Project description:Anaerobic ammonium-oxidising (anammox) bacteria, members of the ‘Candidatus Brocadiaceae’ family, play an important role in the nitrogen cycle and are estimated to be responsible for about half of the oceanic nitrogen loss to the atmosphere. Anammox bacteria combine ammonium with nitrite and produce dinitrogen gas via the intermediates nitric oxide and hydrazine (anammox reaction) while nitrate is formed as a by-product. These reactions take place in a specialized, membrane-bound compartment called the anammoxosome. Therefore, the substrates ammonium, nitrite and product nitrate have to cross the outer-, cytoplasmic- and anammoxosome membranes to enter or exit the anammoxosome. The genomes of all anammox species harbour multiple copies of ammonium-, nitrite- and nitrate transporter genes. Here we investigated how the distinct genes for ammonium-, nitrite- and nitrate- transport were expressed during substrate limitation in membrane bioreactors. Transcriptome analysis of Kuenenia stuttgartiensis planktonic cells under ammonium-limitation showed that three of the seven ammonium transporter genes and one of the six nitrite transporter genes were significantly upregulated, while another ammonium and nitrite transporter gene were downregulated in nitrite limited growth conditions. The two nitrate transporters were expressed to similar levels in both conditions. In addition, genes encoding enzymes involved in the anammox reaction were differentially expressed, with those using nitrite as a substrate being upregulated under nitrite limited growth and those using ammonium as a substrate being upregulated during ammonium limitation. Taken together, these results give a first insight in the potential role of the multiple nutrient transporters in regulating transport of substrates and products in and out of the compartmentalized anammox cell.
Project description:The manufacturing of autologous chimaeric antigen receptor (CAR) T cells largely relies either on fed-batch and manual processes that often lack environmental monitoring and control or on bioreactors that cannot be easily scaled out to meet patient demands. Here we show that human primary T cells can be activated, transduced and expanded to high densities in a 2 ml automated closed-system microfluidic bioreactor to produce viable anti-CD19 CAR T cells (specifically, more than 60 million CAR T cells from donor cells derived from patients with lymphoma and more than 200 million CAR T cells from healthy donors). The in vitro secretion of cytokines, the short-term cytotoxic activity and the long-term persistence and proliferation of the cell products, as well as their in vivo anti-leukaemic activity, were comparable to those of T cells produced in a gas-permeable well. The manufacturing-process intensification enabled by the miniaturized perfusable bioreactor may facilitate the analysis of the growth and metabolic states of CAR T cells during ex vivo culture, the high-throughput optimization of cell-manufacturing processes and the scale-out of cell-therapy manufacturing.
Project description:Human cells require pH regulation to maintain physiological function, yet the molecular consequences of acidic environments remain incompletely understood. Here, we employ a gas-only bioreactor to control pH, oxygen, and temperature. Integrated Omics analyses reveal that acidic pH induces a glycolytic metabolic shift, suppresses proliferation, and promotes accumulation of lactate and oncometabolites alongside mitochondrial dysfunction. Acidic conditions increase reactive oxygen species (ROS) and activate inflammatory and immune pathways, leading to heteroplasmic enrichment of a pathogenic mitochondrial mutation. Acidic pH depletes intracellular NAD⁺, partly driven by PARP1 activation. Restoring NAD⁺ through nicotinamide mononucleotide (NMN) supplementation partially rescues proliferation and stress-associated transcription, while elevating NAD+ levels by NMN or PARP1 inhibition reverses heteroplasmic enrichment of mutant mitochondrial DNA. These findings underscore the role of pH homeostasis in coordinating metabolism, redox balance, and immune signaling, and identify NAD⁺ metabolism as a mechanistic link between acidic microenvironments, mitochondrial genome instability, and immune–metabolic remodeling.
Project description:D-galactose orally intake ameliorate DNCB-induced atopic dermatitis by modulating microbiota composition and quorum sensing. The increased abundance of bacteroidetes and decreased abundance of firmicutes was confirmed. By D-galactose treatment, Bacteroides population was increased and prevotella, ruminococcus was decreased which is related to atopic dermatitis.
