Beyond butyrate: microbial fiber metabolism supporting colonic epithelial homeostasis.

Human gut bacteria produce metabolites that support energy and carbon metabolism of colonic epithelial cells. While butyrate is commonly considered the primary fuel, it alone cannot meet all the carbon requirements for cellular synthetic functions. Glucose, delivered via circulation or microbial metabolism, serves as a universal carbon source for synthetic processes like DNA, RNA, protein, and lipid production. Detailed knowledge of epithelial carbon and energy metabolism is particularly relevant for epithelial regeneration in digestive and metabolic diseases, such as inflammatory bowel disease and type 2 diabetes. Here, we review the production and role of different colonic microbial metabolites in energy and carbon metabolism of colonocytes, also critically evaluating the common perception that butyrate is the preferred fuel.

Differential and organ-specific functions of organic solute transporter α and ß in experimental cholestasis.

Background & Aims: Organic solute transporter (OST) subunits OSTα and OSTß facilitate bile acid efflux from the enterocyte into the portal circulation. Patients with deficiency of OSTα or OSTß display considerable variation in the level of bile acid malabsorption, chronic diarrhea, and signs of cholestasis. Herein, we generated and characterized a mouse model of OSTß deficiency. Methods: Ostß -/- mice were generated using CRISR/Cas9 and compared to wild-type and Ostα -/- mice. OSTß was re-expressed in livers of Ostß -/- mice using adeno-associated virus serotype 8 vectors. Cholestasis was induced in both models by bile duct ligation (BDL) or 3.5-diethoxycarbonyl-1.4-dihydrocollidine (DDC) feeding. Results: Similar to Ostα -/- mice, Ostß -/- mice exhibited elongated small intestines with blunted villi and increased crypt depth. Increased expression levels of ileal Fgf15, and decreased Asbt expression in Ostß -/- mice indicate the accumulation of bile acids in the enterocyte. In contrast to Ostα -/- mice, induction of cholestasis in Ostß -/- mice by BDL or DDC diet led to lower survival rates and severe body weight loss, but an improved liver phenotype. Restoration of hepatic Ostß expression via adeno-associated virus-mediated overexpression did not rescue the phenotype of Ostß -/- mice. Conclusions: OSTß is pivotal for bile acid transport in the ileum and its deficiency leads to an intestinal phenotype similar to Ostα -/- mice, but it exerts distinct effects on survival and the liver phenotype, independent of its expression in the liver. Our findings provide insights into the variable clinical presentation of patients with OSTα and OSTß deficiencies. Lay summary: Organic solute transporter (OST) subunits OSTα and OSTß together facilitate the efflux of conjugated bile acids into the portal circulation. Ostα knockout mice have longer and thicker small intestines and are largely protected against experimental cholestatic liver injury. Herein, we generated and characterized Ostß knockout mice for the first time. Ostα and Ostß knockout mice shared a similar phenotype under normal conditions. However, in cholestasis, Ostß knockout mice had a worsened overall phenotype which indicates a separate and specific role of OSTß, possibly as an interacting partner of other intestinal proteins.

Incorporation of Toll-Like Receptor Ligands and Inflammasome Stimuli in GM3 Liposomes to Induce Dendritic Cell Maturation and T Cell Responses.

Cancer vaccination aims to activate immunity towards cancer cells and can be achieved by delivery of cancer antigens together with immune stimulatory adjuvants to antigen presenting cells (APC). APC maturation and antigen processing is a subsequent prerequisite for T cell priming and anti-tumor immunity. In order to specifically target APC, nanoparticles, such as liposomes, can be used for the delivery of antigen and adjuvant. We have previously shown that liposomal inclusion of the ganglioside GM3, an endogenous ligand for CD169, led to robust uptake by CD169-expressing APC and resulted in strong immune responses when supplemented with a soluble adjuvant. To minimize the adverse effects related to a soluble adjuvant, immune stimulatory molecules can be incorporated in liposomes to achieve targeted delivery of both antigen and adjuvant. In this study, we incorporated TLR4 (MPLA) or TLR7/8 (3M-052) ligands in combination with inflammasome stimuli, 1-palmitoyl-2-glutaryl-sn-glycero-3-phosphocholine (PGPC) or muramyl dipeptide (MDP), into GM3 liposomes. Incorporation of TLR and inflammasome ligands did not interfere with the uptake of GM3 liposomes by CD169-expressing cells. GM3 liposomes containing a TLR ligand efficiently matured human and mouse dendritic cells in vitro and in vivo, while inclusion of PGPC or MDP had minor effects on maturation. Immunization with MPLA-containing GM3 liposomes containing an immunogenic synthetic long peptide stimulated CD4+ and CD8+ T cell responses, but additional incorporation of either PGPC or MDP did not translate into stronger immune responses. In conclusion, our study indicates that TLRL-containing GM3 liposomes are effective vectors to induce DC maturation and T cell priming and thus provide guidance for further selection of liposomal components to optimally stimulate anti-cancer immune responses.

Marker-free lineage tracing reveals an environment-instructed clonogenic hierarchy in pancreatic cancer.

Effective treatments for pancreatic ductal adenocarcinoma (PDAC) are lacking, and targeted agents have demonstrated limited efficacy. It has been speculated that a rare population of cancer stem cells (CSCs) drives growth, therapy resistance, and rapid metastatic progression in PDAC. These CSCs demonstrate high clonogenicity in vitro and tumorigenic potential in vivo. However, their relevance in established PDAC tissue has not been determined. Here, we use marker-independent stochastic clonal labeling, combined with quantitative modeling of tumor expansion, to uncover PDAC tissue growth dynamics. We find that in contrast to the CSC model, all PDAC cells display clonogenic potential in situ. Furthermore, the proximity to activated cancer-associated fibroblasts determines tumor cell clonogenicity. This means that the microenvironment is dominant in defining the clonogenic activity of PDAC cells. Indeed, manipulating the stroma by Hedgehog pathway inhibition alters the tumor growth mode, revealing that tumor-stroma crosstalk shapes tumor growth dynamics and clonal architecture.