Understanding glioblastoma stromal barriers against NK cell attack using tri-culture 3D spheroid model.

Glioblastoma multiforme (GBM), a highly aggressive tumor type with a dismal survival rate, has a poor outcome which is at least partly attributed to the crosstalk between cancer cells and cells from the tumor microenvironment such as astrocytes and microglia. We aimed to decipher the effect of these cells on GBM progression and on cell-based therapies using 3D co-cultures. Co-culturing of glioblastoma cells with patient-derived astrocytes or microglia or both formed dense and heterogeneous spheroids. Both, astrocytes and microglia, enhanced the spheroid growth rate and formed a physical barrier for macromolecules penetration, while only astrocytes enhanced the migration. Interestingly bi-/tri-cultured spheroids showed significant resistance against NK-92 cells, likely attributed to dense stroma and induced expression of immunosuppressive genes such as IDO1 or PTGES2. Altogether, our novel 3D GBM spheroid model recapitulates the cell-to-cell interactions of human glioblastoma and can serve as a suitable platform for evaluating cancer therapeutics.

Subversion of a family of antimicrobial proteins by Salmonella enterica.

Salmonella enterica is a food-borne pathogen able to cause a wide spectrum of diseases ranging from mild gastroenteritis to systemic infections. During almost all stages of the infection process Salmonella is likely to be exposed to a wide variety of host-derived antimicrobial peptides (AMPs). AMPs are important components of the innate immune response which integrate within the bacterial membrane, thus forming pores which lead ultimately to bacterial killing. In contrast to other AMPs Bactericidal/Permeability-increasing Protein (BPI) displayed only weak bacteriostatic or bactericidal effects towards Salmonella enterica sv. Typhimurium (STM) cultures. Surprisingly, we found that sub-antimicrobial concentrations of BPI fold-containing (BPIF) superfamily members mediated adhesion of STM depending on pre-formed type 1 fimbriae. BPIF proteins directly bind to type 1 fimbriae through mannose-containing oligosaccharide modifications. Fimbriae decorated with BPIF proteins exhibit extended binding specificity, allowing for bacterial adhesion on a greater variety of abiotic and biotic surfaces likely promoting host colonization. Further, fimbriae significantly contributed to the resistance against BPI, probably through sequestration of the AMP before membrane interaction. In conclusion, functional subversion of innate immune proteins of the BPIF family through binding to fimbriae promotes Salmonella virulence by survival of host defense and promotion of host colonization.

Formation of vivianite in digested sludge and its controlling factors in municipal wastewater treatment.

Engineering solutions to recover phosphorus from municipal wastewater are required to close the anthropogenic phosphorus cycle. After chemical phosphorus elimination by iron, the ferrous iron­phosphorus mineral vivianite forms in digested sludge, and its separation is being researched at the pilot scale. In this study, sludge samples from 16 wastewater treatment plants (WWTPs) demonstrated that phosphorus bound to biomass and redox-sensitive iron in activated sludge was transformed into other phosphorus binding forms, including vivianite, during digestion. Vivianite quantity was approximated using X-ray diffraction and two sequential extractions. These three independent methods of approximating vivianite quantity were closely related confirming their relationship to the vivianite content in the samples. The digested sludge from three WWTPs exhibited comparatively high levels of vivianite-bound phosphorus approximated between 31 % and 51 % of total phosphorus. The controlling factors of vivianite formation were investigated in order to enhance its formation in digested sludge and increase the amount of phosphorus recoverable as vivianite. They were identified using single and multivariate correlation (MLR), considering the sludge properties, sludge composition, and process parameters within the operating range of the 16 WWTPs. Increasing iron content was verified as the primary predictor of significantly increased vivianite formation (MLR: p < 0.001). In addition, increasing sulphur content was found to be an additional significant factor that decreased vivianite formation (MLR: p < 0.05). Furthermore, a comparison of plants using sulphur-free (FeCl2 and FeCl3) and sulphur-containing (FeSO4 and FeClSO4) precipitants indicated that the latter could increase the sulphur content in digested sludge (one-tailed Welch two-sample t-test: t(14.6) = 2.3, p = 0.02). Thus, by increasing the sulphur content, the use of sulphur-comprising precipitants may counteract vivianite formation, whereas sulphur-free precipitants may facilitate it and, hence, promote vivianite recovery.

Endotoxin contamination alters macrophage-cancer cell interaction and therapeutic efficacy in pre-clinical 3D in vitro models.

The rapid developments in biofabrication, in particular 3D bioprinting, in the recent years have facilitated the need for novel biomaterials that aim to replicate the target tissue in great detail. The presence of endotoxins in these biomaterials is often an overlooked problem. In pre-clinical 3D in vitro models, endotoxins can have significant influence on cell behavior and credibility of the model. In this study we demonstrate the effects of high levels of endotoxins in commercially-available gelatin on the macrophage-cancer cell crosstalk in a 3D bioprinted co-culture model. First, it is demonstrated that, while presenting the same mechanical and structural stimuli, high levels of endotoxin can have significant influence on the metabolic activity of macrophages and cancer cells. Furthermore, this study shows that high endotoxin contamination causes a strong inflammatory reaction in macrophages and significantly inhibits the effects of a paracrine macrophage-cancer cell co-culture. At last, it is demonstrated that the differences in endotoxin levels can drastically alter the efficacy of novel macrophage modulating immunotherapies, AS1517499 and 3-methyladenine. Altogether, this study shows that endotoxin contamination in biomaterials can significantly alter intra- and intercellular communication and thereby drug efficacy, which might lead to misinterpretation of the potency and safety of the tested compounds.

Lipoxygenases in chronic liver diseases: current insights and future perspectives.

Chronic liver diseases (CLDs) caused by viral infections, alcohol/drug abuse, or metabolic disorders affect millions of people globally and have increased mortality owing to the lack of approved therapies. Lipoxygenases (LOXs) are a family of multifaceted enzymes that are responsible for the oxidation of polyunsaturated fatty acids (PUFAs) and are implicated in the pathogenesis of multiple disorders including liver diseases. This review describes the three main LOX signaling pathways - 5-, 12-, and 15-LOX - and their involvement in CLDs. We also provide recent insights and future perspectives on LOX-related hepatic pathophysiology, and discuss the potential of LOXs and LOX-derived metabolites as diagnostic biomarkers and therapeutic targets in CLDs.

Transformation of redox-sensitive to redox-stable iron-bound phosphorus in anoxic lake sediments under laboratory conditions.

Phosphorus (P) can be retained in mineral association with ferrous iron (Fe) as vivianite, Fe(II)3(PO4)2 ∙ 8 H2O, in lake sediments. The mineral is formed and remains stable under anoxic non-sulphidogenic conditions and, therefore, acts as a long-term P sink. In laboratory experiments under anoxic conditions, we investigated whether P adsorbed to amorphous Fe(III)-hydroxide functioned as a precursor phase of vivianite when added to different sediments as a treatment. The untreated sediments served as controls and were naturally Fe-rich (559 µmol/g DW) and Fe-poor (219 µmol/g DW), respectively. The solid P binding forms analysed by sequential extraction and X-ray diffraction were related to coinciding pore water analyses and the bacterial community compositions of the sediments by bacterial 16S rRNA gene amplicon sequencing. In the treatments, within a period of 40 d, 70 % of the redox-sensitive Fe(III)-P was transformed into redox-stable P, which contained vivianite. The mineral was supersaturated in the pore water, but the presence of Fe(III)-P functioning as a precursor was sufficient for measurable vivianite formation. The composition of the microbial community did not differ significantly (PERMANOVA, p = 0.09) between treatment and control of the naturally Fe-rich sediment. In the naturally Fe-poor sediment, the microbial community changed significantly (PERMANOVA, p = 0.001) in response to the addition of Fe(III)-P to the sediment. The freshly formed redox-stable P was not retransferred to a redox-sensitive compound by aeration for 24 h until 90 % O2 saturation was reached in the sediment slurry. We conclude that 1) Fe(III)-hydroxide bound P, resulting from oxic conditions at the sediment-water interface, is immobilised during anoxic conditions and stable even after re-oxygenation; 2) the process is feasible within the time scales of anoxic lake stratification periods; and 3) in relatively Fe-poor lakes, Fe dosing can provide excess Fe to form the precursor.

Sediment core data reconstruct the management history and usage of a heavily modified urban lake in Berlin, Germany.

Urban surface waters face several stressors associated with industry and urban water management. Over much of the past century, the wastewater treatment in Berlin, Germany, relied on inefficient sewage farms, which resulted in severe eutrophication and sediment contamination in the recipient surface waterbodies. A prominent example is Lake Tegel, where a multitude of management measures were applied in the last decades for the purpose of ecosystem restoration. In this study, we analyzed sediment cores of three lakes with X-ray fluorescence spectroscopy: Lake Tegel, Lake Großer Wannsee, which is environmentally similar but has a different management history, and Lake Userin, which serves as a reference located in a nature protection area. Multivariate statistical methods (principal component analysis, k-means clustering, and self-organizing maps) were used to assess the sediment quality and to reconstruct the management history of Lake Tegel. Principal component analysis established two main gradients of sediment composition: heavy metals and lithogenic elements. The impact of the management measures was visualized in the lake sediment composition changing from high abundance of heavy metals and reducing redox conditions to less-impacted sediments in recent layers. The clustering techniques suggested heterogeneity among sites within Lake Tegel that probably reflect urban water management measures. The abundance of heavy metals in recent lake sediments of Lake Tegel is similar to a lake with low urban impact and is lower than in Lake Großer Wannsee suggesting that the management measures were successful in the reduction of heavy metals, which are still a threat for surface waters worldwide.