ABSTRACT
Microglia are resident inflammatory cells of the CNS and have important roles in development, homeostasis and a variety of neurologic and psychiatric diseases. Difficulties in procuring human microglia have limited their study and hampered the clinical translation of microglia-based treatments shown to be effective in animal disease models. Here we report the differentiation of human induced pluripotent stem cells (iPSC) into microglia-like cells by exposure to defined factors and co-culture with astrocytes. These iPSC-derived microglia have the phenotype, gene expression profile and functional properties of brain-isolated microglia. Murine iPSC-derived microglia generated using a similar protocol have equivalent efficacy to primary brain-isolated microglia in treatment of murine syngeneic intracranial malignant gliomas. The ability to generate human microglia facilitates the further study of this important CNS cell type and raises the possibility of their use in personalized medicine applications.
Subject(s)
Cell Differentiation/physiology , Induced Pluripotent Stem Cells/physiology , Microglia/metabolism , Microglia/physiology , Animals , Astrocytes/cytology , Cell Movement , Coculture Techniques , Cytokines/metabolism , Gene Expression Profiling , Gene Knock-In Techniques , Glioma/therapy , Humans , Induced Pluripotent Stem Cells/cytology , Mice , Microglia/transplantation , Phagocytosis/physiology , Reactive Oxygen Species/metabolism , Survival Analysis , Xenograft Model Antitumor AssaysABSTRACT
Neotyphodium species are fungal endophytes best known for their protection of grass hosts and production of bioactive metabolites including ergot alkaloids. Perennial ryegrass-Neotyphodium sp. Lp1 symbiota that have altered ergot alkaloid profiles (resulting from knockouts in two different endophyte genes) were fed, along with controls, to rabbits to test the effects of ergot alkaloids on food preference and satiety. Interestingly, rabbits dramatically preferred plants that were endophyte-infected but free of ergot alkaloids over endophyte-free plants (P = 0.01). Accumulation of ergot alkaloids of the clavine class counteracted the added appeal of endophyte-infected plants. In satiety tests, consumption of ergovaline (the ultimate ergot pathway product in wild-type endophyte), but not of several other ergot alkaloids, during an initial meal had a negative effect on subsequent rabbit chow consumption (P < 0.05). The data indicate that clavines were sufficient to reduce the appeal of endophyte-infected grasses, whereas only ergovaline reduced appetite.
Subject(s)
Ergot Alkaloids/pharmacology , Food Preferences/drug effects , Hypocreales/genetics , Lolium/microbiology , Satiation/drug effects , Animals , Diet , Ergot Alkaloids/analysis , Ergot Alkaloids/genetics , Female , Gene Deletion , Hypocreales/metabolism , Lolium/chemistry , Plant Leaves/chemistry , Rabbits , SymbiosisABSTRACT
Lysergyl peptide synthetase 1 catalyzes the assembly of toxic ergopeptines from activated D-lysergic acid and three amino acids. The gene encoding this enzyme in the endophytic fungus Neotyphodium lolii was analyzed and compared to a homologous gene from the ergot fungus Claviceps purpurea. Each gene contained two introns, which were found in the same relative position within two modules of the gene. The 5' ends of the two genes were unusually divergent. Signature sequences determining substrate specificity were similar in adenylation domains that recognized identical amino acids but differed within the adenylation domain for the amino acid that varies between the major ergopeptines of the two fungi. Homologues were detected in several related endophytic fungi; the tall fescue endophyte Neotyphodium coenophialum contained a divergent, second copy of the gene. Our results provide new information on the structure and distribution of this important peptide synthetase involved in ergot alkaloid biosynthesis.
