TNFa Protein Levels Research Articles

Effect of PM2.5 exposure on gestational hypertension, fetal size in preeclampsia-like rats.

Studies have shown intriguing associations between gestational PM2.5 exposure and preeclampsia (PE), as well as fetal growth restriction (FGR). This study investigated the impact of PM2.5 exposure on gestational hypertension and fetal outcome in a preeclampsia-like rat model. Pregnant Sprague Dawley rats were exposed to either filtered (FA) or PM2.5-contaminated air during the whole pregnancy period. A PE-like rat model was established by intraperitoneal injection of L-NAME (300mg/kg) from gestational day (GD) 12 to until GD20. Systolic blood pressure (SBP), weight gain, pup weight and placental weight were measured. The percentages of rat Treg/Th17 cells and Th17-related cytokines were examined by flow cytometry. Gene expression profiles were analyzed by microarray, and the expression of differentially expressed genes was validated by qRT-PCR. The results showed that maternal PM2.5 exposure had no effect on SBP but was associated with low birth weight (LBW) and a higher labyrinth/basal zone ratio. The percentages of splenic Th17 cells from the PM2.5 group of PE-like rats were higher than those from the FA or PM2.5 groups of healthy controls. A significantly decreased Treg/Th17 cell ratio was found in the PM2.5 group of PE-like rats. The mRNA expression of Foxp3 was downregulated, while the mRNA expression of RORα and RORγτ was upregulated after PM2.5 exposure. Furthermore, we observed that both the mRNA and protein levels of TNF-a, CCL2, CCL3 and CCR1 increased in the PM2.5 groups. Our study suggested that systemic inflammation may contribute to the development of FGR associated with PM2.5 exposure throughout pregnancy.

Evidence for Necroptosis as a Mechanism of Islet Amyloid–Induced Beta-Cell Death

Islet amyloid deposition occurs in the majority of individuals with type 2 diabetes (T2D) and contributes to beta-cell death by mechanisms which are not fully understood. Necroptosis is an immunogenic form of cell death dependent on tumor necrosis factor receptor 1 (TNFR1) signaling, receptor interacting protein kinase 3 (RIPK3) oligomerization, and mixed lineage kinase domain-like (MLKL) phosphorylation. We previously showed mRNA and protein levels of TNFa, a TNFR1 ligand, are increased in islets by amyloid deposition in vitro and in vivo, raising the possibility that amyloid formation induces necroptosis. Thus, we sought to determine whether the components of necroptosis are present in beta cells, and if islet amyloid-induced TNFa expression elicits beta-cell necroptosis. We observed protein expression of TNFR1, RIPK3, and MLKL in the INS-1 beta-cell line, indicating necroptosis components are present in beta cells. In human islet amyloid polypeptide (hIAPP) transgenic mouse islets cultured in 16.7 mM glucose, an in vitro model of islet amyloidosis, we found necroptosis signaling to be increased compared to amyloid-free wild type (WT) islets. This manifested as increased oligomerization of RIPK3 protein (trimers: 2.7 ± 0.2 [hIAPP] vs. 1.0 [WT], n=3, p<0.01) and phosphorylation of the RIPK3 target MLKL (1.8 ± 0.3 vs. 1.0, n=3, p=0.06), without a change in TNFR1 protein expression (1.1 ± 0.1 vs. 1.0, n=2, p=0.74). As further support that amyloid induces immunogenic cell death, immune cell mRNA abundance was increased in vivo in amyloid containing hIAPP transgenic islets vs. WT islets (macrophages: Emr1, 1.6 ± 0.2 [hIAPP] vs. 1.1 ± 0.1 [WT], n=4, p=0.04, T cells: Cd3, 2.7 ± 0.5 vs. 1.0 ± 0.2, n=4, p=0.03, and B cells: Cd20, 2.3 ± 0.2 vs. 1.2 ± 0.5, n=2, p=0.17). These data suggest that necroptosis may be a previously unrecognized mechanism of beta-cell death in response to islet amyloid formation. Further studies are needed to determine whether inhibition of necroptosis signaling may reduce beta-cell death in T2D. Disclosure A.T. Templin: None. M.F. Hogan: None. N. Esser: None. S. Zraika: None. R.L. Hull: Research Support; Self; Boehringer Ingelheim Pharmaceuticals, Inc., Eli Lilly and Company. S.E. Kahn: Advisory Panel; Self; Boehringer Ingelheim GmbH, Elcelyx Therapeutics, Inc., Eli Lilly and Company, Intarcia Therapeutics, Inc., Janssen Research & Development, Merck & Co., Inc., Novo Nordisk A/S.

In vitro Testing of Experimental and Commercial Bracket Bonding Materials

The aims of our study were to assess the cytotoxic effect of five orthodontic bonding materials, in vitro, on HUVECs to study the induction of apoptosis and inflammatory response generated to assess the shear-bond strength of the 5 tested materials in order to quantify their performance. Standardized samples from each material were obtained and incubated with HUVEC cells for 24 and 72 h immersed in complete medium. Cell viability was determined by means of MTS method. Active caspase 3 and TNFa protein levels were measured through ELISA techniques. The shear-bond strength was tested on 60 extracted premolars which were bonded with the same type of bracket, using the 5 different materials. Statistical analysis Student T-tests, Chi-square and Anova tests were used for results interpretation. Cell viability was decreased with material exposure in a time dependent manner. All materials exerted cytotoxic effects, the experimental materials showed a significantly higher decrease in cell viability at the 72 h reading. Shear Bond strength was superior for the resin commercial bracket-bonding materials. The study shows that orthodontic adhesives� cytotoxicity and physical performance is related to their chemical properties and proves that all orthodontic practitioners should use freely their material of choice on condition they are aware of all its� properties.

414 Identification of sebocytes as a novel source for adipokines within human skin

412 Characterization of the immunomodulatory effects of sebocytes on macrophages M Lovaszi, M Mattii, K Eyerich, D Kovacs, C Zouboulis, M Stahle, S Eyerich and D Torocsik 1 Department of Dermatology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary, 2 ZAUM e Center for Allergy and Environment, Technische Universitat and Helmholtz Center Munich, Munich, Germany, 3 Department of Dermatology and Allergy, Technische Universitat Munich, Munich, Germany, 4 Departments of Dermatology, Venereology, Allergology and Immunology, Dessau Medical Center, Dessau, Germany and 5 Unit of Dermatology and Venereology, Department of Medicine, Karolinska lnstitutet, Karolinska University Hospital, Stockholm, Sweden Macrophages surround the sebaceous glands both in the healthy and the inflamed pilosebaceous unit; however, the reason for this phenomenon and the possible role of sebocytes in it has not yet been investigated. By performing immunohistochemistry on healthy biopsies, we could demonstrate that macrophages in the close proximity of sebaceous glands are exclusively alternatively activated (CD163/FXIII-A/CD206/CD209). To investigate if sebocytes contribute to the differentiation, polarization and function of macrophages, human peripheral blood monocytes were differentiated and activated in the presence of either supernatant from the human SZ95 sebocyte cell line, or major sebum lipid components such as oleic-, linoleic-, palmitic-, stearicacids and squalene. Our results showed that with the secretion of CXCL8, sebocytes could exert a chemoattractant effect towards monocytes, while sebocytes derived lipids promoted monocyte differentiation into alternatively activated macrophages as marked by the up-regulation of CD206, CD209 and FXIII-A. Moreover, detection of the produced IL-1b, IL-6 and TNFa protein levels by Propionibacterium acnes activated macrophages revealed a selective inflammatory effect for the various sebum lipids. Our results altogether suggest a role for sebaceous glands in initiating and modulating innate immune responses via their proteins and lipids that are of possible pathologic and therapeutic relevance. 413 Epithelial resident and infiltrating dendritic cells amplify active and resolved psoriasis inflammation E Martini, M Wiken, S Cheuk, A Smed Sorensen, M Stahle and L Eidsmo Dept. of Medicine, Solna, Karolinska Institutet, Stockholm, Sweden In psoriasis, scaly skin lesions are caused by interactions between activated keratinocytes, T cells and dendritic cells present in the skin. Here we investigate the inflammatory profiles of skin resident Langerhans cells (LCs) and infiltrating epitdermal dendritic cells (eDCs) to determine if they have the potential to steer epithelial pathology in psoriasis. A population of LangerinCD11cCD1c eDCs with variable levels of CD1a and CD14 was strictly confined to epidermis in active psoriasis lesions and could not be detected in non lesional or resolved psoriasis or healthy skin. LCs and eDCs sorted from active psoriasis displayed shared expression of genes associated with T cell activation and neutrophil attraction. Although LCs from psoriatic lesions stimulated with Toll-like receptor (TLR) ligands produced the pro-inflammatory cytokines IL-1 beta and IL-23, eDCs were the major producer of these cytokines in the epidermis, and displayed comparable cytokine production to dDCs. In resolved psoriasis eDCs are absent but LCs promptly responded to TLR stimulation with increased IL-23 production. Our results show that infiltrating eDCs together with LCs strongly contribute to the pro-inflammatory environment within the epidermal compartment of the skin. In contrast, LCs alone show the capacity to activate pathogenic tissue resident T cells in resolved psoriasis.

412 Characterization of the immunomodulatory effects of sebocytes on macrophages

412 Characterization of the immunomodulatory effects of sebocytes on macrophages M Lovaszi, M Mattii, K Eyerich, D Kovacs, C Zouboulis, M Stahle, S Eyerich and D Torocsik 1 Department of Dermatology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary, 2 ZAUM e Center for Allergy and Environment, Technische Universitat and Helmholtz Center Munich, Munich, Germany, 3 Department of Dermatology and Allergy, Technische Universitat Munich, Munich, Germany, 4 Departments of Dermatology, Venereology, Allergology and Immunology, Dessau Medical Center, Dessau, Germany and 5 Unit of Dermatology and Venereology, Department of Medicine, Karolinska lnstitutet, Karolinska University Hospital, Stockholm, Sweden Macrophages surround the sebaceous glands both in the healthy and the inflamed pilosebaceous unit; however, the reason for this phenomenon and the possible role of sebocytes in it has not yet been investigated. By performing immunohistochemistry on healthy biopsies, we could demonstrate that macrophages in the close proximity of sebaceous glands are exclusively alternatively activated (CD163/FXIII-A/CD206/CD209). To investigate if sebocytes contribute to the differentiation, polarization and function of macrophages, human peripheral blood monocytes were differentiated and activated in the presence of either supernatant from the human SZ95 sebocyte cell line, or major sebum lipid components such as oleic-, linoleic-, palmitic-, stearicacids and squalene. Our results showed that with the secretion of CXCL8, sebocytes could exert a chemoattractant effect towards monocytes, while sebocytes derived lipids promoted monocyte differentiation into alternatively activated macrophages as marked by the up-regulation of CD206, CD209 and FXIII-A. Moreover, detection of the produced IL-1b, IL-6 and TNFa protein levels by Propionibacterium acnes activated macrophages revealed a selective inflammatory effect for the various sebum lipids. Our results altogether suggest a role for sebaceous glands in initiating and modulating innate immune responses via their proteins and lipids that are of possible pathologic and therapeutic relevance. 413 Epithelial resident and infiltrating dendritic cells amplify active and resolved psoriasis inflammation E Martini, M Wiken, S Cheuk, A Smed Sorensen, M Stahle and L Eidsmo Dept. of Medicine, Solna, Karolinska Institutet, Stockholm, Sweden In psoriasis, scaly skin lesions are caused by interactions between activated keratinocytes, T cells and dendritic cells present in the skin. Here we investigate the inflammatory profiles of skin resident Langerhans cells (LCs) and infiltrating epitdermal dendritic cells (eDCs) to determine if they have the potential to steer epithelial pathology in psoriasis. A population of LangerinCD11cCD1c eDCs with variable levels of CD1a and CD14 was strictly confined to epidermis in active psoriasis lesions and could not be detected in non lesional or resolved psoriasis or healthy skin. LCs and eDCs sorted from active psoriasis displayed shared expression of genes associated with T cell activation and neutrophil attraction. Although LCs from psoriatic lesions stimulated with Toll-like receptor (TLR) ligands produced the pro-inflammatory cytokines IL-1 beta and IL-23, eDCs were the major producer of these cytokines in the epidermis, and displayed comparable cytokine production to dDCs. In resolved psoriasis eDCs are absent but LCs promptly responded to TLR stimulation with increased IL-23 production. Our results show that infiltrating eDCs together with LCs strongly contribute to the pro-inflammatory environment within the epidermal compartment of the skin. In contrast, LCs alone show the capacity to activate pathogenic tissue resident T cells in resolved psoriasis.

Lung Injuries Induced by Gastric Acid Aspiration Are Attenuated by Exogenous Surfactant during Ex Vivo Reconditioning in Pigs

Purpose Gastric acid aspiration is a common reason for rejecting lungs for transplantation as it is associated with increased risks of graft dysfunction. We investigated Ex vivo lung perfusion (EVLP) reconditioning on lung injuries due to gastric acid aspiration and effects of surfactant administration. Methods and Materials 30 piglets were allocated to 5 groups. Aspiration lung injury (LI) was induced by infusing 1 ml/kg of autologous gastric juice into the left lower lobe (LLL). 24 hours later, the lungs were studied (LI group) or reconditioned using EVLP for 4 hours without additional treatment (LI-EVLP group) or after surfactant lavage (curosurf 100 mg/kg) in the LLL (Surf-EVLP group). Sham animals were studied 24 h after saline infusion and Sham-EVLP animals 24 h after saline infusion followed by 4 h of EVLP. Gross anatomy, hemodynamics, and aerodynamics were evaluated; neutrophil and bacterial counts in bronchoalveolar lavage (BAL) fluid and blood were determined. LLLs were evaluated by tissue wet/dry ratio; histology (semi-quantitative severity score); apoptotic cell using TUNEL; and IL-1, IL-6, IL-8, IL-10, and TNFa protein levels. Results There were no significant differences between the Sham and Sham-EVLP groups. Compared to Sham, LI animals had irreversible atelectasis, a higher lung infection rate (P 2 (P=0.0006), higher IL1 (P=0.022) and IL8 (P=0.006) levels and apoptotic cell percentage (P=0.007), and worse histology severity score (P Conclusions Although 4 hours EVLP worsened LI due to gastric acid aspiration, when preceded by local infusion of surfactant it allowed complete PaO2 and partial histology recovery.

Green tea extract protects against early alcohol-induced liver injury in rats.

Oxidants have been shown to be involved in alcohol-induced liver injury. This study was designed to test the hypothesis that the antioxidant polyphenolic extract of green tea, comprised predominantly of epigallocatechin gallate, protects against early alcohol-induced liver injury in rats. Male Wistar rats were fed high-fat liquid diets with or without ethanol (10-14 g kg(-1) day(-1)) and green tea (300 mg kg(-1) day(-1)) continuously for 4 weeks using an intragastric enteral feeding protocol. Mean body weight gains (approximately 4 g/day) were not significantly different between treatment groups, and green tea extract did not the affect average concentration or the cycling of urine ethanol concentrations (0-550 mg dl(-1) day(-1)). After 4 weeks, serum ALT levels were increased significantly about 4-fold over control values (35+/-3 IU/l) by enteral ethanol (114+/-18); inclusion of green tea extract in the diet significantly blunted this increase (65+/-10). Enteral ethanol also caused severe fatty accumulation, mild inflammation, and necrosis in the liver. While not affecting fat accumulation or inflammation, green tea extract significantly blunted increases in necrosis caused by ethanol. Furthermore, ethanol significantly increased the accumulation of protein adducts of 4-hydroxynonenal, a product of lipid peroxidation and an index of oxidative stress; green tea extract blocked this effect almost completely. TNFalpha protein levels were increased in liver by alcohol; this phenomenon was also blunted by green tea extract. These results indicate that simple dietary antioxidants, such as those found in green tea, prevent early alcohol-induced liver injury, most likely by preventing oxidative stress.