| 1. |
- Erdei, Borbala, et al.
(författare)
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SSF of steam-pretreated wheat straw with the addition of saccharified or fermented wheat meal in integrated bioethanol production
- 2013
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Ingår i: Biotechnology for Biofuels. - : Springer Science and Business Media LLC. - 1754-6834. ; 6
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Tidskriftsartikel (refereegranskat)abstract
- Background: Integration of second-generation (2G) bioethanol production with existing first-generation (1G) production may facilitate commercial production of ethanol from cellulosic material. Since 2G hydrolysates have a low sugar concentration and 1G streams often have to be diluted prior to fermentation, mixing of streams is beneficial. Improved ethanol concentrations in the 2G production process lowers energy demand in distillation, improves overall energy efficiency and thus lower production cost. There is also a potential to reach higher ethanol yields, which is required in economically feasible ethanol production. Integrated process scenarios with addition of saccharified wheat meal (SWM) or fermented wheat meal (FWM) were investigated in simultaneous saccharification and (co-)fermentation (SSF or SSCF) of steam-pretreated wheat straw, while the possibility of recovering the valuable protein-rich fibre residue from the wheat was also studied. Results: The addition of SWM to SSF of steam-pretreated wheat straw, using commercially used dried baker's yeast, S. cerevisiae, resulted in ethanol concentrations of about 60 g/L, equivalent to ethanol yields of about 90% of the theoretical. The addition of FWM in batch mode SSF was toxic to baker's yeast, due to the ethanol content of FWM, resulting in a very low yield and high accumulation of glucose. The addition of FWM in fed-batch mode still caused a slight accumulation of glucose, but the ethanol concentration was fairly high, 51.2 g/L, corresponding to an ethanol yield of 90%, based on the amount of glucose added. In batch mode of SSCF using the xylose-fermenting, genetically modified S. cerevisiae strain KE6-12, no improvement was observed in ethanol yield or concentration, compared with baker's yeast, despite the increased xylose utilization, probably due to the considerable increase in glycerol production. A slight increase in xylose consumption was seen when glucose from SWM was fed at a low feed rate, after 48 hours, compared with batch SSCF. However, the ethanol yield and concentration remained in the same range as in batch mode. Conclusion: Ethanol concentrations of about 6% (w/v) were obtained, which will result in a significant reduction in the cost of downstream processing, compared with SSF of the lignocellulosic substrate alone. As an additional benefit, it is also possible to recover the protein-rich residue from the SWM in the process configurations presented, providing a valuable co-product.
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| 2. |
- Hancz, Dóra, et al.
(författare)
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Inhibition of inflammasome-dependent interleukin 1β production by streptococcal NAD+-glycohydrolase : Evidence for extracellular activity
- 2017
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Ingår i: mBio. - 2161-2129. ; 8:4
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Tidskriftsartikel (refereegranskat)abstract
- Group A Streptococcus (GAS) is a common human pathogen and the etiologic agent of a large number of diseases ranging from mild, self-limiting infections to invasive life-threatening conditions. Two prominent virulence factors of this bacterium are the genetically and functionally linked pore-forming toxin streptolysin O (SLO) and its cotoxin NAD+-glycohydrolase (NADase). Overexpression of these toxins has been linked to increased bacterial virulence and is correlated with invasive GAS disease. NADase can be translocated into host cells by a SLO-dependent mechanism, and cytosolic NADase has been assigned multiple properties such as protection of intracellularly located GAS bacteria and induction of host cell death through energy depletion. Here, we used a set of isogenic GAS mutants and a macrophage infection model and report that streptococcal NADase inhibits the innate immune response by decreasing inflammasome-dependent interleukin 1β (IL-1β) release from infected macrophages. Regulation of IL-1β was independent of phagocytosis and ensued also under conditions not allowing SLO-dependent translocation of NADase into the host cell cytosol. Thus, our data indicate that NADase not only acts intracellularly but also has an immune regulatory function in the extracellular niche. IMPORTANCE In the mid-1980s, the incidence and severity of invasive infections caused by serotype M1 GAS suddenly increased. The results of genomic analyses suggested that this increase was due to the spread of clonal bacterial strains and identified a recombination event leading to enhanced production of the SLO and NADase toxins in these strains. However, despite its apparent importance in GAS pathogenesis, the function of NADase remains poorly understood. In this study, we demonstrate that NADase inhibits inflammasome-dependent IL-1β release from infected macrophages. While previously described functions of NADase pertain to its role upon SLO-mediated translocation into the host cell cytosol, our data suggest that the immune regulatory function of NADase is exerted by nontranslocated enzyme, identifying a previously unrecognized extracellular niche for NADase functionality. This immune regulatory property of extracellular NADase adds another possible explanation to how increased secretion of NADase correlates with bacterial virulence.
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| 3. |
- Hancz, Dora
(författare)
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Regulation of inflammasomes by host- and pathogen-derived factors
- 2019
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The term inflammasome was first coined in 2002 and describes an intracellular multiprotein complex mediating inflammation by inducing the release of pro-inflammatory cytokines (IL-1β and IL-18) and a lytic form of cell death called pyroptosis. Since then, an impressive amount of research has revealed many details about the mechanisms of inflammasome activation as well as the functional relevance of inflammasome activation in different contexts. Yet, the continuously expanding inflammasome field has still many open, unresolved questions to answer and there is a need for further knowledge to help us navigate the sometimes confusing and challenging complexity of inflammasomes.Inflammasomes have been suggested to have a crucial role in host defense against pathogens. Their significance in host protection is highlighted by a large number of studies identifying immune evasion strategies interfering with inflammasomes. However, these complexes can also be detrimental for the host. Signaling leading to inflammasome activation requires tight regulation, as uncontrolled activation can promote inflammatory, metabolic as well as neurodegenerative disorders. The work presented in this thesis is focused on how inflammasomes and the subsequent release of IL-1β are regulated by different host- and pathogen-derived factors.In Paper I and II, we investigated the effect of two structurally and functionally related virulence factors of the major human pathogen Group A Streptococcus (streptolysin O and NAD+-glycohydrolase) linked to invasive streptococcal diseases, on NLRP3 inflammasome activation. We identified two immune evasion strategies, both down-regulating GAS-induced IL-1β levels. In Paper I, we demonstrate that extracellularly located NAD+-glycohydrolase inhibits the secretion of inflammasome-dependent IL-1β from infected macrophages without effecting inflammasome assembly. In Paper II, we established a role for streptolysin O in promoting the ubiquitination and subsequent degradation of pro-IL-1β. We propose that both immune evasion strategies could limit IL-1β-induced inflammation and immune cell recruitment to the site of infection, potentially benefiting bacterial survival and/or spread.In Paper III, we propose a role for extracellular histone H4 in increasing the inflammatory potential of macrophages by licensing them for a previouslyunknown, TLR-mediated inflammasome activation. TLR-mediated inflammasome activation leads to IL-1β secretion but proceeds without IL-18 release or pyroptosis, supporting recent findings suggesting that inflammasome effector functions can be uncoupled from each other. We suggest that upon tissue injury, histone H4-licensed macrophages may exhibit a lowered inflammatory threshold and thus react to a single microbial challenge with inflammasome activation to fight the infection. However, macrophages continuously releasing IL-1β in response to TLR stimulation may also contribute to an undesired extended inflammatory response under conditions such as sepsis or sterile inflammation, disorders with known pathological role for both extracellular histones and IL-1β.Overall, studies presented in this thesis expand our understanding on the mechanism of inflammasome activation as well as the regulation of IL-1β levels in infectious as well as inflammatory settings.
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| 4. |
- Hancz, Dóra, et al.
(författare)
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Streptolysin O induces the ubiquitination and degradation of pro-IL-1β
- 2019
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Ingår i: Journal of Innate Immunity. - : S. Karger AG. - 1662-811X .- 1662-8128. ; 11:6, s. 457-468
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Tidskriftsartikel (refereegranskat)abstract
- Group A Streptococcus (GAS) is a common and versatile human pathogen causing a variety of diseases. One of the many virulence factors of GAS is the secreted pore-forming cytotoxin streptolysin O (SLO), which has been ascribed multiple properties, including inflammasome activation leading to release of the potent inflammatory cytokine IL-1β from infected macrophages. IL-1β is synthesized as an inactive pro-form, which is activated intracellularly through proteolytic cleavage. Here, we use a macrophage infection model to show that SLO specifically induces ubiquitination and degradation of pro-IL-1β. Ubiquitination was dependent on SLO being released from the infecting bacterium, and pore formation by SLO was required but not sufficient for the induction of ubiquitination. Our data provide evidence for a novel SLO-mediated mechanism of immune regulation, emphasizing the importance of this pore-forming toxin in bacterial virulence and pathogenesis.
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| 5. |
- Riva, Matteo, et al.
(författare)
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Induction of NFκB responses by the S100A9 protein is TLR4-dependent.
- 2012
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Ingår i: Immunology. - : Wiley. - 0019-2805. ; 137:2, s. 172-182
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Tidskriftsartikel (refereegranskat)abstract
- Interactions between danger and pathogen-associated molecular patterns (DAMP and PAMP) and pattern recognition receptors such as Toll-like receptors (TLRs) are critical for the regulation of the inflammatory process via activation of NFκB and cytokine secretion. In this report, we investigated the capacity of LPS-free S100A9 (DAMP) protein to activate human and mouse cells compared to lipoprotein-free LPS (PAMP). Firstly, we showed that LPS and S100A9 were able to increase NFκB activity followed by increased cytokine and nitric oxide (NO) secretion both in human THP-1 cells and mouse bone marrow-derived dendritic cells (BM-DC). Surprisingly, although S100A9 triggered a weaker cytokine response compared to LPS, we found that S100A9 more potently induced IκBα degradation and hence NFkB activation. Both the S100A9- and LPS-induced response was completely absent in TLR4 knock-out mice, while it was only slightly affected in RAGE knock-out mice. Also, we showed that LPS and S100A9 NFkB induction were strongly reduced in the presence of specific inhibitors of TLR-signaling. Chloroquine reduced S100A9 but not LPS signaling, indicating that S100A9 may need to be internalized in order to be fully active as a TLR4 inducer. This was confirmed using A488-labeled S100A9 that was internalized in THP-1 cells, showing a raise in fluorescence after 30 minutes at 37°C. Chloroquine treatment significantly reduced the fluorescence. In summary, our data indicate that both human and mouse S100A9 are TLR4 agonists. Importantly, S100A9 induced stronger NFκB activation albeit weaker cytokine secretion than LPS, suggesting that S100A9 and LPS activated NFκB in a qualitative distinct manner. © 2012 The Authors. Immunology © 2012 Blackwell Publishing Ltd, Immunology.
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| 6. |
- Valfridsson, Christine, et al.
(författare)
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Detection of Inflammasome Activation in Murine Bone Marrow-Derived Macrophages Infected with Group A Streptococcus
- 2023
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Ingår i: Methods in Molecular Biology. - 1064-3745 .- 1940-6029. ; 2674, s. 261-282
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Bokkapitel (refereegranskat)abstract
- Inflammasomes are large multiprotein complexes that assemble mainly in innate immune cells after detection of microbial or sterile insults. Activation of inflammasomes is a key proinflammatory event during infection, and many pathogens have evolved specific evasion mechanisms to evade or inhibit inflammasome activation. One such pathogen is the common bacterium group A Streptococcus (GAS), which causes a wide range of diseases of varying severity. GAS secretes a multitude of virulence factors whereof the pore-forming protein streptolysin O (SLO) is the main inflammasome activation determinant. Here we provide a protocol for reliable evaluation of inflammasome activation in murine bone marrow-derived macrophages (BMDM) infected with GAS, including instructions for generating BMDMs and growing the bacterium. This protocol can easily be modified to other bacterial pathogens, or human macrophages.
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