Visualization of mitochondria with green fluorescent protein in cultured fibroblasts from patients with mitochondrial diseases

cDNAs for green fluorescent protein (GFP) and for a GFP fusion protein containing the presequence of human ornithine transcarbamylase (pOTC-GFP) were transfected into cultured human fibroblasts. GFP cDNA gave diffuse fluorescence throughout the cytoplasm and the nucleus, whereas pOTC-GFP cDNA gave mitochondria-associated fluorescence. Fluorescent mitochondrial structures could be classified into five patterns: thread-like mitochondria, fine thread-like ones, rod-like ones, granular ones, and granular ones with weak cytosolic fluorescence. pOTC-GFP mutants resulted in a loss of mitochondrial fluorescence and an appearance of weak fluorescence throughout the cytoplasm. pOTC-GFP cDNA was transfected into fibroblasts from patients with various mitochondrial diseases. Higher ratios of fibroblasts with granular mitochondria and those with fine thread-like ones were observed in a patient with Reye's syndrome and a patient with Kearns-Sayre syndrome. Weak cytosolic fluorescence was sometimes observed in fibroblasts from these patients. This method will be useful to analyze mitochondrial structural alterations and disorders of mitochondrial protein import.     

High performance 35 nm gate length CMOS with NO oxynitride gate dielectric and Ni salicide

The 35 nm gate length CMOS devices with oxynitride gate dielectric and Ni salicide have been fabricated to study the feasibility of higher performance operation. Nitrogen concentration in gate oxynitride was optimized to reduce gate current I/sub g/ and to prevent boron penetration in the pFET. The thermal budget in the middle of the line (MOL) process was reduced enough to realize shallower junction depth in the S/D extension regions and to suppress gate poly-Si depletion. Finally, the current drives of 676 /spl mu/A//spl mu/m in nFET and 272 /spl mu/A//spl mu/m in pFET at V/sub dd/=0.85 V (at I/sub off/=100 nA//spl mu/m) were achieved and they are the best values for 35 nm gate length CMOS reported to date.     

A Novel Artificially Humanized Anti-Cripto-1 Antibody Suppressing Cancer Cell Growth

Cripto-1 is a member of the EGF-CFC/FRL1/Cryptic family and is involved in embryonic development and carcinogenesis. We designed a novel anti-Cripto-1 artificial antibody and assessed the recognition to the antigen and the potential to suppress the growth of cancer stem cells. First, single chain antibody clones were isolated by bio-panning with the affinity to recombinant Cripto-1 protein from our original phage-display library. Then, the variable regions of heavy chain VH and light chain VL in each clone were fused to constant regions of heavy chain CH and light chain CL regions respectively. These fused genes were expressed in ExpiCHO-S cells to produce artificial humanized antibodies against Cripto-1. After evaluation of the expression levels, one clone was selected and the anti-Cripto-1 antibody was produced and purified. The purified antibody showed affinity to recombinant Cripto-1 at 1.1 pmol and immunoreactivity to cancer tissues and cell lines. The antibody was available to detect the immunoreactivity in tissue microarrays of malignant tumors as well as in Cripto-1 overexpressing cells. Simultaneously, the antibody exhibited the potential to suppress the growth of human colon cancer derived GEO cells overexpressing Cripto-1 with IC₅₀ at approximately 110 nM. The artificially humanized antibody is proposed to be a good candidate to target cancer cells overexpressing Cripto-1.     

Preparation and properties of graft and block copolymers of poly(p-phenylene terephthalamide) with polybutadiene

Graft copolymers of polybutadiene (PBD) onto poly(p-phenylene terephthalamide) (PPTA) were prepared by the nucleophilic substitution of N-metalated PPTA with telechelic PBD having bromide end groups. Block copolymers were synthesized by the condensation reaction of telechelic PBD having acid chloride end groups with amino-group-terminated PPTA. The structure of these copolymers was identified by IR spectra. Graft and block copolymers contained PBD segments up to 85 wt % and 45 wt %, respectively. Thermomechanical analyses (TMA) proved the existence of distinctive primary absorption peak corresponding with T_g of PBD for both graft and block copolymers. The T_g's of both types of the copolymers were further ascertained by the DSC curves. TMA curves suggested that the microphase separation occurred between PPTA and PBD. The incorporation of PPTA segments into PBD increased the decomposition temperature compared with the blend polymer composed of PPTA and PBD with the same composition.     

Structural significance of the alkyl substituent at the C-4 of (+)-trans-verbenyl acetate in sex pheromonal activity of the American cockroach

The significance of the alkyl group at the C-4 of (+)-trans-verbenyl acetate, which is the sex pheromone mimic of the American cockroach, was investigated. Seven alcohols possessing an ethyl, propyl, or dimethyl group at this position of the 6,6-dimethylbicyclo[3.1.1]heptane skeleton were synthesized and evaluated by behavioral assay. All of the alcohols were inactive, while three of four acetates of the 2-alcohols induced sexual behavior in male cockroaches at the 0.02 or 0.5 mg dosage level, either of which is many orders of magnitude higher than the threshold level of the natural sex pheromones (10^–8 mg). Among the acetates, the compounds with a methyl group or an -oriented ethyl group at C-4 showed the highest activity. The results are discussed in terms of spatial requirements of the molecules for interactions with the receptor.Studies on the sex pheromone mimic of the American cockroach, (+)-trans-verbenyl acetate. Part VIII. For Part VII, seeComp. Biochem. Physiol.,70A: 229–234 (1981).     

Preparation of poly(p-phenylene terephthalamide)/poly(vinyl chloride) molecular composite and its thermal and mechanical properties

Poly(p-phenylene terephthalamide) (PPTA) was blended with poly(vinyl chloride) (PVC) by solution-blending method. PPTA was metalated for dissolving in dimethyl sulfoxide. Dimethyl sulfoxide was used as a common solvent. In PPTA/PVC composite, PPTA accelerated the thermal degradation of PVC. PPTA molecules are aggregated as microfibrillar form in PVC matrix. Such microfibrils are dispersed homogeneously in PVC matrix, according to polarizing microscopic observation. The average diameter of the microfibrils becomes smaller in the composite with lower content of PPTA. In the surface region of PPTA microfibrils the intermolecular hydrogen bonds between C? Cl of PVC and N? H of PPTA are formed. Young's modulus and the yield stress at room temperature were higher in the composites than those in PVC. The modulus of the composites was higher, especially at the high temperatures above their glass transition temperatures, than that in PVC. The temperature dependence of modulus can be calculated by using the mechanical model equivalent to the quasi-3-dimensional microfibrillar model which will be approximately applied to the composite structure. It becomes apparent that the modulus of the PPTA microfibrils evaluated by using the mechanical model is higher in the higher molecular weight PPTA.     

Two-stage nuclear demagnetization refrigerator

Two-stage nuclear demagnetization has been performed using PrCu₆ and Cu as coolants. The Cu nuclear stage reached temperatures as low as 10 K with a rate of temperature rise of less than a few K/h. The corresponding conduction electron temperature is estimated to be less than 50 K. A nuclear orientation thermometer of Al ⁵⁴Mn has been successfully used to measure temperatures down to 10 K.Supported by the Japan Society for the Promotion of Science.     

Simultaneous STM and UHV electron microscope observation of silicon nanowires extracted from Si(111) surface

A miniaturized scanning tunnelling microscope (STM) was fitted in a side-entry holder of an ultra-high vacuum electron microscope. The clean Si(111)7 x 7 surface was observed by both STM and reflection electron microscopy (REM) at atomic resolution. The tungsten tips were often rounded off upon tip-approach with a constant current, through a gentle touch with the sample surface. The apices of such rounded tips had radii of several tens of granometre with widths of about 3 x 3 nm. Atomically resolved STM of the Si(111)7 x 7 surface was obtainable when an atom or an atomic cluster sits on the tip surface. The rounded tips were used for fabrication of Si nanowires by the touch-and-away operation of the tip. The nanowires grew longer at higher substrate temperature and they reached as long as several tens of nanometre at 700 degrees C. The nanowire had many twins and the (111) twin lamellae were stacked in the direction of the wire axis. In another case, the twin planes were oblique to the wire axis so that the (112) direction was nearly parallel to the wire axis.     

The operation mechanism of a charge modulation device (CMD) imagesensor

The operational mechanism of the charge modulation device (CMD) as an image sensor is presented. A newly developed device simulator, the transistor analysis program for imagers calculating non-steady-state equations (TRINE) is used in this analysis. Comparison of the calculated results with measured data shows that the model predictions are consistent with measured electrical and optical characteristics within a range of 20%. The operational mechanism has been clarified, and the accuracy of the method proves that it is effective in quantitative device design. The discretization scheme of the cylindrical coordinate system used is also shown     

A case of gastric cancer with peritonitis carcinomatosa and malignant biliary stenosis treated by intraperitoneal CDDP and MMC with UFT therapy

A 55-year-old man, who complained of vomiting, was diagnosed as having a Borrmann type 3 gastric cancer (T3N3P2H0M0: Stage IVb). He was treated by distal gastrectomy. After four months, PTCD was applied for malignant biliary stenosis due to lymphnode metastasis. Since ascites due to peritonitis carcinomatosa developed about six months after operation, UFT therapy combined with CDDP 50 mg and MMC 10 mg intraperitoneally were performed. The patient became gradually less aware of subjective symptoms after chemotherapy. Three months later, he enjoyed a good general condition and all of the fixed tubes were removed. This chemotherapy seemed effective to support his quality of life.