title: A defined, controlled culture system for primary bovine chromaffin progenitors reveals novel biomarkers and modulators.
abstract: We present a method to efficiently culture primary chromaffin progenitors from the adult bovine adrenal medulla in a defined, serum-free monolayer system. Tissue is dissociated and plated for expansion under support by the mitogen basic fibroblast growth factor (bFGF). The cultures, although not homogenous, contain a subpopulation of cells expressing the neural stem cell marker Hes3 that also propagate. In addition, Hes3 is also expressed in the adult adrenal medulla from where the tissue is taken. Differentiation is induced by bFGF withdrawal and switching to Neurobasal medium containing B27. Following differentiation, Hes3 expression is lost, and cells acquire morphologies and biomarker expression patterns of chromaffin cells and dopaminergic neurons. We tested the effect of different treatments that we previously showed regulate Hes3 expression and cell number in cultures of fetal and adult rodent neural stem cells. Treatment of the cultures with a combination of Delta4, Angiopoietin2, and a Janus kinase inhibitor increases cell number during the expansion phase without significantly affecting catecholamine content levels. Treatment with cholera toxin does not significantly affect cell number but reduces the ratio of epinephrine to norepinephrine content and increases the dopamine content relative to total catecholamines. These data suggest that this defined culture system can be used for target identification in drug discovery programs and that the transcription factor Hes3 may serve as a new biomarker of putative adrenomedullary chromaffin progenitor cells.
authors.affiliation: Department of Internal Medicine III and Institute of Clinical Chemistry and Laboratory Medicine, University Clinic Carl-Gustav Carus, University of Dresden, Dresden, Germany; Center for Regenerative Therapies Dresden, Dresden, Germany.
authors.forename: Jimmy
authors.initials: J
authors.lastname: Masjkur
authors.affiliation: Department of Internal Medicine III and Institute of Clinical Chemistry and Laboratory Medicine, University Clinic Carl-Gustav Carus, University of Dresden, Dresden, Germany; Center for Regenerative Therapies Dresden, Dresden, Germany.
authors.forename: Ian
authors.initials: I
authors.lastname: Levenfus
authors.affiliation: Department of Internal Medicine III and Institute of Clinical Chemistry and Laboratory Medicine, University Clinic Carl-Gustav Carus, University of Dresden, Dresden, Germany; Center for Regenerative Therapies Dresden, Dresden, Germany.
authors.forename: Sven
authors.initials: S
authors.lastname: Lange
authors.affiliation: Department of Internal Medicine III and Institute of Clinical Chemistry and Laboratory Medicine, University Clinic Carl-Gustav Carus, University of Dresden, Dresden, Germany; Center for Regenerative Therapies Dresden, Dresden, Germany.
authors.forename: Carina
authors.initials: C
authors.lastname: Arps-Forker
authors.affiliation: Department of Internal Medicine III and Institute of Clinical Chemistry and Laboratory Medicine, University Clinic Carl-Gustav Carus, University of Dresden, Dresden, Germany; Center for Regenerative Therapies Dresden, Dresden, Germany.
authors.forename: Steve
authors.initials: S
authors.lastname: Poser
authors.affiliation: Department of Internal Medicine III and Institute of Clinical Chemistry and Laboratory Medicine, University Clinic Carl-Gustav Carus, University of Dresden, Dresden, Germany; Center for Regenerative Therapies Dresden, Dresden, Germany.
authors.forename: Nan
authors.initials: N
authors.lastname: Qin
authors.affiliation: Department of Internal Medicine III and Institute of Clinical Chemistry and Laboratory Medicine, University Clinic Carl-Gustav Carus, University of Dresden, Dresden, Germany; Center for Regenerative Therapies Dresden, Dresden, Germany.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Vukicevic
authors.affiliation: Department of Internal Medicine III and Institute of Clinical Chemistry and Laboratory Medicine, University Clinic Carl-Gustav Carus, University of Dresden, Dresden, Germany; Center for Regenerative Therapies Dresden, Dresden, Germany.
authors.forename: Triantafyllos
authors.initials: T
authors.lastname: Chavakis
authors.affiliation: Department of Internal Medicine III and Institute of Clinical Chemistry and Laboratory Medicine, University Clinic Carl-Gustav Carus, University of Dresden, Dresden, Germany; Center for Regenerative Therapies Dresden, Dresden, Germany.
authors.forename: Graeme
authors.initials: G
authors.lastname: Eisenhofer
authors.affiliation: Department of Internal Medicine III and Institute of Clinical Chemistry and Laboratory Medicine, University Clinic Carl-Gustav Carus, University of Dresden, Dresden, Germany; Center for Regenerative Therapies Dresden, Dresden, Germany.
authors.forename: Stefan R
authors.initials: SR
authors.lastname: Bornstein
authors.affiliation: Department of Internal Medicine III and Institute of Clinical Chemistry and Laboratory Medicine, University Clinic Carl-Gustav Carus, University of Dresden, Dresden, Germany; Center for Regenerative Therapies Dresden, Dresden, Germany.
authors.forename: Monika
authors.initials: M
authors.lastname: Ehrhart-Bornstein
authors.affiliation: Department of Internal Medicine III and Institute of Clinical Chemistry and Laboratory Medicine, University Clinic Carl-Gustav Carus, University of Dresden, Dresden, Germany; Center for Regenerative Therapies Dresden, Dresden, Germany andreas.theotokis@uniklinikum-dresden.de.
authors.forename: Andreas
authors.initials: A
authors.lastname: Androutsellis-Theotokis
mesh_terms: D000313:Adrenal Medulla / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D042702:Angiopoietin-2 / Q000494:pharmacology; D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002395:Catecholamines / Q000378:metabolism; D002417:Cattle; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D049109:Cell Proliferation; D002478:Cells, Cultured; D002772:Cholera Toxin / Q000494:pharmacology; D019439:Chromaffin Cells / Q000187:drug effects / Q000378:metabolism*; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D047908:Intracellular Signaling Peptides and Proteins / Q000494:pharmacology; D053612:Janus Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D008565:Membrane Proteins / Q000494:pharmacology; D058953:Neural Stem Cells / Q000187:drug effects / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D015398:Signal Transduction; D013997:Time Factors; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
chemical_list: D042702:Angiopoietin-2; D015415:Biomarkers; D002395:Catecholamines; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D047428:Protein Kinase Inhibitors; D014157:Transcription Factors; C064045:delta protein; D016222:Fibroblast Growth Factor 2; D002772:Cholera Toxin; D053612:Janus Kinases
keywords: Cell culture; Differentiation; Self-renewal; Signal transduction