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The interleukin-17 (IL-17) family of cytokines has emerged as a critical player in inflammatory diseases. Among them, IL-25 has been shown to be important in allergic inflammation and protection against parasitic infection. Here we have demonstrated that IL-17B, a poorly understood cytokine, functions to inhibit IL-25-driven inflammation. IL-17B and IL-25, both binding to the interleukin-17 receptor B (IL-17RB), were upregulated in their expression after acute colonic inflammation. Individual inhibition of these cytokines revealed opposing functions in colon inflammation: IL-25 was pathogenic but IL-17B was protective. Similarly opposing phenotypes were observed in Citrobacter rodentium infection and allergic asthma. Moreover, IL-25 was found to promote IL-6 production from colon epithelial cells, which was inhibited by IL-17B. Therefore, our data demonstrate that IL-17B is an anti-inflammatory cytokine in the IL-17 family., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Contact hypersensitivity (CHS) has been widely used to study cutaneous immune responses, as a prototype of delayed-type hypersensitivity. Although natural killer T (NKT) cells have been assumed to have an important role in CHS, their role is controversial. Here, we report the role of NKT cells in the sensitization phase of CHS, by promoting the survival and maturation of dendritic cells (DCs) in the draining lymph nodes (LNs). The CHS response was attenuated with Cd1d1(-/-) and Traj18(-/-) BALB/c mice in which NKT cells were absent. In the draining LNs, the number of effector T cells and cytokine production were significantly reduced with NKT cell-deficient mice. NKT cells activated and colocalized with DCs in the draining LNs after sensitization. The number of migrated and mature DCs was reduced in NKT cell-deficient mice 72 hours after FITC application. In in vitro experiments, activated NKT cells enhanced bone marrow-derived DC (BMDC) survivability via tumor necrosis factor (TNF) production from BMDCs. In addition, TNF production from BMDCs was partially suppressed by the neutralizing anti-CD54 or CD154 antibodies. Our data demonstrate that DC-NKT interaction has a pivotal role in the sensitization phase of CHS.

NKT cells are characterized by their expression of an NKT-cell-specific invariant antigen-receptor α chain encoded by Vα14Jα18 gene segments. These NKT cells bridge the innate and acquired immune systems to mediate effective and augmented responses; however, the limited number of NKT cells in vivo hampers their analysis. Here, two lines of induced pluripotent stem cell-derived mice (NKT-iPSC-derived mice) were generated by reprogramming of mature NKT cells, where one harbors both rearranged Vα14Jα18 and Vβ7 genes and the other carries rearranged Vα14Jα18 on both alleles but germline Vβ loci. The analysis of NKT-iPSC-derived mice showed a significant increase in NKT cell numbers with relatively normal frequencies of functional subsets, but significantly enhanced in some cases, and acquired functional NKT cell maturation in peripheral lymphoid organs. NKT-iPSC-derived mice also showed normal development of other immune cells except for the absence of γδT cells and disturbed development of conventional CD4 αβT cells. These results suggest that the NKT-iPSC-derived mice are a better model for NKT cell development and function study rather than transgenic mouse models reported previously and also that the presence of a pre-rearranged Vα14Jα18 in the natural chromosomal context favors the developmental fate of NKT cells., (© The Author 2014. Published by Oxford University Press on behalf of The Japanese Society for Immunology.)

Immunological memory has been regarded as a unique feature of the adaptive immune response mediated in an antigen-specific manner by T and B lymphocytes. However, natural killer (NK) cells and γδT cells, which traditionally are classified as innate immune cells, have been shown in recent studies to have hallmark features of memory cells. Invariant NKT cell (iNKT cell)-mediated antitumor effects indicate that iNKT cells are activated in vivo by vaccination with iNKT cell ligand-loaded CD1d(+) cells, but not by vaccination with unbound NKT cell ligand. In such models, it previously was thought that the numbers of IFN-γ-producing cells in the spleen returned to the basal level around 1 wk after the vaccination. In the current study, we demonstrate the surprising presence of effector memory-like iNKT cells in the lung. We found long-term antitumor activity in the lungs of mice was enhanced after vaccination with iNKT cell ligand-loaded dendritic cells. Further analyses showed that the KLRG1(+) (Killer cell lectin-like receptor subfamily G, member 1-positive) iNKT cells coexpressing CD49d and granzyme A persisted for several months and displayed a potent secondary response to cognate antigen. Finally, analyses of CDR3β by RNA deep sequencing demonstrated that some particular KLRG1(+) iNKT-cell clones accumulated, suggesting the selection of certain T-cell receptor repertoires by an antigen. The current findings identifying effector memory-like KLRG1(+) iNKT cells in the lung could result in a paradigm shift regarding the basis of newly developed extrathymic iNKT cells and could contribute to the future development of antitumor immunotherapy by uniquely energizing iNKT cells.

We report a novel monofilament slip knot technique for bronchial ligation and for visualization of the anatomic plane during lung segmentectomy. After threading the bronchus, a slip knot is made outside the thorax. During lung ventilation, one end of the string is pulled, and the knot slips to reach the bronchus without a knot-pusher. Bronchial ligation is then performed to block the outflow of segmental air while the segment remains expanded, whereas the other segments become collapsed. This technique allows identification of the anatomic intersegmental plane, facilitating thoracoscopic anatomic lung segmentectomy., (Copyright © 2014 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

Pulmonary segmentectomy is generally acknowledged to be more technically complex than lobectomy. Three-dimensional computed tomography (3D CT) angiography is useful for understanding the pulmonary arterial and venous branching, as well as planning the surgery to secure adequate surgical margins. Comprehension of the intersegmental and intrasegmental veins makes the parenchymal dissection easier. To visualize the segmental border, creation of an inflation-deflation line by using a method of inflating the affected segment has become the standard in small-sized lung cancer surgery. Various modifications to create the segmental demarcation line have been devised to accurately perform the segmentectomy procedure.

Background: The commensal yeast Candida albicans is a major cause of invasive fungal infections. Despite treatment with antifungal agents, the mortality rate attributed to these types of infection is high. Although numerous cases have been reported regarding a poor outcome for patients with bacterial and C. albicans coinfection, the mechanisms by which the coinfecting bacteria exacerbate the C. albicans infection remain elusive., Methods and Results: We evaluated how glycolipid-mediated activation of invariant natural killer T (iNKT) cells affects the clearance of C. albicans. Surprisingly, C. albicans-infected, glycolipid-treated mice exhibited significantly lower survival rates, increased fungal burden, and higher interleukin (IL)-6 production in the kidneys compared with control mice. Glycolipid-induced exacerbation of C. albicans infection was not observed in interferon-gamma knockout (IFN-γKO) mice. In the C. albicans-infected, glycolipid-treated mice, the number of neutrophils in the blood and bone marrow dramatically decreased in an IFN-γ-dependent manner. Furthermore, mice that were coinfected with C. albicans and nonfermentative gram-negative commensal bacteria exhibited increased fungal burden and inflammatory cytokine production in the kidneys that were dependent on IFN-γ and iNKT cells., Conclusions: Our results indicate that coinfecting commensal bacteria exacerbate C. albicans infection through IFN-γ produced, in part, by iNKT cells.

RCAI-147 is one of the hydroxylated analogues of KRN7000 which is known as a ligand for the activation of CD1d mediated invariant natural killer T cells (iNKT cells) and releases both T helper 1 (Th1) cytokines such as IFN-γ and T helper 2 (Th2) cytokines such as IL-4. KRN7000 has been anticipated as an antitumor drug or an adjuvant for viral infection such as influenza, because of its strong secretion of IFN-γ. In an interesting twist, it has been obvious in our previous paper that RCAI-147 induces much more Th2 cytokines (IL-4) than Th1 cytokines (IFN-γ) from iNKT cells compared to KRN7000, and shows fairly good result in the experimental autoimmune encephalomyelitis (EAE) test. Therefore, synthesis of RCAI-172 (C6-OH epimer of RCAI-147) was attempted to examine the biological activity. As a result, RCAI-172 was synthesized and its biological activity biased to Th2 response largely compared to that of KRN7000. However, this level decreased to approximately 61% compared to that of RCAI-147. And the clinical score of RCAI-172 for EAE suppression was disappointing. There exist seven chiral centers in the aglycon part of RCAI-172, and even though the change of configuration is just one position (C6-OH), the effect on both Th1/Th2 response and EAE test is fairly large., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

A simple zero-velocity method to determine the particle magnetic susceptibility by measuring the magnetophoretic velocity was proposed. The principle is that the magnetophoretic velocity of a particle in a liquid medium must be zero when the magnetic susceptibilities of the medium and the particle are equal, or the gravity force and the magnetophoretic force are balanced. By changing the medium magnetic susceptibility and measuring the magnetophoretic velocity of a particle, the particle magnetic susceptibility was determined from the medium magnetic susceptibility under the zero-velocity condition. The feasibility of the method was demonstrated for polystyrene particles using a Dy(III) solution in the horizontal migration mode and different organic solvents in the vertical migration mode.

Macrophages play a critical role in chronic inflammation and metabolic diseases. We identified a longer splice variant of ubiquitin specific protease (USP) 2-69 as a novel molecule that modulates pathways implicated in metabolic disorders. Expression levels of aP2/FABP4 and PAI-1/SERPINE1 genes were increased by 4- and 1.8-fold, respectively, after short hairpin RNA-mediated knockdown (KD) of the USP2 gene, and such expression was alleviated by overexpression of USP2-69 in human myeloid cell lines. Supernatants derived from USP2-KD cells induced IL6 (∼6-fold) and SAA3 (∼15-fold) in 3T3-L1 adipocytes to suggest the anti-inflammatory properties of USP2. In addition, we observed a 30% decrease in the number of macrophages in mesenteric adipose tissue derived from USP2-69 transgenic mice fed a high-fat diet for 14 wk compared with that in their C57BL/6 littermates (P<0.01), which was consistent with a ∼40% decrease in transcription of aP2 and PAI-1. The aP2 locus exhibited elevated chromatin accessibility (>2.1-fold), methylation of histone H3 lysine 4 (>4.5-fold), and acetylation of histone H4 (>2.5-fold) in USP2-KD cells. Transfection of isopeptidase-mutated USP2-69 did not alter chromatin conformation on the aP2 locus in USP2-KD cells. Our results suggest that USP2-69 suppresses meta-inflammatory molecules involved in the development of type-2 diabetes.

Pancreatic islet transplantation is an attractive therapy for the treatment of insulin-dependent diabetes mellitus. However, the low efficiency of this procedure necessitating sequential transplantations of islets with the use of 2-3 donors for a single recipient, mainly due to the early loss of transplanted islets, hampers its clinical application. Previously, we have shown in mice that a large amount of HMGB1 is released from islets soon after their transplantation and that this triggers innate immune rejection with activation of DC, NKT cells and neutrophils to produce IFN-γ, ultimately leading to the early loss of transplanted islets. Thus, HMGB1 release plays an initial pivotal role in this process; however, its mechanism remains unclear. Here we demonstrate that release of HMGB1 from transplanted islets is due to hypoxic damage resulting from Ca(2+) influx into β cells through the Na(+) /Ca(2+) exchanger (NCX). Moreover, the hypoxia-induced β cell damage was prevented by pretreatment with an NCX-specific inhibitor prior to transplantation, resulting in protection and long-term survival of transplanted mouse and human islets when grafted into mice. These findings suggest a novel strategy with potentially great impact to improve the efficiency of islet transplantation in clinical settings by targeting donor islets rather than recipients., (© Copyright 2013 The American Society of Transplantation and the American Society of Transplant Surgeons.)

In another hospital, a 71-year-old man had been pointed out to have an abnormal shadow in the left lung field on chest radiograph at a medical checkup. Computed tomography( CT) scan revealed a mass both in the S1+2 and in the S10 of the left lung. The mass in the S1+2 was diagnosed as squamous cell carcinoma by bronchoscopy. CT scan revealed a swelling lymph node at the opposite side. He was diagnosed as having lung cancer (cT2N3M1:stage IV) and was given chemotherapy. The S1+2-mass showed a partial response and the S10-nodule was stable disease. He was then changed to gefitinib administration due to severe side effect of the chemotherapy. The side of S10-nodule once reduced, however, it grew up again after 16 months treatment with gefitinib. He was then referred to our hospital for surgery, and bisegmentectomy with mediastinal dissection was performed. He had no complications after the operation, and is well without recurrence for 8 years after the operation.

Previous direct observations of the J aggregates of diprotonated 5,10,15,20-tetraphenyl-21H,23H-porphine (H4TPP(2+)) formed at the dodecane-water interface in a centrifugal liquid membrane (CLM) cell using conventional CD spectroscopy have shown the existence of circular dichroic signals with a bisignated shape whose sign depends on the rotation direction of the cell. Herein we demonstrate that the determination of the optical Mueller matrix with a two-photoelastic modulator generalized ellipsometer (2-MGE), working in transmission mode, along with the assumption of a two superimposed twin Mueller matrices model (two opposite interfaces in the cylindrical rotation cell) allows us to infer the CD spectra due to overlapping linear polarizations from J aggregates at the front and back liquid-liquid interfaces in the rotating cell. The rotation direction dependence of the CD spectra was thoroughly interpreted by the present model considering the sign and the magnitude of the orientation angle between the front and back J aggregates. The present analysis should help us to understand the optical chirality due to the structural changes in supramolecular and macromolecular systems under flow shear forces as well as changes in birefringence.

Objectives: The diagnosis of small lung nodules has increased in recent years; limited resection and minimally invasive surgery are highly desirable in patients with these lesions. While wedge resection may be curative for small lung nodules, the technique is sometimes difficult to perform when the tumour nodule is near the pulmonary hilum. In such situations, either anatomical segmentectomy or subsegmentectomy can obtain an adequate surgical margin; port-access thoracoscopic surgery is the preferred type of minimally invasive surgery. Three-dimensional (3D) computed tomography (CT) simulations are reportedly useful in planning and performing thoracoscopic surgery. We use 3D CT simulation to aid thoracoscopic segmentectomy for small lung nodules and subsegmentectomy for even smaller nodules and conduct here a retrospective evaluation of the clinical results of subsegmentectomy. We present our technique for 3D CT simulation-assisted port-access thoracoscopic subsegmentectomy in the superior segment of the left lower lobe., Methods: Between July 2008 and June 2012, 15 patients underwent port-access thoracoscopic subsegmentectomy. We evaluated the pathological diagnoses, the tumour sizes, the indications, the operative times and the volumes of blood loss., Results: Seven patients were diagnosed with lung cancer (LC) and eight had metastatic lung tumours (MLT). The median tumour size was 12 mm. The indication for using this surgical technique was to secure surgical margins in 13 patients (LC, 6; MLT, 7) and because of poor surgical risk in two patients (LC, 1; MLT, 1). The mean surgical time was 166 min and the median blood loss was 19 ml. There were no recurrences., Conclusions: Port-access thoracoscopic lung subsegmentectomy using 3D CT simulation can be safely performed and is able to secure adequate surgical margins.

We synthesized ten new analogs of 6'-modified KRN7000 (A): RCAI-58, 61, 64, 83, 85-87, 113, 119, and 125. They could be synthesized by α-selective galactosylation of ceramide 9 with the 6-modified D-galactopyranosyl fluorides (8a-8f) or L-arabinopyranosyl fluoride (17), or by etherification of the known alcohol 19. Bioassay of the ten analogs demonstrated that RCAI-61 (1, 6'-O-methylated analog of A) was the most potent immunostimulant among them, and could induce the production of a large amount of IFN-γ even at a low concentration in mice in vivo., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

We have constructed an experimental setup for Faraday rotation dispersion imaging and demonstrated the performance of a novel imaging principle. By using a pulsed magnetic field and a polarized light synchronized to the magnetic field, quantitative Faraday rotation images of diamagnetic organic liquids in glass capillaries were observed. Nonaromatic hydrocarbons, benzene derivatives, and naphthalene derivatives were clearly distinguished by the Faraday rotation images due to the difference in Verdet constants. From the wavelength dispersion of the Faraday rotation images in the visible region, it was found that the resonance wavelength in the UV region, which was estimated based on the Faraday B-term, could be used as characteristic parameters for the imaging of the liquids. Furthermore, simultaneous acquisition of Faraday rotation image and natural optical rotation image was demonstrated for chiral organic liquids.

Laser photophoretic migration behavior of human blood cells in saline solution was investigated under the irradiation of Nd:YAG laser beam (532 nm) in the absence and the presence of the flow in a fused silica capillary. Red blood cells (RBC) were migrated faster than white blood cells (WBC) and blood pellets to the direction of propagation of laser light. The observed photophoretic velocity of RBC was about 11 times faster than those of others. This was understood from the larger photophoretic efficiency of RBC than that of WBC, which was simulated based on the Mie scattering theory. Furthermore, it was found that, during the photophoretic migration, RBCs spontaneously orientated parallel to the migration direction so as to reduce the drag force. Finally, it was demonstrated that RBC and WBC were separated in a micro-channel flow system by the laser photophoresis., (Copyright © 2013 Elsevier B.V. All rights reserved.)

KRN7000 is one of the α-galactosylceramides, which has a 2-hexacosanoylamino-3,4-dihydroxyoctadecyl group. This compound, known as a ligand for the activation of CD1d mediated invariant natural killer T cells (iNKT cells) which release both T helper 1 (Th1) cytokines such as IFNγ and Th2 cytokines such as IL-4, has been anticipated as an antitumor drug, because of its strong secretion of IFNγ. This time, we focused on the hydroxylated analogues of KRN7000 which could be thought of as increasing hydrophilicity and showing bias to Th2 cytokine (IL-4) secretion. Therefore, they may become the drugs for autoimmune diseases for the following reasons: (i) compound OCH, one of the α-galactosylceramide analogues with a shorter sphingosine chain than KRN7000, increases hydrophilicity relative to KRN7000; and (ii) OCH is known to induce much more Th2 cytokines (IL-4) than Th1 cytokines from iNKT cells compared to KRN7000. Naturally, OCH has become one of the candidate drugs for autoimmune diseases. The more hydroxylated derivatives of KRN7000 are anticipated to induce Th2 bias. Therefore, eight analogues with 1-4 excess hydroxyl groups on the lipid chain of KRN7000 were synthesized to examine if they behave in the same way as OCH. As a result, three out of eight compounds biased largely to IL-4 secretion, and their effectiveness for experimental autoimmune encephalomyelitis (EAE) was examined. It was recognized that two compounds (†)RCAI-147/-160 showed good suppression of EAE symptoms., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

We constructed an apparatus to measure the wavelength dispersion of the Faraday rotation in the visible region, and determined the Verdet constants of diamagnetic organic liquids, including aliphatic compounds, benzene derivatives, and naphthalene derivatives. These three groups were easily distinguished by the magnitudes of their Verdet constants. Based on the theory developed by Serber, we determined the enhancing effect of π*←π transitions on the visible-light Faraday rotation angles observed for aromatic compounds. Furthermore, we propose a novel approach for simultaneously observing Faraday rotation dispersion and natural optical rotatory dispersion.

From the aerial part of Boerhaavia erecta L., three new rotenoids (3, 8, 10) and two new coumaronochromonoids (6, 11) were isolated, together with ten known compounds. The structure of the new compounds was established by one dimensional (1D)- and 2D-NMR spectroscopy, as well as high resolution-electrospray ionization (HR-ESI)-MS analysis. The absolute configuration of compound 11 was determined by UV circular dichroism spectroscopy. Compounds were evaluated for their cytotoxic activity against HeLa (human epithelial carcinoma), NCI-H460 (human lung cancer) and MCF-7 (human breast cancer) cell lines at the concentration of 100 µg/mL. Rotenoids 3, 4 and 5 showed a strong cytotoxic activity against HeLa cell line and rotenoids 5 and 8 showed good activity against MCF-7 cell line.

During the past decade, methods for the continuous separation of microparticles with microaction forces have rapidly advanced. Various action forces have been used in designs of both microchannel and capillary continuous separation systems, which depend on properties such as conductivity, permittivity, absorptivity, refractive index, magnetic susceptibility, and compressibility. Particle migration velocity has been used to characterize the particles. Biological cells have been the most interesting targets of these continuous separation methods.

The total internal reflection laser fluorescence microscope method was used to observe the lateral diffusion of a single octadecylrhodamine B (C(18)RB) molecule at the toluene-water interface. The interfacial diffusion constant of single fluorescent cation C(18)RB(+) was obtained from the maximum residence time in a small observation area with pH <2. For pH >3, the maximum residence time was remarkably shortened, indicating that single fluorescent zwitterion C(18)RB(±) rapidly converted to the nonfluorescent lactone at the interface. The lactonization rate was completed within 0.13 ms at the toluene-water interface but slowed to 67 ms at an interface saturated with dimyristoylphosphatidylcholine.

We report a resected case of intrahepatic cholangiocarcinoma (ICC) with a cholangiolocellular carcinoma (CoCC) component. A 77-year-old man had been followed up regularly for chronic hepatitis C for 20 years. In April 2008, a hepatic tumor of 30-mm diameter was detected in segment 2 by computed tomography scan. The tumor was enhanced in the early phase and the enhancement was prolonged. He was treated with transcatheter arterial embolization following the diagnosis of hepatocellular carcinoma (HCC). He had a local recurrence in the same segment (S2) in March 2009, and transcatheter arterial embolization and radiofrequency ablation were performed. In March 2011, abdominal dynamic computed tomography and magnetic resonance imaging showed a sectoral lesion in S2/4 with early and prolonged enhancement. Following the preoperative diagnosis of local recurrence of HCC, he underwent left lobectomy of the liver. Histopathologically, strongly atypical glandular epithelial cells and mucin production were observed, and these cells grew invasively into the liver parenchyma. Furthermore, tumor cells in the peripheral area grew and replaced adjacent hepatic cells, and anastomosing cavities of glands were found on the gitter stein. For these reasons, the histopathological diagnosis was ICC with a CoCC component.

Goblet cell carcinoid (GCC) tumor of the appendix is rare, and the prognosis is considered poor compared to classical carcinoid tumor. We present a case of a 60-year old woman who underwent appendectomy for acute appendicitis. Histopathological studies revealed a GCC tumor with submucosal invasion. Based on the malignancy of this tumor and the possible risk of regional lymph node metastases, we performed laparoscopic ileocecectomy with lymph node dissection. Histologically, no residual tumor or lymph node metastases were found. The patient remains well without any sign of recurrence.

An 81-year-old man complaining of abdominal pain underwent ultrasonography at another hospital that revealed gallbladder swelling, and he was admitted to our hospital for further examination. CT and ERCP revealed a papillary tumor extending from the body to the fundus, leading to the diagnosis of gallbladder cancer. Other examinations showed no sign of invasion or lymph node metastases. We performed full-thickness cholecystectomy. The histopathological diagnosis was mucin-producing papillary adenocarcinoma(T1N0M0, Stage I) of the gallbladder.

A new magnetophoresis method to determine the magnetic susceptibility of single nano/microparticles was developed by applying Brownian motion analysis to determine the size of the particle. This method could measure simultaneously both the magnetophoretic velocity and the radius of the identical single nano/microparticles, which are necessary for the determination of the magnetic susceptibility of the particle. The advantage of this method was demonstrated by the measurement of the diamagnetic susceptibilities of polystyrene particles 500 nm-3 μm in diameter in a paramagnetic 0.5 M manganese(II) chloride solution under a high magnetic field gradient of 5180 T(2) m(-1) generated by a small magnetic circuit.

The J-aggregate of diprotonated tetraphenylporphyrin (H(4)TPP(2+)) formed at the dodecane-water interface showed circular dichroism spectra corresponding to the chirality of 2-alkyl alcohols, longer than 2-butanol, added to the dodecane phase. The phenomenon suggested the preferential interaction between the nano-sized J-aggregates and the chiral alcohols at the interface, and provided a potential use of the J-nanoaggregate as a chiral recognition probe.

RCAI-84, 91, and 105-108 (1-6), the analogs of KRN7000 (A) with a ureido or a thioureido linkage instead of a carboxamido bond, were synthesized to examine their immunostimulatory activity against mouse lymphocytes. According to their bioassay, the ureido analog of KRN7000 [RCAI-105 (1)] and its 6'-O-methylated derivative [RCAI-106 (4)] induced a large amount of IFN-γ in mice in vivo. The hexadecyl ureido analog [RCAI-84 (2)] was comparable to KRN7000 in its bioactivity. The octylureido [RCAI-107 (3)], 5-phenylpentylureido [RCAI-108 (5)], and thioureido [RCAI-91 (6)] analogs were almost inactive., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

A new two-dimensional micro-flow magnetophoresis device was constructed in a superconducting magnet (10 T) using triangular shaped pole pieces, which could apply a magnetic strength, B(dB/dx), in the range of ca. 0-14,000 T(2) m(-1) across a capillary cell. Polystyrene particles with diameters of 1, 3, and 6 μm were used as test samples in a paramagnetic medium of 1 M MnCl(2) to evaluate the performance of this method. Microparticles migrated across the capillary along the edge of the pole pieces, and then flowed through the gap in the pole piece at a position defined as the migration distance, depending on the magnetic susceptibility and the size of particles as well as the flow rate. The most effective flow rate to exhibit the largest resolution among the particles was theoretically predicted and experimentally confirmed. By this method, the magnetic susceptibilities of individual deoxygenated and non-deoxygenated red blood cells were measured from the relative migration distance.

Invariant natural killer T (iNKT) cells are characterized by the expression of an invariant Vα14-Jα18 paired with Vβ8/7/2 in mice, and Vα24-Jα18 with Vβ11 in humans, that recognizes glycolipids, such as α-galactosylceramide (α-GalCer), presented on the MHC class I-like molecule, CD1d. iNKT cells act as innate T lymphocytes and serve as a bridge between the innate and acquired immune systems. iNKT cells augment anti-tumor responses by producing IFN-γ, which acts on NK cells to eliminate MHC-non-restricted (MHC(-)) target tumor cells, and on CD8(+) cytotoxic T lymphocytes to directly kill MHC-restricted (MHC(+)) tumor cells. Thus, when iNKT cells are activated by α-GalCer-pulsed dendritic cells, both MHC(-) and MHC(+) tumor cells can be effectively eliminated. Both of these tumor cell types are simultaneously present in cancer patients, and at present iNKT cells are only the cell type capable of eliminating them. Based on these findings, we have developed iNKT cell-targeted adjuvant immunotherapies with strong anti-tumor activity in humans. However, two-thirds of patients were ineligible for this therapy due to the limited numbers of iNKT cells in their bodies. In order to overcome the problem in cancer patients, we successfully established a method to generate iNKT cells with adjuvant activity from embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). In this review, we would like to outline the clinical potential for iNKT cells derived from ESCs and iPSCs for cancer immunotherapy, and the technical hurdles that must be overcome if we achieve effective ESC/iPSC-mediated cancer therapies.

There is heterogeneity in invariant natural killer T (iNKT) cells based on the expression of CD4 and the IL-17 receptor B (IL-17RB), a receptor for IL-25 which is a key factor in T(H)2 immunity. However, the development pathway and precise function of these iNKT cell subtypes remain unknown. IL-17RB⁺iNKT cells are present in the thymic CD44⁺/⁻ NK1.1⁻ population and develop normally even in the absence of IL-15, which is required for maturation and homeostasis of IL-17RB⁻iNKT cells producing IFN-γ. These results suggest that iNKT cells contain at least two subtypes, IL-17RB⁺ and IL-17RB⁻ subsets. The IL-17RB⁺iNKT subtypes can be further divided into two subtypes on the basis of CD4 expression both in the thymus and in the periphery. CD4⁺ IL-17RB⁺iNKT cells produce T(H)2 (IL-13), T(H)9 (IL-9 and IL-10), and T(H)17 (IL-17A and IL-22) cytokines in response to IL-25 in an E4BP4-dependent fashion, whereas CD4⁻ IL-17RB⁺iNKT cells are a retinoic acid receptor-related orphan receptor (ROR)γt⁺ subset producing T(H)17 cytokines upon stimulation with IL-23 in an E4BP4-independent fashion. These IL-17RB⁺iNKT cell subtypes are abundantly present in the lung in the steady state and mediate the pathogenesis in virus-induced airway hyperreactivity (AHR). In this study we demonstrated that the IL-17RB⁺iNKT cell subsets develop distinct from classical iNKT cell developmental stages in the thymus and play important roles in the pathogenesis of airway diseases., Competing Interests: The authors have declared that no competing interests exist.

The amyloid fibril of bovine insulin and its renaturing intermediates were studied by using Raman optical activity (ROA). In the spectrum of the amyloid, the sharp +/- ROA couplet of amide I band characteristic of the β-sheet-rich proteins was observed, together with a sharp peak at 1271 cm(-1) characteristic of a turn structure. The shoulder ROA peak of the native insulin at ~ 1340 cm(-1), which was assigned to the hydrated α-helix, was not observed in the amyloid, suggesting that the hydrated α-helix was converted to the parallel β-sheet structure in the amyloid. Recovery of the amyloid to the native state was also monitored by ROA. The intermediate states showed distinct features from the amyloid or native ones. The intermediates did not show a characteristic ROA peak of the poly(L-proline) II helix at ~ 1318 cm(-1). The hydrated α-helix ROA peak was not recovered in the intermediate states. In a process of the amyloid formation, at first the hydrated α-helix of the native insulin is converted to a specific partially unfolded structure, and then, it was converted to the parallel β-sheet structure with many turns., (Copyright © 2011 Wiley Periodicals, Inc.)

The magnetic-force quartz crystal microbalance (QCM) method was attempted for the dynamic evaluation of chemical interactions. Thiol-modified magnetic particles bound to a QCM gold electrode of less than three layers were pulled by a magnetic force, and the dissociation dynamics of the magnetic particles was measured based on the change in the weight as a function of time under different temperatures. From an analysis of the dissociated layers and the activation energy for the dissociation, the chemical interactions between particles were evaluated.

Sublobar resection and the optimum approach for small-sized lung cancer remain controversial. We defined the criteria for the selection of cT1aN0M0 patients for thoracoscopic lung surgery and the criteria for other patients for open lung surgery in 2004. Limited resection for small-sized lung cancer was performed when the diameter of the solid lesion was less than 20% the diameter of lesions of groundglass opacity( GGO). The 5-year overall survival and 5-year disease-free survival( 5Y-DFS) was 89.9% and 91.7%, respectively. The 5Y-DFS according to the pathological stage was as follows:stage IA, 96.5%;IB, 100%;IIA, 50%;IIIA, 66.7%;and stage IV, 100%. According to computed tomography (CT) findings, the 5Y-DFS was as follows:pure GGO lesion, 100%;mixed GGO lesion, 100%;and solid lesion, 84.6%. Lymph node metastases and recurrence were noticed only in patients with a solid lesion. Thus, patients with GGO lesions are considered good candidates for thoracoscopic sublobar resection. In addition, thoracoscopic segmentectomy was performed in cT1bN0M0 patients with GGO lesions, and good prognostic results were obtained;therefore, these patients are also considered good candidates for the procedure.

Candida albicans is a major cause of invasive fungal infections. Mortality attributable to candidemia is very high, even when patients are treated with adequate antifungal agents. Therefore, it is important to investigate the mechanisms of immune response to C. albicans. Invariant natural killer T (iNKT) cells, innate lymphocytes that express an invariant T cell receptor α chain, participate in the response to various microbes, including two fungal pathogens, Aspergillus fumigatus and Cryptococcus neoformans. However, it is unknown whether iNKT cells play a role in the immune response to C. albicans. In this study, we have investigated the role of iNKT cells in the host defense against systemic C. albicans infection in mice. We compared the survival and fungal clearance between control mice and Jα18KO mice, which specifically lack iNKT cells, after intravenous C. albicans infection. There was no difference in the survival and fungal burden in the kidneys of the control and Jα18KO mice. Furthermore, production of inflammatory cytokines in several organs during C. albicans infection did not significantly differ between these two groups. These results suggest that iNKT cells play a minor role in controlling systemic C. albicans infections in mice.

Malignant melanoma of the anorectal region is rare, and the prognosis is considered to be poor. We present a case of long-term survival in a 56-year-old patient with primary malignant melanoma in the anorectal area, who complained of anal bleeding. Barium enema showed an elevated lesion in the anorectal region. Colonoscopy revealed a 3 cm sessile tumor with focal pigmentation, and a satellite nodule, 1 cm in diameter. Based on diagnosis of malignant melanoma by biopsy, abdominoperineal resection with lateral node dissection was performed. Pathologically the tumor remained in the mucosa, and no lymph node metastasis was found. This patient refused any adjuvant chemotherapy after the operation, and remains well without any sign of recurrence for seven years.

Raman optical activity (ROA) spectroscopy is used to investigate the backbone conformation of valinomycin in methanol and dioxane solution. Experimental Raman and ROA spectral differences are interpreted by using density functional calculations, molecular dynamics, and Cartesian tensor transfer. Of the several conformers with different numbers of intramolecular hydrogen bonds which were preselected by calculations of relative energies, the dominant ones are identified on the basis of ROA. To separate the backbone signal from that of the side chains, conformational search for the isopropyl residues is performed for each backbone conformer. In dioxane, the most populated conformer does not exhibit C(3) symmetry, but adopts a distorted "bracelet" structure, similar to a crystal structure. This complements previous NMR spectroscopic results that could not distinguish the nonsymmetric structures. In methanol, a different, "propeller" conformer is indicated by ROA, which has three loops resembling a standard β-turn peptide motif. Molecular dynamics simulations suggest that the propeller structure is very flexible in methanol. Spectra simulated for geometries not having the β-turn do not agree with experiment. On the basis of these results, a distinct +/- ROA couplet at ∼1335/1317 cm(-1) observed in the extended amide III region is assigned to a turn in the valinomycin backbone., (Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The immunoregulatory cytokine interleukin 10 (IL-10) is expressed mainly by T helper type 2 (T(H)2) cells but also by T(H)1 cells during chronic infection. Here we observed plasticity in the expression of IL-10 and IL-13 after chronic T(H)1 stimulation; furthermore, the expression of Il10 and Il13 was regulated by the transcription factor E4BP4. Chronically stimulated E4BP4-deficient (Nfil3(-/-); called 'E4bp4(-/-)' here) T(H)1 cells, regulatory T cells (T(reg) cells) and natural killer T cells (NKT cells) had attenuated expression of IL-10 and IL-13. Enforced expression of E4bp4 initiated the production of IL-10 and IL-13 by conventional T(H)1 cells. E4bp4(-/-) T(H)2 cells showed impairment of IL-10 production with no effect on IL-13. Our results indicate that E4BP4 has multiple functions in controlling the plasticity of IL-13 in T(H)1 cells and IL-10 in T(H)1 cells, T(H)2 cells, T(reg) cells and NKT cells.

Application of magnetic field on the separation and analysis of nano/microparticles is a growing subject in analytical separation chemistry. The migration phenomenon of a particle under inhomogeneous magnetic field is called magnetophoresis. The migration velocity depends on the magnetic susceptibility and the size of a particle. Therefore, magnetophoresis allows us to determine the magnetic susceptibility of particles, and to separate particles based on the magnetic properties. Magnetic separation of ferromagnetic particles in liquid has been utilized for a long time. For example, a high gradient magnetic separation under the non-uniform magnetic field generated by ferromagnetic mesh has been utilized in a wide region from chemical industry to bioscience. Recent progress on magnetic nanoparticles and microfluidic devices has made it possible to extend the range of application. Furthermore, it has been demonstrated that the very sensitive measurement of the magnetic susceptibility of microparticles can be performed by observing magnetophoretic velocity. In this review, we mainly introduce novel separation and detection methods based on magnetophoresis, which have been invented in this decade, and then new principles of particle migration under magnetic field are presented., (Copyright © 2011 Elsevier B.V. All rights reserved.)

The effect of static magnetic fields on the budding of single yeast cells was investigated using a magnetic circuit that was capable of generating a strong magnetic field (2.93 T) and gradient (6100 T²  m⁻¹). Saccharomyces cerevisiae yeast cells were grown in an aqueous YPD agar in a silica capillary under either a homogeneous or inhomogeneous static magnetic field. Although the size of budding yeast cells was only slightly affected by the magnetic fields after 4 h, the budding angle was clearly affected by the direction of the homogeneous and inhomogeneous magnetic fields. In the homogeneous magnetic field, the budding direction of daughter yeast cells was mainly oriented in the direction of magnetic field B. However, when subjected to the inhomogeneous magnetic field, the daughter yeast cells tended to bud along the axis of capillary flow in regions where the magnetic gradient, estimated by B(dB/dx), were high. Based on the present experimental results, the possible mechanism for the magnetic effect on the budding direction of daughter yeast cells is theoretically discussed., (Copyright © 2010 Wiley-Liss, Inc.)

The formation and structure of the potassium complex with valinomycin in solution were studied by means of Raman and Raman optical activity (ROA) spectroscopy. The complexation caused significant spectral changes, particularly in the region 1200-1400 cm(-1). The experimental spectra were interpreted using first principles computations. A complete computational conformational search combined with the spectral analysis revealed the arrangement of the isopropyl side chains in the complex. From a total of 6579 unique conformers two predominant ones were confirmed in the solution by ROA. A third one was predicted theoretically, but its population in the experiment could be estimated only roughly. The most populated conformer does not exhibit C(3) symmetry, and is different from that present in the crystal and the NMR-derived structure. Molecular dynamics techniques were used to estimate the molecular flexibility and its effect on the spectra. Density functional computations and Cartesian coordinate transfer (CCT) techniques provided the ROA and Raman spectral shapes and intensities well comparable with the experiment. The polar solvent (methanol) environment modeled with a polarizable continuum model (PCM) leads to rather minor changes in the conformer populations and vibrational properties as compared to vacuum computations, due to the hydrophobic character of the complex. Additional computational experiments suggest that the vibrational interactions determining the ROA spectra are quite local, which contributes to the good spatial resolution of the method. A reduction of the noise in the experimental spectra as well as increased precision of the simulations is desirable for the further exploration of the potential of the ROA spectroscopy for biomolecular studies in the future.

This is a case of a 72-year-old male whose chest computed tomography (CT) revealed a 2.0 x 1.6 cm anterior mediastinal solid tumor during follow-up of an abnormal shadow of the lung. The tumor increased its size during preoperative follow-up, and multilocular cyst was also observed. Radical thymectomy was performed, and histopathologically the tumor was diagnosed as thymic basaloid carcinoma. Thymic basaloid carcinoma is a rare tumor and is often associated with multilocular thymic cyst. There are only 32 cases reported both locally and internationally. Surgical resection is the general treatment for this disease. Adjuvant radiotherapy can be considered in cases of incomplete resection and invasive tumor. In our case, no recurrence of the tumor was noted 12 months post-operative. Generally, the malignancy of thymic basaloid carcinomas are regarded as low-grade compared with other thymic carcinomas, however, since mortality and recurrence have been reported, careful follow-up is required.

Alpha-Galactosylceramide (alphaGalCer, KRN7000) has been identified as a modulator of immunological processes through its capacity to bind iNKT cells mediated by CD1d molecules. Some analogues in while the amide group in alphaGalCer is replaced with ester or ether groups were synthesized from d-arabinitol or l-ribose to evaluate their ability to activate iNKT cells. Ester analogues 30a, 31a, and 61 showed activity for IFNgamma and IL-4 production of iNKT cells, while ether (31b) and 4-methoxy ester (76) analogues of alpha-galactosylceramide were not active for iNKT cells., (Copyright 2010 Elsevier Ltd. All rights reserved.)

NKT cells demonstrate antitumor activity when activated to produce Th1 cytokines by DCs loaded with alpha-galactosylceramide, the prototypic NKT cell-activating glycolipid antigen. However, most patients do not have sufficient numbers of NKT cells to induce an effective immune response in this context, indicating a need for a source of NKT cells that could be used to supplement the endogenous cell population. Induced pluripotent stem cells (iPSCs) hold tremendous potential for cell-replacement therapy, but whether it is possible to generate functionally competent NKT cells from iPSCs has not been rigorously assessed. In this study, we successfully derived iPSCs both from embryonic fibroblasts from mice harboring functional NKT cell-specific rearranged T cell receptor loci in the germline and from splenic NKT cells from WT adult mice. These iPSCs could be differentiated into NKT cells in vitro and secreted large amounts of the Th1 cytokine IFN-gamma. Importantly, iPSC-derived NKT cells recapitulated the known adjuvant effects of natural NKT cells and suppressed tumor growth in vivo. These studies demonstrate the feasibility of expanding functionally competent NKT cells via an iPSC phase, an approach that may be adapted for NKT cell-targeted therapy in humans.

Using the electromagnetophoretic buoyancy for a microparticle in a silica capillary containing an electrolyte solution, the dynamic force dissociation kinetics of the interaction between the mannan polysaccharide on a yeast cell surface and lectins bound to the silica capillary wall have been studied by using the two different modes of the increasing force mode and the constant force mode. Lectins used in the study were concanavalin A (Con A), Hippeastrum hybrid lectin (HHL), Galanthus nivalis lectin (GNL) and Narcissus pseudonarcissus lectin (NPL). The dynamic force measurement by the two different modes gave the spontaneous dissociation rate constant, k(off), of the polysaccharide-lectin binding and the critical increment of bond length at the transition state, Deltax. It was found that the value of k(off) for Con A was the smallest among the lectins studied, due to the strongest binding interaction. It was also confirmed that the magnetophoretic pulling force decreased the transition free energy just like an enzyme.

Background: There have been few case reports of 3rd-line chemotherapy for gastric cancer. So we reported the results of CPT-11 therapy as the 3rd-line chemotherapy for gastric cancer., Patients and Methods: 549 cases underwent gastrectomy from Jan. 2004 to Aug. 2007 in our hospital. In 76 of these cases, which underwent non-curative resection or evidenced a recurrence until July 2009, were analyzed in this study. CPT -11 3rd-line chemotherapy was administered to 11 cases., Results: The mean survival time of non-curative or recurrent cases was 16.9 months. Mean survival times of the non-chemotherapy group, the group administered only 1st-line chemotherapy, the group administered until 3rd-line chemotherapy, the group administered 3rd-line chemotherapy were 7.9 , 11.3 , 21.4 and 28.9 months, respectively(p=0.000 ). Adverse effects occurred in 90.9% of 3rd-line CPT-11, however, all cases were categorized in GradeI., Conclusion: The group administered 3rd-line chemotherapy survived the longest. It is probably correct to administer 3rd-line chemotherapy, if the patient maintains a good performance status.

The effects of unidirectional pulling forces on covalently bridged cysteamine between superparamagnetic particles and Ag nanoparticles (NPs) were studied with SERS spectroscopy. With an increase in the pulling force from 0 to 100 pN per magnetic particle, the nu(C-S)(Trans)/nu(C-S)(Gauche) intensity ratio was increased from 0.6 to 1.08, the Raman frequency of nu(C-S)(Trans) was shifted from 716 to 719 cm(-1), and the Raman bands associated with the amide groups were diminished. From these observations, it was concluded that the magnetic forces induced the extension of distance between the magnetic particle surface and the Ag NP surface and the rotational isomerization equilibrium of SC-CN was shifted from the gauche to the trans conformation with the longer molecular length.