Sustained activation of NLRP3 inflammasome and release of neutrophil extracellular traps (NETs) impair wound healing of diabetic foot ulcers (DFUs). Our previous study reported that milk fat globule epidermal growth factor VIII (MFG-E8) attenuates tissue damage in systemic lupus erythematosus. However, the functional effect of MFG-E8 on "NLRP3 inflammasome-NETs" inflammatory loop in wound healing of diabetes is not completely elucidated. In this study, neutrophils from DFU patients are susceptible to undergo NETosis, releasing more NETs. The circulating levels of NET components neutrophil elastase and proteinase 3 and inflammatory cytokines IL-1β and IL-18 were significantly elevated in DFU patients compared with healthy controls or diabetic patients, in spite of higher levels of MFG-E8 in DFU patients. In Mfge8-/- diabetic mice, skin wound displayed exaggerated inflammatory response, including leukocyte infiltration, excessive activation of NLRP3 inflammasome (release of higher IL-1β, IL-18, and TNF-α), largely lodged NETs, resulting in poor angiogenesis and wound closure. When stimulated with high-dose glucose or IL-18, MFG-E8-deficient neutrophils release more NETs than WT neutrophils. After administration of recombinant MFG-E8, IL-18-primed NETosis of WT or Mfge8-/- neutrophils was significantly inhibited. Furthermore, NET and mCRAMP (component of NETs, the murine equivalent of cathelicidin LL-37 in human)-mediated activation of NLRP3 inflammasome and production of IL-1β/IL-18 were significantly elevated in Mfge8-/- macrophages compared with WT macrophages, which were also significantly dampened by the administration of rmMFG-E8. Therefore, our study demonstrated that as inhibitor of the "NLRP3 inflammasome-NETs" inflammatory loop, exogenous rMFG-E8 improves angiogenesis and accelerates wound healing, highlighting possible therapeutic potential for DFUs.
Recently, autoimmune mechanisms and pulmonary epithelial cells have attracted attention in chronic obstructive pulmonary disease (COPD). Circulating antibodies against human bronchial epithelial cells (anti-HBEC) bind to bronchial epithelial antigens and induce bronchial epithelial cell damage. This study aimed to detect the expression of IgG, IgM, and IgA anti-HBEC in patients with COPD.The association of gender, age, body mass index (BMI), and pulmonary function with the presence of IgG, IgA, and IgM anti-HBEC in the plasma was determined in 170 patients with COPD and 150 age-matched healthy controls. Circulating IgG, IgA, and IgM anti-HBEC were detected by indirect immunofluorescence (IIF).Positive IgG anti-HBEC was seen in 34/170 (20.00%) COPD and 11/150 (7.33%) healthy controls (p < 0.001) (1:100 dilution); positive IgA anti-HBEC were presented in 50/170 (29.41%) COPD and 13/150 (8.67%) healthy controls (p < 0.0001) (1:40 dilution); 19/170 (11.19%) COPD and 10/150 (6.67%) healthy controls exhibited positive IgM anti-HBEC (p > 0.05) (1:40 dilution). The positive IgG and IgA anti-HBEC COPD patients were mostly classified as GOLD (Global Initiative for Chronic Obstructive Lung Disease) III and GOLD IV. The positive IgA anti-HBEC COPD patients had lower BMI than healthy controls (p < 0.05).Our results suggest that an autoimmune component associated with bronchial epithelial cell damage is possibly involved in COPD and the presence of IgG and IgA anti-HBEC correlated with the GOLD stage of COPD. Therefore, our studies indicate that IgG and IgA anti-HBEC may associate with the disease severity of COPD.
Rationale: The effects of hookah smoke on respiratory epithelia have not been well characterized.Objectives: To characterize and compare the effects of hookah tobacco smoke and conventional cigarette smoke on the epigenome and transcriptome of human respiratory epithelia.Methods: Normal human small airway epithelial cells and cyclin-dependent kinase 4/human telomerase reverse transcriptase–immortalized human bronchial epithelial cells were cultured for 5 days in normal media in the presence or absence of water pipe condensates or cigarette smoke condensates under relevant exposure conditions. CyQUANT assay (Thermo Fisher Scientific), RNA sequencing, quantitative reverse transcriptase–polymerase chain reaction, and Western blotting techniques were used to examine/compare the effects of hookah and cigarette smoke on cell proliferation, messenger RNA/microRNA expression, and global histone marks.Results: Water pipe condensates– and cigarette smoke condensates–mediated, dose-dependent growth-inhibitory effects in cultured respiratory epithelial cells were measured. Under conditions mediating equipotent mild to moderate growth-inhibitory effects, water pipe condensates as well as cigarette smoke condensates decreased histone H4 lysine 15 acetylation (H4K16ac) and histone H4 lysine 20 trimethylation (H4K20me3) levels in small airway epithelial cells and human bronchial epithelial cells; these histone alterations are putative “hallmarks of cancer.” RNA sequencing analysis demonstrated that cigarette smoke condensates and water pipe condensates mediated dose-dependent alterations in gene expression in small airway epithelial cells and human bronchial epithelial cells. A total of 873 genes were commonly regulated (fold change >2) by water pipe condensates (1.0 mg/ml) and cigarette smoke condensates (0.05 mg/ml) in small airway epithelial cells, whereas a total of 1,577 genes were commonly regulated by water pipe condensates (1.0 mg/ml) and cigarette smoke condensates (0.05 mg/ml) in human bronchial epithelial cells. Ingenuity Pathway Analysis (QIAGEN Bioinformatics) of the 100 genes commonly regulated by water pipe condensates and cigarette smoke condensates in small airway epithelial cells and human bronchial epithelial cells demonstrated that the top three activated upstream regulators are cigarette smoke (z-score = 2.577), benzo[a]pyrene (z-score = 2.905), and nuclear factor (erythroid-derived 2)-like 2 (NFE2L2, Nrf2) (z-score = 2.531). Analysis of downstream effects indicated that growth of epithelial tissue was predicted to be increased (z-score = 2.813; P < 0.001). The top three canonical pathways included xenobiotic metabolism signaling, aryl hydrocarbon receptor signaling, and nicotine degradation III.Conclusions: These preliminary findings strongly suggest that hookah tobacco is not a safe alternative to cigarettes, and warrant further evaluation of the epigenomic effects of hookah smoke in human respiratory epithelia using our established laboratory models and correlative analyses in hookah smokers.
Smoking is considered to be the main source of indoor pollution, and it has been identified as an important environmental factor contributing to asthma onset. We know that T helper 2 (Th2) response plays a crucial role in the process of asthma disease. We have investigated the reaction of cigarette smoke extract (CSE) on Th polarization which is controlled by dendritic cells (DCs). Stimulated by CSE, immature DCs from murine bone marrow showed upregulated levels of TIM4. Cocultured with CD4+ T cells, stimulated DCs increased the ratio of IL-4+ versus IFN-γ+ of CD4+ T cells. This suggests a differentiation towards Th2 response. Moreover, antibodies against TIM4 reversed the upexpression of the IL-4+/IFN-γ+ ratio provoked by CSE, indicating that the Th2 polarization which was induced by CSE is via TIM4 mechanisms. CSE could activate mitogen-activated protein kinase pathways like ERK and p38. Upregulation of TIM4 expression by CSE stimulation was found to be inhibited by an ERK inhibitor but not p38 and JNK. In conclusion, DC-induced Th2 polarization is a hallmark of CSE allergy, and this aspect can be explained by CSE-induced TIM4 expression.
Very thin heat spreaders made of materials with high in-plane thermal conductivity are desirable for thermal solutions to eliminate hot spots on packages of highly converged handheld electronic devices with shrinking form factors. Graphite is used commercially in these applications because of its unique anisotropic properties of high in-plane thermal conductivity and low thru-thickness thermal conductivity. This paper presents application-level thermal performance tests on graphite based spreaders attached to a plastic substrate. Two graphite materials, provided by GrafTech International Holdings Inc., with different thicknesses and in-plane thermal conductivities, were tested. Two types of spreader constructions were also compared. The thermal performance tests emulate the typical heat dissipation scenario for handheld device packages; a constant and uniform heat flux on a very limited area on one side of the spreader and a natural convection environment on the other side. The thermal resistance of the spreader assembly is examined as a function of the different materials and constructions. Additionally, samples were subjected to three types of accelerated life tests (ALT). The thermal performance of these samples before and after ALT were compared to determine the effects of the ALT sequences. Statistical procedures including Analysis of variance and the paired student t test were used to analyze the data.
Abnormal features of the systemic lupus erythematosus (SLE)-derived neutrophils, promoted aberrant immune response, have inspired new studies of the induction of autoimmunity and the development of organ damage in SLE. In this study, we explore the effect of milk fat globule-EGF factor 8 (MFG-E8) on the aberrant nitrification features in pristane-induced lupus. SLE patients and mice with pristane-induced lupus develop autoantibodies associated with MFG-E8 overproduction. However, the deletion of MFG-E8 leads to uncontrolled early pulmonary and peritoneal inflammation and tissue damage in mice with pristane-induced lupus. Consistent with these findings, MFG-E8-deficient mice that are exposed to pristane show enhanced neutrophil accumulation and increased neutrophil death, including apoptosis, necrosis and NETosis, as well as impaired phagocytosis of macrophages. The consequences are the expansion of diffuse pulmonary hemorrhage, increased anti-nuclear antibody, anti-dsDNA antibody and anti-neutrophil cytoplasmic antibody levels, and enhanced immune complexes deposition and neutrophil extracellular traps (NETs) formation in the lung and kidney tissues of MFG-E8-deficient mice exposed to pristane. In patients with SLE and mice with pristane-induced lupus, neutrophil accumulation is elevated, which depends on higher expression of the surface receptor CXCR2. After pretreatment with recombinant MFG-E8, the surface expression of CXCR2 on neutrophil is downregulated, and the MFG-E8 deletion increase CXCR2 expression by ~40%. These studies indicate that MFG-E8 reduces neutrophil migration and NETosis via downregulating surface CXCR2 expression in parallel with its role in the phagocytosis of apoptotic neutrophils, suggesting that MFG-E8 may serve as a therapeutic agent for attenuating the early inflammatory responses of SLE and protect patients from lupus-related damage.
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