Two Distinct Subsets Are Identified from the Peritoneal Myeloid Mononuclear Cells Expressing both CD11c and CD115

To this date, the criteria to distinguish peritoneal macrophages and dendritic cells (DCs) are not clear. Here we delineate the subsets of myeloid mononuclear cells in the mouse peritoneal cavity. Considering phenotypical, functional, and ontogenic features, peritoneal myeloid mononuclear cells are divided into 5 subsets: large peritoneal macrophages (LPMs), small peritoneal macrophages (SPMs), DCs, and 2 MHCII⁺CD11c⁺CD115⁺ subpopulations (i.e., MHCII⁺CD11c⁺CD115⁺CD14⁻CD206⁻ and MHCII⁺CD11c⁺CD115⁺CD14⁺CD206⁺). Among them, 2 subsets of competent Ag presenting cells are demonstrated with distinct functional characteristics, one being DCs and the other being MHCII⁺CD11c⁺CD115⁺CD14⁻CD206⁻ cells. DCs are able to promote fully activated T cells and superior in expanding cytokine producing inflammatory T cells, whereas MHCII⁺CD11c⁺CD115⁺CD14⁻CD206⁻ cells generate partially activated T cells and possess a greater ability to induce Treg under TGF-β and retinoic acid conditions. While the development of DCs and MHCII⁺CD11c⁺CD115⁺CD14⁻CD206⁻ cells are responsive to the treatment of FLT3 ligand and GM-CSF, the number of LPMs, SPMs, and MHCII⁺CD11c⁺CD115⁺CD14⁺CD206⁺ cells are only influenced by the injection of GM-CSF. In addition, the analysis of gene expression profiles among MHCII⁺ peritoneal myeloid mononuclear cells reveals that MHCII⁺CD11c⁺CD115⁺CD14⁺CD206⁺ cells share high similarity with SPMs, whereas MHCII⁺CD11c⁺CD115⁺CD14⁻CD206⁻ cells are related to peritoneal DC2s. Collectively, our study identifies 2 distinct subpopulations of MHCII⁺CD11c⁺CD115⁺ cells, 1) MHCII⁺CD11c⁺CD115⁺CD14⁻CD206⁻ cells closely related to peritoneal DC2s and 2) MHCII⁺CD11c⁺CD115⁺CD14⁺CD206⁺ cells to SPMs.

Extended Culture of Bone Marrow with Granulocyte Macrophage-Colony Stimulating Factor Generates Immunosuppressive Cells

Bone marrow-derived dendritic cells (BM-DCs) are generated from bone marrow (BM) cells cultured with granulocyte macrophage-colony stimulating factor (GM-CSF) for a week. In this study we investigated the effect of duration on the BM culture with GM-CSF. Within several months, the cells in the BM culture gradually expressed homogeneous levels of CD11c and major histocompatibility complex II on surface, and they became unable to stimulate allogeneic naïve T cells in mixed lymphocyte reaction (MLR). In addition, when the BM culture were sustained for 32 wk or longer, the BM cells acquired ability to suppress the proliferation of allogeneic T cells in MLR as well as the response of ovalbumin-specific OT-I transgenic T cells in antigen-dependent manner. We found that, except for programmed death-ligand 1, most cell surface molecules were expressed lower in the BM cells cultured with GM-CSF for the extended duration. These results indicate that BM cells in the extended culture with GM-CSF undergo 2 distinct steps of functional change; first, they lose the immunostimulatory capacity; and next, they gain the immunosuppressive ability.

TCF4-Targeting miR-124 is Differentially Expressed amongst Dendritic Cell Subsets

Dendritic cells (DCs) are professional antigen-presenting cells that sample their environment and present antigens to naive T lymphocytes for the subsequent antigen-specific immune responses. DCs exist in a range of distinct subpopulations including plasmacytoid DCs (pDCs) and classical DCs (cDCs), with the latter consisting of the cDC1 and cDC2 lineages. Although the roles of DC-specific transcription factors across the DC subsets have become understood, the posttranscriptional mechanisms that regulate DC development are yet to be elucidated. MicroRNAs (miRNAs) are pivotal posttranscriptional regulators of gene expression in a myriad of biological processes, but their contribution to the immune system is just beginning to surface. In this study, our in-house probe collection was screened to identify miRNAs possibly involved in DC development and function by targeting the transcripts of relevant mouse transcription factors. Examination of DC subsets from the culture of mouse bone marrow with Flt3 ligand identified high expression of miR-124 which was able to target the transcript of TCF4, a transcription factor critical for the development and homeostasis of pDCs. Further expression profiling of mouse DC subsets isolated from in vitro culture as well as via ex vivo purification demonstrated that miR-124 was outstandingly expressed in CD24+ cDC1 cells compared to in pDCs and CD172alpha+ cDC2 cells. These results imply that miR-124 is likely involved in the processes of DC subset development by posttranscriptional regulation of a transcription factor(s).

Modified Method of Multilocus Sequence Typing (MLST) for Serotyping in Salmonella Species

Multilocus sequence typing (MLST) of Salmonella is useful method for replacing serotyping using antisera but is limited by difficulties associated with in polymerase chain reaction (PCR). We optimized the PCR reaction, especially annealing temperature and extension time (94degrees C for 2 min; 40 cycles at 94degrees C for 30 sec, 56.8degrees C for 1 min, 72degrees C for 2 min; and 72degrees C for 10 min). The degradation of PCR product by thermostable nucleases was inhibited by using template DNAs treated proteinase K or purified by a commercialized preparation kit. The resulting modified MLST was used as accurate and fast typing method.

Modified Mandibulotomy Technique to Reduce Postoperative Complications: 5-Year Results

PURPOSE: To review the 5-year outcomes of our modified mandibulotomy technique. Retrospective review of a tertiary level oral cancer center. MATERIALS AND METHODS: During a 5-year period, 30 patients who had a uniform surgical technique consisting of a lower lip-splitting, modified stair-step osteotomy with thin saw blade and osteotome after plate-precontouring and combination fixation with monocortical osteosynthesis (miniplate) and bicortical osteosynthesis (maxiplate and bicortical screws), with at least 14 months postoperative follow-up, were selected and reviewed retrospectively. RESULTS: There were 8 women and 22 men with an average age of 56.5 years. All the patients involved malignancies were squamous cell carcinoma. The main primary sites of the those who underwent a mandibulotomy were the tonsil, the base of tongue, the oral tongue, the retromolar pad area, and others. Others included buccal cheek, floor of mouth, and soft palate. 23 patients received postoperative radiation therapy, and among whom 8 patients also received chemotherapy. Total four (13%) mandibulotomy-related complications occurred, only two (6.7%) requiring additional operation under general anesthesia. CONCLUSION: Our modified mandibulotomy meets the criteria for an ideal mandibulotomy technique relatively well because it requires no intermaxillary fixation, can precise preserve the occlusion in a precise way, allows early function, requires no secondary procedures, and has few complications.

Contradictory Effects of Superoxide and Hydrogen Peroxide on KCa3.1 in Human Endothelial Cells

Reactive oxygen species (ROS) are generated in various cells, including vascular smooth muscle and endothelial cells, and regulate ion channel functions. KCa3.1 plays an important role in endothelial functions. However, the effects of superoxide and hydrogen peroxide radicals on the expression of this ion channel in the endothelium remain unclear. In this study, we examined the effects of ROS donors on KCa3.1 expression and the K+ current in primary cultured human umbilical vein endothelial cells (HUVECs). The hydrogen peroxide donor, tert-butyl hydroperoxide (TBHP), upregulated KCa3.1 expression, while the superoxide donors, xanthine/xanthine oxidase mixture (X/XO) and lysopho-sphatidylcholine (LPC), downregulated its expression, in a concentration-dependent manner. These ROS donor effects were prevented by antioxidants or superoxide dismustase. Phosphorylated extracellular signal-regulated kinase (pERK) was upregulated by TBHP and downregulated by X/XO. In addition, repressor element-1-silencing transcription factor (REST) was downregulated by TBHP, and upregulated by X/XO. Furthermore, KCa3.1 current, which was activated by clamping cells with 1 microM Ca2+ and applying the KCa3.1 activator 1-ethyl-2-benzimidazolinone, was further augmented by TBHP, and inhibited by X/XO. These effects were prevented by antioxidants. The results suggest that hydrogen peroxide increases KCa3.1 expression by upregulating pERK and downregulating REST, and augments the K+ current. On the other hand, superoxide reduces KCa3.1 expression by downregulating pERK and upregulating REST, and inhibits the K+ current. ROS thereby play a key role in both physiological and pathological processes in endothelial cells by regulating KCa3.1 and endothelial function.

Idiopathic Localized Lipodystrophy of Subcutaneous Fat after Cyst Enucleation of Jaw: Report of 3 Cases

Keratoameloblastoma of the maxilla: a case report

A keratoameloblastoma is a histologically variant of the ameloblastoma group, which varies in size and contains keratin material in the fibrous connective tissue among cystic lesions. A keratoameloblastoma is a rare disease with only 13 cases reported in the literature since Pindborg's first report in 1970. A 41-year-old man visited, complaining of pus discharged from the right maxilla. He had been diagnosed with an odontogenic keratocyst and was treated with cyst enucleation in the past. The clinical and radiology examination found evidence of recurrence and finally diagnosed him with keratoameloblastoma after enucleation and biopsy. This report discusses the clinical, radiological and histological characteristics of keratoameloblastoma and its treatment. In addition, we report another case of keratoameloblastoma that had transformed from an odontogenic keratocyst.

Detection of antibodies against glucose 6-phosphate isomerase in synovial fluid of rheumatoid arthritis using surface plasmon resonance (BIAcore)

We have used a surface plasmon resonance biosensor (SPR, BIACORE 2000) to detect antibodies against glucose 6-phosphate isomerase (GPI) in synovial fluids of rheumatoid arthritis (RA) and osteoarthritis (OA). Recombinant human GPI proteins fused with or without NusA were expressed in E. coli, purified to homogeneity and immobilized in flow cells of CM5 sensor chips. The flow cells immobilized with NusA protein or bovine serum albumin were used to monitor non-specific binding. Synovial fluid samples from RA patients showed a significantly higher level of binding to recombinant GPI proteins than samples from OA patients. Proteins which bound to the recombinant GPI proteins were confirmed to be immunoglobulin through the administration of anti-human immunoglobulin. NusA fusion protein was excellent for this assay because of a low background binding activity in the SPR analysis and its advantage of increased solubility in recombinant protein production. These results suggested a useful utilization of recombinant NusA-GPI fusion protein for the detection of autoantibodies against GPI in RA patients.