Tomato E8 Encodes a C-27 Hydroxylase in Metabolic Detoxification of α-Tomatine during Fruit Ripening.

Tomato (Solanum lycopersicum) contains α-tomatine, a steroidal glycoalkaloid that contributes to the plant defense against pathogens and herbivores through its bitter taste and toxicity. It accumulates at high levels in all the plant tissues, especially in leaves and immature green fruits, whereas it decreases during fruit ripening through metabolic conversion to the nontoxic esculeoside A, which accumulates in the mature red fruit. This study aimed to identify the gene encoding a C-27 hydroxylase that is a key enzyme in the metabolic conversion of α-tomatine to esculeoside A. The E8 gene, encoding a 2-oxoglutalate-dependent dioxygenase, is well known as an inducible gene in response to ethylene during fruit ripening. The recombinant E8 was found to catalyze the C-27 hydroxylation of lycoperoside C to produce prosapogenin A and is designated as Sl27DOX. The ripe fruit of E8/Sl27DOX-silenced transgenic tomato plants accumulated lycoperoside C and exhibited decreased esculeoside A levels compared with the wild-type (WT) plants. Furthermore, E8/Sl27DOX deletion in tomato accessions resulted in higher lycoperoside C levels in ripe fruits than in WT plants. Thus, E8/Sl27DOX functions as a C-27 hydroxylase of lycoperoside C in the metabolic detoxification of α-tomatine during tomato fruit ripening, and the efficient detoxification by E8/27DOX may provide an advantage in the domestication of cultivated tomatoes.

Identification of α-Tomatine 23-Hydroxylase Involved in the Detoxification of a Bitter Glycoalkaloid.

Tomato plants (Solanum lycopersicum) contain steroidal glycoalkaloid α-tomatine, which functions as a chemical barrier to pathogens and predators. α-Tomatine accumulates in all tissues and at particularly high levels in leaves and immature green fruits. The compound is toxic and causes a bitter taste, but its presence decreases through metabolic conversion to nontoxic esculeoside A during fruit ripening. This study identifies the gene encoding a 23-hydroxylase of α-tomatine, which is a key to this process. Some 2-oxoglutarate-dependent dioxygenases were selected as candidates for the metabolic enzyme, and Solyc02g062460, designated Sl23DOX, was found to encode α-tomatine 23-hydroxylase. Biochemical analysis of the recombinant Sl23DOX protein demonstrated that it catalyzes the 23-hydroxylation of α-tomatine and the product spontaneously isomerizes to neorickiioside B, which is an intermediate in α-tomatine metabolism that appears during ripening. Leaves of transgenic tomato plants overexpressing Sl23DOX accumulated not only neorickiioside B but also another intermediate, lycoperoside C (23-O-acetylated neorickiioside B). Furthermore, the ripe fruits of Sl23DOX-silenced transgenic tomato plants contained lower levels of esculeoside A but substantially accumulated α-tomatine. Thus, Sl23DOX functions as α-tomatine 23-hydroxylase during the metabolic processing of toxic α-tomatine in tomato fruit ripening and is a key enzyme in the domestication of cultivated tomatoes.

Identification of a 3ß-Hydroxysteroid Dehydrogenase/ 3-Ketosteroid Reductase Involved in α-Tomatine Biosynthesis in Tomato.

α-Tomatine and dehydrotomatine are major steroidal glycoalkaloids (SGAs) that accumulate in the mature green fruits, leaves and flowers of tomato (Solanum lycopersicum), and function as defensive compounds against bacteria, fungi, insects and animals. The aglycone of dehydrotomatine is dehydrotomatidine (5,6-dehydrogenated tomatidine, having the Δ5,6 double bond; the dehydro-type). The aglycone of α-tomatine is tomatidine (having a single bond between C5 and C6; the dihydro-type), which is believed to be derived from dehydrotomatidine via four reaction steps: C3 oxidation, isomerization, C5 reduction and C3 reduction; however, these conversion processes remain uncharacterized. In the present study, we demonstrate that a short-chain alcohol dehydrogenase/reductase designated Sl3ßHSD is involved in the conversion of dehydrotomatidine to tomatidine in tomato. Sl3ßHSD1 expression was observed to be high in the flowers, leaves and mature green fruits of tomato, in which high amounts of α-tomatine are accumulated. Biochemical analysis of the recombinant Sl3ßHSD1 protein revealed that Sl3ßHSD1 catalyzes the C3 oxidation of dehydrotomatidine to form tomatid-4-en-3-one and also catalyzes the NADH-dependent C3 reduction of a 3-ketosteroid (tomatid-3-one) to form tomatidine. Furthermore, during co-incubation of Sl3ßHSD1 with SlS5αR1 (steroid 5α-reductase) the four reaction steps converting dehydrotomatidine to tomatidine were completed. Sl3ßHSD1-silenced transgenic tomato plants accumulated dehydrotomatine, with corresponding decreases in α-tomatine content. Furthermore, the constitutive expression of Sl3ßHSD1 in potato hairy roots resulted in the conversion of potato SGAs to the dihydro-type SGAs. These results demonstrate that Sl3ßHSD1 is a key enzyme involved in the conversion processes from dehydrotomatidine to tomatidine in α-tomatine biosynthesis.

Efficient Regeneration of Human Vα24+ Invariant Natural Killer T Cells and Their Anti-Tumor Activity In Vivo.

Reprogramming of antigen-specific T lymphocytes into induced pluripotent stem cells (iPSCs) and their subsequent re-differentiation has enabled expansion of functional T lymphocytes in vitro, thus opening up new approaches for immunotherapy of cancer and other diseases. In this study, we have established a robust protocol to reprogram human invariant NKT (Vα24+ iNKT) cells, which have been shown to act as cellular adjuvants and thus exert anti-tumor activity in mice and humans, and to re-differentiate the iNKT cell-derived iPSCs into functional iNKT cells. These iPSC-derived iNKT cells (iPS-Vα24+ iNKT cells) can be activated by ligand-pulsed dendritic cells (DCs) and produce a large amount of interferon-γ upon activation, as much as parental Vα24+ iNKT cells, but exhibit even better cytotoxic activity against various tumor cell lines. The iPS-Vα24+ iNKT cells possess significant anti-tumor activity in tumor-bearing mice and can activate autologous NK cells upon activation by ligand-pulsed DCs in the NOG mouse model in vivo, further extending their therapeutic potential. This study thus provides a first proof of concept for the clinical application of human iPS-Vα24+ iNKT cells for cancer immunotherapy. Stem Cells 2016;34:2852-2860.

B and T lymphocyte attenuator inhibits LPS-induced endotoxic shock by suppressing Toll-like receptor 4 signaling in innate immune cells.

Although innate immune responses are necessary for the initiation of acquired immune responses and the subsequent successful elimination of pathogens, excessive responses occasionally result in lethal endotoxic shock accompanied by a cytokine storm. B and T lymphocyte attenuator (BTLA), a coinhibitory receptor with similarities to cytotoxic T-lymphocyte antigen (CTLA)-4 and programmed death (PD)-1, is expressed in not only B and T cells but also dendritic cells (DCs) and macrophages (Mϕs). Recently, several studies have reported that BTLA-deficient (BTLA(-/-)) mice show enhanced pathogen clearance compared with WT mice in early phase of infections. However, the roles of BTLA expressed on innate cells in overwhelming and uncontrolled immune responses remain unclear. Here, we found that BTLA(-/-) mice were highly susceptible to LPS-induced endotoxic shock. LPS-induced TNF-α and IL-12 production in DCs and Mϕs was significantly enhanced in BTLA(-/-) mice. BTLA(-/-) DCs also produced high levels of TNF-α on stimulation with Pam3CSK4 but not poly(I:C) or CpG, suggesting that BTLA functions as an inhibitory molecule on Toll-like receptor signaling at cell surface but not endosome. Moreover, BTLA(-/-) DCs showed enhanced MyD88- and toll/IL-1R domain-containing adaptor inducing IFN (TRIF)-dependent signaling on LPS stimulation, which is associated with impaired accumulation of Src homology 2-containing protein tyrosine phosphatase in lipid rafts. Finally, we found that an agonistic anti-BTLA antibody rescued mice from LPS-induced endotoxic shock, even if the antibody was given to mice that had developed a sign of endotoxic shock. These results suggest that BTLA directly inhibits LPS responses in DCs and Mϕs and that agonistic agents for BTLA might have therapeutic potential for LPS-induced endotoxic shock.

Th2-type inflammation instructs inflammatory dendritic cells to induce airway hyperreactivity.

Dendritic cells (DCs) play critical roles in determining the fate of CD4⁺ T cells. Among DC sub-populations, monocyte-derived inflammatory DCs (iDCs) have been shown to play an important role in the induction of adaptive immune responses under inflammatory conditions. Although previous studies have shown that DCs have an indispensable role in the induction of allergic airway inflammation and airway hyperreactivity (AHR) in murine asthma models, the precise roles of iDCs in the asthmatic responses remain largely unknown. We show here that T(h)2 cell-mediated inflammation in murine asthma models induces the expression of some markers of alternatively activated macrophage such as arginase 1 and resistin-like molecule-α in iDCs by a mechanism depending on the intrinsic expression of STAT6. In contrast, T(h)1 cell-mediated inflammation induces iDCs to express TNF-α and inducible nitric oxide synthase (iNOS), markers of TNF-α- and iNOS-producing DCs. Moreover, we show that iDCs under a T(h)2 environment play an important role in the induction of AHR, independently of allergic airway inflammation. Our results thus indicate the importance of iDCs in the induction of AHR as downstream effector cells in T(h)2 cell-mediated asthmatic responses.

Efficacy and safety of IgPro20, a subcutaneous immunoglobulin, in Japanese patients with primary immunodeficiency diseases.

PURPOSE: Intravenous (IVIG) and subcutaneous (SCIG) immunoglobulin infusions are widely used for the treatment of patients with primary immunodeficiency (PID) worldwide. This prospective, multicenter, open-label, single-arm Phase III study evaluated the efficacy, tolerability, and safety of IgPro20 (Hizentra®; L-proline-stabilized 20 % human SCIG) in adult and pediatric Japanese patients with PID. METHODS: Patients received three IVIG infusions at 3-4-week intervals followed by a dose-equivalent switch to weekly SCIG infusions. A 12-week wash-in/wash-out period was followed by a 12-week SCIG efficacy period. The primary efficacy endpoint was the comparison of total serum IgG trough levels during the IVIG and SCIG efficacy periods by calculating the geometric mean ratio (GMR). RESULTS: The GMR of IgG trough levels on SCIG versus IVIG was 1.09 (2-sided 90% confidence interval: 1.06-1.13). No serious bacterial infections were reported. Eleven patients (52.4%) had infectious episodes with an overall rate of 2.98 infections/patient/year; 7 patients (33.3%) missed school/work/daycare due to infection (3.48 days/patient/year). Sixteen patients (76.2%) were treated with antibiotics for an adverse event (AE; 47.6%) or prophylaxis (23.8%), resulting in 167.42 days/patient/year of antibiotic use. During SCIG treatment, 24 patients (96.0%) had 269 AEs (0.461 AEs per/infusion) including local reactions as the most common AE (20 patients, 80.0%). Local tolerability of IgPro20 was assessed as "very good" or "good" after 85.4% of SCIG infusions. One patient (4.0%) experienced a serious AE of moderate severity (bacterial infection) that was considered unrelated to study medication. CONCLUSION: IgPro20 was effective and well tolerated in Japanese patients with PID.

Effect of Dapagliflozin on Serum Uric Acid Levels in Patients with Advanced Chronic Kidney Disease.

Objective Sodium-glucose cotransporter 2 (SGLT2) inhibitors, which are hypoglycemic agents, have been shown to be cardioprotective through a variety of mechanisms, and the effect of lowering uric acid (UA) levels may be one of the mechanisms. In the present retrospective study, we investigated the changes in serum UA levels in patients with chronic kidney disease (CKD) treated with SGLT2 inhibitors. Methods We included 31 patients with CKD who were newly started on dapagliflozin for renal protection and evaluated trends in various parameters, including serum UA levels and UA excretion from urine. Results The patients' median age was 71 years old, 20 patients were men, 7 patients had diabetes, and the median estimated glomerular filtration rate was 33.9 mL/min/1.73 m2 (CKD stage 3: 21 cases, stage 4: 10 cases). The differences in UA and fractional excretion of UA after three weeks and three months of prescription showed significantly decreased UA values and an increased fractional excretion of UA. Conclusion Our findings suggest that dapagliflozin can lower serum UA levels via increased UA excretion, even in patients with advanced CKD.

Protocol for cortical-wide field-of-view two-photon imaging with quick neonatal adeno-associated virus injection.

We recently established a simple and versatile adeno-associated virus (AAV) induction approach that enables dense (>90% labeled neurons) and cortical-wide Ca2+ sensor expression. Here, we describe the stepwise protocol for neonatal AAV injection of a Ca2+ sensor. We also detail the steps for subsequent craniotomy to generate a chronic cranial window, followed by wide-field two-photon Ca2+ imaging in an awake mouse. This protocol serves as an alternative to the use of transgenic animals and offers translatable options for cortical-wide experiments. For complete details on the use and execution of this protocol, please refer to Ota et al. (2021).

Pain-like behavior in mice can be induced by the environmental context in which the pain stimulus was previously given.

It has been known for decades that classical conditioning influences pain perception. However, the precise mechanism of pain modified by conditioning remains unclear, partly because of the lack of dedicated behavioral tests. In the present study, we aimed to develop a new method to detect conditioned pain using mice that were injected with formalin as an unconditioned nociceptive stimulus into the hind paw repetitively under a neutral environment. On the test day, the mice exhibited a pain-like behavior without the application of a pain stimulus in the environment. These results demonstrate that a conditioned nociceptive response can be induced by exposure alone to the environmental context in which the pain was previously experienced. The conditioned nociceptive response was sustained for at least 2 weeks. Furthermore, the conditioned nociceptive response was reduced by fentanyl but not by ibuprofen, pregabalin or fluvoxamine. This method may be useful for studying the mechanisms of irritable chronic pain and for the development of novel therapeutic strategies.

Fast, cell-resolution, contiguous-wide two-photon imaging to reveal functional network architectures across multi-modal cortical areas.

Fast and wide field-of-view imaging with single-cell resolution, high signal-to-noise ratio, and no optical aberrations have the potential to inspire new avenues of investigations in biology. However, such imaging is challenging because of the inevitable tradeoffs among these parameters. Here, we overcome these tradeoffs by combining a resonant scanning system, a large objective with low magnification and high numerical aperture, and highly sensitive large-aperture photodetectors. The result is a practically aberration-free, fast-scanning high optical invariant two-photon microscopy (FASHIO-2PM) that enables calcium imaging from a large network composed of ∼16,000 neurons at 7.5 Hz from a 9 mm2 contiguous image plane, including more than 10 sensory-motor and higher-order areas of the cerebral cortex in awake mice. Network analysis based on single-cell activities revealed that the brain exhibits small-world rather than scale-free behavior. The FASHIO-2PM is expected to enable studies on biological dynamics by simultaneously monitoring macroscopic activities and their compositional elements.

Alpha-amylase production is induced by sulfuric acid in rice aleurone cells.

The hydrolytic enzyme alpha-amylase (EC 3.2.1.1) is produced mainly in aleurone cells of germinating cereals, and the phytohormone gibberellin (GA) is essential for its induction. However, in rice (Oryza sativa L.), sulfuric acid (H(2)SO(4)) induces alpha-amylase production in aleurone tissue even in the absence of GA. Here, the pre-treatment of rice aleurone cells with H(2)SO(4) and incubation in water induced alpha-amylase activity, as if the cells had been incubated in GA solution.

Whole-body MRI for detecting metastatic bone tumor: diagnostic value of diffusion-weighted images.

PURPOSE: We assessed the diagnostic value of whole body magnetic resonance (MR) imaging (WB-MRI) using diffusion-weighted images (DWI) for detecting bone metastasis and compared it with that of skeletal scintigraphy (SS). MATERIALS AND METHODS: Thirty patients with malignancies (breast cancer, 17 patients; prostate cancer, 9; and one patient each, thyroid cancer, liposarcoma, leiomyosarcoma, and extraskeletal Ewing sarcoma) underwent both WB-MRI and SS to detect bone metastasis. All patients were followed more than 6 months by MR imaging, SS, or computed tomographic (CT) examination. For WB-MRI, patients were placed in feet-first supine position with table-top extender and quadrature body coil. We acquired DWI (axial plane from lower neck to proximal femur) (single shot short TI inversion-recovery [STIR]: repetition time [TR] 6243/echo time [TE] 59/inversion time [TI] 180 ms; b value: 600 s/mm(2); 5-mm slice thickness; 112 x 112 matrix), T(1)-weighted fast spin echo (T(1)WI), and STIR (sagittal plane of total spine images and coronal plane of whole body images) images. Four blinded readers independently and separately interpreted images of combined MR sequences of T(1)WI+STIR (session 1) and T(1)WI+STIR+DWI (session 2). RESULTS: In 10 of 30 patients, we detected a total of 52 metastatic bone lesions; in the other 20, follow-up examinations confirmed no metastatic bone lesions. For these 52 lesions, for session 2, the mean sensitivity was 96% and the positive predictive value (PPV) was 98%. Those values were superior to those of session 1 (sensitivity: 88%; PPV: 95%) and those of SS (sensitivity: 96%; PPV: 94%). CONCLUSION: WB-MRI that included DWI was useful for detecting bone metastasis.

Whole body MRI for detecting metastatic bone tumor: comparison with bone scintigrams.

PURPOSE: To compare the effectiveness of whole body MRI (WB-MRI [magnetic resonance imaging]) and bone scintigram (BS) at detecting bone metastasis. MATERIALS AND METHODS: WB-MRI was performed on 16 patients for detecting bone metastasis (6 breast carcinoma, 7 prostatic carcinoma, 1 renal cell carcinoma [RCC], 1 hepatocellular carcinoma [HCC], and 1 primary unknown). BS was also performed in all cases. Patients were placed on a table top extender (Philips Medical Systems). The maximal longitudinal field of view (FOV) was 200 cm. At first, the total spine was imaged in the sagittal plane with a three-station approach for two image sets (fast spin-echo [SE] T1-weighted images [T1WI] and short tau inversion recovery [STIR] images). The whole body was then imaged in the coronal plane with a seven-station approach for two image sets (fast field echo [FFE] T1WI and STIR). Total examination time, including patient positioning, was within 40 min. Three independent radiologists interpreted the imaging data. RESULTS: WB-MRI identified 5 cases of 24 lesions as bone metastasis, while BS identified 3 cases of 25 lesions. Concordance between WB-MRI and BS was seen in 3 cases of 22 lesions (81%). For two cases of 2 lesions, which were identified only with WB-MRI, the lesions were located in the sacrum and thoracic spine. For one case of 3 lesions, which was identified only with BS, the lesions were located in the skull and rib. CONCLUSION: WB-MRI was an excellent method for screening bone metastasis, especially the vertebral body.

Distinctiveness of the genomic sequence of Shiga toxin 2-converting phage isolated from Escherichia coli O157:H7 Okayama strain as compared to other Shiga toxin 2-converting phages.

Shiga toxin 2-converting phage was isolated from Escherichia coli O157:H7 associated with an outbreak that occurred in Okayama, Japan in 1996 (M. Watarai, T. Sato, M. Kobayashi, T. Shimizu, S. Yamasaki, T. Tobe, C. Sasakawa and Y. Takeda, Infect. Immun. 61 (1998) 3210-3204). In this study, we analyzed the complete nucleotide sequence of Shiga toxin 2-converting phage, designated Stx2phi-I, and compared it with three recently reported Stx2-phage genomes. Stx2phi-I consisted of 61,765 bp, which included 166 open reading frames. When compared to 933W, VT2-Sakai and VT2-Sa phages, six characteristic regions (regions I-VI) were found in the Stx2 phage genomes although overall homology was more than 95% between these phages. Stx2phi-I exhibited remarkable differences in these regions as compared with VT-2 Sakai and VT2-Sa genes but not with 933W phage. Characteristic repeat sequences were found in regions I-IV where the genes responsible for the construction of head and tail are located. Regions V and VI, which are the most distinct portion in the entire phage genome were located in the upstream and downstream regions of the Stx2 operons that are responsible for the immunity and replication, and host lysis. These data indicated that Stx2phi-I is less homologous to VT2-Sakai and VT2-Sa phages, despite these three phages being found in the strains isolated at the almost same time in the same geographic region but closely related to 933W phage which was found in the E. coli O157 strain 933W isolated 14 years ago in a different geographic area.

Cost-minimization analysis of IgPro20, a subcutaneous immunoglobulin, in Japanese patients with primary immunodeficiency.

PURPOSE: IgPro20, Hizentra(®) an L-proline-stabilized 20% human subcutaneous immunoglobulin (SCIG), has been shown in a Phase III pivotal study to be well tolerated and efficacious in adult and pediatric Japanese patients with primary immunodeficiency. Economic aspects of SCIG treatment in comparison with previous intravenous immunoglobulin (IVIG) therapy were analyzed in this Phase III study in Japan. METHODS: Twenty-four Japanese patients with primary immunodeficiency on IVIG treatment were switched to IgPro20 at an equivalent dose (full analysis set). The study consisted of a screening period, an IVIG treatment period with 3 planned infusions every 3 or 4 weeks, a 12-week SCIG wash-in and wash-out period, and a 12-week SCIG efficacy period. The difference in medical cost and productivity loss resulting from changes in hospital frequency between the SCIG and IVIG treatment was evaluated. Information about treatment cost was collected as part of the Life Quality Index questionnaire. In addition, productivity loss and hospital-related absenteeism were evaluated. FINDINGS: Life Quality Index scores for all domains were higher with SCIG than with IVIG in this patient population. In the full analysis set, the mean (SD) Life Quality Index score of the Costs domain increased from 45.1 (26.34) at Week 1 (IVIG period) to 71.9 (18.52) at Week 24 (end of the SCIG efficacy period), representing a mean change of 26.74 and a large score improvement effect size (1.01). Median productivity loss was reduced by 60% from baseline to Weeks 12 and 24. This resulted in a reduction in costs of JPY 10,875 per patient per month at Weeks 12 and 24. Subcutaneous treatment with IgPro20 also reduced hospital-related absenteeism. The number of patients, parents, or guardians who were not absent from work or housework duties and had no reduction in working time increased from 4 (17.4%) at Week 1 to 9 (39.1%) at Week 24. Similar results were obtained in the per-protocol set (n = 21). IMPLICATIONS: Switching from IVIG to SCIG reduced markedly productivity loss and hospital-related absenteeism. The reduction in hospital visit frequency due to the use of home-based IgG therapy enabled by the change in administration route is expected to produce an important pharmacoeconomic benefit in Japan. Study Code: ZLB06_002CR, ClinicalTrials.gov identifier: NCT01199705.

Skin temperature responses to cold stress in patients with severe motor and intellectual disabilities.

Patients with severe motor and intellectual disabilities (SMID) often suffer from autonomic nervous system disturbances. At the same time, the caregivers of patients with SMID face challenges to understand the patients' chronic health problems effectively by simply observing them. Therefore, recognizing specific symptoms is important to improve support for SMID. We investigated the autonomic nervous function in patients with SMID with skin vasomotor responses to cold stimuli. The relationship of the results of cold stress and autonomic symptoms observed by the main caretakers was also examined. We analyzed 38 patients with SMID. Their hand skin temperature was measured before and after cold stimuli using infrared thermography. A 'distal-dorsal difference' (DDD) at baseline, and the recovery rate of the second fingertip and dorsum were calculated. All main caregivers filled out questionnaires evaluating autonomic symptoms. The recovery rate of the second fingertip and dorsum after cold stimuli was lower than 80% in 64% and 60% patients, respectively. The baseline DDD was greater than 1 °C in 84% of the patients. A DDD>1 °C was associated with a reduced recovery rate. All caregivers recognized some autonomic-related symptoms. Patients with constipation or snoring demonstrated a reduced recovery rate. However, none of the observed symptoms can predict the presence of a reduced rate with cold stimuli in a statistically significant way. This study showed excessive sympathetic nerve activities in patients with SMID. The baseline DDD could be a valuable parameter accessing their microvascular circulation. To improve the life of a person with SMID, accessing autonomic function using a noninvasive method, such as thermography is warranted without directly observed symptoms.

Chemical discrimination and aggressiveness via cuticular hydrocarbons in a supercolony-forming ant, Formica yessensis.

BACKGROUND: Territorial boundaries between conspecific social insect colonies are maintained through nestmate recognition systems. However, in supercolony-forming ants, which have developed an extraordinary social organization style known as unicoloniality, a single supercolony extends across large geographic distance. The underlying mechanism is considered to involve less frequent occurrence of intraspecific aggressive behaviors, while maintaining interspecific competition. Thus, we examined whether the supercolony-forming species, Formica yessensis has a nestmate recognition system similar to that of the multicolonial species, Camponotus japonicus with respect to the cuticular hydrocarbon-sensitive sensillum (CHC sensillum), which responds only to non-nestmate CHCs. We further investigated whether the sensory system reflects on the apparent reduced aggression between non-nestmates typical to unicolonial species. METHODOLOGY/PRINCIPAL FINDINGS: F. yessensis constructs supercolonies comprising numerous nests and constitutes the largest supercolonies in Japan. We compared the within-colony or between-colonies' (1) similarity in CHC profiles, the nestmate recognition cues, (2) levels of the CHC sensillar response, (3) levels of aggression between workers, as correlated with geographic distances between nests, and (4) their genetic relatedness. Workers from nests within the supercolony revealed a greater similarity of CHC profiles compared to workers from colonies outside it. Total response of the active CHC sensilla stimulated with conspecific alien CHCs did not increase as much as in case of C. japonicus, suggesting that discrimination of conspecific workers at the peripheral system is limited. It was particularly limited among workers within a supercolony, but was fully expressed for allospecific workers. CONCLUSIONS/SIGNIFICANCE: We demonstrate that chemical discrimination between nestmates and non-nestmates in F. yessensis was not clear cut, probably because this species has only subtle intraspecific differences in the CHC pattern that typify within a supercolony. Such an incomplete chemical discrimination via the CHC sensilla is thus an important factor contributing to decreased occurrence of intraspecific aggressive behavior especially within a supercolony.

Murine induced pluripotent stem cells can be derived from and differentiate into natural killer T cells.

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.