Ontogeny and Age Distribution of Specific Antigens and Surface Immunoglobulins on the Lymphocyte Membranes of Chick Embryos

鶏の胚の胸腺,BF及び脾臓中のT+リンパ球,B+リンパ球,sIg+リンパ球及びsIgM+リンパ球の比率の加齢に伴う変化を経時的に追求した. 胸腺では,胚齢11日目にT+リンパ球及びB+リンパ球が,胚齢12日目にｓIg+リンパ球及びsIgM+リンパ球が発生した. 一方BFでは,胚齢10日目にT+リンパ球が,胚齢11日目にB+リンパ球,sIg+リンパ球及びsIgM+リンパ球が発生した. 胸腺では,胚齢15日目からT+リンパ球の比率が,一方BFでは胚齢17日目からB+リンパ球の比率が急激に上昇することから,この時点より真の分化が始まると考えられる. BFでは,ｓIg+リンパ球及びsIgM+リンパ球が胚齢11日目に発生してから胚齢16日目まで,ほぼ同じ値で推移していることから,胚齢16日目までBリンパ球のsIgはIgMのみであると考えられる. BFでは,胚齢16日目以降,sIg+リンパ球の比率がsIgM+リンパ球の比率を大幅に上回ることから,胚齢16日目前後に,Bリンパ球のsIgでは,IgMから他のクラスのlgへの転換(スイッチ)が始まると考えられる

Ontogeny and Age Distribution of Specific Antigens and Surface Immunoglobulins on the Lymphocyte Membranes of Chick Embryos

鶏の胚の胸腺,BF及び脾臓中のT+リンパ球,B+リンパ球,sIg+リンパ球及びsIgM+リンパ球の比率の加齢に伴う変化を経時的に追求した. 胸腺では,胚齢11日目にT+リンパ球及びB+リンパ球が,胚齢12日目にｓIg+リンパ球及びsIgM+リンパ球が発生した. 一方BFでは,胚齢10日目にT+リンパ球が,胚齢11日目にB+リンパ球,sIg+リンパ球及びsIgM+リンパ球が発生した. 胸腺では,胚齢15日目からT+リンパ球の比率が,一方BFでは胚齢17日目からB+リンパ球の比率が急激に上昇することから,この時点より真の分化が始まると考えられる. BFでは,ｓIg+リンパ球及びsIgM+リンパ球が胚齢11日目に発生してから胚齢16日目まで,ほぼ同じ値で推移していることから,胚齢16日目までBリンパ球のsIgはIgMのみであると考えられる. BFでは,胚齢16日目以降,sIg+リンパ球の比率がsIgM+リンパ球の比率を大幅に上回ることから,胚齢16日目前後に,Bリンパ球のsIgでは,IgMから他のクラスのlgへの転換(スイッチ)が始まると考えられる

Cholesteryl Pullulan Encapsulated TNF-α Nanoparticles Are an Effective Mucosal Vaccine Adjuvant against Influenza Virus

We encapsulated tumor necrosis factor-α (TNF-α), a major proinflammatory cytokine, into cholesteryl pullulan (CHP) to prepare TNF/CHP nanoparticles. In this report, we describe the immune-enhancing capability of the nanoparticles to act as a vaccine adjuvant. TNF/CHP nanoparticles showed excellent storage stability and enhanced host immune responses to external immunogens. The nanoparticles were effective via the nasal route of administration for inducing systemic IgG1 as well as mucosal IgA. We applied the nanoparticles in a model experimental influenza virus infection to investigate their adjuvant ability. TNF/CHP nanoparticles combined with a conventional split vaccine protected mice via nasal administration against a lethal challenge of A/PR/8/34 (H1N1) influenza virus. Mechanistic studies showed that the nanoparticles enhanced antigen uptake by dendritic cells (DCs) and moderately induced the expression of inflammation-related genes in nasopharynx lymphoid tissue (NALT), leading to the activation of both B and T cells. Preliminary safety study revealed no severe toxicity to TNF/CHP nanoparticles. Slight-to-moderate influences in nasal mucosa were observed only in the repeated administration and they seemed to be reversible. Our data show that TNF/CHP nanoparticles effectively enhance both humoral and cellular immunity and could be a potential adjuvant for vaccines against infectious diseases, especially in the mucosa

NK-4 exerts selective regulatory effects on the activation and function of allergy-related Th2 cells

<div><p>NK-4 is the main component of the antiallergic drug Lumin, which has been in popular usage since the early 1950s. In this study, we examined whether NK-4 exerts a regulatory effect on the activation and effector function of Th2 cells. NK-4 inhibited IL-4 production by anti-CD3ε mAb-stimulated BALB/c mouse spleen cells, whereas NK-4 had little effect on IFN-γ production. IL-4 and IL-5 secretion by anti-CD3ε mAb- or antigen-stimulated Th2 cells (D10.G4.1) was abrogated by NK-4 without affecting cell numbers, whereas IFN-γ secretion by activated Th1 cells was unchanged. Mechanistic analysis revealed that NK-4 inhibited mRNA expression of the Th2-associated transcription factors GATA-3 and NFATc1 in anti-CD3ε mAb-stimulated D10.G4.1 cells. Regarding the regulation of Th2 cell effector functions, NK-4 inhibited the secretion of eotaxin and thymus and activation-regulated chemokine (TARC) by normal human dermal fibroblasts in response to IL-4 and/or TNF-α. NK-4 achieved TARC attenuation comparable to what is observed with suplatast tosilate, an antiallergic drug that selectively inhibits Th2 cytokine production, at 14-fold lower concentrations of suplatast tosilate. Dexamethasone increased TARC production by 2.2- to 2.6-fold of control cultures. NK-4 successfully inhibited the STAT6 signaling pathway, suggesting a potential mechanism for down-regulating chemokines expression. In addition, NK-4 abrogated IL-4-driven modulation of cytokine production profile in human monocytic THP-1 cells from proinflammatory to anti-inflammatory response, as seen in the inverted ratio of TNF-α to IL-10 produced in response to LPS. These results suggest that NK-4 could prevent IL-4-driven polarization to alternatively activated macrophages, which are proposed to have pathogenic roles in allergic asthma. The importance of Th2 cytokines and chemokines in the development and progression of type 2 inflammatory disorders has been highlighted by recent advance in our understanding the immunological mechanism underlying allergic disease. Our results support the use of NK-4 as a reasonable therapeutic option to alleviate Th2-mediated allergic inflammation.</p></div

NK-4 suppresses the STAT6 signaling pathway in NHDF stimulated with IL-4 and TNF-α.

<p>NHDF were grown in confluent monolayer cultures in 12-well plates and were stimulated with 10 ng/ml IL-4 and 5 ng/ml TNF-α in the presence or absence of varying concentrations of NK-4 for 15 min. Phosphorylation of STAT6 was determined by immunoblotting whole-cell lysates using specific antibodies against the phosphorylated or total STAT6 protein. A representative blot is shown (A). The optical density ratio of phospho-STAT6 to total STAT6 is shown (B). Data from three independent experiments were combined and expressed as the means ± SD. **<i>p</i> < 0.01 compared with control cultures. Original uncropped and unadjusted Western blots were provided as supplementary files (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0199666#pone.0199666.s001" target="_blank">S1 Fig</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0199666#pone.0199666.s002" target="_blank">S2 Fig</a>).</p

NK-4 down-regulates secretion of TARC by NHDF stimulated with both IL-4 and TNF-α.

<p>NHDF were stimulated with 10 ng/ml IL-4 and 5 ng/ml TNF-α in the presence or absence (control) of varying concentrations of NK-4, dexamethasone or FK-506 (A) for 48 h at 37°C. In experiments comparing NK-4 with suplatast tosilate (B), NHDF were exposed to test articles for 15 min before stimulation with IL-4 and TNF-α. TARC levels in the cultures of NHDF without stimulation were below detectable limits. Results are the means ± S.D. of triplicate cultures. Results are representative of three independent experiments with similar results. **<i>p</i> < 0.01 compared with control cultures.</p

Chemical structure of NK-4.

<p>Chemical structure of NK-4.</p

NK-4 selectively down-regulates Th2 cytokine production by anti-CD3ε mAb-stimulated established Th2 cells.

<p>Th1 clone #4 (A and B) and Th2 clone D10.G4.1 cells (C to E) (2.5 x 10⁴ cells/well) were stimulated with immobilized anti-CD3ε mAb (8 μg/ml) in the presence or absence (control) of varying concentrations of NK-4 for 48 h at 37°C in 96-well plates. Concentrations of IFN-γ (A), IL-4 (C) and IL-5 (D) in culture supernatants were measured by ELISA. Cell numbers of #4 (B) and D10.G4.1 (E) were determined by cell counting kit-8. Results are the means ± S.D. of triplicate cultures. Results are representative of three independent experiments with similar results. *<i>p</i> < 0.05, **<i>p</i> < 0.01 compared with control cultures.</p

Primers used for quantitative real-time PCR analysis.

<p>Primers used for quantitative real-time PCR analysis.</p