Glucose-6-phosphate transporter mediates macrophage proliferation and functions by regulating glycolysis and mitochondrial respiration.

Glycogen storage disease type Ib (GSD-Ib), caused by a deficiency in glucose-6-phosphate transporter (G6PT), is characterized by disrupted glucose homeostasis, inflammatory bowel disease, neutropenia, and neutrophil dysfunction. The purpose of this study was to investigate the role of G6PT on macrophage functions and metabolism. Peritoneal macrophages of G6pt-/- mice were lower in number and their effector functions including migration, superoxide production, and phagocytosis were impaired. To investigate the underlying mechanisms of macrophage dysfunction, the G6PT gene was mutated in porcine alveolar macrophage 3D4/31 cells using the CRISPR/Cas9 technology. The G6PT-deficient macrophages exhibited significant decline in cell growth, bactericidal activity, and antiviral response. These phenotypes are associated with the impaired glycolysis and mitochondrial oxidative phosphorylation. We therefore propose that the G6PT-mediated metabolism is essential for effector functions of macrophage, the immune deficiencies observed in GSD-Ib extend beyond neutropenia and neutrophil dysfunction, and future therapeutic targets aimed both the neutrophils and macrophages may be necessary.

Dudleya brittonii extract promotes survival rate and M2-like metabolic change in porcine 3D4/31 alveolar macrophages.

OBJECTIVE: Although alveolar macrophages play a key role in the respiratory immunity of livestock, but studies on the mechanism of differentiation and survival of alveolar macrophages are lacking. Therefore, we undertook to investigate changes in the lipid metabolism and survival rate, using 3D4/31 macrophages and Dudleya brittonii which has been used as a traditional asthma treatment. METHODS: 3D4/31 macrophages were used as the in vitro porcine alveolar macrophages model. The cells were activated by exposure to Phorbol 12-Myristate 13-Acetate (PMA). D. Brittonii extraction was performed with distilled water. For evaluating the cell survival rate, we performed the water-soluble tetrazolium salt (WST) cell viability assay and growth curve analysis. To confirm cell death, cell cycle and intracellular reactive oxygen species levels were measured using flow cytometric analysis by applying fluorescence dye dichlorofluorescein diacetate (DCFDA) and propidium iodide (PI). Furthermore, we also evaluated cellular lipid accumulation with Oil Red O staining, and fatty acid synthesis related genes expression levels using quantitative PCR (qPCR) with SYBR green dye. Glycolysis, fatty acid oxidation, and tricarboxylic acid (TCA) cycle related gene expression levels were measured using qPCR after exposure to Dudleya brittonii extract (DB) for 12 h. RESULTS: Reactive oxygen species (ROS) production and cell death were induced by PMA treatment, and exposure to DB reduced the PMA induced downregulation of cell survival. PMA and DB treatments upregulated the lipid accumulation, with corresponding increase in the acetyl-CoA carboxylase alpha (ACACA), fatty acid synthase (FASN) mRNA expressions. DB-PMA co-treatment reduced the glycolysis genes expression, but increased the expressions of fatty acid oxidation and TCA cycle genes. CONCLUSION: This study provides new insights and directions for further researches relating to the immunity of porcine respiratory system, by employing a model based on alveolar macrophages and natural materials.