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Protein induced by vitamin K absence or antagonist II (PIVKA-II) is a possible hepatocellular carcinoma (HCC) specific marker, however it may also be generated by a limited number of stomach malignancies. PIVKA-II-producing stomach cancer has been recorded in 16 instances so far, two of which were reported by us and all of which were found in Japan. In virtually all instances, serum alpha-fetoprotein (AFP) levels are also elevated. In roughly 80% of instances, liver metastasis is found, and portal vein tumor thrombus is detected in roughly 20% of cases. It's often difficult to tell the difference between a metastatic liver tumor and HCC. Almost all cases appear to be advanced stomach cancer macroscopically. In many cases, a hepatoid pattern is present, as well as a moderately to poorly differentiated adenocarcinoma component. Immunohistochemical staining is frequently used to confirm the generation of PIVKA-II and AFP. The presence of liver metastasis influences treatment and prognosis, and individuals with liver metastasis have a poor prognosis. PIVKA-II may be generated by tumor cells during hepatocellular metaplasia.

Aging is considered to be a functional retardation of continuous xenobiotic responses over a lifetime after the developmental period; thus, the effects of ionizing radiation over a lifetime may be somewhat accounted for by a modifier of aging effects. This study was conducted to evaluate the possible/synergic effects of radiation during aging by determining cell-cycle parameters of hematopoietic stem cells/hematopoietic progenitor cells (HSCs/HPCs), such as the percent of cells in cycling, the generation doubling time, and the cumulative cycling-cell fraction, by bromodeoxyuridine-ultraviolet assay, which enables the determination of their cycling capacity in vivo. Colony-forming progenitor cells, such as colony-forming unit (CFU)-granulocyte/macrophage (GM), CFU in the spleen on day 9 (CFU-S9), and CFU-S on day 13 (CFU-S13) for mature, less mature, and immature HPCs, respectively, were evaluated in young and old mice (6 weeks and 21 months of age, respectively) with or without 2-Gy whole-body irradiation and a 4-week recovery period. Then, cell-cycle parameters were evaluated and compared. As a result, the generation doubling time of all types of HPC was prolonged by the irradiation in both young and old mouse groups, except that of CFU-S13 in old mice, which showed acceleration of the cell cycle following the irradiation. In addition, only CFU-S13 in irradiated old mice showed a significant increase in the cumulative cycling-cell-fraction ratio. Significant changes due to the effects of aging and irradiation on HPCs were observed only in the immature HPCs, i.e., the cell cycle of immature HPCs was suppressed by aging without irradiation and was, in contrast, accelerated as the cells recovered from radiation-induced damage. This suggests that the mechanisms of peripheral blood recovery after 2-Gy whole-body irradiation are markedly different between young and old mice, although 21-month-old mice showed almost the same level of recovery as the young mice.

The number of murine mature blood cells recovered within 6 weeks after 2-Gy whole-body irradiation at 6 weeks of age, whereas in the case of the undifferentiated hematopoietic stem/progenitor cell (HSC/HPC) compartment [cells in the lineage-negative, c-kit-positive and stem-cell-antigen-1-positive (LKS) fraction], the numerical differences between mice with and without irradiation remained more than a year, but conclusively the cells showed numerical recovery. When mice were exposed to radiation at 6 months of age, acute damages of mature blood cells were rather milder probably because of their maturation with age; but again, cells in the LKS fraction were specifically damaged, and their numerical recovery was significantly delayed probably as a result of LKS-specific cellular damages. Interestingly, in contrast to the recovery of the number of cells in the LKS fraction, their quality was not recovered, which was quantitatively assessed on the basis of oxidative-stress-related fluorescence intensity. To investigate why the recovery in the number of cells in the LKS fraction was delayed, expression levels of genes related to cellular proliferation and apoptosis of cells in the bone marrow and LKS fraction were analyzed by real-time polymerase chain reaction (RT-PCR). In the case of 21-month-old mice after radiation exposure, Ccnd1, PiK3r1 and Fyn were overexpressed solely in cells in the LKS fraction. Because Ccnd1and PiK3r1 upregulated by aging were further upregulated by radiation, single-dose radiation seemed to induce the acceleration of aging, which is related to the essential biological responses during aging based on a lifetime-dependent relationship between a living creature and xenobiotic materials. Copyright © 2014 John Wiley & Sons, Ltd.

Hypoxia induces innumerable changes in humans and other animals, including an increase in peripheral red blood cells (polycythemia) caused by the activation of erythropoiesis mediated by increased erythropoietin (EPO) production. However, the elevation of EPO is limited and levels return to normal ranges under normoxia within 5–7 days of exposure to hypoxia, whereas polycythemia continues for as long as hypoxia persists. We investigated erythropoiesis in bone marrow and spleens from mouse models of long-term normobaric hypoxia (10 % O2) to clarify the mechanism of prolonged polycythemia in chronic hypoxia. The numbers of erythroid colony-forming units (CFU-E) in the spleen remarkably increased along with elevated serum EPO levels indicating the activation of erythropoiesis during the first 7 days of hypoxia. After 14 days of hypoxia, the numbers of CFU-E returned to normoxic levels, whereas polycythemia persisted for >140 days. Flow cytometry revealed a prolonged increase in the numbers of TER119-positive cells (erythroid cells derived from pro-erythroblasts through mature erythrocyte stages), especially the TER119 (high) CD71 (high) population, in bone marrow. The numbers of annexin-V-positive cells among the TER119-positive cells particularly declined under chronic hypoxia, suggesting that the numbers of apoptotic cells decrease during erythroid cell maturation. Furthermore, RT-PCR analysis showed that the RNA expression of BMP-4 and stem cell factor that reduces apoptotic changes during erythroid cell proliferation and maturation was increased in bone marrow under hypoxia. These findings indicated that decreased apoptosis of erythroid cells during erythropoiesis contributes to polycythemia in mice during chronic exposure to long-term hypoxia.

Background and aim The cytokeratin (CK)7(-) /CK20(+) immunoprofile is characteristic of colorectal carcinoma (CRC), although CK7(+) or CK20(-) phenotypes are occasionally encountered, particularly in histologically variant CRCs. We analyzed CK7/CK20 profiles in variant CRCs in association with clinicopathologic parameters and prognosis. Methods CK expression in well- and moderately differentiated adenocarcinoma (WMDA) (n = 63), poorly differentiated adenocarcinoma (PDA) (n = 91), mucinous adenocarcinoma (MUA) (n = 81), signet-ring cell carcinoma (SRCC) (n = 15), undifferentiated carcinoma (UDC) (n = 12), and adenosquamous carcinoma (n = 2) was analyzed using immunohistochemistry. Cut-off scores were set at 1% for CK7 and 25% for CK20 using the receiver operating characteristic curve analysis of PDA. Association between CK20(-) and better prognosis in PDA was validated in the second cohort (n = 66). Results CK7/CK20 immunoprofiling revealed a predominant CK7(-) /CK20(+) profile in WMDA, MUA, and SRCC, while the majority of UDC was characterized by a CK7(-) /CK20(-) profile. The CK7/CK20 profile in PDA was variable. Contingency table analysis revealed that CK expression was not significantly associated with any clinicopathologic parameters in WMDA, PDA, and MUA. However, survival analysis demonstrated that CK20(-) was significantly associated with better prognosis in PDA. Although CK20(-) was significantly associated with mismatch repair deficiency in PDA, it was an independent prognostic factor in multivariate analysis. Finally, we confirmed that CK20 status, determined using a 25% cut-off score, was a significant prognostic parameter in the second PDA cohort. Conclusions CK20 status may be used as a prognostic predictor of PDA.

An age-related decline in immune functions is referred to as immunosenescence. Mast cells play an important role in the immune system. However, it has not yet been determined if aging may affect mast-cell development. In the present study, we examined the age-related change in mast-cell development after myeloablation with 5-fluorouracil (5-FU) in senescence accelerated mice (SAMP1), which exhibit senescence-mimicking stromal cell impairment after 30 weeks of age. We found that aged mice with stromal cell impairment (30-36 weeks old) showed a lower recovery of the number of femoral mast-cell progenitors (colony-forming unit [CFU]-mast) (64% of steady state), whereas young mice (8-12 weeks old) showed a higher recovery (122% of steady state). Stromal cells influence mast-cell development by producing positive regulators such as stem cell factor (SCF) and negative regulators such as transforming growth factor-beta (TGF-β). The ratio of the gene expression of SCF to that of TGF-β (SCF/TGF-β βratio) indicates the balance of positive and negative regulation of mast-cell development. SCF/TGF-β βratio increased in both the young and aged mice after 5-FU treatment. However, the SCF/TGF-β βratio rapidly decreased in aged mice, whereas it remained high in young mice. The number of femoral CFU-mast in the S-phase after 5-FU treatment reflects the activation of positive-dominant regulation for mast-cell development by stromal cells. Aged mice showed lower recovery of the number of femoral CFU-mast in the S-phase (47% of steady state), whereas young mice showed a higher recovery (205% of steady state). These results suggest that mast-cell development declines with aging due to stromal-cell functional impairment, which contributes to immunosenescence.

Hideki Hirakata, Kosaku Nitta, Masaaki Inaba, Tetsuo Shoji, Hideki Fujii, Shuzo Kobayashi, Kaoru Tabei, Nobuhiko Joki, Hiroki Hase, Masato Nishimura, Shigeyuki Ozaki, Yuji Ikari, Yoshitaka Kumada, Kazuhiko Tsuruya, Shouichi Fujimoto, Tohru Inoue, Hiroyoshi Yokoi, Sumio Hirata, Kazuaki Shimamoto, Kiyotaka Kugiyama, Takashi Akiba, Kunitoshi Iseki, Yoshiharu Tsubakihara, Tadashi Tomo, and Tadao Akizawa

Surface chemical properties of a phospholipid, dilauroylphosphatidylcholine (DLPC), in two ionic liquids (ILs), 1-butyl-3-methylimidazolium tetrafluoroborate (bmimBF(4)) and hexafluorophosphate (bmimPF(6)), were investigated by means of surface tension, dynamic light-scattering, and freeze-fracture transmission electron microscopy. It was found that DLPC shows finite solubility in the ILs and spontaneously forms vesicles with size distribution around 400 nm in diameter above the critical vesicular concentration (CVC) of 0.040 wt.% (in bmimBF(4)) and approx. 0.08 wt.% (in bmimPF(6)). Other than the CVC value, anion specificity of the ILs was also seen in the temperature effect on the vesicular aggregation; that is, a temperature-induced reversible aggregation was observed in bmimPF(6), but not in bmimBF(4). The differences in the vesicular stability against the temperature-induced aggregation could be attributed to differences in the interaction between anion species of the ILs and zwitterionic phosphatidylcholine head groups. The apparent molecular area occupied by DLPC at the air/solution interface was estimated to be 0.37 nm(2) in bmimBF(4) and 0.20 nm(2) in bmimPF(6) by applying the Gibbs adsorption equation. These values are much smaller than the molecular area of 0.69 nm(2) reported for the hydrated DLPC bilayer of lamellar liquid-crystalline phase. This result is not consistent with the traditional Gibbs adsorption model, but can be interpreted in terms of a picture for the surface adsorption of soluble amphiphiles proposed by Moroi et al. Differential scanning calorimetric study is also reported regarding the phase transition behavior of DLPC bilayer solvated by the ILs.

Anemia induced by inflammation is well known to be more serious in the elderly than in non-elderly adults; however, the reason why this is so remains unclear. Neopterin produced by monocytes during inflammation promotes myelopoiesis but suppresses B-lymphopoiesis and erythropoiesis, by activating stromal cells in mice. Here, age-related changes in the erythropoietic response to neopterin were determined using senescence accelerated mice (SAMP1) with senescence stromal-cell impairment. Intravenous injection of neopterin into young mice (8–12 weeks old) resulted in a decrease in erythroid progenitor cell number in the bone marrow (BM), concomitant with an increase in myeloid progenitor cell number over one week. Intravenous injection of neopterin into aged mice (30–36 weeks old) resulted in a prolonged decrease in erythroid progenitor cell number in the BM over three weeks and a limited increase in myeloid progenitor cell number over one day. Neopterin treatment induced a decrease in serum erythropoietin concentrations in young mice but not in aged mice. The gene expression of tumor necrosis factor α (TNF- α), a negative regulator of erythropoiesis, was up-regulated in the BM of both young and aged mice, and the degree of TNF- α up-regulation was the same in both groups. The gene expression of interleukin (IL)-11, a positive regulator of erythropoiesis, was also up-regulated over one day in both young and aged mice. However, IL-11 gene expression remained up-regulated thereafter in young mice, whereas it was rapidly down-regulated in aged mice. These data suggest that prolonged suppression of erythropoiesis in aged mice may be due to a decrease in the production of positive regulators rather than to an increase in the production of negative regulators. Our combined data suggest that age-related impairment of stromal cells induces serious anemia in the elderly during inflammation.

Surface adsorption and aggregation behavior of polyoxyethylene (POE)-type nonionic surfactants in a room temperature ionic liquid, 1-butyl-3-methylimidazolium hexafluorophosphate (bmimPF6), were investigated by means of surface tension, 1H NMR, and dynamic light-scattering (DLS) measurements. The maximum surface excess in bmimPF6 is much lesser than those observed for aqueous solution, reflecting a much weaker solvophobicity exhibited by the surfactant hydrocarbon chain in the ionic liquid compared with the hydrophobicity experienced in water. Compared with another ionic liquid with different anion species, 1-butyl-3-methylimidazolium terafluoroborate (bmimBF4), the surfactant solution in bmimPF6 brings about higher critical micelle concentration (cmc), smaller micellar aggregation number, and weaker dependence of cmc on the surfactant hydrocarbon chain length. This indicates that the solvophobic interaction between surfactant hydrocarbon chains acts more weakly in bmimPF6 than in bmimBF4. The weaker solvophobic effect in bmimPF6 is also appreciable in the thermodynamic parameters for micelle formation derived from temperature dependence of cmc, and would be attributed to weaker attractive interaction between bmim cation and PF 6 - ion compared with that between bmim cation and BF 4 - ion. Present results give an insight into a better understanding of the importance of anion species for the properties of ionic liquids as a solvent to support a self-assembly of amphiphilic molecules.

Self-aggregation of polyoxyethylene (POE)-type nonionic surfactants in ionic liquids, 1-butyl- and 1-ethyl-3-methylimidazolium trifluoromethanesulfonate (bmimCF3SO3 and emimCF3SO3), was investigated by means of 1H-NMR chemical shift, dynamic light-scattering (DLS), and surface tension measurements. The surfactants showed no definite aggregate formation in bmimCF3SO3. This shows a remarkable contrast to the previous observation in bmimBF4 and bmimPF6, and demonstrates an importance of anion species to determine the property of ionic liquids as a solvent to support the self-assembly of amphiphilic compounds. On the other hand, the surfactants formed micelles in emimCF3SO3, which shows an importance of alkyl chain attached to imidazolium ring to determine the solvophobic interaction between surfactant hydrocarbon chains in imidazolium-based ionic liquids. The low solvophobicity of the surfactants to the ionic liquid composed of imidazolium cation with long alkyl chain is attributed to an affinity of the surfactant hydrocarbon chain to the imidazolium alkyl chain. The values of micellization parameters and surface adsorption parameters obtained for the surfactant solutions in emimCF3SO3 are reported.

症例は68歳，男性．不消化性の下痢，曖気の精査加療目的で入院となった．腹部CTで下行結腸～脾彎曲にかけて13cm大の腫瘤性病変を認めた．下部消化管内視鏡検査では下行結腸に隆起を主体とする5型病変を認め，ファイバーは潰瘍底より容易に胃内へと挿入された．腫瘍からの生検で高分化型腺癌と診断された．胃結腸瘻を形成した進行下行結腸癌と診断し，左半結腸切除術，D3リンパ節郭清，胃全摘，膵尾部・脾臓合併切除術を施行した．最終診断はmuc，si，n0，ly0，v0，ow(－)，aw(－)，stageIIで，手術根治度Aであった．われわれは内視鏡が通過することで診断できた胃結腸瘻を形成した下行結腸癌の1例を経験したので，自験例を加えた本邦報告25例の集計および文献的考察を行った．

Phase behaviors of the binary mixtures composed of ethylene carbonate (EC) and aliphatic alcohols, ω-phenyl alcohols, and alkylbenzenes were investigated. In addition, heat of solution of EC into these organic solvents was measured. The EC/methanol and EC/ethanol systems gave homogeneous solution at the temperature above their liquidus lines, while the mixtures of EC and alcohols with longer alkyl chain showed a miscibility gap in a liquid phase and provided the monotectic-type phase diagram. The liquid–liquid phase separation region expanded with the increase in the alkyl chain length. A similar phase behavior was also observed for the mixtures of EC and alkylbenzenes. On the other hand, the EC mixtures with ω-phenyl alcohols showed no miscibility gap in a liquid phase at least up to 4-phenylbutan-1-ol which has C4 alkyl chain intervening between phenyl and hydroxyl groups. This result demonstrates that both of the hydroxyl and phenyl groups act to facilitate the mixing of aliphatic compounds with EC. The phase behavior of these EC mixtures was analyzed applying the modified regular solution model in which the pair interaction energy was regarded as free energy. The model calculation with the use of heat of solution of EC at infinite dilution as the pair interaction enthalpy reproduced well both of the experimentally obtained liquidus line and mutual solubility curve as well as monotectic point.

症例は62歳，男性．37年前に十二指腸潰瘍に対して胃切除術の既往がある．1カ月前から嘔気・嘔吐を認めるようになり近医受診した．腹部CTで吻合部に腫瘤影と輸入脚の著明な拡張を認めた．同日の上部消化管内視鏡検査では吻合部に全周性の3型腫瘍を認め，残胃癌による輸入脚閉塞症と診断し緊急手術を施行した．前回手術はBillroth-II法の結腸後経路で再建されており，Treitz靱帯から吻合部の空腸は暗赤色を呈し著明に拡張していた．残胃全摘術，膵尾部・脾臓合併切除術を施行した．治癒切除であったが，術後3カ月で胃癌死された．残胃癌による輸入脚閉塞症の本邦論文報告7例について，集計および文献的考察を行った．

The impact of endocrine disruptors, and specifically the low-dose issue, involves interdisciplinary sciences. Thus, in the past these topics have been published widely in the toxicology area. Owing to recent developments in biology, including the whole-genome reading program, the mechanisms underlying the low-dose issue have been clarified. These mechanisms have been found to involve stochastic and probabilistic receptor-mediated adverse effects induced by endocrine disruptors. The effects thought to be induced by low doses of endocrine-disrupting chemicals remain disputed, and the underlying mechanisms remain poorly understood. Three independent factors, each only recently identified and never before encountered in the history of toxicological studies, are associated with what is termed the ‘low-dose issue’. First, toxicological risk has been estimated only by extrapolation of adverse phenotypes from high-dose effects and thus provides no reliable information on low-dose effects observed at the right time under experimental paradigm with sufficient sensitivity. Second, toxicity is based on disturbances of homeostatic regulation, a largely unexplored area in toxicology. Third, toxicity is based on stochastic and probabilistic xenobiotic response, a new field of toxicology that is specifically linked to low-dose and less-frequent events. To resolve the low-dose issue whether it causes effects or whether effects observed at low-doses should be considered ‘adverse’ – or both, each of these factors needs to be addressed. Copyright © 2010 John Wiley & Sons, Ltd.

An aqueous phase behavior was investigated for ionic liquid-related cationic gemini surfactant, [C12im-4-C12im][Br2], in which two dodecylimidazolium parts are linked by a spacer composed of four methylene units. Differential scanning calorimetry was used to detect phase boundaries among various phases, from which a temperature−composition (T−X) phase diagram was constructed. Lyotropic liquid-crystalline phases appearing in this mixture were characterized by means of polarized optical microscopy. It was revealed that (1) two types of liquid-crystalline phases, lamellar ( $$ {{\hbox{L}}_\alpha } $$ ) and hexagonal (H1), are formed; (2) H1 phase coexists with micellar solution (L1) over a wide concentration range from a few wt.% to ca. 80 wt.% of the surfactant; and (3) the hydrated solid with the composition of [C12im-4-C12im][Br2]·2H2O exhibits incongruent melting point at which the hydrated solid decomposes into H1 phase and unhydrated surfactant solid.

Neopterin is produced by monocytes and is a useful biomarker for inflammation. We found previously that neopterin enhanced myelopoiesis but suppressed B-lymphopoiesis triggered by the positive and negative regulations of cytokines produced by stromal cells in mice. The effects of neopterin on erythropoiesis during the enhancement of myelopoiesis were determined in the present study using C57BL/6J mice. The intravenous injection of neopterin into mice resulted in a prolonged decrease in the number of femoral erythroid progenitor cells (BFU-Es and CFU-Es), whereas the number of femoral myeloid progenitor cells (CFU-GMs) was increased. Interestingly, the oscillatory changes in the number of erythroid progenitor cells were reciprocal to those of myeloid progenitor cells. The expression of Cdc42, a regulator of the balance between erythropoiesis and myelopoiesis, was down-regulated, implying that the suppression of erythropoiesis is due to myelopoietic predominance. Furthermore, the expression of SDF-1 in stromal cells, a negative regulator of erythropoiesis, was up-regulated. These results suggest that neopterin facilitates myelopoiesis in the bone marrow by suppressing erythropoiesis, thereby contributing to the potential up-regulation of inflammatory process.

In this study, Trp53-deficient and wild-type mice of both C57BL/6 and C3H/He strains were exposed to benzene (33, 100, and 300 ppm; 6h/day, 5 days/week for 26 weeks) and then observed for lifetime. As results, first, the incidence of nonthymic lymphomas in C57BL/6 mice and acute myeloid leukemias (AMLs) in C3H/He mice showed linear responses at the lower exposure level in Trp53-deficient mice; second, the incidence of thymic lymphomas in C57BL/6 mice and nonthymic lymphomas in C3H/He mice increased without a plateau-like ceiling; thus, the former equivocal induction of hematopoietic neoplasms (HPNs) in the case of low-dose benzene exposure was assumed to be based on the DNA repair potential in wild-type mice, and the latter limited increase in HPNs in the case of high-dose benzene exposure was considered to be due to excessive apoptosis in wild-type mice. Concerning the incidence of AMLs, though a dose of 300 ppm benzene inhalation induced 9% AMLs in wild-type C3H/He mice-AML-prone, it induced AMLs in 38% of Trp53-deficient C3H/He mice. Because AMLs were also observed in Trp53-deficient mice, including in the C57BL/6 mice, benzene exposure may also be a potent inducer of AMLs in mice with some strain differences. In the present study, to elucidate the hematopoietic stem cell-specific, aryl hydrocarbon-receptor-related low-dose adverse effect, global gene expression in the bone marrow was analyzed at 28 days after 2-week-intermittent exposure to 150 mg/kg b.w. benzene, by gavage, i.e., equivalent to the above inhalation protocol with 300 ppm. We observed two conceptually different gene expression profiles; "common gene profiles" (CGPs) shared among mice in each group, and "stochastic gene profiles" (SGPs), i.e., unique union genes from one individual mouse to another. The CGPs of the experimental group and the SGPs of each individual mouse were separately characterized by individual assay. Concerning the CGPs, reciprocal strain differences between C3H/He and C57BL/6 mice in expression gene profiles, both plausible for leukemogenesis, were identified; namely, dominant downmodulations of Sltm and Cryl1, related to suppression of apoptosis and genomic instability in C3H/He mice, respectively, and dominant downmodulations of Atrx/rad54 and Kdm2a, related to a decrease in DNA repair and genomic instability, respectively, in C57BL/6 mice. These findings imply that these reciprocal gene expression differences induced by benzene exposure may lead each strain to undergo different hematopoietic neoplastic pathways. In contrast, each individual mouse often shows a unique SGP. SGPs often include transcription factors, which regulate reciprocal signaling pathways including further SGPs. Among them, apoptosis-related genes expressed in C57BL/6 mice and those in C3H/He mice were attributable to different combinations of SGPs. Such stochastic case-by-case gene expression may be in good agreement with the individual and strain differences observed following benzene exposure. Because gene chip microarray techniques can elucidate stochastic changes in gene expression profiles, possible stochastic toxicology and its future role are discussed.

The melting behavior of polyethyleneglycol dodecyl ethers (C 12 E 6 , C 12 E 7 , and C 12 E 8 ) in a room temperature ionic liquid, 1-butyl-3-methylimidazolium tetrafluoroborate (bmimBF 4 ), was investigated by means of differential scanning calorimetry (DSC). The melting temperature as a function of the surfactant concentration, combined with the cmc curve and cloud point curve, provided phase diagrams for the surfactant/bmimBF 4 mixtures in solvent-rich region. The characteristic feature of the mixtures is an existence of the Krafft temperature which is usually not observed with aqueous solutions of nonionic surfactants. The heat of fusion as a function of the surfactant concentration provided the interaction energy between the surfactant and bmimBF 4 . The interaction energy shows a linear dependence on the length of polyoxyethylene (POE) chain of the surfactants, which suggests that the solvation takes place around the POE chain.

This research focused on three major questions regarding benzene-induced hematopoietic neoplasms (HPNs). First, why are HPNs induced equivocally and at only threshold level with low-dose benzene exposure despite the significant genotoxicity of benzene even at low doses both in experiments and in epidemiology? Second, why is there no linear increase in incidence at high-dose exposure despite a lower acute toxicity (LD50 > 1000 mg/kg body weight; WHO, 2003, Benzene in drinking-water. Background document for development of WHO Guidelines for Drinking-Water Quality)? Third, why are particular acute myeloid leukemias (AMLs) not commonly observed in mice, although AMLs are frequently observed in human cases of occupational exposure to benzene? In this study, we hypothesized that the threshold-like equivocal induction of HPNs at low-dose benzene exposure is based on DNA repair potential in wild-type mice and that the limited increase in HPNs at a high-dose exposure is due to excessive apoptosis in wild-type mice. To determine whether Trp53 deficiency satisfies the above hypotheses by eliminating or reducing DNA repair and by allowing cells to escape apoptosis, we evaluated the incidence of benzene-induced HPNs in Trp53-deficient C57BL/6 mice with specific regard to AMLs. We also used C3H/He mice, AML prone, with Trp53 deficiency to explore whether a higher incidence of AMLs on benzene exposure might explain the above human-murine differences. As a result, heterozygous Trp53-deficient mice of both strains showed a nonthreshold response of the incidence of HPNs at the lower dose, whereas both strains showed an increasing HPN incidence up to 100% with increasing benzene exposure dose, including AMLs, that developed 38% of heterozygous Trp53-deficient C3H/He mice compared to only 9% of wild-type mice exposed to the high dose. The detection of AMLs in heterozygous Trp53-deficient mice, even in the C57BL/6 strain, implies that benzene may be a potent inducer of AMLs also in mice with some strain differences.

Studying the biological functions of the aryl hydrocarbon receptor (AhR) other than its function in xenobiotic drug metabolism may answer the questions as to why AhR orthologues have long been conserved phylogenically widely in the animal kingdom, and why homologues have diverged from nonvertebrate species such as nematodes and drosophila to all the vertebrate species. In this review, we focused on the mechanism of longevity possibly derived from evolution of AhRs and compared the functional difference of hematopoietic progenitors between wild-type (AhR(+/+)) mice and AhR-deficiencies (AhR(+/-), AhR(-/-)). Particular advantages found in wild-type mice compared with AhR-deficiencies were as follows: first, higher antioxidative function in the hematopoietic microenvironment with low oxidative tension seemed to have developed with the evolution of AhR; second, primitive hematopoietic progenitor-cell-specific deceleration and dormancy of cell-cycle regulation may have developed also with AhR evolution, which keeps hematopoietic progenitor cell compartment dormant without extinction by continuous differentiation; third, the consequent evolution of genomic stabilization with a longer lifespan in wild-type mice developed with the evolution of AhR. Experimentally, mice showed a significant extension of lifespan in a gene-dosage-dependent manner with a delayed onset of leukemogenicity. Another possible additional advantage observed in wild-type mice, the mechanism of which is not yet clarified, is an improved efficiency of fertilization in wild-type mice as compared with AhR-deficiencies, which seems to have developed with the evolution of AhR. Four advantages altogether, including the anti-aging feature mentioned above may have induced the AhR molecule to diverge various of species in the animal kingdom.

The cloud point phenomenon has been investigated for the solutions of polyoxyethylene (POE)-type nonionic surfactants (C(12)E(5), C(12)E(6), C(12)E(7), C(10)E(6), and C(14)E(6)) in 1-butyl-3-methylimidazolium tetrafluoroborate (bmimBF(4)), a typical room temperature ionic liquid (RTIL). The cloud point, T(c), increases with the elongation of the POE chain, while decreases with the increase in the hydrocarbon chain length. This demonstrates that the solvophilicity/solvophobicity of the surfactants in RTIL comes from POE chain/hydrocarbon chain. When compared with an aqueous system, the chain length dependence of T(c) is larger for the RTIL system regarding both POE and hydrocarbon chains; in particular, hydrocarbon chain length affects T(c) much more strongly in the RTIL system than in equivalent aqueous systems. In a similar fashion to the much-studied aqueous systems, the micellar growth is also observed in this RTIL solvent as the temperature approaches T(c). The cloud point curves have been analyzed using a Flory-Huggins-type model based on phase separation in polymer solutions.

Previously, we found an aryl hydrocarbon receptor (AhR)-transmitted benzene-induced hematotoxicity; that is, AhR-knockout (KO) mice did not show any hematotoxicity after benzene exposure [Yoon, B.I., Hirabayashi, Y., Kawasaki, Y., Kodama, Y., Kaneko, T., Kanno, J., Kim, D.Y., Fujii-Kuriyama, Y., Inoue, T., 2002. Aryl hydrocarbon receptor mediates benzene-induced hematotoxicity. Toxicol. Sci. 70, 150-156]. Furthermore, our preliminary study showed a significant attenuation of benzene-induced hematopoietic toxicity by AhR expression, when the bone marrow (BM) of mice was repopulated with AhR-KO BM cells [Hirabayashi, Y., Yoon, B.I., Li, G., Fujii-Kuriyama, Y., Kaneko, T., Kanno, J., Inoue, T., 2005a. Benzene-induced hematopoietic toxicity transmitted by AhR in the wild-type mouse was negated by repopulation of AhR deficient bone marrow cells. Organohalogen Comp. 67, 2280-2283]. In this study, benzene-induced hematotoxicity and its nullification by AhR-KO BM cells were further precisely reevaluated including the duration of the effect after benzene treatment and recovery after the cessation of exposure. Exposure routes, namely, intraperitoneal (i.p.) injection used in our previous study and intragastric (i.g.) administration used in this study, were also compared in terms of their toxicologic outcomes. From the results of this study, mice that had been lethally irradiated and repopulated with BM cells from AhR-KO mice essentially did not show any benzene-induced hematotoxicity. The AhR-KO BM cells nullified benzene-induced toxicities in notably different hematopoietic endpoints between the i.p. treatment and the i.g. treatment; however, the number of granulo-macrophage colony-forming unit in vitro (CFU-GM) was a common target parameter, the benzene-induced toxicity of which was nullified by the AhR-KO BM cells.

Irradiation of mice at doses of 1-1.5 Gy induced a predominant regeneration of the B-cell lineage but suppressed the regeneration of the myeloid lineage. The mechanisms underlying such reciprocal regulation of regeneration and the relationship between the two lineages remain unclear. Because the predominant regeneration of the B-cell lineage observed is considered to depend on the stromal cell function, and because the impairment of such stromal function may nullify such reciprocal responses, mouse models of senescent stromal cell impairment (SCI) and the less senescent stage of SCI (non-SCI) were compared to elucidate the mechanisms underlying the reciprocal regulation of both lineages after radiation exposure. In non-SCI mice irradiated with 1 Gy, the numbers of B-lymphocyte progenitor (CFU-preB) and granulocyte-macrophage progenitor (CFU-GM) cells in the bone marrow decreased rapidly during the first 24 h. Then the number of CFU-preB cells in the bone marrow promptly recovered from the nadir and exceeded the pretreatment level, whereas that of CFU-GM cells remained lower than the pretreatment level. The expression of genes encoding positive regulators of the B-lymphoid lineage [interleukin (IL)10, Flt3 ligand and IL7] was up-regulated; in contrast, expression of the positive regulators of the myeloid lineage [granulocyte macrophage colony-stimulating factor (GM-CSF) and stem cell factor (SCF)] was down-regulated. In SCI mice irradiated with 1 Gy, the oscillatory changes in the numbers of femoral CFU-preB and CFU-GM cells and in the expression levels of cytokine genes were less marked than those in the non-SCI mice. These results thus imply that the reciprocal regeneration depends on the up-regulation of IL10, Flt3 ligand and IL7 expression and the down-regulation of GM-CSF and SCF expression in the bone marrow, possibly depending on the hematopoietic microenvironment.

Aqueous solutions of long-chain imidazolium ionic liquids have been investigated by surface tension and steady-state fluorescence measurements at room temperature (298 K). The micelle aggregation number (Nagg) was obtained by pyrene fluorescence quenching method. From the surface tension data, critical micelle concentration (cmc), surface tension at the cmc (γcmc), adsorption efficiency (pC20), and effectiveness of surface tension reduction (Πcmc), were determined. Moreover, applying the Gibbs adsorption isotherm, maximum surface excess concentration (Γmax) and minimum surface area per molecule (Amin) at the air–water interface were estimated. The effect of sodium halides, NaCl, NaBr, and NaI, on the surface activity was also investigated. The addition of salts decreases significantly both cmc and γcmc, and the dependence of the salt effect on the anion species is analogous for the case of conventional ionic surfactants. Due to the bulkiness of the imidazolium head group, the microenvironment in long-chain imidazolium ionic liquid micelles exhibits higher polarity compared with the corresponding CnTAB micelles, and the micelle aggregation numbers (Nagg) are smaller than those of CnTAB.

The study examined the feasibility of screening for hepatotoxicity by an in vitro gene expression analysis using rat primary hepatocytes and Affymetrix Rat Toxicology U34 arrays. Hepatocytes were exposed for 6 or 24 h to eight drugs, with different mechanisms of hepatotoxicity, at one third of the cytotoxic concentration TC50, i.e. acetaminophen, cyclophosphamide, clofibrate, chlorpromazine, lithocholic acid, cisplatin, diclofenac and disulfiram. The types of transcriptional changes observed in this study were generally consistent with previously reported in vivo data, although there were some differences. In hierarchical cluster analysis, drugs formed clusters depending on their mode of toxicity against cells. The number of transcripts affected by the cholestatic hepatotoxicants (lithocholic acid and chlorpromazine) or the drugs that rarely cause of hepatotoxicity (cisplatin, diclofenac and disulfiram) were limited compared with the other drugs (acetaminophen, clobifibrate and cyclophosphamide), where they did not induce transcriptional changes apparently related to toxicity. It is concluded that in vitro gene expression analysis of hepatocytes using microarray is a useful tool for evaluating the toxicological profile of drugs and in screening for the direct toxicity of drugs against hepatocytes.

Purification of perfluoro-n-alkanoic acids (C(n)F(2n+1)COOH, n=7, 9, 11, 13, 15 and 17) was made by repeated recrystallizations from n-hexane/acetone mixed solvent, and their purity was found to be more than 99.5% by GC-MS, NMR, and elemental analysis. The thermal behaviors such as melting point and enthalpy change of fusion were investigated using differential scanning calorimetry (DSC). The melting point monotonously increased with increasing carbon number (n) of the acids, while the enthalpy change showed irregularity at n=14. The crystal structure of these acids was found to be dependent upon solvent used for recrystallization; that is, the acids recrystallized from the above solvent becomes more stable energetically, indicating their higher enthalpy change of fusion than that of the solidified acids from fused ones. The solid state was also found to vary depending upon the thermal history, indicating that a few crystal structures of the solid state are quite similar energetically. The melting points (T(m)) of perfluoro-n-alkanoic acids are higher than those of corresponding n-alkanoic acids, and the difference in T(m) increases with increasing carbon number in the acids.

Since malignant tumors are life-threatening, the death rate from these diseases is high, and existing therapies have limited effectiveness, it is desired to provide new effective anticancer drugs to tumor patients sooner. However, there is no guideline regarding non-clinical safety studies on the development of anticancer drugs required for the first in human clinical trials and for the approval applications in Japan. Then, the Ministry of Health, Labour and Welfare (MHLW) established the collaboration group including regulatory, academic and industrial scientists to prepare the guideline on the non-clinical safety evaluation of anticancer drugs in 2004. As a guide for basic concept of non-clinical safety studies on anticancer drugs, the "Points to Consider" document was prepared by this group in 2007.

Electrical conductivity was measured for aqueous solutions of long-chain imidazolium ionic liquids (IL), 1-alkyl-3-methylimidazolium bromides with C(12)-C(16) alkyl chains. The break points appeared in specific conductivity (kappa) vs concentration (c) plot indicates that the molecular aggregates, i.e., micelles, are formed in aqueous solutions of these IL species. The critical micelle concentration (cmc) determined from the kappa vs c plot is somewhat lower than those for typical cationic surfactants, alkyltrimethylammonium bromides with the same hydrocarbon chain length. The electrical conductivity data were analyzed according to the mixed electrolyte model of micellar solution, and the aggregation number, n, and the degree of counter ion binding, beta, were estimated. The n values of the present ILs are somewhat smaller than those reported for alkyltrimethylammonium bromides, which may be attributed to bulkiness of the cationic head group of the IL species. The thermodynamic parameters for micelle formation of the present ILs were estimated using the values of cmc and beta as a function of temperature. The contribution of entropy term to the micelle formation is superior to that of enthalpy term below about 30 degrees C, and it becomes opposite at higher temperature. This coincides with the picture drawn for the micelle formation of conventional ionic surfactants.

The surface behaviors of 3-nonylphenoxy-2-hydroxyl-propyl-trimethyl-ammonium bromide (RTAB), a cationic surfactant, are examined by dynamic surface tensions (DSTs), π–A isotherms and molecular dynamics simulations as a function of pH. While the DST remains almost constant when the pH of the solutions changes from 7.0 to 3.0, which decreases remarkably in alkaline solutions. The results of π–A isotherms indicate that RTAB molecule has a much larger cross-sectional area at the air–water interface in alkaline solutions than that in acidic or neutral solutions. Molecular dynamics (MD) simulation studies revealed that the hydrocarbon chains of the surfactant molecules adsorbed at the air–water interface become conformationally more complex under alkaline solutions than in neutral or acidic conditions. These abnormal surface behaviors of RTAB in alkaline solutions are explained by assuming the binding of a hydroxide ion to the hydroxyl group in RTAB molecule.

Membrane channel connexin (Cx) forms gap junctions that are implicated in the homeostatic regulation of multicellular systems; thus, hematopoietic cells were assumed not to express Cxs. However, hematopoietic progenitors organize a multicellular system during the primitive stage; thus, the aim of the present study was to determine whether Cx32, a member of the Cx family, may function during the primitive steady-state hematopoiesis in the bone marrow (BM). First, the numbers of mononuclear cells in the peripheral blood and various hematopoietic progenitor compartments in the BM decreased in Cx32-knockout (KO) mice. Second, on the contrary, the number of primitive hematopoietic progenitor cells, specifically the Lin(-)/c-kit(+)/Scal(+ )fraction, the KSL progenitor cell compartment, also increased in Cx32-KO mice. Third, expression of Cx32 was detected in Lin(-)/c-kit(+) hematopoietic progenitor cells of wild-type mice (0.27% in the BM), whereas it was not detected in unfractionated wild-type BM cells. Furthermore, cell-cycle analysis of the fractionated KSL compartment from Cx32-KO BM showed a higher ratio in the G(2)/M fraction. Taken together, all these results imply that Cx32 is expressed solely in the primitive stem cell compartment, which maintains the stemness of the cells, i.e., being quiescent and noncycling; and once Cx32 is knocked out, these progenitor cells are expected to enter the cell cycle, followed by proliferation and differentiation for maintaining the number of peripheral blood cells.

Mice exposed to a lethal dose of radiation were repopulated with heterozygous p53(+/-) (TRP53(+/-)) bone marrow cells and then exposed to doses of 1, 3 and 5 Gy 1 month later. This resulted in the transplanted bone marrow-specific diseases other than competitively induced nonhematopoietic neoplasms. Interestingly, the present study showed a high frequency of stem cell leukemia, i.e., leukemias characterized by a lack of differentiation due also to p53 deficiency, even after 5 Gy irradiation. The frequencies of stem cell leukemias (and those of total hematopoietic malignancies) were 16% (24%) at 1 Gy and 45% (75%) at 3 Gy. Furthermore, markedly high incidences of stem cell leukemias were observed at 5 Gy in p53(+/-) mice, i.e., 87% (100%) in the transplantation assay and 60% (83.3%) in the whole-body assay, whereas a conventional whole-body assay induced only 14% in wild-type mice. The high incidence of stem cell leukemias observed in this study using heterozygous p53-deficient mice agrees with results of a previous study of homozygous p53-deficient mice and is consistent with the high frequency of loss of heterozygosity in the p53 wild-type allele observed in leukemias. This suggests that the target cells for radiation-induced stem cell leukemias may be p53-deficient hematopoietic stem cells.

Thermolysis of 3,3-di-tert-butylthiirane-2-thione S-oxide 4 gave di-tert-butyl ketone 5 and carbon disulfide (CS2). Treatment of α-dithiolactone S-oxide 4 with excess m-CPBA or triphenylphosphine gave di-tert-butylthioketene S-oxide 3 almost quantitatively. Treatment of 4 with (η2-ethylene)bis(triphenylphosphine)platinum(0) 13 or tetrakis(triphenylphosphine) palladium 17 resulted in the formation of thiolato sulfinato–platinum complex 15 or palladium complex 18, respectively. The structure of complex 15 was determined by X-ray crystallographic analysis.

Aqueous solutions of three kinds of surface active ionic liquids composed of the 1-alkyl-3-methylimidazolium cation have been investigated by means of surface tension and electrical conductivity measurements at room temperature (298 K). The surface tension measurements provided a series of parameters, including critical micelle concentration (cmc), surface tension at the cmc (gammacmc), adsorption efficiency (pC20), and effectiveness of surface tension reduction (Picmc). In addition, with application of the Gibbs adsorption isotherm, maximum surface excess concentration (Gammamax) and minimum surface area/molecule (Amin) at the air-water interface were estimated. The effect of sodium halides, NaCl, NaBr, and NaI, on the surface activity was also investigated. It was found that both the pC20 and the Picmc were rather larger than those reported for traditional ionic surfactants and the cmc values were somewhat lower than those for typical cationic surfactants, alkyltrimethylammonium bromides, and comparable to typical anionic surfactants, sodium alkyl sulfates. These results demonstrate that the surface activity of long-chained imidazolium IL is somewhat superior to that of conventional ionic surfactants.

The National Science Museum (NSM) in Japan has recently completed a project using different types of visitor-oriented digital technologies. With sponsorship from the Japan Society for the Promotion of Science (JSPS), the NSM team carried out a four-year study to examine how digital technologies can be used to enhance as well as educationally enrich a visit to a science and natural history museum. Two types of devices, a handheld audio guide and an integrated circuit card, were tested in the museum. The results of the project are described along with suggestions for designers and educators who are seeking to add similar devices to their museums.

Phase behavior of aqueous mixture of imidazolium ionic liquid, 1-dodecyl-3-methylimidazolium bromide, was investigated by means of differential scanning calorimetry and polarized optical microscopy. The mixture forms two types of lyotropic liquid-crystalline gels, one is composed of lamellar phase and the other is of hexagonal phase. T – X phase diagram of the mixture was constructed, which defines the regions of various phases appearing in this mixture.

Objective Reactive oxygen species (ROS), generated following benzene exposure, are considered to trigger the development of hematopoietic neoplasms, although little supporting evidence has been found. In this study, we examined whether the experimental elimination of ROS generated following benzene exposure prevents the development of benzene-induced hematopoietic disorders to clarify the mechanism underlying the development of benzene-induced hematopoietic disorders. Methods C57BL/6 mice, overexpressing human thioredoxin (h-Trx-Tg), were used to examine the possible nullification of ROS induction following benzene exposure. The experimental group was exposed to 300 ppm benzene 6 hours/day, 5 days/week, for 26 weeks, and lifetime observation followed by molecular and histopathological examinations were carried out. Results The present study using h-Trx-Tg mice showed a complete suppression of the development of thymic lymphoma induced by benzene inhalation (0% in h-Trx-Tg vs 30% in wild-type (Wt) mice). This was associated with a 48% decrease in the incidence of clastogenic micronucleated reticulocyte induction in the h-Trx-Tg mice compared with the Wt control after 2 weeks of inhalation. As underlying mechanisms, the attenuation of oxidative stress was accompanied by a complete abrogation of hemato-lymphoid toxicity, as shown by the upregulation of the activity of superoxide-dismutase, and a consequently stable ROS level, as determined by cell sorting using 2′, 7′-dichlorodihydrofluorescein diacetate, along with a significant attenuation of the overexpression of a cell cycle–dependent kinase inhibitor, p21. Conclusion The attenuation of benzene-induced oxidative stress and that of the consequent lymphomagenesis were observed for the first time, and these indicate a role of oxidative stress in benzene-induced clastogenesis and lymphomagenesis. (These attenuations were not seen in nonthymic lymphomas, and no leukemias developed in C57BL/6 used in this study.) During the constitutive overexpression of h- Trx , the expression of aryl-hydrocarbon receptor in h-Trx-Tg mice was downregulated, which may also contribute to the attenuation.

Neopterin is produced by monocytes and is a useful biomarker of inflammatory responses. We found that neopterin enhances granulopoiesis, but suppresses B-lymphopoiesis triggered by the positive and negative regulations of cytokines produced by stromal cells in mice. In this study, neopterin was found to regulate mast cell development, which was confirmed in the mouse model of senescent stromal-cell impairment (SCI). In non-SCI mice (= less senescent stage of SCI mice), neopterin decreased the number of colonies of IL-3-dependent mast-cell progenitor cells (CFU-mast) from unfractionated bone-marrow cells, but not that from the lineage-negative bone-marrow cell population without stromal cells in a semisolid in vitro system. Neopterin increased the gene expression and protein production of TGF-β, a negative regulator of CFU-mast, in cultured stromal cells, indicating that neopterin suppressed CFU-mast colony formation by inducing TGF-β in stromal cells. In contrast to this in vitro study, in vivo treatment with neopterin did not significantly up-regulate TGF-β. The intravenous injection of neopterin into mice decreased the number of femoral CFU-mast and the expression level of the gene for stem cell factor (SCF), a positive regulator of CFU-mast, whereas the number of splenic CFU-mast and SCF gene expression level increased. In SCI mice, the in vivo and in vitro responses of mast cell development and cytokine gene expression level to neopterin treatment were less marked than those in non-SCI mice. These results suggest that, firstly, neopterin augments the splenic pool of CFU-mast by the production of SCF, and secondly, such neopterin function becomes impaired during senescence because of an impaired stromal-cell function, resulting in the down-modulation of host-defense mechanisms.

Bone marrow (BM) functions as the primary hematopoietic tissue throughout adult life by providing a microenvironment for the proliferation, differentiation, and retention of hematopoietic stem cells and progenitors. We describe novel roles for oncostatin M (OSM) in the BM hematopoietic microenvironment. Hematopoietic progenitor activity in OSM-deficient mice was reduced in BM but elevated in the spleen and peripheral blood. The level of circulating granulocyte colony-stimulating factor (G-CSF) was increased, whereas that of stromal cell-derived factor 1 (SDF-1) was decreased in OSM-deficient mice. Moreover, the ability of OSM-deficient BM stromal cells to support hematopoiesis in vitro was significantly reduced. These results indicate that OSM plays a unique role in hematopoiesis by maintaining the proper microenvironment for BM hematopoiesis; it also retains hematopoietic progenitors in BM by regulating G-CSF and SDF-1 levels.

Previous fate mapping analysis, using Cre recombinase driven by the Mesp1 locus, revealed that Mesp1 is expressed in almost all of the precursors of the cardiovascular system, including the endothelium, endocardium, myocardium, and epicardium. Mesp1-nonexpressing cells were found to be restricted to the outflow tract cushion and along the interventricular septum (IVS), which is a location that is suggestive of specialized cardiac conduction system (CCS). In our current study, we examined the identity of these IVS cells by using the pattern of beta-galactosidase activity in CCS-lacZ mice. In addition, by crossing Mesp1-Cre and floxed GFP reporter mice with CCS-lacZ mice, we have calculated that approximately 20% of the ventricular CCS within the IVS corresponds to Mesp1-nonexpressing cells. These data suggest that the ventricular CCS is of heterocellular origin. Furthermore, we indicate a possibility that a population of the cells that contribute to the ventricular CCS might be distinguished at an early stage of development.

Objectives: Lymph node metastasis is one of the determining factors of a poor prognosis for colorectal cancer. Recent studies have reported that cancer cells can promote lymphangiogenesis and that chemokine receptors expressed by cancer cells might play a role in metastasis. In this study, we examined the correlation between the expression of vascular endothelial growth factor (VEGF) C, the chemokine receptor CXCR4 and lymph node metastasis in colorectal cancer. Methods: One hundred and sixty-one consecutive patients who underwent resection at our department were studied. Lymph node metastasis was observed in 69 cases (43%) and lymphatic involvement was present in 105 cases (65%). Immunohistochemical staining was performed using antibodies for VEGF-C and CXCR4. Moreover, lymphatic vessel density (LVD) was evaluated within the tumor by immunostaining with a D2-40 antibody. Results: VEGF-C expression was found in 81 cases (50%) and CXCR4 expression in 87 cases (54%). Regarding the correlation between nodal metastasis and the expression of CXCR4 and VEGF-C, the incidence of nodal metastasis was significantly (p < 0.01) higher in patients with CXCR4-positive tumors than in those with CXCR4-negative tumors. In addition, a significant correlation was observed between CXCR4 and VEGF-C expression and lymphatic invasion (p < 0.01). LVD was significantly higher in VEGF-C-positive tumors compared with VEGF-C-negative tumors. However, there was no significant correlation between LVD and CXCR4 expression. Using multivariate analysis, VEGF-C, CXCR4, lymphatic invasion and wall invasion were found to be independent risk factors for lymph node metastasis. Conclusions: This study suggests that although the mechanism that promoted lymph node metastasis was different between VEGF-C and CXCR4, both VEGF-C and CXCR4 contributed to lymphatic involvement and nodal metastasis in colorectal cancer.