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Cdc42, a Rho family low molecular weight G protein, has important roles in various cell functions, including cytoskeletal rearrangement, cell adhesion and cell proliferation and differentiation. To investigate the involvement of Cdc42 in the activities of vascular endothelial cells, we generated Cdc42 conditional knockout mice in which Cdc42 was time -specifically deficient in vascular endothelial cells (Cdc42 ​fl/fl; VE-Cad CreERT: Cdc42 cKO). When the Cdc42 gene was deleted after birth, Cdc42 cKO mice were smaller than the control mice, and died between postnatal day 8 (P8) and P10. Necropsy findings confirmed that these mice had various pathological aberrances in the vessels of most organs, such as blood flow congestion and blood cell invasion. Electron microscopic observations also revealed that capillary endothelial cells were detached from the basement membrane as well as phagocytosis of dead endothelial cells induced by macrophages. Moreover, vascular sprouting from aortic rings induced by VEGF-A was diminished in samples from the Cdc42 cKO mice because of an endothelial cell proliferation defect. These results suggest that Cdc42 in vascular endothelial cells has important roles in blood vessel formation after birth.

AIM The TRICOLORE trial previously demonstrated that S-1 and irinotecan plus bevacizumab was non-inferior, based on progression-free survival (PFS), to 5-fluorouracil, leucovorin and oxaliplatin (mFOLFOX6)/capecitabine and oxaliplatin (CapeOX) plus bevacizumab as first-line chemotherapy for metastatic colorectal cancer (mCRC). Overall survival (OS) data were immature at the time of the primary analysis. METHODS In total, 487 patients from 53 institutions with previously untreated mCRC were randomly assigned (1:1) to receive either mFOLFOX6/CapeOX plus bevacizumab (control group) or S-1 and irinotecan plus bevacizumab (experimental group; 3- or 4-week regimen). The final OS data were analysed from follow-up data collected until 30th September 2017. RESULTS With a median follow-up period of 48.7 months, median survival times were 32.6 and 34.3 months (hazard ratio [HR]: 0.89, 95% confidence interval [CI]: 0.72-1.10, P = 0.293) and median PFS durations were 10.8 and 14.0 months in the control and experimental groups, respectively (HR: 0.86, 95% CI: 0.71-1.04, P

Nephronectin (Npnt), an extracellular matrix protein, is a ligand for integrin α8β1 and is involved in the development of various organs, such as the kidneys, bones, liver, and muscles. Previously, we found that Npnt expression was inhibited by various cytokines including transforming growth factor-β (Tgf-β) and oncostatin M (Osm). Fibroblast growth factor (Fgf)-2, otherwise known as basic Fgf, also plays important roles in skeletal development and postnatal osteogenesis. In this study, Npnt expression was found to be suppressed by Fgf-2 in MC3T3-E1 cells, an osteoblast-like cell line, in a dose- and time-dependent manners. Furthermore, Fgf-2-mediated Npnt mRNA suppression was shown to involve the Jun N-terminal kinase (JNK) and phosphoinositide-3 kinase (PI3K) pathways. Together, our results suggest that FGF-2 suppresses Npnt gene expression via JNK and PI3K pathways.

The laser ablation (LA) method is an effective technique for quantitative analysis. In the present work, a new LA system was developed for the high-sensitivity analysis of metal materials using inductively coupled plasma mass spectrometry (ICP-MS). This system consists of a high-frequency Q-switched laser and 2 scanning mirrors for scanning the ablation spot in an adequately large area of the specimen without vacant spaces. The influence of elemental fractionation (non-stoichiometric generation of vapor species) can be eliminated by repetitive irradiation of this pattern on the same area. Particles generated with an average laser power of 0.6 W with the developed LA system gave intensity and stability substantially similar to that of a 500 microg/ml solution steel sample in solution ICP-MS. The analytical performance of the developed LA-ICP-MS was compared with that of a solution ICP-MS using NIST steel SRMs. The performance of the newly-developed system is comparable to that of conventional solution ICP-MS in both accuracy and precision. The correlation coefficients between the contents and the intensity ratios to Fe were over 0.99 for most elements. The relative standard deviation (RSD) obtained by LA-ICP-MS revealed that this system can analyze iron samples with good precision. The results of ultra trace level analysis of high-purity iron showed that developed LA-ICP-MS is capable of analyzing ppm concentration levels with a 20 - 30 ppb level standard deviation. The detection limit was on the order of 10 ppb for most elements.

誘導結合プラズマ質量分析（ICP-MS）装置を用いた金属材料の高感度な直接分析を目的として，新たなレーザーアブレーション（LA）システムを開発した．装置は繰り返し周波数の高いQスイッチレーザーと，試料表面の適切な大きさの領域に対して隙間なくビームの集光スポットを走査させるための2枚のスキャンミラーを基本構成とする．この方法で，同一領域に対してレーザー照射を何度も繰り返すことにより，気化微粒子の選択蒸発を解消した．0.6 Wの平均出力で鉄鋼試料にレーザーを照射し，生成微粒子をICP-MSに導入した場合に検出されるFeのイオン強度とその変動は，500 μg/mlの鉄鋼溶解試料を噴霧法で導入した場合とほぼ同等のレベルであった．NIST製の鉄鋼認証標準物質を用い，開発したLA-ICP-MSの分析特性を溶液ICP-MSと比較した．ほとんどの元素について，Feとのイオン強度比と濃度の相関係数は0.99以上であり，繰り返しの分析精度も良好であった．開発したLA-ICP-MSでは，高純度鉄中のppmレベルの含有成分をおおむね3％ 以下の相対標準偏差で分析することができ，多くの元素について検出下限は数十ppb以下であった．

A previous study reported that a nondifferentiating myeloid leukemia cell line produced differentiation-inhibiting factors. One of the factors was purified as a homologue of the nm23 genes. Thenm23 genes were overexpressed in acute myelogenous leukemia (AML) cells, and a higher level of nm23 gene expression was correlated with a poor prognosis in AML. The present study determined the plasma levels of nm23-H1 protein by enzyme-linked immunosorbent assay and assessed the association between this level and the clinical outcome in 102 patients with AML. The plasma concentration of nm23-H1 was higher in patients with AML than in normal controls (P = .0001). Plasma nm23-H1 levels were correlated with the product of the intracellular nm23 messenger RNA (mRNA) level and the white blood cell count, but not with the mRNA level alone. Therefore, nm23-H1 plasma levels probably depend on the total mass of leukemic cells overexpressing the nm23-H1 gene. Overall survival was lower in patients with higher plasma nm23-H1 levels than in those with lower levels. Multivariate analysis using the Cox proportional hazard model showed that elevated plasma nm23-H1 levels significantly contributed to the prognosis of AML patients. Furthermore, the plasma nm23-H1 levels were investigated in 70 patients with other hematologic neoplasms, including 6 with acute lymphoblastic leukemia, 13 with chronic myelogenous leukemia, and 12 with myelodysplastic syndrome. Plasma nm23-H1 levels were significantly higher in all of these hematologic neoplasms than in normal controls. Increased plasma levels of nm23-H1 may have prognostic value in these hematologic malignancies as well as in AML.

The differentiation inhibitory factor nm23 can inhibit the differentiation of murine and human myeloid leukemia cells. We recently reported that nm23 genes were overexpressed in acute myelogenous leukemia (AML), and a higher level of nm23-H1expression was correlated with a poor prognosis in AML, especially in AML-M5 (acute monocytic leukemia). To evaluate the importance ofnm23 expression as a prognostic factor in AML, we compared it with other putative prognostic factors in AML. An analysis of the correlation between nm23 expression and the clinical parameters of 110 patients with AML demonstrated that increased nm23-H1mRNA levels were associated with resistance to initial chemotherapy and with reduced overall survival. Multivariate analysis using Cox's proportional hazard model also showed that elevated nm23-H1mRNA levels significantly contributed to the prognosis of patients with AML. Especially in AML-M5, nm23-H1 status was the most important prognostic factor. Furthermore, to determine whether we can apply the results observed in AML to other hematologic malignancies, we investigated the relative levels of nm23-H1 and nm23-H2transcripts in 149 patients with hematologic neoplasms, including 110 with de novo AML, 9 with de novo acute lymphoblastic leukemia, 14 with myelodysplastic syndrome, 16 with chronic myelogenous leukemia (CML), and 5 normal subjects by the reverse transcriptase-polymerase chain reaction. Expression of nm23-H1 was significantly higher in all the hematologic neoplasms, except CML in chronic phase, than in normal blood cells. nm23 may have a prognostic effect in these hematologic malignancies as well as in AML.

硝酸イナンは電気化学的に還元されにくい陰イオンであるが,多価金属イオンの共存下では,ポーラログラフ的に明瞭な還元波を示すようになる。本研究では,ランタニドイオン共存下での硝酸イオンの触媒的還元波について,その還元機構を明らかにするために種々の検討を加えた。まず,硝酸イオン濃度やランタニドイオン濃度,pH,共存陰イオンの種類,そして共存陽イオンの種類を変化させて,直流ポーラログラフ測定を中心に,各種の電気化学的測定を行った。その結果,この触媒的還元は,還元電位付近での電極表面へのランタニド化学種の吸着と,その後の吸着層を通しての硝酸イオンへの電子移動の二つのステップからなることが明らかになった。また,pHの増大につれて還元電位が正側にシフトする現象は,ランタニドイオンの加水分解によって生じる多核ヒドロキソ錯体の生成と,それにともなって起こる吸着層の負電荷の増大によるものと考えられた。定電位電解生成物の分析結果は,硝酸イオンの還元がNO3-+6e-+8H+→NH30H++2H2Oのように進行することを示唆した。

Inductively coupled plasma mass spectrometry using a laser ablation technique has been developed for the analysis of small inclusions of fused silica. A Q-switched pulsed Nd: YAG laser is fired at interesting sites of sample pieces with sizes of 10-40mm. Ablated particles were introduced directly into the ICP. Preablation with the laser pulses was effective in removing the surface contamination; thus, 1-min preablation could be conducted at interesting sites before data acquisition. The intensity ratios of interesting elements to matrix element with corrections using the relative sensitivity factors obtained by a solution introduction method provide some semi-quantitative data concerning the samples. The established procedure has been applied to the analysis of black inclusions of fused silica. The results have indicated that the major constituent of the inclusions was Fe (oxides).

レーザーアブレーション(LA)/ICP-MS法の深さ方向の分析特性を調べた.検討には,膜厚1.5μmのSnめっき鋼板を用い,レーザー光のデフォーカスの影響や出力,発振周波数の影響について調べた.Snめっき鋼板については,焦点位置から10mm程度レンズに近づけてデフォーカスさせたときが最適で,200mJ以下の低出力,1Hz以下の低発振周波数でμmレベルの深さ分解能が得られた.又,サンプリング速度は対象となる物質により異なり,検討した7種の金属のうちCu以外の金属については,サンプリング速度と沸点が逆比例の関係にあった.LA/ICP-MSの応用として,TiN膜及び塗装鋼板の深さ方向分析を試みた.

A flow injection (FI) technique has been applied to the trace analysis of metal samples using inductively coupled plasma mass spectrometry (ICP-MS). Parameters such as carrier flow rate, injection volume and sample concentration has a great influence on the sensitivity, precision and analysis time of the FI/ICP-MS. A small volume (240μl) of nickel-base alloy solutions containing up to 0.75% (m/v) was introduced into an ICP-MS instrument by the flow of carrier solution. The optimized procedure was applied to the determination of Tl, Pb and Bi in standard nickel-base alloys of the National Institute of Standards and Technology. The result was compared with that of the continuous sample nebulization method. The FI method gave improved precision as well as high sensitivity. The precision ranged from 2.0 to 8.2 (RSD, %) for the elements containing 0.25-11.7ppm. Detection limits were 0.001-0.04ppm.

Cytogenetic and molecular-genetic characteristics in peripheral T cell lymphoma (PTL) have not been well defined, except for those in adult T cell leukemia/lymphoma (ATL/L). Translocations and inversions involving a chromosome band 14q32 were extremely common abnormalities reported in PTL and ATL/L. We studied the involvement of TCL1, a recently isolated gene located in 14q32.1, in tumor tissues from 20 patients with PTL including three with 14q32 translocations by two color fluorescent in situ hybridization (FISH) using two cosmid probes flanking the TCL1 gene. The two cosmid signals were separated in none of them, but much increased in number in one tumor without 14q32 translocation, indicating that the TCL1 genomic region was amplified in this tumor. Reverse transcription-polymerase chain reaction (RT-PCR), however, failed to detect the TCL1 transcript in the tumor. These findings suggest that an oncogene other than TCL1 may be located in 14q32.1, and its amplification may be involved in the neoplastic process of PTL.

Differentiation inhibitory factor (nm23 protein) inhibited the induction of differentiation of mouse myeloid leukemia M1 and WEHI-3BD+ and human erythroleukemia HEL, KU812, and K562 cells. Block of differentiation may be associated with the aggressive behavior of leukemia. To examine the role of nm23 in human myeloid leukemia, we investigated the relative levels of nm23-H1, nm23-H2, and c-myc transcripts in 42 patients with acute myelogenous leukemia (AML), and in 5 with chronic myelogenous leukemia at chronic phase by reverse transcriptase polymerase chain reaction. The expression of nm23-H1 and -H2 but not of c-myc in AML was significantly higher than that in normal blood cells. Among AMLs, acute monocytic leukemia (presentation with AML-M5 morphology) was especially associated with elevated nm23-H1 and -H2 mRNA levels. On the other hand, the elevated levels of c-myc expression in AML-M5 were less evident. An analysis of correlation between nm23 expression and clinicopathological parameters showed that resistance to initial chemotherapy is associated with increased nm23-H1 mRNA levels and that a high initial white blood cell count is associated with increased nm23-H2 mRNA levels. Elevated nm23-H1 mRNA levels were associated with significantly reduced the overall survival of AML, especially of AML-M5 patients. The present results indicate that nm23-H1 and -H2 are overexpressed in AML and especially nm23-H1 gene expression predicts the prognosis of AML, especially of AML-M5.

Un systeme d'ablation par laser YAG a ete developpe pour l'analyse directe des aciers par spectrometrie ICP-AES (le laser permet d'obtenir des particules finies de taille

The interaction of granulocyte-colony stimulating factor (G-CSF) and retinoic acid (RA) in proliferation and differentiation of acute promyelocytic leukemia (APL) cells was examined. G-CSF stimulated proliferation of APL cells at concentrations of 0.1 to 50 ng/ml in a dose dependent manner. More than 10(-8) M RA induced granulocytic differentiation of APL cells. Although G-CSF induced lysozyme activities in APL cells, it alone did not induce terminal differentiation of APL cells. G-CSF significantly enhanced the RA-induced granulocytic differentiation of APL cells in vitro. Enhancement by G-CSF was not due to the prolongation of survival of RA-induced differentiated cells, but the differentiation-inducing effects of G-CSF might be evident only in the presence of RA. Since G-CSF has a potential to induce the granulocytic differentiation of myeloid leukemia cells, G-CSF in combination with RA may be applicable in differentiation induction therapy for some types of myeloid leukemia.

The results of a preliminary study on direct analysis of solid metals and nonconductive samples by inductively coupled plasma mass spectrometry (ICP-MS) are described. The aerosols ablated from solid samples by a ruby laser are introduced into an inductively coupled plasma, with detection of the resulting ions by a mass spectrometer. With conventional solution introduction, polyatomic species from mineral acids and solvents prevent the accurate determination of some trace analytes. However, using laser ablation, these background peaks are greatly reduced. The relative sensitivity factors for the laser ablation method are seen to vary from unity, with volatile elements such as Pb and Bi being more sensitive. Detection limits of rare-earth elements, Th and U in silicate rocks are excellent (0.02-0.9ppm at the effective integrating time of 1.4s), being better than those of the examined elements in steel (1.7-29ppm at the effective integrating time of 0.6s).

鉄鋼試料中の副・微量成分の迅速定量法として,ルビーレーザーによる固体試料の直接サンプリング/ICP-AES法を検討した.レーザー照射により生成したエアロゾル試料の搬送条件及びICP測定条件について最適化した後,鉄鋼試料の分析に本法を適用した.Si,Mn,P,Ni,Cr,Cu,Mo,V,Ti,Co,Nb,B,As,Snについて,正確さの良い検量線が得られた.鉄鋼試料の日常分析法であるスパーク発光分光法と比較し,検量線の直線性が良く,ダイナミックレンジが広い利点がある.しかし,分析精度はスパーク発光分光法より劣る元素が多い.

粉体試料の迅速分析法として,ルビーレーザーによる固体試料の直接サンプリング/ICP-AES法を確立した.粉体試料は加圧成形によりペレットとするか(ブリケット法),あるいは,四ホウ酸ナトリウムと融解してガラスビードとして(ガラスビード法),レーザー照射に供した.鉄鋼精錬スラグの分析に本法を適用したところ,ブリケット法では,今回検討したSi, Al, Mg, Fe, Mn, Ti, Pについて良好な精度と正確さが得られた.又,ガラスビード法はブリケット法に比し,迅速性・適用性は劣るものの,主成分の定量において精度・正確さを向上することができる.