Severe fever with thrombocytopenia syndrome virus (SFTSV) is a novel phlebovirus responsible for causing an emerging zoonotic disease. We previously established subclones from SFTSV strain YG1 based on differences in low-pH-dependent cell fusion activities and found two amino acid substitutions, Y328H and R624W, in the envelope glycoprotein (GP) of high fusion subclones. In this study, we show that transiently expressed GP with the R624W mutation, but not the Y328H mutation, induced cell fusion under acidic conditions. GP possessing either tryptophan, serine, glycine or aspartic acid at position 624 induced cell fusion, whereas GP possessing basic amino acids such as arginine or lysine did not induce cell fusion. These results indicated that the amino acid at position 624 has an important role for inducing low-pH-dependent cell fusion.
Pituitary adenylyl cyclase-activating polypeptide (PACAP) is a bioactive peptide with diverse effects in the nervous system. The present study investigated whether stimulation of PACAP receptors (PACAPRs) induces responses in neurons and satellite cells of the superior cervical ganglia (SCG), with special reference to intracellular Ca2+ ([Ca2+]i) changes. The expression of PACAPRs in SCG was detected by reverse transcription-PCR. PACAP type 1 receptor (PAC1R), vasoactive intestinal peptide receptor type (VPAC)1R, and VPAC2R transcripts were expressed in SCG, with PAC1R showing the highest levels. Confocal microscopy analysis revealed that PACAP38 and PACAP27 induced an increase in [Ca2+]i in SCG, first in satellite cells and subsequently in neurons. Neither extracellular Ca2+ removal nor Ca2+ channel blockade affected the PACAP38-induced increase in [Ca2+]i in satellite cells; however, this was partly inhibited in neurons. U73122 or xestospongin C treatment completely and partly abrogated [Ca2+]i changes in satellite cells and in neurons, respectively, whereas VPAC1R and VPAC2R agonists increased [Ca2+]i in satellite cells only. This is the first report demonstrating the expression of PACAPRs specifically, VPAC1 and VPAC2 in SCG and providing evidence for PACAP38-induced [Ca2+]i changes in both satellite cells and neurons via Ca2+ mobilization.
Mucosal-associated invariant T (MAIT) cells are innate-like T cells involved in anti-bacterial immunity. Recent studies have demonstrated that MAIT cells might be implicated in inflammatory bowel diseases (IBDs), but their precise function in IBD remains to be elucidated. We investigated the possible involvement of MAIT cells in the immunopathogenesis of IBDs. Heparinized peripheral blood and biopsy specimens of the colon were collected from 25 patients with ulcerative colitis (UC), 15 patients with Crohn’s disease (CD), and 19 heathy individuals. Lymphocytes were isolated from the blood and colon, and then MAIT cells were analyzed by flow cytometry. The frequency of MAIT cells was significantly lower in the blood of IBD patients compared to healthy donors and significantly higher in the inflamed colons compared to healthy colons (P = 0.001). Among the IBD patients, the frequency of MAIT cells in the blood and colon was correlated with disease activities. In vitro activated MAIT cells from IBD patients secreted significantly more tumor necrosis factor-α and interleukin-17 than those from healthy donors. These findings indicate that MAIT cells are activated in IBD patients, and their accumulation in the inflamed mucosa is correlated with disease activities.
Since osteoblastic activities are believed to be coupled with osteoclasts, we have attempted to histologically verify which of the distinct cellular circumstances, the presence of osteoclasts themselves or bone resorption by osteoclasts, is essential for coupled osteoblastic activity, by examining c-fos−/− or c-src−/− mice. Osteopetrotic c-fos deficient (c-fos−/−) mice have no osteoclasts, while c-src deficient (c-src−/−) mice, another osteopetrotic model, develop dysfunctional osteoclasts due to a lack of ruffled borders. c-fos−/− mice possessed no tartrate-resistant acid phosphatase (TRAPase)-reactive osteoclasts, and showed very weak tissue nonspecific alkaline phosphatase (TNALPase)-reactive mature osteoblasts. In contrast, c-src−/− mice had many TNALPase-positive osteoblasts and TRAPase-reactive osteoclasts. Interestingly, the parallel layers of TRAPase-reactive/osteopontin-positive cement lines were observed in the superficial region of c-src−/− bone matrix. This indicates the possibility that in c-src−/− mice, osteoblasts were activated to deposit new bone matrices on the surfaces that osteoclasts previously passed along, even without bone resorption. Transmission electron microscopy demonstrated cell-to-cell contacts between mature osteoblasts and neighboring ruffled border-less osteoclasts, and osteoid including many mineralized nodules in c-src−/− mice. Thus, it seems likely that osteoblastic activities would be maintained in the presence of osteoclasts, even if they are dysfunctional.
We previously generated induced pluripotent stem (iPS) cells from human dental pulp cells of deciduous teeth. Neural crest cells (NCCs) play a vital role in the development of the oral and maxillofacial region. Therefore, NCCs represent a cell source for bone, cartilage, and tooth-related tissue engineering. In this study, we examined whether iPS cells are capable of differentiating into NCCs through modification of the human embryonic stem cell protocol. First, iPS cells were dissociated into single cells and then reaggregated in low-cell-adhesion plates with neural induction medium for 8 days in suspension culture to form neurospheres. The neurospheres were transferred to fibronectin-coated dishes and formed rosette structures. The migrated cells from the rosettes abundantly expressed NCC markers, as evidenced by real-time polymerase chain reaction, immunofluorescence, and flow cytometric analysis. Furthermore, the migrated cells exhibited the ability to differentiate into neural crest lineage cells in vitro. They also exhibited tissue-forming potential in vivo, differentiating into bone and cartilage. Collectively, the migrated cells had similar characteristics to those of NCCs. These results suggest that human dental pulp cell-derived iPS cells are capable of differentiating into NCCs. Therefore, iPS cell-derived NCCs represent cell sources for bone and cartilage tissue engineering.
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