Soluble mediators in the function of the epidermal-immune-neuro unit in the skin

Skin is the largest, environmentally exposed (barrier) organ, capable of integrating various signals into effective defensive responses. The functional significance of interactions among the epidermis and the immune and nervous systems in regulating and maintaining skin barrier function is only now becoming recognized in relation to skin pathophysiology. This review focuses on newly described pathways that involve soluble mediator-mediated crosstalk between these compartments. Dysregulation of these connections can lead to chronic inflammatory diseases and/or pathologic conditions associated with chronic pain or itch

Alum Activates the Bovine NLRP3 Inflammasome

peer-reviewedThere has been a move away from vaccines composed of whole or inactivated antigens toward subunit-based vaccines, which although safe, provide less immunological protection. As a result, the use of adjuvants to enhance and direct adaptive immune responses has become the focus of much targeted bovine vaccine research. However, the mechanisms by which adjuvants work to enhance immunological protection in many cases remains unclear, although this knowledge is critical to the rational design of effective next generation vaccines. This study aimed to investigate the mechanisms by which alum, a commonly used adjuvant in bovine vaccines, enhances IL-1β secretion in bovine peripheral blood mononuclear cells (PBMCs). Unlike the case with human PBMCs, alum promoted IL-1β secretion in a subset of bovine PBMCs without priming with a toll-like receptor agonist. This suggests that PBMCs from some cattle are primed to produce this potent inflammatory cytokine and western blotting confirmed the presence of preexisting pro-IL-1β in PBMCs from a subset of 8-month-old cattle. To address the mechanism underlying alum-induced IL-1β secretion, specific inhibitors identified that alum mediates lysosomal disruption which subsequently activates the assembly of an NLRP3, ASC, caspase-1, and potentially caspase-8 containing complex. These components form an inflammasome, which mediates alum-induced IL-1β secretion in bovine PBMCs. Given the demonstrated role of the NLRP3 inflammasome in regulating adaptive immunity in murine systems, these results will inform further targeted research into the potential of inflammasome activation for rational vaccine design in cattle

Regulation of innate and adaptive immunity by the vaccine adjuvant alum

THESIS 10273While many licensed vaccines consist of whole or inactivated pathogens, there is a move toward vaccines based on purified antigens which although safer are generally less immunogenic and therefore require adjuvants to trigger protective immunity. Alum, the most common adjuvant, has a record of successful use in vaccines, where an antibody-mediated immune response can confer protective immunity. However, alum is a poor inducer of cellular immune responses. The mechanism underlying this selective enhancement of humoral responses is still not well understood. Here, to gain an improved insight into its mode of action, innate immune responses to alum and their impact on adaptive immune responses were studied

Effects of the combination of BH3 mimetic and MEK inhibitor on acute myeloid leukemia cells

Poszukiwanie skutecznej terapii przeciwnowotworowej to jeden z największych problemów, z którymi zmaga się współczesna medycyna. Wiedza na temat szlaków odpowiedzialnych za powstawanie i proliferację komórek nowotworowych oraz czynników prowadzących do zaburzenia prawidłowego funkcjonowania komórki, pozwala na rozwój terapii celowanej. Obecnie stosuje się wiele związków, zarówno syntetycznych, jak i pochodzenia naturalnego, wpływających na szlaki przekazywania sygnału i białka odpowiedzialne za utrzymanie prawidłowego funkcjonowania komórki, w celu zwiększenia skuteczności oraz indywidualizacji terapii przeciwnowotworowej.Celem niniejszych badań było określenie wpływu mimetyku BH3, obatoklaksu oraz inhibitora kinazy MEK, PD98059, na komórki ostrej białaczki promielocytowej HL-60. Komórki eksponowano na działanie obatoklaksu w stężeniu 0.1 µM i 0.5 µM, PD98059 w stężeniach 25 µM i 50 µM oraz kombinacji tych związków. Analizy wykonywano w dwóch punktach czasowych, 24- i 48 godzin od podania związków. Stosując techniki spektrofotometryczne, cytometryczne,mikroskopowe oraz metodę Coultera dokonano oceny żywotności, liczby i objętości komórek, przebiegu cyklu komórkowego wraz z analizą populacji sub-G1 oraz wpływu testowanych związków na indukowanie apoptozy i nekrozy komórek białaczkowych. Wykazano, że zastosowanie obatoklaksu w skojarzeniu z PD98059 zwiększa aktywność cytotoksyczną związków i efektywność indukowania programowanej śmierci w komórkach HL-60. Wyniki przedstawione w pracy wskazują na synergistyczne działanie przeciwbiałaczkowe obatoklaksu i PD98059. Uzyskane wyniki skłaniają do prowadzenia dalszych badań nad zastosowaniem mimetyków BH3 i inhibitorów MEK w terapii ostrej białaczki szpikowej.Nowadays, the search for effective anti-cancer therapy is one of the biggest problems modern medicine is facing. Knowledge about the pathways responsible for the formation and proliferation of cancer cells and factors leading to impaired cellular function, allows the development of targeted therapy. Currently, many compounds of both synthetic and natural origin are used that affect the signaling pathways and proteins responsible for maintaining normal cell function, in order to increase the effectiveness and individualization of anti-cancer therapy. The purpose of this study was to determine the effect of BH3 mimetic, obatoclax and MEK inhibitor, PD98059, on acute promyelocytic leukemia HL-60 cells. Cells were exposed to obatoclax at concentrations of 0.1 μM and 0.5 μM, PD98059 at concentrations of 25 μM and 50 μM, and combinations of these compounds. The analyzes were performed at two time points, 24- and 48 hours after treatment with the tested compounds. Using the spectrophotometric, cytometric, microscopic and Coulter methods, the cell viability, number and volume, the cell cycle distribution with sub-G1 population analysis and the effect of tested compounds on the induction of apoptosis and necrosis of leukemia cells, were performed. It has been shown that the use of obatoclax in combination with PD98059 increases the cytotoxic activity of compounds and their effectiveness in inducing programmed cell death in HL-60 cells. The results presented in this study indicate the synergistic anti-leukemic activity of the obatoclax and PD98059. The obtained results encourage further research using BH3 mimetics and MEK inhibitors in the treatment of acute myeloid leukemia

Assessment of the hydrocarbon potential of copper-bearing shale from southern part of the Fore-Sudetic Monocline

Głównym celem pracy była próba określenia możliwości generacji węglowodorów z poziomu łupku miedzionośnego na podstawie wyników badań pirolitycznych. W celu oceny materii organicznej 13 próbek rdzeniowych pochodzących z rejonu złóż rud miedzi w południowej części monokliny przedsudeckiej poddano badaniom pirolitycznym Rock-Eval. Dodatkowo próbki skalne badano również przy użyciu pirolizy wysokotemperaturowej sprzężonej z kapilarną chromatografią gazową (Py-GC/FID) i studiowano pod kątem zmienności trzech grup węglowodorowych: lekkich – C1–C9, ciekłych – C10–C15 oraz ciężkich – powyżej C15+. W przypadku każdej próbki została zastosowana jednoetapowa piroliza, prowadzona w programowanej temperaturze 500°C przez 0,4 min. Uzyskane wyniki były interpretowane kompleksowo i pozwoliły na rozróżnienie charakteru generowanych produktów na podstawie dystrybucji węglowodorów otrzymanych w wyniku pirolizy substancji organicznej zawartej w skale. Do określenia potencjalnych możliwości generacyjnych badanych łupków miedzionośnych wykorzystano zarówno udział procentowy poszczególnych frakcji, jak też bezwymiarowy wskaźnik uzysku (liczony z analizy Py-GC/FID). Na podstawie wyników badań pirolitycznych Rock-Eval oraz zbieżnych z nimi wartości wskaźników uzysku Py-GC/FID stwierdzono doskonały potencjał węglowodorowy dla wybranych próbek pochodzących z poziomu łupku miedzionośnego. Stopień dojrzałości substancji organicznej wyrażony poprzez parametr Tmax w przypadku większości badanych próbek z poziomu łupku miedzionośnego mieści się w zakresie okna ropnego, co potwierdziły również badania Py-GC/FID. Dodatkowo o możliwości generacji węglowodorów świadczy fakt występowania wysokiej zawartości ekstrahowalnej substancji organicznej oraz bardzo wysoka wartość parametru S2 (nawet do 41,88 mg HC/g skały). Analiza wyników badań geochemicznych pod kątem oceny potencjału generacyjnego poziomu łupku miedzionośnego udowodniła, że poziom ten nie powinien być pomijany jako potencjalnie macierzysty przy rozpatrywaniu źródła napełniania pułapek węglowodorów w systemie naftowym Niżu Polskiego.The main objective of the research was to attempt to determine the possibility of generating hydrocarbon from the copperbearing shales based on the results of pyrolysis tests. In order to assess the organic matter, 13 core samples from the copper ore deposit region of the southern part of the Fore-Sudetic monocline were subjected to Rock-Eval pyrolysis tests. In addition, rock samples were also tested using high-temperature pyrolysis coupled with capillary gas chromatography (Py-GC/FID) and studied for the variability of three hydrocarbon groups: light C1–C9, liquid C10–C15 and heavy above C15+. For each sample, a one-stage pyrolysis was used, carried out at a programmed temperature of 500°C for 0.4 min. The obtained results were interpreted comprehensively and enabled the nature of the generated products to be distinguished on the basis of the distribution of hydrocarbons obtained as a result of pyrolysis of organic matter contained in the rock. To determine the potential generation capacity of the tested copper-bearing shales, both the percentage share of individual fractions and the dimensionless recovery index (calculated from the Py-GC/FID analysis) were used. Based on the results of the Rock-Eval pyrolysis tests and the Py-GC/FID yield indicators, an excellent hydrocarbon potential was found for selected samples from the copper-bearing shale level. The degree of maturity of the organic substance expressed by the Tmax parameter for most of the tested samples from the copper-bearing shale horizon is within the oil window, which was also confirmed by the Py-GC/FID tests. In addition, the possibility of hydrocarbons generation is evidenced by the presence of a high content of extractable organic matter and a very high value of the S2 parameter (up to 41.88 mg HC/g of rock). The analysis of the results of geochemical research in terms of the assessment of the generation potential of the copper-bearing shales proved that this horizon should not be overlooked as a potential source when considering filling hydrocarbon traps in the oil system of the Polish Lowlands

Dendritic Cell-Specific Role for Pellino2 as a Mediator of TLR9 Signaling Pathway

Ubiquitination regulates immune signaling, and multiple E3 ubiquitin ligases have been studied in the context of their role in immunity. Despite this progress, the physiological roles of the Pellino E3 ubiquitin ligases, especially Pellino2, in immune regulation remain largely unknown. Accordingly, this study aimed to elucidate the role of Pellino2 in murine dendritic cells (DCs). In this study, we reveal a critical role of Pellino2 in regulation of the proinflammatory response following TLR9 stimulation. Pellino2-deficient murine DCs show impaired secretion of IL-6 and IL-12. Loss of Pellino2 does not affect TLR9-induced activation of NF-κB or MAPKs, pathways that drive expression of IL-6 and IL-12. Furthermore, DCs from Pellino2-deficient mice show impaired production of type I IFN following endosomal TLR9 activation, and it partly mediates a feed-forward loop of IFN-β that promotes IL-12 production in DCs. We also observe that Pellino2 in murine DCs is downregulated following TLR9 stimulation, and its overexpression induces upregulation of both IFN-β and IL-12, demonstrating the sufficiency of Pellino2 in driving these responses. This suggests that Pellino2 is critical for executing TLR9 signaling, with its expression being tightly regulated to prevent excessive inflammatory response. Overall, this study highlights a (to our knowledge) novel role for Pellino2 in regulating DC functions and further supports important roles for Pellino proteins in mediating and controlling immunity

Bile acids induce IL-1α1\alpha and drive NLRP3 inflammasome-independent production of IL-1β1\beta in murine dendritic cells

Bile acids are amphipathic molecules that are synthesized from cholesterol in the liver and facilitate intestinal absorption of lipids and nutrients. They are released into the small intestine upon ingestion of a meal where intestinal bacteria can modify primary into secondary bile acids. Bile acids are cytotoxic at high concentrations and have been associated with inflammatory diseases such as liver inflammation and Barrett’s Oesophagus. Although bile acids induce pro-inflammatory signalling, their role in inducing innate immune cytokines and inflammation has not been fully explored to date. Here we demonstrate that the bile acids, deoxycholic acid (DCA) and chenodeoxycholic acid (CDCA) induce IL-$1\alpha$ and IL-$1\beta$ secretion in vitro in primed bone marrow derived dendritic cells (BMDCs). The secretion of IL-$1\beta$ was found not to require expression of NLRP3, ASC or caspase-1 activity; we can’t rule out all inflammasomes. Furthermore, DCA and CDCA were shown to induce the recruitment of neutrophils and monocytes to the site of injection an intraperitoneal model of inflammation. This study further underlines a mechanistic role for bile acids in the pathogenesis of inflammatory diseases through stimulating the production of pro-inflammatory cytokines and recruitment of innate immune cells

Lazy neutrophils : a lack of DGAT1 reduces the chemotactic activity of mouse neutrophils

Background Neutrophils are key players in the innate immune system, actively migrating to sites of inflammation in the highly energetic process of chemotaxis. In this study, we focus on the role of acyl-CoA: diacylglycerol acyltransferase 1 (DGAT1), an enzyme that catalyzes the synthesis of triglycerides, the major form of stored energy, in neutrophil chemotaxis. Methods and results Using a mouse model of psoriasis, we show that DGAT1-deficiency reduces energy-demanding neutrophil infiltration to the site of inflammation, but this inhibition is not caused by decreased glycolysis and reduced ATP production by neutrophils lacking DGAT1. Flow cytometry and immunohistochemistry analysis demonstrate that DGAT1 also does not influence lipid accumulation in lipid droplets during inflammation. Interestingly, as has been shown previously, a lack of DGAT1 leads to an increase in the concentration of retinoic acid, and here, using real-time PCR and publicly-available next-generation RNA sequencing datasets, we show the upregulation of retinoic acid-responsive genes in Dgat1KO neutrophils. Furthermore, supplementation of WT neutrophils with exogenous retinoic acid mimics DGAT1-deficiency in the inhibition of neutrophil chemotaxis in in vitro transwell assay. Conclusions These results suggest that impaired skin infiltration by neutrophils in Dgat1KO mice is a result of the inhibitory action of an increased concentration of retinoic acid, rather than impaired lipid metabolism in DGAT1-deficient mice