SummaryDendritic cells (DCs) are professional antigen-presenting cells capable of initiating primary/adaptive immune responses and tolerance. DC functions are regulated by their state of maturation. However, the molecular pathways leading to DC development and maturation remain poorly understood. We attempted to determine whether inhibition of nuclear factor kappa B (NFkB), which is one of the pivotal pathways underlying these processes, could induce immunophenotypic and functional changes in lipopolysaccharideinduced mature DCs derived from murine bone marrow. A comparative in vitro study of five clinically used drugs that are known to inhibit NF-kB demonstrated that azithromycin, a macrolide antibiotic, significantly inhibited expression of co-stimulatory molecules (CD40 and CD86) and major histocompatibility complex (MHC) class II by DCs. It also reduced Toll-like receptor 4 expression, interleukin-12 production and the allostimulatory capacity of DCs. These data suggest that azithromycin, as not only an NF-kB inhibitor but also an antibiotic, has potential as a novel drug for manipulation of allogeneic responses.

The immunologic effects of developmental exposure to noninherited maternal Ags (NIMAs) are quite variable. Both tolerizing influence and inducing alloreaction have been observed on clinical transplantation. The role of minor histocompatibility Ags (MiHAs) in NIMA effects is unknown. MiHA is either matched or mismatched in NIMA-mismatched transplantation because a donor of the transplantation is usually limited to a family member. To exclude the participation of MiHA in a NIMA effect for MHC (H-2) is clinically relevant because mismatched MiHA may induce severe alloreaction. The aim of this study is to understand the mechanism of NIMA effects in MHC-mismatched, MiHA-matched hematopoietic stem cell transplantation. Although all offsprings are exposed to the maternal Ags, the NIMA effect for the H-2 Ag was not evident. However, they exhibit two distinct reactivities, low and high responder, to NIMA in utero and during nursing depending on the degree of maternal microchimerism. Low responders survived longer with less graft-versus-host disease. These reactivities were correlated with Foxp3 expression of peripheral blood CD4+CD25+ cells after graft-versus-host disease induction and the number of IFN-γ–producing cells stimulated with NIMA pretransplantation. These observations are clinically relevant and suggest that it is possible to predict the immunological tolerance to NIMA.

Objective This study assessed maternal cytomegalovirus antibodies, and the occurrence of primary and congenital cytomegalovirus infections, and risk factors of congenital infection after a maternal primary infection. Study design We included 19,435 pregnant women in Japan, who were tested for serum cytomegalovirus antibodies before 20 gestational weeks. Immunoglobulin (Ig) G avidity was evaluated in women with both IgG and IgM antibodies; tests were repeated at ≥28 gestational weeks among women without IgG and IgM antibodies. Result Primary and congenital infections were 162 and 23 cases, respectively. The risk ratios for congenital infection were 8.18 (95% confidence interval: 2.44–27.40) in teenage versus older women, and 2.25 (95% confidence interval: 1.28–3.94) in parity ≥ 2 versus parity ≤ 1. Of 22 live birth congenital infection cases, three had abnormal neurological findings. Conclusion We demonstrated teenage and parity ≥ 2 pregnant women as risk factors of post-primary congenital infection.

Natural killer (NK) cells acquire effector function through a licensing process and exert anti-leukemia/tumor effect. However, there is no means to promote a licensing effect of allogeneic NK cells other than cytomegalovirus reactivation-induced licensing in allogeneic hematopoietic stem cell transplantation in human. In mice, a licensing process is mediated by Ly49 receptors which recognize self-major histocompatibility complex class I. The distribution of four Ly49 receptors showed similar pattern in congenic mice, B10, B10.BR, and B10.D2, which have B10 background. Forty Gy-irradiated 2 × 106 B10.D2 cells including splenocytes, peripheral blood mononuclear cells in untreated mice, or granulocyte colony-stimulating factor treated mice were injected intraperitoneally into B10 mice. We found that murine NK cells were effectively licensed by intraperitoneal injection of donor neutrophils with its corresponding NK receptor ligand in B10 mice as a recipient and B10.D2 as a donor. Mechanistic studies revealed that NK cells showed the upregulation of intracellular interferon-γ and CD107a expression as markers of NK cell activation. Moreover, enriched neutrophils enhanced licensing effect of NK cells; meanwhile, licensing effect was diminished by depletion of neutrophils. Collectively, injection of neutrophils induced NK cell licensing (activation) via NK receptor ligand interaction.

SummaryAcute graft-versus-host disease (GVHD) following allogeneic bone marrow transplantation (BMT) is initiated by donor T lymphocytes that recognize histocompatibility antigens presented by recipient dendritic cells (DCs). Current approaches to reduce GVHD are focused on suppressing donor T lymphocyte responses to alloantigens. However, these strategies may be inadequate in the setting of allogeneic transplants (particularly histoincompatible transplants), may increase the risk of tumour relapse and are associated with high rates of opportunistic infections. We hypothesized that inhibition of recipient DCs might suppress GVHD. We recently demonstrated in vitro that azithromycin, a macrolide antibiotic, also acts as a nuclear factor (NF)-kB inhibitor of murine DCs and inhibits their maturation and functions, including allogeneic responses. We investigated whether azithromycin could prevent alloreactions in a murine histoincompatibility model. Oral administration of azithromycin to recipient mice for 5 days during majorhistoincompatible BMT suppressed lethal GVHD significantly, whereas ex-vivo lymphocyte function was not affected by the drug. These data suggest that azithromycin has potential as a novel prophylactic drug for lethal GVHD.

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ABSTRACTcytokine SFC in patients following HSCT and correlated the results with clinical findings. Furthermore, expression of CD29 (b1 integrin) on monocytes was correlated with plasma levels of fibronectin in patients with active chronic GVHD. Design and Methods PatientsFifty-seven patients who underwent allogeneic HSCT between 1995 and 2009 were included in this study. The patients' demographic data are shown in Table 1. Sixteen patients with chronic GVHD were grouped into mild (n=7), moderate (n=8), and severe (n=1) categories based on the NIH chronic GVHD consensus criteria. 5 According to Sarantopoulos's definition of chronic GVHD activity, patients were subclassified into those with no, active, and inactive chronic GVHD. 28 Patients who never developed chronic GVHD (n=41) were designated as "no chronic GVHD". Patients with active chronic GVHD (n=14) were more likely to be receiving immunosuppressive therapy. Inactive chronic GVHD (n=10) was determined by clinical assessment and included patients who had achieved a complete response to immunosuppressive therapy at the time of analysis. The number of patients with active and inactive chronic GVHD overlapped during the clinical course. Disease activity was assessed without knowledge of the laboratory results. The median time of the analysis was 48 (range, 4-204), 52 (5-142), and 55 (4-151) months after transplantation for patients with no, active, and inactive chronic GVHD, respectively.This study was conducted according to the principles expressed in the Declaration of Helsinki and approved by the Institutional Ethics Committee Review Board at Mie University Hospital. The study was registered with the national regulatory authority (UMIN-Clinical Trials Registry). All participants or their guardians provided written informed consent for the collection of samples and subsequent analyses. Diagnosis of chronic graft-versus-host disease and processing of samplesThe diagnosis of chronic GVHD requires the presence of at least one diagnostic manifestation of the disease or at least one distinctive manifestation, with the diagnosis confirmed by pertinent biopsy, laboratory tests, or radiology in the same or another organ.5,29 Diagnostic manifestations of chronic GVHD were found in the skin, nails, mouth, eyes, lungs, gastrointestinal tract, and liver. The grade of chronic GVHD was determined according to NIH consensus criteria. 5,30 Peripheral blood mononuclear cells (PBMC) were obtained from patients with no evidence of infection, and the diagnoses were confirmed by laboratory testing or radiology, at least 100 days after allogeneic HSCT. The total numbers of white blood cells, lymphocytes, and monocytes were analyzed by an automatic blood cell counter (Sysmex K4500, Toa Medical Electronics, Tokyo, Japan). Flow cytometry and cell separationCells were stained with fluorescein-conjugated monoclonal antibodies to human anti-CD3, CD4, CD8, CD11c, CD16, CD19, CD25, CD29, CD33, CD56, CD123 (BD Biosciences, San Jose, CA, USA), CD14 (Beckman Coulter, CA, USA), CD68, interle...

Targeting protein for Xenopus kinesin-like protein 2 (TPX2) is upregulated in various tumors, and several studies have demonstrated the role of TPX2 as a prognostic marker in cancer. However, the function of TPX2 in neuroblastoma (NB) has not been completely elucidated. In the present study, the clinical significance and functional role of TPX2 in NB was investigated. The Therapeutically Applicable Research to Generate Effective Treatments (TARGET)-NB dataset was used. A total of 43 patients with NB were enrolled in the present study as the validation set. After evaluating the prognostic role of TPX2, the combined predictive effect of TPX2 and MYCN proto-oncogene bHLH transcription factor (MYCN) gene amplification was assessed. Double immunofluorescence staining for TPX2 and N-Myc was used to analyze colocalization, and multiple cell function tests were performed by means of in vitro experiments to elucidate the functional role of TPX2 using RNA interference technology in NB cell lines. In both the TARGET-NB set and the validation set, it was found that upregulated of TPX2 was significantly associated with poor overall survival (OS) in patients with NB. The expression of TPX2 was higher in NB patients with MYCN gene amplification, and NB patients with high TPX2 expression and MYCN gene amplification had the poorest OS compared with patients with low TPX2 expression or a single copy of MYCN.In vitro experiments indicated that TPX2 positively regulated cell proliferation and the cell cycle, and promoted cell survival by increasing the resistance to apoptosis. The colocalization of TPX2 with N-Myc in NB cells and tissue was observed. The findings of the present study indicate that TPX2 plays an oncogenic role in NB development and may be a potential prognostic indicator in patients with NB.