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Background Premature infants often require oxygen supplementation and, therefore, are exposed to oxidative stress. Following oxygen exposure, preterm infants frequently develop chronic lung disease and have a significantly increased risk of asthma. Objective We sought to identify the underlying mechanisms by which neonatal hyperoxia promotes asthma development. Methods Mice were exposed to neonatal hyperoxia followed by a period of room air recovery. A group of mice was also intranasally exposed to house dust mite antigen. Assessments were performed at various time points for evaluation of airway hyperresponsiveness, eosinophilia, mucus production, inflammatory gene expression, and T H and group 2 innate lymphoid cell (ILC2) responses. Sera from term- and preterm-born infants were also collected and levels of IL-33 and type 2 cytokines were measured. Results Neonatal hyperoxia induced asthma-like features including airway hyperresponsiveness, mucus hyperplasia, airway eosinophilia, and type 2 pulmonary inflammation. In addition, neonatal hyperoxia promoted allergic T H responses to house dust mite exposure. Elevated IL-33 levels and ILC2 responses were observed in the lungs most likely due to oxidative stress caused by neonatal hyperoxia. IL-33 receptor signaling and ILC2s were vital for the induction of asthma-like features following neonatal hyperoxia. Serum IL-33 levels correlated significantly with serum levels of IL-5 and IL-13 but not IL-4 in preterm infants. Conclusions These data demonstrate that an axis involving IL-33 and ILC2s is important for the development of asthma-like features following neonatal hyperoxia and suggest therapeutic potential for targeting IL-33, ILC2s, and oxidative stress to prevent and/or treat asthma development related to prematurity.

Summary Tissue macrophages self-renew during homeostasis and produce inflammatory mediators upon microbial infection. We examined the relationship between proliferative and inflammatory properties of tissue macrophages by defining the impact of the Wnt/β-catenin pathway, a central regulator of self-renewal, in alveolar macrophages (AMs). Activation of β-catenin by Wnt ligand inhibited AM proliferation and stemness, but promoted inflammatory activity. In a murine influenza viral pneumonia model, β-catenin-mediated AM inflammatory activity promoted acute host morbidity; in contrast, AM proliferation enabled repopulation of reparative AMs and tissue recovery following viral clearance. Mechanistically, Wnt treatment promoted β-catenin-HIF-1α interaction and glycolysis-dependent inflammation while suppressing mitochondrial metabolism and thereby, AM proliferation. Differential HIF-1α activities distinguished proliferative and inflammatory AMs in vivo. This β-catenin-HIF-1α axis was conserved in human AMs and enhanced HIF-1α expression associated with macrophage inflammation in COVID-19 patients. Thus, inflammatory and reparative activities of lung macrophages are regulated by β-catenin-HIF-1α signaling, with implications for the treatment of severe respiratory diseases.

The accuracy of a bioassay based on smartphone-integrated fluorescent biosensors has been limited due to the occurrence of false signals from non-specific reactions as well as a high background and low signal-to-noise ratios for complementary metal oxide semiconductor image sensors. To overcome this problem, we demonstrate dual-wavelength fluorescent detection of biomolecules with high accuracy. Fluorescent intensity can be quantified using dual wavelengths simultaneously, where one decreases and the other increases, as the target analytes bind to the split capture and detection aptamer probes. To do this, we performed smartphone imaging-based fluorescence microscopy using a microarray platform on a substrate with metal-enhanced fluorescence (MEF) using Ag film and Al2O3 nano-spacer. The results showed that the sensitivity and specificity of the dual-wavelength fluorescent quantitative assay for the target biomolecule 17-β-estradiol in water were significantly increased through the elimination of false signals. The detection limit was 1pg/mL and the area under the receiver operating characteristic curve of the proposed assay (0.922) was comparable to that of an enzyme-linked immunosorbent assay (0.956) from statistical accuracy tests using spiked wastewater samples. This novel method has great potential as an accurate point-of-care testing technology based on mobile platforms for clinical diagnostics and environmental monitoring.

Probe-mediated fluorescence biosensing methods based on spectrophotometry still have limitations such as detection inaccuracy caused by the occurrence of false signals and lack of simultaneous qualitative and quantitative read-outs with an ultra-low detection limit. Herein, we describe a novel seesawed fluorescence detection strategy based on dual-colour imaging-based quantitation in which the green fluorescence of the capture aptamer decreases and the red fluorescence of the detection aptamer increases simultaneously upon their respective interactions with the target biomolecule. This approach enhances detection accuracy through facilitating identification of probable false-positives in biological samples. Furthermore, combining the seesawed detection scheme with three-dimensional imaging of fluorescence signal enhanced by highly vertical ZnO nanorods increases signal-to-noise ratio, which addresses the limited performance of digital cameras and, in turn, enhances sensitivity and dynamic range. This simple, robust, scalable, imaging-based and label-free fluorescence method allows highly specific and sensitive quantification of biomolecules with excellent reliability.

Background: Panax ginseng (i.e., ginseng) root is extensively used in traditional oriental medicine. It is a modern pharmaceutical reagent for preventing various human diseases such as cancer. Ginsenosides—the major active components of ginseng—exhibit immunomodulatory effects. However, the mechanism and function underlying such effects are not fully elucidated, especially in human monocytes and dendritic cells (DCs). Methods: We investigated the immunomodulatory effect of ginsenosides from Panax ginseng root on CD14+ monocytes purified from human adult peripheral blood mononuclear cells (PBMCs) and on their differentiation into DCs that affect CD4+ T cell activity. Results: After treatment with ginsenoside fractions, monocyte levels of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-10 increased through phosphorylation of extracellular signal-regulated kinase (ERK)1/2 and c-Jun N-terminal kinase (JNK), but not p38 mitogen-activated protein kinase (MAPK). After treatment with ginsenoside fractions, TNF-α production and phosphorylation of ERK1/2 and JNK decreased in lipopolysaccharide (LPS)-sensitized monocytes. We confirmed that DCs derived from CD14+ monocytes in the presence of ginsenoside fractions (Gin-DCs) contained decreased levels of the costimulatory molecules CD80 and CD86. The expression of these costimulatory molecules decreased in LPS-treated DCs exposed to ginsenoside fractions, compared to their expression in LPS-treated DCs in the absence of ginsenoside fractions. Furthermore, LPS-treated Gin-DCs could not induce proliferation and interferon gamma (IFN-γ) production by CD4+ T cells with the coculture of Gin-DCs with CD4+ T cells. Conclusion: These results suggest that ginsenoside fractions from the ginseng root suppress cytokine production and maturation of LPS-treated DCs and downregulate CD4+ T cells.

Cell-mediated and acute vascular rejections remain to be one of the primary hurdles to achieve successful xenotransplantation. Fas ligand is known to be an important molecule for the formation of 'immune-privileged' condition and dendritic cells treated with dexamethasone (Dex-DCs) acting like tolerogenic DCs (tDCs) which are known to protect transplanted cells and organs from unwanted immune responses. The present study investigated the possibility that porcine fibroblasts expressing human Fas ligand (PhF) together with human Dex-DCs could induce prolonged survival of porcine fibroblasts in vitro. PhF was collected from an ear of human Fas ligand transgenic porcine and cell-line was established by MGEM Inc. PhF labeled with CFSE co-cultured with human peripheral blood mononuclear cells (hPBMCs) were examined with respect to induction of tolerance and cell death when co-cultured with Dex-DCs for 3days. PhF induced the apoptosis in hPBMCs, especially CD4(+) T cells. Dex-DCs showed significant (P

Lipoteichoic acid (LTA) is a major virulence factor of Gram-positive bacteria including Staphylococcus aureus. Despite its pivotal role in causing sepsis, the systemic immune responses to LTA in human cells are poorly understood. Here, we produced highly-pure and structurally-intact LTA from S. aureus and examined the gene expression profile of LTA-stimulated human peripheral blood mononuclear cells (PBMCs). The LTA preparation did not contain any detectable biologically-active impurities and stimulated Toll-like receptor 2. Protein expression profiling using a cytokine array kit and ELISA revealed expression of MCP-1/CCL2, IL-6, and IL-1β. We performed transcriptional profiling of PBMCs in response to S. aureus LTA using an Affymetrix genechip microarray. A total of 208 genes were significantly (fold change>1.5 and P

To understand the interaction of dendritic cells (DCs) with cancer cells, we investigated molecular changes in DCs following co-culture with cancer cells. DCs co-cultured with Jurkat cancer cells showed remarkable down-regulation of MHC class I molecules, while DCs co-cultured with MCF-7 cancer cells showed minimal changes. Interestingly, down-regulation of MHC class I on DCs was not observed upon treatment with Jurkat cell lysate or culture supernatant, suggesting the importance of direct cell-cell interactions. The expressions of CD40, CD80, CD83, MHC class II, and IL-12p40 on DCs co-cultured with Jurkat cells were only slightly affected. In contrast, DCs co-cultured with MCF-7 cells showed increased expressions of CD80, CD83, CD86, and IL-12p40. Furthermore, DCs co-cultured with Jurkat cells showed a down-regulation of low molecular weight polypeptides (LMP) 7, and of transporter associated with antigen processing (TAP) 1 and 2 at the mRNA expression level. LMP7, TAP2 and β2-microglobulin (β2M) were also down-regulated at the protein level. We further demonstrated how altered expression of MHC class I on DCs caused by co-culture with cancer cells affected autologous CD8(+) T cells, using the model MHC class I-presented HSV antigen. We found that DCs that had been HSV-treated and co-cultured with Jurkat cells showed a reduced potency to activate CD8(+) T cells. In contrast, HSV-treated DCs that had been co-cultured with MCF-7 cells induced activation of CD8(+) T cells, including high expression of CD25, CD69, granzyme B and cytokines, TNF-α and IFN-γ.

In the present study, we examined the effects of ginsenoside Re (Re) on cytokine expression, cytokine-dependent autophagy and cell survival in human CD4(+) T cells. When CD4(+) T cells isolated from human peripheral blood were treated with Re, LC3 and monodansylcadaverine (MDC), representative markers of autophagy, were decreased in a dose-dependent manner. Interestingly, Re suppressed the production of interferon-gamma (IFN-gamma) and immunity-related GTPase family M (IRGM) in CD4(+) T cells whereas no changes in other autophagy-related signaling molecules (ERK, p38 and AKT-mTOR-p70S6k) were found. Concomitantly, we observed that Re increased the proliferation of CD4(+) T cells with decreased cell death. Our results demonstrate that ginsenoside Re enhanced viability of CD4(+) T cells through the regulation of IFN-gamma-dependent autophagy activity.

Resistin is an adipokine whose physiologic role in obesity, type II diabetes mellitus, and inflammatory diseases has been a subject of debate because while it is expressed in adipocytes and adipose tissue in mouse, it is expressed in leukocytes, such as macrophages, in human. In the present study, we attempt to define the effect of resistin on human dendritic cells (DCs) derived from CD14 + monocytes. When DCs were stimulated with lipoteichoic acid (LTA) and treated with various concentrations of resistin, antigen-uptake process and the endocytic capacity of DCs were decreased. It is intriguing that resistin attenuated cytokine production in LTA-primed DCs. Consequently, T cell activity was reduced when lymphocytes were mixed with Staphylococcus aureus -primed autologous DCs treated with resistin compared to S. aureus -primed DCs without resistin. Our results suggest that resistin interferes with the efficacy of immune responses activated by Gram-positive bacterial infection in human DCs.