the other hand, apparent viscosity increases in the region, apart from a mold wall by more than half of fiber length, because of high fiber concentration, and plug flow-like velocity distribution is generated. (ii) Pressure gradient increases toward downstream in low injection rate, because of cooling of mixture, whereas it decreases in high injection rate, because of viscosity decrease due to shear heating. (iii) As fiber length decreases, pressure gradient decreases. (iv) Fiber orientation distribution changes considerably with injection rate change.

We have previously established and reported a novel monoclonal antibody (mAb), U5A2-13, which recognizes a phenotypically similar population of natural killer (NK)-like T cells. Using U5A2-13 mAb, we now describe the functional properties of U5A2-13(+) T cells in both NK1.1-positive or -negative mouse strains. Similar to NK1.1(+) T cells, hepatic U5A2-13(+) T cells of C57BL/6 (NK1.1(+) strain) mice, but not U5A2-13(-) T cells, could be induced to produce large amounts of IL-4 and IFN-gamma by stimulation with glycolipid alpha-galactosylceramide (alpha-GalCer) present on dendritic cells (DC) in a dose-dependent manner. The abundant production of these cytokines from U5A2-13(+) T cells of BALB/c (NK1.1(-) strain) mice is similar to that noted in C57BL/6 mice. Cytokine production by cultures stimulated with DC of beta2-microglobulin-deficient mice was significantly less than that of cultures stimulated with DC of intact mice. Overall, U5A2-13(+) T cells recognize alpha-GalCer presented by CD1d, indicating that U5A2-13(+) T cells can be used to analyze NK-like T cell function in various strains of mice.

Both the therapeutic and preventative effects of a murine T-cell line, tMK-2, with T-cell receptor (TCR) alpha/beta positive and CD4-/8- double negative (DN) phenotype against autochthonously tumors induced by subcutaneous (s.c.) injection of 3-methylcholanthrene (MC) were examined. Complete regression of the tumor was observed when administration of tMK-2 cells was begun on tumors 5 mm in diameter. The tumor mass in five out of five mice was reduced in size after the administration of tMK-2 cells regardless of the routes of administration: s.c. injection of tMK-2 cells (5 x 10(7) cells) once a week around tumors, intraperitoneal (i.p.) injection (5 x 10(7) cells), or intravenous (i.v.) injection (1 x 10(7) cells). The tumors regressed to the status of a scar within 1 month of initial injection, and this status was maintained throughout the remainder of the 3 months period of tMK-2 cell injection. One month after discontinuation of tMK-2 cell administration, the diameter of the tumors had not increased regardless of the route of injection. The control groups consisted of either untreated mice, mice with i.v. injection of 1 microg of recombinant murine interleukin (IL)-12 once a week, or mice with s.c. injection of autologous splenocytes (5 x 10(7)) from BALB/c mice once a week. Continuous growth of tumors was observed in each group and all control mice died due to bleeding ulcerations of the tumors. Tumor development was effectively prevented when tMK-2 cells were administrated 1 week after the s.c. injection of MC. In the groups receiving s.c., i.p., and i.v. injection of tMK-2 cells, no MC-induced tumors developed, whereas four out of five of the control mice developed autochthonous tumors. The tMK-2 cells also exerted in vitro NK-like cytotoxic activity, and their killing activity was strongly increased in the presence of both IL-2 and IL-12. These results suggest that the injected T-cells with TCR alpha/beta positive and CD4- /8- DN phenotype and NK-like activity are important in the therapy as well as the prevention of tumor development.

Immunosuppressive potentials of the blockade of intercellular adhesion molecule-1 (ICAM)-1/leukocyte function-associated antigen 1 (LFA-1) were examined in a murine islet allotransplantation model by using blocking monoclonal antibodies (MAbs) against these molecules. Isolated islets from ICR mice were transplanted into the renal subcapsular space of streptozotocin-induced diabetic C57BL/6 mice. Antibodies were administered immediately after transplantation at a dose of 100 micrograms/mouse/d for 3 or 7 d. In non-treated mice, islet grafts were rejected within 16 d, but the treatment with an anti-ICAM-1 MAb (KAT-1) alone, with anti-LFA-1 MAb (KBA) alone, or with both MAbs significantly prolonged the graft survival. In particular, the combination of KAT-1 and KBA in a 7-d course produced a marked prolongation and induced indefinite graft survivals over 100 d in 88% of recipients. Expression of cytokine transcripts within the islet allografts was analyzed by reverse transcriptase polymerase chain reaction (RT-PCR). In the mice treated with KAT-1 and KBA, the transcripts for Th1 cytokines (interleukin 2 [IL-2] and interferon gamma [IFN-gamma]) were not detected, but the expression of Th2 cytokines (IL-4 and IL-10) was enhanced and persisted over 140 d. In contrast, Th1 cytokines were dominantly expressed in the grafts from untreated mice. These results indicate that administration of anti-ICAM-1 and/or anti-LFA-1 MAbs prolongs murine islet allograft survival potentially by indicating a Th2 deviation.

In order to induce tumor specific cytotoxicity, the poorly immunogenic murine colon cancer cell colon26 was transfected with murine IL-2 cDNA and/or IL-12 cDNA and their anti-tumor effects were investigated. Double transfectants produced murine IL-2 and murine IL-12, the same as single transfectants. Intraperitoneal administration of double transfectants inhibited pulmonary metastasis of colon26 inoculated intravenously to a stronger degree than that of single transfectants. Splenocytes from mice administered double transfectants intraperitonealy showed higher cytolytic activity against colon26 than those from mice administered single transfectants, and also showed cytolytic activity against murine B16-BL6 melanoma. In the NK cell-depleted mice, double transfectants inhibited pulmonary metastasis from the control markedly, but could not do completely, the same as in the NK cell-reserved mice. The difference of the metastatic colonies between NK cell-depleted mice and the control was much greater than that between NK cell-depleted mice and NK cell-reserved mice. These results suggested that cytotoxic T lymphocytes might participate in this anti-tumor effect.

Capillary flow experiments were performed to evaluate the slip property of injection moldable ceramic compounds of Al2O3, and the following results were obtained. Firstly, slip velocity us was obtained by applying the Mooney method, and the effects of material temperature, particle size and binder composition on the slip velocity were evaluated. Secondly, it was found that the slip velocity changed significantly, when counter pressure was applied at the exit of the capillary. Then, the slip velocity was expressed by an empirical equation considering the pressure effect. Thirdly, an accurate viscosity equation was obtained, based on the pressure dependent slip behavior. Lastly, the flow behavior in a capillary was discussed in consideration of the slip behavior, and the application of counter pressure was found to be effective for realizing high strain rate flow.

Generation of molding defects in ceramic injection molding, e. g. melt front asperity or surface roughness of an injected part, has been examined by using radial flow in a circular mold cavity. The following results were obtained. Firstly, it was found that an injected part is separated into 2 regions: region I with rough surface and region II with smooth and fine surface. The rough surface of region I results from a rough melt front, which moves to the mold surface and forms the surface of an injected part in the early filling stage. Secondly, front asperity is generated by the blowout of voids at the front surface, which are mixed into the material and by jetting flow. Voids in the sprue can be prevented by applying counter pressure. An injection molding method using a film gate with proper gap and a side gate is proposed. In this method, high pressure is applied to the filling material in the sprue to reduce voids, and it becomes possible to mold parts with smooth surfaces by removing voids in the sprue and jetting flow in the mold cavity.

Transparent conducting thin fillns made of Indium oxide (In2O3) have been prepared by a mechanical process conposed of rolling' and oxidizing stages without vacuum procesSeS. The In ingot was rolled to approximately 5μm in thich first, and then the In sheet was pretreated by corona discharge in the air. Finally, the sheet was annealed in a furnace in the temperature range from 300°C to 700°C. The corona discharge pretreatment was found to have an important role to oxidize the In sheets due to the introduction of the oxygen species through micro pits formed by electron and ion bombardments. The transmission coefficient of the In2O3 film prepared by this process was 40%, when the corona discharge time was 3 h and heat treatment temperature was 700°C. The resistivity of this ITO film was 8.0 × l0 -5Ω·m according to the conventional four-probe measurement.

CD40-CD40-ligand (CD154) interactions play a critical role in immune activation. Using replication defective adenovirus encoding mouse CD154 (Ad-CD154), we modified human chronic lymphocytic leukemia B cells to express a functional ligand for CD40. This not only induces expression of immune accessory molecules on the infected cell, but also allows it to trans-activate noninfected bystander leukemia B cells. Also, factors that impair the antigen-presenting capacity of leukemia B cells are downmodulated. Ad-CD154- infected leukemia cells are highly effective stimulators in mixed lymphocyte reactions and can induce generation of cytotoxic T lymphocytes specific for autologous nonmodified leukemia cells. As such, Ad-CD154 can induce a host antileukemia response that may have therapeutic potential.

Selective growth of aluminum nitride (AlN) and zinc oxide (ZnO) has been performed by an Ar-laser-enhanced reactive vapor deposition method on Si (100) substrates. The substrate was selectively heated by laser irradiation. Al and Zn were evaporated from Ta crucibles by an electron gun in NH3 and O2 atmospheres, respectively. The metal vapor reacted with atmospheric gas on the substrate surface due to laser heating, when the temperature of the laser-irradiated part was higher than 450°C for AlN and 230°C for ZnO. Not only the substrate temperature, but also the thickness of the Al and Zn layers, which are deposited during the time interval between the first laser irradiation and subsequent laser irradiation, was an important factor for realizing the selective growth. The critical thickness was found to be 0.2 nm and 6 nm for AlN and ZnO, respectively. Two-dimensional selective growth and three-dimensional micro free-form fabrication of AlN were performed under these conditions using a two-dimensional laser scanning setup. The AlN layer was successfully deposited in accordance with a laser drawing pattern, even when the drawing pattern changed during synthesis.

The fiber volume fraction and fiber orientation play an important role in both the rheological analysis of molten polymer and mechanical properties of a finished part in injection molding. Many works have been devoted to the motion of a fiber, rheology of suspension and fiber-fiber interaction for dilute and semi-dilute systems, but less for concentrated suspension, and also, less works to the interaction between fiber and die surface. In this paper, equations are derived to describe the motion of fibers for concentrated suspension basing on the equilibrium of force and torque acted on each fiber in a simple shear flow, where fiber-die surface interaction is numerically analyzed, and fiber-fiber interaction is considered by modifying Tuckers' model. It is found that fiber volume fraction near die surface decreases and fiber orientation distribution along the flow direction is obtained. It results in decrease of viscosity and increase of velocity gradient near die surface.

A neural model is proposed for the spatiotemporal properties of simple cells in the visual cortex. In the model, several cortical cells are arranged on a ring, with mutual excitatory or inhibitory connections. The cells also receive excitatory inputs either from lagged and nonlagged cells of the lateral geniculate nucleus in one setting or from nonlagged cells in the other. Computer simulation shows that the cortical cells have spatiotemporally inseparable receptive fields in the former setting and separable fields in the latter; spatial profiles at a given time in the spatiotemporal fields are described with a Gabor function whose phase parameter varies regularly from 0 to 2 pi with rotation along the ring; the inseparable cells have directional selectivity as observed physiologically.

Experimental autoimmune uveoretinitis (EAU) induced by immunization with interphotoreceptor retinoid-binding protein (IRBP), a retinal self antigen, has been regarded to be a typical T helper type 1 (Th1)-mediated inflammatory disease. In this study, we examined the effect of a neutralizing monoclonal antibody (mAb) to interleukin-12 (IL-12), which has been known to play a critical role in the Th1 differentiation, on the development of EAU. While 9 of 13 control mice developed EAU by the immunization with IRBP, none of 12 mice developed EAU when given anti-IL-12 mAb 1 day before immunization. These mice did not develop EAU even after a rechallenge with IRBP on day 30, indicating that a protective mechanism had been established by the anti-IL-12 treatment. The proliferative response of splenocytes to IRBP in vitro was not significantly impaired, but the production of IL-2 and IFN-gamma was greatly reduced by the anti-IL-12 treatment. Moreover, production of IL-5 and expression of IL-4 mRNA were increased by the anti-IL-12 treatment. Consistently, IgG2a anti-IRBP serum antibodies were decreased and IgG1 were increased. Administration of a neutralizing anti-IL-4 mAb at the time of IRBP rechallenge reversed the protection established by the anti-IL-12 treatment at the primary immunization. These results indicate that the anti-IL-12 treatment at the IRBP priming not only prevented the development of pathogenic Th1 cells, but also induced suppressive Th2 cells that protect the animals from further challenge with the same antigen.

In recent years, research on resistance to chemotherapy and radiotherapy in cancer treatment has come under the spotlight, and researchers have also begun investigating the relationship between resistance and cancer stem cells. Cancer stem cells are assumed to be present in esophageal cancer, but experimental methods for identification and culture of these cells have not yet been established. To solve this problem, we created spheroids using a NanoCulture® Plate (NCP) for 3-dimensional (3-D) cell culture, which was designed as a means for experimentally reproducing the 3-D structures found in the body. We investigated the potential for induction of cancer stem cells from esophageal cancer cells. Using flow cytometry we analyzed the expression of surface antigen markers CD44, CD133, CD338 (ABCG2), CD318 (CDCP1), and CD326 (EpCAM), which are known cancer stem cell markers. None of these surface antigen markers showed enhanced expression in 3-D cultured cells. We then analyzed aldehyde dehydrogenase (ALDH) enzymatic activity using the ALDEFLUOR reagent, which can identify immature cells such as stem cells and precursor cells. 3-D-cultured cells were strongly positive for ALDH enzyme activity. We also analyzed the expression of the stem cell-related genes Sox-2, Nanog, Oct3/4, and Lin28 using RT-PCR. Expression of Sox-2, Nanog, and Lin28 was enhanced. Analysis of expression of the hypoxic surface antigen marker carbonic anhydrase-9 (CA-9), which is an indicator of cancer stem cell induction and maintenance, revealed that CA-9 expression was enhanced, suggesting that hypoxia had been induced. Comparison of cancer drug resistance using cisplatin and doxorubicin in 3-D-cultured esophageal cancer cells showed that cancer drug resistance had increased. These results indicate that 3-D culture of esophageal squamous cell carcinoma lines is a useful method for inducing cancer stem cells.

The p40 subunit of interleukin 12 (IL-12p40) has been known to act as an IL-12 antagonist in vitro. We here describe the immunosuppressive effect of IL-12p40 in vivo. A murine myoblast cell line, C2C12, was transduced with retro-virus vectors carrying the lacZ gene as a marker and the IL-12p40 gene. IL-12p40 secreted from the transfectant inhibited the IL-12-induced interferon gamma (IFN-gamma) production by splenocytes in vitro. Survival of C2C12 transplanted into allogeneic recipients was substantially prolonged when transduced with IL-12p40. Cytokine (IL-2 and IFN-gamma) production and cytotoxic T lymphocyte induction against allogeneic C2C12 were impaired in the recipients transplanted with the IL-12p40 transfectant. Delayed-type hypersensitivity response against C2C12 was also diminished in the IL-12p40 recipients. Furthermore, serum antibodies against beta-galactosidase of the T-helper 1-dependent isotypes (IgG2 and IgG3) were decreased in the IL-12p40 recipients. These results indicate that locally produced IL-12p40 exerts a potent immunosuppressive effect on T-helper 1-mediated immune responses that lead to allograft rejection. Therefore, IL-12p40 gene transduction would be useful for preventing the rejection of allografts and genetically modified own cells that are transduced with potentially antigenic molecules in gene therapy.

Micro flow marks have been found to occur in injection molding of polycarbonate. Their minimum depth is ˜ 5 nm on the mold smooth surface. Such marks seem to be formed when the molten polymer near the mold wall at the advancing front is cooled to the no-flow temperature of resin before it comes into contact with the mold wall, preventing the precision transcription of the mold smooth surface. We investigated the molding factors affecting their occurrence by heat transfer analysis. It was found that the size of micro flow marks is related to the melt temperature, mold temperature, no-flow temperature, thermal diffusivity of the resin, and time pitch formation of flow mark ridges and valleys. The relationships between those factors and the sizes of micro flow marks are clarified quantitatively, and the guidelines for precision transcription in injection molding are formulated.

Adhesion molecules of the integrin family, including very late activation antigens (VLA), have been implicated in various cellular functions. In this study, we investigated the contribution of integrin-mediated interaction with ECM proteins to the cytokine gene expression in human chrondrocytes. Human articular chronrdocytes expressed VLA-1, -2, -3 and -5 on the cell surface, and could adhere to various ECM proteins, especially to fibronectin (FN). Futhermore, the production of GM-CSF and IL-6 was potently induced by culturing chondrocytes on immobilized FN. This stimulative effect of FN was completely inhibited by an anti-integrin α5 chain mAb, as well as by anti-integrin β1 chain mAbs. These results indicate an important role of the VLA-5-mediated interaction with FN in regulating inflammatory cytokine production by human articular chondrocytes.

The beta 1 integrin subfamily, alternatively called very late activation antigen (VLA), has been implicated in various cellular functions. In this study, we generated a mAb against the mouse beta 1 subunit (CD29) to examine the functional property of mouse VLA proteins. After immunization with affinity-purified mouse VLA-4 (alpha 4 beta 1), a hamster mAb, HM beta 1-1, was established by screening mAb that reacted with alpha 4-negative neuroblastoma C1300. The antigen defined by HM beta 1-1 was widely distributed in various mouse cell lines and HM beta 1-1 immunoprecipitated a 110-120 kDa protein common to VLA-1 and VLA-6, indicating that HM beta 1-1 recognizes the beta 1 subunit of mouse integrins. We then examined the inhibitory effect of HM beta 1-1 on VLA-dependent cell adhesion and activation. HM beta 1-1 blocked the adhesion of mouse tumor cell lines to extracellular matrix proteins including collagen, laminin and fibronectin. Moreover, splenic T cell proliferation induced by anti-CD3 mAb and allogeneic mixed lymphocyte response were strongly inhibited by HM beta 1-1 in combination with an anti-LFA-1 mAb. We conclude that HM beta 1-1 reactive with mouse CD29 can inhibit VLA-dependent cellular functions and, thus, would be useful for studying the physiological role of beta 1 integrins in vivo.

In injection molding of substrates for optical discs, precise transcription of pits and grooves is very important. For high-density optical discs, molding transcription for minute grooves with widths on the sub-micron level is required. In this paper, in order to clarify the relationship between molding conditions and transcription quantitatively, transcription of minute rectangular grooves (width 0.55 μm, depth 70 nm) in injection molding of polycarbonate resin was studied. The relationship between molding conditions and the transcription heights was verified. And, by introducing the equivalent thickness of the vitrified layer concept, a transcription model related to the minute groove width is proposed. This equivalent thickness of the vitrified layer is estimated to be 100 to 150 nm at the point of filling completion under a certain molding condition. Also, using the above description of the equivalent thickness of the vitrified layer, it is possible to estimate the extent of transcription under the various molding conditions.

Alpinists who challenge Mt. Everest need adaptation to hypoxia before the attack of Mt. Everest. Although this adaptation is important for the success of climbing Mt. Everest, the molecular mechanism on the adaptation to hypoxia is not well understood. In order to clarify this mechanism, we investigated hypoxia-induced gene expressions specific for top alpinists using microarray analyses. We report here that heme oxygenase-1 (HO-1) is significantly higher in the blood of top alpinist compared with non-alpinists. Although HO-1 expression of non-alpinists is also up-regulated in response to hypoxia, HO-1 level of the top alpinists are constitutively higher than that of non-alpinists. Serial examinations of HO-1 in one top alpinist revealed that the higher expression of HO-1 is maintained in high-level several months after the attack of top mountains. Taken together with the biochemical function of HO-1 that catalyzes heme into CO and bilirubin, HO-1 expression may improve the circulation and compensate with oxidative tissue damages induced by hypoxia. These data also suggest that peripheral blood has the memory on hypoxia independent of antigens by maintaining the high-level of HO-1 expression in top alpinists, which merits the rapid adaptation to hypoxia for 8000m climbing.