Development and implementation of a significantly low-cost 3D bioprinter using recycled scrap material

The field of 3D bioengineering proposes to effectively contribute to the manufacture of artificial multicellular organ/tissues and the understanding of complex cellular mechanisms. In this regard, 3D cell cultures comprise a promising bioengineering possibility for the alternative treatment of organ function loss, potentially improving patient life expectancies. Patients with end-stage disease, for example, could benefit from treatment until organ transplantation or even undergo organ function restoration. Currently, 3D bioprinters can produce tissues such as trachea cartilage or artificial skin. Most low-cost 3D bioprinters are built from fused deposition modeling 3D printer frames modified for the deposition of biologically compatible material, ranging between $13.000,00 and$300.000,00. Furthermore, the cost of consumables should also be considered as they, can range from $3,85 and$100.000,00 per gram, making biomaterials expensive, hindering bioprinting access. In this context, our report describes the first prototype of a significantly low-cost 3D bioprinter built from recycled scrap metal and off-the-shelf electronics. We demonstrate the functionalized process and methodology proof of concept and aim to test it in different biological tissue scaffolds in the future, using affordable materials and open-source methodologies, thus democratizing the state of the art of this technology

SOCS2-Induced Proteasome-Dependent TRAF6 Degradation: A Common Anti-Inflammatory Pathway for Control of Innate Immune Responses

Pattern recognition receptors and receptors for pro-inflammatory cytokines provide critical signals to drive the development of protective immunity to infection. Therefore, counter-regulatory pathways are required to ensure that overwhelming inflammation harm host tissues. Previously, we showed that lipoxins modulate immune response during infection, restraining inflammation during infectious diseases in an Aryl hydrocarbon receptor (AhR)/suppressors of cytokine signaling (SOCS)2-dependent-manner. Recently, Indoleamine-pyrrole 2,3- dioxygenase (IDO)-derived tryptophan metabolites, including L-kynurenine, were also shown to be involved in several counter-regulatory mechanisms. Herein, we addressed whether the intracellular molecular events induced by lipoxins mediating control of innate immune signaling are part of a common regulatory pathway also shared by L-kynurenine exposure. We demonstrate that Tumor necrosis factor receptor-associated factor (TRAF)6 – member of a family of adapter molecules that couple the TNF receptor and interleukin-1 receptor/Toll-like receptor families to intracellular signaling events essential for the development of immune responses – is targeted by both lipoxins and L-kynurenine via an AhR/SOCS2-dependent pathway. Furthermore, we show that LXA4- and L-kynurenine-induced AhR activation, its subsequent nuclear translocation, leading SOCS2 expression and TRAF6 Lys47-linked poly-ubiquitination and proteosome-mediated degradation of the adapter proteins. The in vitro consequences of such molecular interactions included inhibition of TLR- and cytokine receptor-driven signal transduction and cytokine production. Subsequently, in vivo proteosome inhibition led to unresponsiveness to lipoxins, as well as to uncontrolled pro-inflammatory reactions and elevated mortality during toxoplasmosis. In summary, our results establish proteasome degradation of TRAF6 as a key molecular target for the anti-inflammatory pathway triggered by lipoxins and L-kynurenine, critical counter-regulatory mediators in the innate and adaptive immune systems

Identification of Candidate Susceptibility and Resistance Genes of Mice Infected with Streptococcus suis Type 2

Streptococcus suis type 2 (SS2) is an important swine pathogen and zoonosis agent. A/J mice are significantly more susceptible than C57BL/6 (B6) mice to SS2 infection, but the genetic basis is largely unknown. Here, alterations in gene expression in SS2 (strain HA9801)-infected mice were identified using Illumina mouse BeadChips. Microarray analysis revealed 3,692 genes differentially expressed in peritoneal macrophages between A/J and B6 mice due to SS2 infection. Between SS2-infected A/J and control A/J mice, 2646 genes were differentially expressed (1469 upregulated; 1177 downregulated). Between SS2-infected B6 and control B6 mice, 1449 genes were differentially expressed (778 upregulated; 671 downregulated). These genes were analyzed for significant Gene Ontology (GO) categories and signaling pathways using the Kyoto Encylopedia of Genes and Genomes (KEGG) database to generate a signaling network. Upregulated genes in A/J and B6 mice were related to response to bacteria, immune response, positive regulation of B cell receptor signaling pathway, type I interferon biosynthesis, defense and inflammatory responses. Additionally, upregulated genes in SS2-infected B6 mice were involved in antigen processing and presentation of exogenous peptides, peptide antigen stabilization, lymphocyte differentiation regulation, positive regulation of monocyte differentiation, antigen receptor-mediated signaling pathway and positive regulation of phagocytosis. Downregulated genes in SS2-infected B6 mice played roles in glycolysis, carbohydrate metabolic process, amino acid metabolism, behavior and muscle regulation. Microarray results were verified by quantitative real-time PCR (qRT-PCR) of 14 representative deregulated genes. Four genes differentially expressed between SS2-infected A/J and B6 mice, toll-like receptor 2 (Tlr2), tumor necrosis factor (Tnf), matrix metalloproteinase 9 (Mmp9) and pentraxin 3 (Ptx3), were previously implicated in the response to S. suis infection. This study identified candidate genes that may influence susceptibility or resistance to SS2 infection in A/J and B6 mice, providing further validation of these models and contributing to understanding of S. suis pathogenic mechanisms

Defoliation and Soil Compaction Jointly Drive Large-Herbivore Grazing Effects on Plants and Soil Arthropods on Clay Soil

In addition to the well-studied impacts of defecation and defoliation, large herbivores also affect plant and arthropod communities through trampling, and the associated soil compaction. Soil compaction can be expected to be particularly important on wet, fine-textured soils. Therefore, we established a full factorial experiment of defoliation (monthly mowing) and soil compaction (using a rammer, annually) on a clay-rich salt marsh at the Dutch coast, aiming to disentangle the importance of these two factors. Additionally, we compared the effects on soil physical properties, plants, and arthropods to those at a nearby cattle-grazed marsh under dry and under waterlogged conditions. Soil physical conditions of the compacted plots were similar to the conditions at cattle-grazed plots, showing decreased soil aeration and increased waterlogging. Soil salinity was doubled by defoliation and quadrupled by combined defoliation and compaction. Cover of the dominant tall grass Elytrigia atherica was decreased by 80% in the defoliated plots, but cover of halophytes only increased under combined defoliation and compaction. Effects on soil micro-arthropods were most severe under waterlogging, showing a fourfold decrease in abundance and a smaller mean body size under compaction. Although the combined treatment of defoliation and trampling indeed proved most similar to the grazed marsh, large discrepancies remained for both plant and soil fauna communities, presumably because of colonization time lags. We conclude that soil compaction and defoliation differently affect plant and arthropod communities in grazed ecosystems, and that the magnitude of their effects depends on herbivore density, productivity, and soil physical properties

Environmental and vegetation controls on the spatial variability of CH4 emission from wet-sedge and tussock tundra ecosystems in the Arctic

Aims Despite multiple studies investigating the environmental controls on CH4 fluxes from arctic tundra ecosystems, the high spatial variability of CH4 emissions is not fully understood. This makes the upscaling of CH4 fluxes from plot to regional scale, particularly challenging. The goal of this study is to refine our knowledge of the spatial variability and controls on CH4 emission from tundra ecosystems. Methods CH4 fluxes were measured in four sites across a variety of wet-sedge and tussock tundra ecosystems in Alaska using chambers and a Los Gatos CO2 and CH4 gas analyser. Results All sites were found to be sources of CH4, with northern sites (in Barrow) showing similar CH4 emission rates to the southernmost site (ca. 300 km south, Ivotuk). Gross primary productivity (GPP), water level and soil temperature were the most important environmental controls on CH4 emission. Greater vascular plant cover was linked with higher CH4 emission, but this increased emission with increased vascular plant cover was much higher (86 %) in the drier sites, than the wettest sites (30 %), suggesting that transport and/or substrate availability were crucial limiting factors for CH4 emission in these tundra ecosystems. Conclusions Overall, this study provides an increased understanding of the fine scale spatial controls on CH4 flux, in particular the key role that plant cover and GPP play in enhancing CH4 emissions from tundra soils