The Tumor Necrosis Factor Superfamily Members TNFSF14 (LIGHT), Lymphotoxin β and Lymphotoxin β Receptor Interact to Regulate Intestinal Inflammation

Over 1.5 million individuals in the United States are afflicted with inflammatory bowel disease (IBD). While the progression of IBD is multifactorial, chronic, unresolved inflammation certainly plays a key role. Additionally, while multiple immune mediators have been shown to affect pathogenesis, a comprehensive understanding of disease progression is lacking. Previous work has demonstrated that a member of the TNF superfamily, TNFSF14 (LIGHT), which is pro-inflammatory in several contexts, surprisingly plays an important role in protection from inflammation in mouse models of colitis, with LIGHT deficient mice having more severe disease pathogenesis. However, LIGHT is a single member of a complex signaling network. It signals through multiple receptors, including herpes virus entry mediator (HVEM) and lymphotoxin beta receptor (LTβR); these two receptors in turn can bind to other ligands. It remains unknown which receptors and competing ligands can mediate or counteract the outcome of LIGHT-signaling during colitis. Here we demonstrate that LIGHT signaling through LTβR, rather than HVEM, plays a critical role in the progression of DSS-induced colitis, as LTβR deficient mice exhibit a more severe disease phenotype. Further, mice deficient in LTαβ do not exhibit differential colitis progression compared to WT mice. However, deletion of both LIGHT and LTαβ, but not deletion of both LTαβ and LTβR, resulted in a reversal of the adverse effects associated with the loss of LIGHT. In sum, the LIGHT/LTαβ/LTβR signaling network contributes to DSS colitis, but there may be additional receptors or indirect effects, and therefore, the relationships between these receptors and ligands remains enigmatic

Autophagy limits proliferation and glycolytic metabolism in acute myeloid leukemia.

Decreased autophagy contributes to malignancies, however it is unclear how autophagy impacts on tumour growth. Acute myeloid leukemia (AML) is an ideal model to address this as (i) patient samples are easily accessible, (ii) the hematopoietic stem and progenitor population (HSPC) where transformation occurs is well characterized, and (iii) loss of the key autophagy gene Atg7 in hematopoietic stem and progenitor cells (HSPCs) leads to a lethal pre-leukemic phenotype in mice. Here we demonstrate that loss of Atg5 results in an identical HSPC phenotype as loss of Atg7, confirming a general role for autophagy in HSPC regulation. Compared to more committed/mature hematopoietic cells, healthy human and mouse HSCs displayed enhanced basal autophagic flux, limiting mitochondrial damage and reactive oxygen species in this long-lived population. Taken together, with our previous findings these data are compatible with autophagy limiting leukemic transformation. In line with this, autophagy gene losses are found within chromosomal regions that are commonly deleted in human AML. Moreover, human AML blasts showed reduced expression of autophagy genes, and displayed decreased autophagic flux with accumulation of unhealthy mitochondria indicating that deficient autophagy may be beneficial to human AML. Crucially, heterozygous loss of autophagy in an MLL-ENL model of AML led to increased proliferation in vitro, a glycolytic shift, and more aggressive leukemias in vivo. With autophagy gene losses also identified in multiple other malignancies, these findings point to low autophagy providing a general advantage for tumour growth

Autophagy-dependent generation of free fatty acids is essential for normal neutrophil differentiation by guiding an energy-metabolic switch

Neutrophils are critical and short lived mediators of innate immunity that require constant replenishment. Their differentiation in the bone marrow requires extensive cytoplasmic and nuclear remodeling, but the processes governing these energy-consuming changes are unknown. While previous studies show that autophagy is required for differentiation of other blood cell lineages, its function during granulopoiesis has remained elusive. Autophagy was described as a critical process in HSC differentiation and in memory- and regulatory T cells, where it prevents excessive glycolysis and maintains lipid metabolic homeostasis. In the myeloid lineage, an essential role for autophagy is to prevent pro-inflammatory macrophage polarization and to limit glycolytic metabolism of acute myeloid leukemia. While these studies provide robust in vivo evidence for the relevance of autophagy in the differentiation of hematopoietic and immune cells, the targets and mechanisms of autophagy remain elusive. Here, we show that metabolism and autophagy are developmentally programmed and essential for neutrophil differentiation in vivo. Atg7 -deficient neutrophil precursors had increased glycolytic activity but impaired mitochondrial respiration, decreased ATP production and accumulated lipid droplets. Inhibiting autophagy-mediated lipid degradation or fatty acid oxidation alone was sufficient to cause defective differentiation, while administration of fatty acids or pyruvate for mitochondrial respiration rescued differentiation in autophagy deficient neutrophil precursors. Together, we show that autophagy-mediated lipolysis provides free fatty acids to support a mitochondrial respiration pathway essential to neutrophil differentiation. Evidence presented here also contributed to studies on autophagy-mediated metabolic homeostasis in HSCs, Treg cells and myeloid leukemia, suggesting that this pathway may act broadly during differentiation.</p

Autophagy-dependent generation of free fatty acids is essential for normal neutrophil differentiation by guiding an energy-metabolic switch

Neutrophils are critical and short lived mediators of innate immunity that require constant replenishment. Their differentiation in the bone marrow requires extensive cytoplasmic and nuclear remodeling, but the processes governing these energy-consuming changes are unknown. While previous studies show that autophagy is required for differentiation of other blood cell lineages, its function during granulopoiesis has remained elusive. Autophagy was described as a critical process in HSC differentiation and in memory- and regulatory T cells, where it prevents excessive glycolysis and maintains lipid metabolic homeostasis. In the myeloid lineage, an essential role for autophagy is to prevent pro-inflammatory macrophage polarization and to limit glycolytic metabolism of acute myeloid leukemia. While these studies provide robust in vivo evidence for the relevance of autophagy in the differentiation of hematopoietic and immune cells, the targets and mechanisms of autophagy remain elusive. Here, we show that metabolism and autophagy are developmentally programmed and essential for neutrophil differentiation in vivo. Atg7 -deficient neutrophil precursors had increased glycolytic activity but impaired mitochondrial respiration, decreased ATP production and accumulated lipid droplets. Inhibiting autophagy-mediated lipid degradation or fatty acid oxidation alone was sufficient to cause defective differentiation, while administration of fatty acids or pyruvate for mitochondrial respiration rescued differentiation in autophagy deficient neutrophil precursors. Together, we show that autophagy-mediated lipolysis provides free fatty acids to support a mitochondrial respiration pathway essential to neutrophil differentiation. Evidence presented here also contributed to studies on autophagy-mediated metabolic homeostasis in HSCs, Treg cells and myeloid leukemia, suggesting that this pathway may act broadly during differentiation.</p

Autophagy preserves hematopoietic stem cells by restraining mTORC1-mediated cellular anabolism

Adult stem cells are long-lived and quiescent with unique metabolic requirements. Macroautophagy/autophagy is a fundamental survival mechanism that allows cells to adapt to metabolic changes by degrading and recycling intracellular components. Here we address why autophagy depletion leads to a drastic loss of the stem cell compartment. Using inducible deletion of autophagy specifically in adult hematopoietic stem cells (HSCs) and in mice chimeric for autophagy-deficient and normal HSCs, we demonstrate that the stem cell loss is cell-intrinsic. Mechanistically, autophagy-deficient HSCs showed higher expression of several amino acid transporters (AAT) when compared to autophagy-competent cells, resulting in increased amino acid (AA) uptake. This was followed by sustained mTOR (mammalian target of rapamycin) activation, with enlarged cell size, glucose uptake and translation, which is detrimental to the quiescent HSCs. mTOR inhibition by rapamycin treatment in vivo was able to rescue autophagy-deficient HSC loss and bone marrow failure and resulted in better reconstitution after transplantation. Our results suggest that targeting mTOR may improve aged stem cell function, promote reprogramming and stem cell transplantation

Metabolic activation and colitis pathogenesis is prevented by lymphotoxin β receptor expression in neutrophils.

Inflammatory bowel disease is characterized by an exacerbated intestinal immune response, but the critical mechanisms regulating immune activation remain incompletely understood. We previously reported that the TNF-superfamily molecule TNFSF14 (LIGHT) is required for preventing severe disease in mouse models of colitis. In addition, deletion of lymphotoxin beta receptor (LTβR), which binds LIGHT, also led to aggravated colitis pathogenesis. Here, we aimed to determine the cell type(s) requiring LTβR and the mechanism critical for exacerbation of colitis. Specific deletion of LTβR in neutrophils (LTβRΔN), but not in several other cell types, was sufficient to induce aggravated colitis and colonic neutrophil accumulation. Mechanistically, RNA-Seq analysis revealed LIGHT-induced suppression of cellular metabolism, and mitochondrial function, that was dependent on LTβR. Functional studies confirmed increased mitochondrial mass and activity, associated with excessive mitochondrial ROS production and elevated glycolysis at steady-state and during colitis. Targeting these metabolic changes rescued exacerbated disease severity. Our results demonstrate that LIGHT signals to LTβR on neutrophils to suppress metabolic activation and thereby prevents exacerbated immune pathogenesis during colitis