Allergic inflammatory memory in human respiratory epithelial progenitor cells.

Barrier tissue dysfunction is a fundamental feature of chronic human inflammatory diseases1. Specialized subsets of epithelial cells-including secretory and ciliated cells-differentiate from basal stem cells to collectively protect the upper airway2-4. Allergic inflammation can develop from persistent activation5 of type 2 immunity6 in the upper airway, resulting in chronic rhinosinusitis, which ranges in severity from rhinitis to severe nasal polyps7. Basal cell hyperplasia is a hallmark of severe disease7-9, but it is not known how these progenitor cells2,10,11 contribute to clinical presentation and barrier tissue dysfunction in humans. Here we profile primary human surgical chronic rhinosinusitis samples (18,036 cells, n = 12) that span the disease spectrum using Seq-Well for massively parallel single-cell RNA sequencing12, report transcriptomes for human respiratory epithelial, immune and stromal cell types and subsets from a type 2 inflammatory disease, and map key mediators. By comparison with nasal scrapings (18,704 cells, n = 9), we define signatures of core, healthy, inflamed and polyp secretory cells. We reveal marked differences between the epithelial compartments of the non-polyp and polyp cellular ecosystems, identifying and validating a global reduction in cellular diversity of polyps characterized by basal cell hyperplasia, concomitant decreases in glandular cells, and phenotypic shifts in secretory cell antimicrobial expression. We detect an aberrant basal progenitor differentiation trajectory in polyps, and propose cell-intrinsic13, epigenetic14,15 and extrinsic factors11,16,17 that lock polyp basal cells into this uncommitted state. Finally, we functionally demonstrate that ex vivo cultured basal cells retain intrinsic memory of IL-4/IL-13 exposure, and test the potential for clinical blockade of the IL-4 receptor α-subunit to modify basal and secretory cell states in vivo. Overall, we find that reduced epithelial diversity stemming from functional shifts in basal cells is a key characteristic of type 2 immune-mediated barrier tissue dysfunction. Our results demonstrate that epithelial stem cells may contribute to the persistence of human disease by serving as repositories for allergic memories.

Type 2 Cysteinyl Leukotriene Receptors Drive IL-33-Dependent Type 2 Immunopathology and Aspirin Sensitivity.

Cysteinyl leukotrienes (cysLTs) facilitate mucosal type 2 immunopathology by incompletely understood mechanisms. Aspirin-exacerbated respiratory disease, a severe asthma subtype, is characterized by exaggerated eosinophilic respiratory inflammation and reactions to aspirin, each involving the marked overproduction of cysLTs. Here we demonstrate that the type 2 cysLT receptor (CysLT2R), which is not targeted by available drugs, is required in two different models to amplify eosinophilic airway inflammation via induced expression of IL-33 by lung epithelial cells. Endogenously generated cysLTs induced eosinophilia and expanded group 2 innate lymphoid cells (ILC2s) in aspirin-exacerbated respiratory disease-like Ptges-/- mice. These responses were mitigated by deletions of either Cysltr2 or leukotriene C4 synthase (Ltc4s). Administrations of either LTC4 (the parent cysLT) or the selective CysLT2R agonist N-methyl LTC4 to allergen sensitized wild-type mice markedly boosted ILC2 expansion and IL-5/IL-13 generation in a CysLT2R-dependent manner. Expansion of ILC2s and IL-5/IL-13 generation reflected CysLT2R-dependent production of IL-33 by alveolar type 2 cells, which engaged in a bilateral feed-forward loop with ILC2s. Deletion of Cysltr1 blunted LTC4-induced ILC2 expansion and eosinophilia but did not alter IL-33 induction. Pharmacological blockade of CysLT2R prior to inhalation challenge of Ptges-/- mice with aspirin blocked IL-33-dependent mast cell activation, mediator release, and changes in lung function. Thus, CysLT2R signaling, IL-33-dependent ILC2 expansion, and IL-33-driven mast cell activation are necessary for induction of type 2 immunopathology and aspirin sensitivity. CysLT2R-targeted drugs may interrupt these processes.

[A Case of Gastric Schwannoma Diagnosed Preoperatively].

The patient was a 52-year-old woman, who was found to have an abnormality in the upper gastrointestinal(UGI)tract, via a contrast-imaging study; she had no symptoms.Computed tomography(CT)revealed a tumor, measuring approximately 100mm in diameter, in the antrum of the stomach.The tumor was diagnosed as gastric schwannoma using endoscopic ultrasonography-guided fine-needle aspiration(EUS-FNA).Preoperative CT revealed multiple lymph adenopathies around the antrum, which led to the suspicion of lymph node metastasis. The patient underwent a laparoscopic partial gastrectomy after the confirmation of the absence of lymph node metastasis by intraoperative rapid diagnosis.

Leukotriene E4 elicits respiratory epithelial cell mucin release through the G-protein-coupled receptor, GPR99.

Cysteinyl leukotrienes (cysLTs), leukotriene C4 (LTC4), LTD4, and LTE4 are proinflammatory lipid mediators with pathobiologic function in asthma. LTE4, the stable cysLT, is a weak agonist for the type 1 and type 2 cysLT receptors (CysLTRs), which constrict airway smooth muscle, but elicits airflow obstruction and pulmonary inflammation in patients with asthma. We recently identified GPR99 as a high-affinity receptor for LTE4 that mediates cutaneous vascular permeability. Here we demonstrate that a single intranasal exposure to extract from the respiratory pathogen Alternaria alternata elicits profound epithelial cell (EpC) mucin release and submucosal swelling in the nasal mucosa of mice that depends on cysLTs, as it is absent in mice deficient in the terminal enzyme for cysLT biosynthesis, LTC4 synthase (LTC4S). These mucosal changes are associated with mast cell (MC) activation and absent in MC-deficient mice, suggesting a role for MCs in control of EpC function. Of the three CysLTRs, only GPR99-deficient mice are fully protected from EpC mucin release and swelling elicited by Alternaria or by intranasal LTE4 GPR99 expression is detected on lung and nasal EpCs, which release mucin to doses of LTE4 one log lower than that required to elicit submucosal swelling. Finally, mice deficient in MCs, LTC4S, or GPR99 have reduced baseline numbers of goblet cells, indicating an additional function in regulating EpC homeostasis. These results demonstrate a novel role for GPR99 among CysLTRs in control of respiratory EpC function and suggest that inhibition of LTE4 and of GPR99 may have therapeutic benefits in asthma.

Gastric washing by distilled water can reduce free gastric cancer cells exfoliated into the stomach lumen.

PURPOSE: Intragastric free cancer cells in patients with gastric cancer have rarely been studied. The purpose of this study was to investigate the detection rate of intragastric free cancer cells in gastric washes using two types of solutions during endoscopic examination. We further clarified risk factors affecting the presence of exfoliated free cancer cells. METHODS: A total of 175 patients with gastric cancer were enrolled. Lactated Ringer's solution (N = 89) or distilled water (DW; N = 86) via endoscopic working channel was sprayed onto the tumor surface, and the resultant fluid was collected for cytological examination. We compared the cancer-cell positivity rate between the two (Ringer and DW) groups. We also tested the correlation between cancer-cell positivity and clinicopathological factors in the Ringer group to identify risk factors for the presence of exfoliated cancer cells. RESULTS: The cancer-cell positivity rate was significantly higher in the Ringer group than that in the DW group (58 vs 6%). Cytomorphology in the Ringer group was well maintained, but not in the DW group. The larger tumor size (≥ 20 mm) and positive lymphatic involvement were significant risk factors of exfoliated free cancer cells. CONCLUSIONS: Cancer cells can be highly exfoliated from the tumor surface into the gastric lumen by endoscopic irrigation in large gastric cancer with lymphatic involvement. Gastric washing by DW can lead to cytoclasis of free cancer cells; therefore, it may minimize the possibility of cancer-cell seeding in procedures carrying potential risks of tumor-cell seeding upon transluminal communication, such as endoscopic full-thickness resection and laparoscopy-endoscopy cooperative surgery.

Phosphoinositide 3-Kinase δ Regulates Dectin-2 Signaling and the Generation of Th2 and Th17 Immunity.

The C-type lectin receptor Dectin-2 can trigger the leukotriene C4 synthase-dependent generation of cysteinyl leukotrienes and the caspase-associated recruitment domain 9- and NF-κB-dependent generation of cytokines, such as IL-23, IL-6, and TNF-α, to promote Th2 and Th17 immunity, respectively. Dectin-2 activation also elicits the type 2 cytokine IL-33, but the mechanism by which Dectin-2 induces these diverse innate mediators is poorly understood. In this study, we identify a common upstream requirement for PI3Kδ activity for the generation of each Dectin-2-dependent mediator elicited by the house dust mite species, Dermatophagoides farinae, using both pharmacologic inhibition and small interfering RNA knockdown of PI3Kδ in bone marrow-derived dendritic cells. PI3Kδ activity depends on spleen tyrosine kinase (Syk) and regulates the activity of protein kinase Cδ, indicating that PI3Kδ is a proximal Syk-dependent signaling intermediate. Inhibition of PI3Kδ also reduces cysteinyl leukotrienes and cytokines elicited by Dectin-2 cross-linking, confirming the importance of this molecule in Dectin-2 signaling. Using an adoptive transfer model, we demonstrate that inhibition of PI3Kδ profoundly reduces the capacity of bone marrow-derived dendritic cells to sensitize recipient mice for Th2 and Th17 pulmonary inflammation in response to D. farinae Furthermore, administration of a PI3Kδ inhibitor during the sensitization of wild-type mice prevents the generation of D. farinae-induced pulmonary inflammation. These results demonstrate that PI3Kδ regulates Dectin-2 signaling and its dendritic cell function.

Therapeutic and Preventive Effects of Osteoclastogenesis Inhibitory Factor on Osteolysis, Proliferation of Mammary Tumor Cell and Induction of Cancer Stem Cells in the Bone Microenvironment.

BACKGROUND: We examined the effects of recombinant human osteoclastogenesis inhibitory factor (hOCIF) on osteolysis, proliferation of mammary tumor cells, and induction of cancer stem cells (CSCs) in the tumor-bone and tumor-subcutaneous microenvironments (TB- and TS-microE). METHODS: Mouse mammary tumor cells were transplanted onto the calvaria or into a subcutaneous lesion of female mice, creating a TB-microE and a TS-microE, and the mice were then treated with hOCIF. To investigate the preventive effects of hOCIF, mice were treated with hOCIF before tumor cell implantation onto the calvaria (Pre), after (Post), and both before and after (Whole). The number of CSCs and cytokine levels were evaluated by IHC and ELISA assay, respectively. RESULTS: hOCIF suppressed osteolysis, and growth of mammary tumors in the TB-microE, but not in the TS-microE. In the Pre, Post, and Whole groups, hOCIF suppressed osteolysis, and cell proliferation. hOCIF increased mouse osteoprotegrin (mOPG) levels in vivo, which suppressed mammary tumor cell proliferation in vitro. These preventive effects were observed in the dose-dependent. hOCIF did not affect the induction of CSCs in either microenvironment. CONCLUSION: While receptor activator of NF-κB ligand (RANKL) targeting therapy may not affect the induction of CSCs, RANKL is a potential target for prevention as well as treatment of breast cancer bone metastasis.

Extracts of Musa basjoo induce growth inhibition and changes in the protein expression of cell cycle control molecules in human colorectal cancer cell lines.

Musa basjoo (MB) is a species of the banana plant belonging to the genus Musa that has been used as a folk medicine. However, evidence-based biological activities and the molecular mechanism of action of MB are unknown. Thus, the aim of the present study was to examine whether the crude dried leaf extracts of MB inhibit the growth of colorectal (HT29 and HCT116) and other types (HepG2, MCF-7 and PC-3) of human cancer cell lines. Crude extracts of MB inhibited the growth of cells with IC50 values of 136 µg/ml (acetone extract, HT29), 51 µg/ml (acetone extract, HCT116), 45 µg/ml (acetone extract, HepG2), 40 µg/ml (acetone extract, MCF-7), 29 µg/ml (acetone extract, PC-3), 175 µg/ml (methanol extract, HT29), 137 µg/ml (methanol extract, HCT116), 102 µg/ml (methanol extract, HepG2), 85 µg/ml (methanol extract, MCF-7), and 85 µg/ml (methanol extract, PC-3) in colony formation assays, and 126 µg/ml (acetone extract, HT29), 68 µg/ml (acetone extract, HCT116), 260 µg/ml (methanol extract, HT29), and 216 µg/ml (methanol extract, HCT116) in MTT assays. Thin layer chromatography analysis revealed the potential existence of aromatic compounds in the acetone extract of MB. Flow cytometric analysis indicated that the percentage of cells in G1 increased, and this was associated with a concomitant decrease of cells in the S and/or G2-M phases of the cell cycle. When colorectal cancer cells were treated with acetone extract of MB, there was a marked decrease in the levels of expression of the cyclin D1, cyclin E, cdk2 and cdk4 proteins and a marked increase in the levels of the expression of the p21CIP1, p27KIP1, and p53 proteins, but those of apoptosis-associated protein PARP did not change. There was a tendency for acetone extract of MB to inhibit xenograft tumor growth in mice. Collectively, the crude extracts of MB contain active components that exert growth inhibition of human cancer cells. This is the first systematic study of the anticancer activity of MB and may broaden insights into the possible clinical approach of specific herbal medicines.

Multigenic system controlling viral systemic infection determined by the interactions between Cucumber mosaic virus genes and quantitative trait loci of soybean cultivars.

Soybean 'Harosoy' is resistant to Cucumber mosaic virus soybean strain C (CMV-SC) and susceptible to CMV-S strain D (CMV-SD). Using enzyme-linked immunosorbent assay and Northern hybridization, we characterized the Harosoy resistance and found that CMV-SC did not spread systemically but was restricted to the inoculated leaves in Harosoy. Harosoy resistance was not controlled by either a dominant or recessive single gene. To dissect this system controlling long-distance movement of CMV in soybean, we constructed infectious cDNA clones of CMV-SC and CMV-SD. Using these constructs and the chimeric RNAs, we demonstrated that two viral components were required for systemic infection by the virus. The region including the entire 2b gene and the 5' region of RNA3 (mainly the 5' untranslated region) together were required. By quantitative trait locus (QTL) analysis using an F(2) population and the F(3) families derived from Harosoy and susceptible 'Nemashirazu', we also showed that at least three QTLs affected systemic infection of CMV in soybean. Our study on Harosoy resistance to CMV-SC revealed an interesting mechanism, in which multiple host and viral genes coordinately controlled viral systemic infection.

Palmitoyl piperidinopiperidine, a novel derivative of 10­hydroxy­2­decenoic acid, as a potent and selective anticancer agent against human colon carcinoma cell lines.

The present study, to the best of our knowledge, is the first systematic study of the inhibitory effects of palmitoyl piperidinopiperidine (PPI; Japan Patent no. 5597427), on colon carcinogenesis. PPI exhibited marked growth inhibitory activity in several human colon carcinoma cell lines, with IC50 values of approximately 0.5­2.2 µM. In silico docking analysis indicated that PPI could bind to the SH2 domain of signal transducer and activator of transcription 3 (STAT3). PPI markedly inhibited the transcriptional activity of the SW837 cell line. Flowcytometric analysis demonstrated that PPI induced an increase in the number of cells in the G1 phase of the cell cycle, and induced sub­G1 fractions of cells at a higher concentration level of PPI. In the HT29 and SW837 cells, western blot analyses exhibited that in whole cell lysates, PPI induced a marked decrease in the expression levels of p­STAT3, but not in the levels of STAT3 in these cells. PPI also induced a marked decrease in the expression levels of both STAT3 and p­STAT3 in the chromatin fraction. In addition, PPI affected the protein expression levels of cyclin D1, p53, Bcl­2, Bcl­xL and vascular endothelial growth factor (VEGF). In the HT29 cells, PPI induced a marked and dose­dependent increase in the expression levels of Bax, cleaved caspase­3, cleaved caspase­7, cleaved caspase­8, cleaved caspase­9 and cleaved poly (ADP­ribose) polymerase (PARP). In animal model systems, PPI inhibited the growth of implanted carcinoma cells, and also induced a significant decrease in the multiplicity of colonic aberrant crypt foci. In addition, a marked and dose­dependent inhibition of angiogenesis of the chick chorioallantoic membrane was observed. As regards the possible molecular mechanisms, it is suggested that the inhibition of STAT3 by PPI may affect the function of molecules that are related to apoptosis, angiogenesis and cell cycle progression, eventually contributing to the PPI­induced growth inhibitory effects.

Isolation and Quantitative Evaluation of Brush Cells from Mouse Tracheas.

Tracheal brush cells are cholinergic chemosensory epithelial cells poised to transmit signals from the airway lumen to the immune and nervous systems. They are part of a family of chemosensory epithelial cells which include tuft cells in the intestinal mucosa, brush cells in the trachea, and solitary chemosensory and microvillous cells in the nasal mucosa. Chemosensory cells in different epithelial compartments share key intracellular markers and a core transcriptional signature, but also display significant transcriptional heterogeneity, likely reflective of the local tissue environment. Isolation of tracheal brush cells from single cell suspensions is required to define the function of these rare epithelial cells in detail, but their isolation is challenging, potentially due to the close interaction between tracheal brush cells and nerve endings or due to airway-specific composition of tight and adherens junctions. Here, we describe a procedure for isolation of brush cells from mouse tracheal epithelium. The method is based on an initial separation of tracheal epithelium from the submucosa, allowing for a subsequent shorter incubation of the epithelial sheet with papain. This procedure offers a rapid and convenient solution for flow cytometric sorting and functional analysis of viable tracheal brush cells.

Fibrillation mechanism of glucagon in the presence of phospholipid bilayers as revealed by 13C solid-state NMR spectroscopy.

Glucagon is a 29 amino acid peptide hormone secreted by pancreatic α-cells and interacts with specific receptors located in various organs. Glucagon tends to form gel-like fibril aggregates that are cytotoxic because they activate apoptotic signaling pathways. To understand mechanism of fibril formation, we investigated the structure and kinetics of glucagon fibril formation using 13C solid-state NMR spectroscopy. In aqueous acetic acid solution at pH 3.3, distorted α-helical structure appeared around Gly4, Leu14, Ala19 and Leu26 in the monomeric form. In contrast, Gly4 and Ala19 were involved in ß-sheet structures in the fibril form. The fibrillation process can be explained by a two-step autocatalytic reaction mechanism in which the first step is a homogeneous nuclear formation (k1), and the second step is an autocatalytic heterogeneous fibrillation process (k2). The rate constants k1 and k2 were separately determined in the acetic acid solution. Fibril formation was further investigated in the presence of lipid bilayers to mimic the physiological condition. We used bicelles which form discoidal nano-particles as the bilayer system and observed that the N-terminal α-helix did not change to ß-sheet when fibrils formed in the presence of bicelles. Rate constant k1 became faster and k2 became slower in the presence of bicelles compared to the case in the absence of bicelles. Our findings reveal that the structure and kinetics of fibril formation by glucagon are altered in the presence of lipid bilayers.

The cysteinyl leukotriene 3 receptor regulates expansion of IL-25-producing airway brush cells leading to type 2 inflammation.

Respiratory epithelial cells (EpCs) orchestrate airway mucosal inflammation in response to diverse environmental stimuli, but how distinct EpC programs are regulated remains poorly understood. Here, we report that inhalation of aeroallergens leads to expansion of airway brush cells (BrCs), specialized chemosensory EpCs and the dominant epithelial source of interleukin-25 (IL-25). BrC expansion was attenuated in mice lacking either LTC4 synthase, the biosynthetic enzyme required for cysteinyl leukotriene (CysLT) generation, or the EpC receptor for leukotriene E4 (LTE4), CysLT3R. LTE4 inhalation was sufficient to elicit CysLT3R-dependent BrC expansion in the murine airway through an IL-25-dependent but STAT6-independent signaling pathway. Last, blockade of IL-25 attenuated both aeroallergen and LTE4-elicited CysLT3R-dependent type 2 lung inflammation. These results demonstrate that CysLT3R senses the endogenously generated lipid ligand LTE4 and regulates airway BrC number and function.

Innate defense regulator IDR-1018 activates human mast cells through G protein-, phospholipase C-, MAPK- and NF-ĸB-sensitive pathways.

Host defense (antimicrobial) peptides not only display antimicrobial activities against numerous pathogens but also exert a broader spectrum of immune-modulating functions. Innate defense regulators (IDRs) are a class of host defense peptides synthetically developed from natural or endogenous cationic host defense peptides. Of the IDRs developed to date, IDR-1018 is more efficient not only in killing bacteria but also in regulating the various functions of macrophages and neutrophils and accelerating the wound healing process. Because mast cells intimately participate in wound healing and a number of host defense peptides involved in wound healing are also known to activate mast cells, this study aimed to investigate the effects of IDR-1018 on mast cell activation. Here, we showed that IDR-1018 induced the degranulation of LAD2 human mast cells and caused their production of leukotrienes, prostaglandins and various cytokines and chemokines, including granulocyte-macrophage colony-stimulating factor, interleukin-8, monocyte chemoattractant protein-1 and -3, macrophage-inflammatory protein-1α and -1ß, and tumor necrosis factor-α. Furthermore, IDR-1018 increased intracellular calcium mobilization and induced mast cell chemotaxis. The mast cell activation was markedly suppressed by pertussis toxin, U-73122, U0126, SB203580, JNK inhibitor II, and NF-κB activation inhibitor II, suggesting the involvement of G-protein, phospholipase C, ERK, p38, JNK and NF-κB pathways, respectively, in IDR-1018-induced mast cell activation. Notably, we confirmed that IDR-1018 caused the phosphorylation of MAPKs and IκB. Altogether, the current study suggests a novel immunomodulatory role of IDR-1018 through its ability to recruit and activate human mast cells at the sites of inflammation and wounds. HIGHLIGHTS: We report that IDR-1018 stimulates various functions of human mast cells. IDR-1018-induced mast cell activation is mediated through G protein, PLC, MAPK and NF-κB pathways. IDR-1018 will be a useful therapeutic agent for wound healing.

Successful treatment of large adenoma extending close to the papilla in the duodenum by laparoscopy-assisted pancreas-sparing duodenectomy.

A 54-year-old man had a 65-mm infrapapillary, circular, and laterally spreading tubular adenoma in the distal second and proximal third parts of the duodenum. The papilla was 15 mm from the proximal margin of the tumor. Because the patient requested organ-preserving laparoscopic surgery, we conducted laparoscopy-assisted pancreas-sparing duodenectomy (LAPSD). LAPSD consists of five major procedures: (i) laparoscopic wide Kocher maneuver and transection of the proximal jejunum; (ii) laparoscopic separation of the duodenum from the pancreas; (iii) creation of a small upper median laparotomy; (iv) extracorporeal completion of the segmental duodenectomy; and (v) extracorporeal intestinal reconstruction. The postoperative course was uneventful, and the patient was discharged on postoperative day 8. Histopathological examination revealed that the circumferential margin of the specimen was negative for tumor cells. LAPSD provided a clear margin without damaging the papilla and eliminated the possibility of peritoneal or port-site seeding of tumor cells because part of the procedure was performed extracorporeally. LAPSD is a useful alternative to pancreatoduodenectomy in patients with a large adenoma extending close to the papilla in the duodenum.

Laparoscopy-assisted transduodenal excision of superficial non-ampullary duodenal epithelial tumors.

INTRODUCTION: Transduodenal excision (transduodenal submucosal dissection) is an alternative to pancreaticoduodenectomy for the treatment of benign and low-grade malignant tumors of the duodenum. However, laparoscopic transduodenal excision or laparoscopy-assisted transduodenal excision (LATDE) of such tumors has been rarely reported. In this paper, we present the preliminary results of LATDE in patients with superficial non-ampullary duodenal epithelial tumors. METHODS: Three patients with superficial non-ampullary duodenal epithelial tumors (mucosal adenocarcinoma, n = 1; tubular adenoma, n = 2) underwent LATDE. LATDE consists of four major procedures: (i) laparoscopic wide Kocher maneuver (mobilization of the pancreaticoduodenum); (ii) extracorporeal approach to the fully mobilized duodenum through the upper median longitudinal incision (4 cm in length); (iii) tumor excision by submucosal dissection under direct vision through longitudinal duodenotomy (4 cm in length); and (iv) hand-sewn closure of the mucosal defect and duodenotomy. RESULTS: LATDE was successfully carried out without any intraoperative or postoperative adverse events. The mean operating time and estimated blood loss were 155 min and 17 mL, respectively. Contrast roentgenography on postoperative day 4 showed neither duodenal deformity nor disturbance of gastroduodenal emptying in any of the patients. CONCLUSIONS: LATDE could eliminate the possibility of peritoneal or port-site seeding of tumor cells because the duodenotomy and tumor excision are performed extracorporeally. The meticulously hand-sewn closures of the mucosal defect and duodenotomy can minimize the possibility of postoperative hemorrhage and/or anastomotic leakage. LATDE is a feasible, safe, and minimally invasive treatment for patients with superficial non-ampullary duodenal epithelial tumors that have no risk of lymph node metastasis in the first and second portions of the duodenum.

The coat protein gene of tobamovirus P 0 pathotype is a determinant for activation of temperature-insensitive L 1a-gene-mediated resistance in Capsicum plants.

Tobamovirus resistance in Capsicum plants, which is mediated by L genes (L(1), L(2), L(3) or L(4)), is known to be temperature sensitive. However, the L(1a ) gene, a newly identified tobamovirus resistance gene that is mapped to the L locus, confers temperature-insensitive resistance against the tobamovirus P(0) pathotype. To identify the viral elicitor that activates the L(1a )-gene-mediated resistance, several chimeric viral genomes were constructed between tobacco mosaic virus-L (P(0) pathotype), paprika mild mottle virus-J (P(1 )pathotype) and pepper mild mottle virus-J (P(1,2) pathotype). Infection patterns of these chimeric viruses in L(1a )-harboring plants revealed that the L(1a )-gene-mediated resistance was activated by the CP of a particular pathotype of tobamovirus, like other L-gene-mediated resistances, but the L(1a )-gene-mediated resistance differs from those conferred by other L genes in terms of temperature sensitivity.

Genetic mapping of the compatibility between a lily isolate of Cucumber mosaic virus and a satellite RNA.

Five isolates of Cucumber mosaic virus (CMV) from Lilium sp. (lily), which were isolated from specimens in Japan, Korea and Taiwan, were unable to support satellite RNA (satRNA) accumulation. In order to map the CMV sequences that are involved in satRNA support, HL-CMV (Japanese lily isolate), Y-CMV (ordinary strain) and Y-satellite RNA (Y-sat) were used as the source material. The pseudorecombinants between Y-CMV and HL-CMV revealed that RNA1 was essential for satRNA replication in lily. The results of chimeric constructs and various mutations showed that two amino acid residues (at positions 876 and 891) in the 1a protein were the determinants for the inability of HL-CMV to support a satRNA. Specifically, Thr at position 876 had a more pronounced effect than Met at position 891. Specific changes in RNA sequence were also detected in the 3' terminus of Y-sat and these particular alterations allowed it to be supported by HL-CMV. It is believed that, through evolution, the adaptation of CMV to lily resulted in the introduction of amino acid changes in the 1a protein, changes that coincidentally affected the ability of lily CMV to support satRNAs.