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Background Inflammatory bowel disease (IBD) represents a group of chronic immune-mediated diseases of the gastrointestinal tract, characterized by recurrent inflammation and consequential damage of the gastrointestinal tract. Histone deacetylases (HDACs) are a family of mostly ubiquitous enzymes that remove acetyl groups from lysines on histone proteins to regulate gene transcription. HDAC6, which is localized to the cytoplasm, appears to be a promising candidate in IBD treatment. The purpose of the current study was to examine the anti-inflammatory effects of CKD-506, a novel histone deacetylase 6 (HDAC6) inhibitor, on human peripheral blood mononuclear cells (PBMCs) and CD4+ T cells and to explore the relationship between CKD-506 and gut epithelial barrier function. Methods Lipopolysaccharide (LPS)-stimulated human PBMCs from inflammatory bowel disease (IBD) patients were treated with CKD-506, and tumor necrosis factor-alpha (TNF-α) expression was measured using enzyme-linked immunosorbent assay. The proliferation of CD4+ T cells from IBD patients was evaluated using flow cytometric analysis. The effects of CKD-506 on gut barrier function in a cell line and colon organoids, with examinations of mRNA production, goblet cell differentiation, and E-cadherin recovery, were investigated using quantitative reverse transcription PCR, immunofluorescence, and FITC-dextran permeability assay. Results TNF-α secretion, a pivotal pro-inflammatory mediator in IBD, of LPS-triggered PBMCs was markedly decreased by CKD-506 treatment in a dose-dependent manner and to a greater extent than by tofacitinib or tubastatin A treatment. E-cadherin mRNA expression and goblet cell differentiation increased significantly and dose-dependently in HT-29 cells in response to CKD-506, and inhibition of E-cadherin loss after TNF-α stimulation was significantly reduced both in HT-29 cells and gut organoids. Caco-2 cells treated with CKD-506 showed a significant reduction of barrier permeability in a dose-dependent manner. Conclusion The present study demonstrated that CKD-506 has anti-inflammatory effects on PBMCs and CD4+ T cells and improved gut barrier function, indicating its potential as a promising and competent candidate for small-molecule medicine for IBD treatment.

The oriental armyworm, Mythimna separata is an important crop pest in eastern Asia. Nocturnal insects, including nocturnal moths, have phototactic behavior to an artificial light source. Phototactic behavior in insects is species-specific in response to different wavelengths of light sources. Our previous study showed that green (520 nm) light emitting diode (LED) light resulted in a significantly higher phototactic behavior in M. separata moths compared to the other wavelength LED lights. The goal of the present study is to investigate the influence of green light illumination on biological characteristics of different developmental stages in M. separata. Our results revealed that when different developmental stages of M. separata were exposed to the green light illumination in a dark period, several biological characteristics in all developmental stages except for egg stage were positively changed, but those of F1 generation M. separata which are next generation of the adults exposed to the green light did not significantly change compared with the control level. These findings suggest that green light illumination at night (or dark period) has a positive effect on the development and longevity of M. separata.

Background HDAC6 is a stress-inducible gene and highly expressed in pathological conditions as well as inflammatory bowel disease. Immuno-modulatory functions of HDAC6 inhibitors are well established and proposed therapeutic effects for autoimmune diseases through regulation of Treg cell function and inflammation. Moreover, HDAC6 inhibitors regulate inflammatory cytokines and chemokines, neutrophil activities, and epithelial regeneration in colitis models. CKD-506, a potent and selective oral HDAC6 inhibitor, is generally safe and well-tolerated in human, and is now investigating the efficacy in patients with rheumatoid arthritis. Herein, for future investigation with IBD, we identified molecular action mechanisms of CKD-506 involved in anti-colitis effects. Methods Mouse peritoneal macrophages or Raw264.7 cells were transfected with HDAC6 overexpression plasmid or empty vector as control. Cells were cultured in the presence or absence of 0.03~3 μM CKD-506, and the expression and production of inflammatory mediators were determined by RT–PCR and ELISA respectively. For reporter assays, Raw264.7 cells were transfected with pNF-kB-luc or pAP-1-luc plasmid and luciferase activity in cell lysates was determined by a luminometer. Signalling molecules in HDAC6 overexpressed cells were checked by immunoblot analysis. For the efficacy test of CKD-506, we used DSS-, TNBS-, Piroxicam (IL-10−/−)-, and adaptive T-cell transfer (RAG1−/−)-mediated colitis animal models. Colitis animals were treated with 1 to 100 mg/kg of CKD-506 and analysed disease activities and inflammatory mediators. Results In vivo, CKD-506 strongly inhibited disease activities in DSS-, TNBS-, Piroxicam-, and adaptive T-cell transfer-mediated colitis. In the chemical-induced colitis model, the expression of cell adhesion molecules and chemokines such as IP-10 and also infiltration of immune cells to colon tissues were reduced in CKD-506 treated mice. In vitro, HDAC6 overexpression strongly induced ROS and NADPH oxidase activity in Raw264.7 cells and CKD-506 significantly and dose-dependently inhibited HDAC6-mediated ROS and NADPH activity. Moreover, CKD-506 inhibited the production of pro-inflammatory cytokines and chemokines which are up-regulated in HDAC6 overexpressed mouse peritoneal macrophages or Raw264.7 cells. In promoter assay, HDAC6 overexpression highly induced NF-kB and AP-1 activity and CKD-506 strongly and dose-dependently inhibited both signalling pathways. Conclusion These data provide insight that CKD-506, a selective HDAC6 inhibitor, has anti-inflammatory and anti-colitis effects through regulation of NF-kB and AP-1 signalling pathway. Therefore, CKD-506 may provide beneficial effects in patients with Crohn’s disease and ulcerative colitis.

The pattern of leukocyte locomotion can be changed in many pathological situations, but its accurate analysis is difficult because of technological limitation. In the present study, by using a newly developed time-lapse videomicroscopic technique, we have analyzed the locomotive patterns of leukocytes in a stable concentration gradient of chemokines. Granulocytes, monocytes, and lymphocytes were purified from adult human peripheral blood. Locomotive behavior of the leukocytes was analyzed by an optical assay using a microchannel producing a stable concentration gradient of chemokines. Videomicroscopic analysis showed distinct locomotive patterns of granulocytes, monocytes, and lymphocytes. Granulocytes were intrinsically motile, vigorously moving in random direction without any chemokine stimulation. Upon stimulation with CXCL8/IL-8, the speed of migration was increased from 13.3 +/- 2.8 to 19.4 +/- 2.5 microm/min (P = 0.002, n = 100) and they moved toward the chemokine, although many cells still wandered very much. Stimulation with CCL5/RANTES or CXCL12/SDF-1alpha induced similar changes in locomotive pattern. On the other hand, most lymphocytes did not polarize or move spontaneously without chemokine stimulation. Stimulation with CXCL12 induced directional migration in 37% of the lymphocytes at a speed of 9.6 +/- 1.6 microm/min (n = 100). The movement pattern of monocytes was similar to that of granulocytes in that they tend to become polarized and move spontaneously, but they moved at a very slow speed ranging from 3.9 to 4.2 microm/min even with chemokine stimulation. The new optical assay may be useful for many diagnostic as well as basic studies.

Thrombopoietin (TPO) is one of the most promising stimulants for ex vivo expansion of haematopoietic stem cells. Previously, we have found that TPO induces a characteristic pattern of apoptosis during ex vivo expansion of human cord blood (CB) CD34+ cells and that the TPO-induced apoptotic cells belong to megakaryocyte (MK) lineage. In this study, we have examined the maturation of MK and platelet production in association with the TPO-induced apoptosis. CD34+ cells, purified from human CB, were expanded in serum-free conditions stimulated with TPO. Apoptosis was confirmed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labelling (TUNEL) assay and electron microscopy (EM). Simultaneous measurement of DNA content and immunophenotyping revealed that the cells with higher DNA content (>8 N) constituted less than 5% of the CD41+ fractions until day 14, implying premature apoptosis of MKs before full polyploidization. Nevertheless, EM observation showed not only platelet territories but also newly produced platelets in which granules and microfilaments could be identified. Furthermore, flow cytometry demonstrated that the platelet fraction expressed P-selectin and an activation motif on GPIIb/IIIa recognized by monoclonal antibody PAC-1 upon stimulation with adenosine diphosphate (ADP). In addition, periodic acid-Schiff (PAS)-positive materials and nonspecific esterase activities could be demonstrated. Therefore, it is suggested that platelet production and the accompanying processes, rather than apoptosis only, be hastened during the ex vivo expansion of CB CD34+ cells when using TPO.

The coat protein (CP) genes of the genomic RNA of two Korean isolates of sweet potato feathery mottle potyvirus (SPFMV), SPFMV-K1 and SPFMV-K2, were cloned and their complete nucleotide sequences were determined. Sequence comparisons of the two Korean isolates showed 97.8% amino acid identity in the CP cistron, and 79.9% to 99.0% identity with those of 6 other known SPFMV strains. Of 74 amino acid changes totally among the SPFMV strains, 39 changes were located at the N-terminal region. Pairwise amino acid sequence comparison revealed sequence similarities of 48.6 to 70.2% between SPFMV and 20 other potyviruses, indicating SPFMV to be a quite distinct species. Multiple alignment of the CP cistrons from other potyviruses showed that most of the conserved amino acid residues of the genus Potyvirus are well preserved in the corresponding locations.

The complete nucleotide sequence of the genomic RNA of odontoglossum ringspot tobamovirus (ORSV) was determined. The RNA genome of ORSV is 6618 nucleotides long and contains five open reading frames (ORFs 1 to 5) coding for proteins of M(r) 126 K, 181 K, 34 K, 18 K and 52 K, respectively. This is the longest RNA of the known viruses of the Tobamovirus genus. The sequences of the ORSV RNA encoded proteins exhibit high homology to the proteins of the members of the Tobamovirus genus. The genomic organization and sequence analysis showed that ORSV is more closely related to tobacco mild green mosaic virus (TMGMV), pepper mild mottle virus (PMMV), tomato mosaic virus (ToMV) and TMV than to cucumber green mottle mosaic virus (CGMMV) and sunn-hemp mosaic virus (SHMV).

We have completed the genomic sequence of a potyvirus, freesia mosaic virus (FreMV), and compared it to those of other known potyviruses. The full-length genome sequence of FreMV consists of 9,489 nucleotides. The large protein contains 3,077 amino acids, with an AUG start codon and UAA stop codon, containing one open reading frame typical of a potyvirus polyprotein. The polyprotein of FreMV-Kr gives rise to eleven proteins (P1, HC-pro, P3, PIPO, 6K1, CI, 6K2, VPg, NIa, NIb and CP), and putative cleavage sites of each protein were identified by sequence comparison to those of other known potyviruses. Phylogenetic analysis of the polyprotein revealed that FreMV-Kr was most closely related to PeMoV and was related to BtMV, BaRMV and PeLMV, which belong to the BCMV subgroup. This is the first information on the complete genome structure of FreMV, and the sequence information clearly supports the status of FreMV as a member of a distinct species in the genus Potyvirus.

The 3′-terminal 1855 nucleotides (nts) of a Korean isolate of odontoglossum ringspot tobamovirus (ORSV-Cy) were cloned and sequenced. The sequence contained two open reading frames, which encode the cell-to-cell movement protein (MP) and coat protein genes, and are 912 nts and 477 nts long, respectively. The MP gene contained a conserved sequence motif of tobamoviruses and putative assembly origin of the viral RNA locating between 1117 nts and 1292 nts from the 3′-end. The 3′ untranslated region (UTR) of the virus comprises 414 nts, includes nine pseudoknots and a tRNA-like structure domain containing aminoacyl acceptor arm and the anticodon hairpin loop coding for histidine.