title: HB-EGF affects astrocyte morphology, proliferation, differentiation, and the expression of intermediate filament proteins.
abstract: Heparin-binding epidermal growth factor-like growth factor (HB-EGF), a vascular-derived trophic factor, belongs to the epidermal growth factor (EGF) family of neuroprotective, hypoxia-inducible proteins released by astrocytes in CNS injuries. It was suggested that HB-EGF can replace fetal calf serum (FCS) in astrocyte cultures. We previously demonstrated that in contrast to standard 2D cell culture systems, Bioactive3D culture system, when used with FCS, minimizes the baseline activation of astrocytes and preserves their complex morphology. Here, we show that HB-EGF induced EGF receptor (EGFR) activation by Y1068 phosphorylation, Mapk/Erk pathway activation, and led to an increase in cell proliferation, more prominent in Bioactive3D than in 2D cultures. HB-EGF changed morphology of 2D and Bioactive3D cultured astrocytes toward a radial glia-like phenotype and induced the expression of intermediate filament and progenitor cell marker protein nestin. Glial fibrillary acidic protein (GFAP) and vimentin protein expression was unaffected. RT-qPCR analysis demonstrated that HB-EGF affected the expression of Notch signaling pathway genes, implying a role for the Notch signaling in HB-EGF-mediated astrocyte response. HB-EGF can be used as a FCS replacement for astrocyte expansion and in vitro experimentation both in 2D and Bioactive3D culture systems; however, caution should be exercised since it appears to induce partial de-differentiation of astrocytes.
authors.affiliation: Center for Brain Repair and Rehabilitation, Department of Clinical Neuroscience and Rehabilitation, Institute of Neuroscience and Physiology, Sahlgrenska Academy at the University of Gothenburg, Gothenburg, Sweden.
authors.forename: Till B
authors.initials: TB
authors.lastname: Puschmann
authors.forename: Carl
authors.initials: C
authors.lastname: Zandén
authors.forename: Isabell
authors.initials: I
authors.lastname: Lebkuechner
authors.forename: Camille
authors.initials: C
authors.lastname: Philippot
authors.forename: Yolanda
authors.initials: Y
authors.lastname: de Pablo
authors.forename: Johan
authors.initials: J
authors.lastname: Liu
authors.forename: Milos
authors.initials: M
authors.lastname: Pekny
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D018929:Cell Culture Techniques / Q000379:methods; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D005260:Female; D005904:Glial Fibrillary Acidic Protein; D066257:Heparin-binding EGF-like Growth Factor; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology / Q000502:physiology*; D007381:Intermediate Filament Proteins / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology*; D008297:Male; D051379:Mice; D057507:Mice, 129 Strain; D008810:Mice, Inbred C57BL; D009419:Nerve Tissue Proteins / Q000378:metabolism; D064231:Nestin / Q000378:metabolism; D051880:Receptors, Notch / Q000378:metabolism; D014746:Vimentin / Q000378:metabolism
chemical_list: D005904:Glial Fibrillary Acidic Protein; C587884:Hbegf protein, mouse; D066257:Heparin-binding EGF-like Growth Factor; D036341:Intercellular Signaling Peptides and Proteins; D007381:Intermediate Filament Proteins; D009419:Nerve Tissue Proteins; C577485:Nes protein, mouse; D064231:Nestin; D051880:Receptors, Notch; D014746:Vimentin; C528900:glial fibrillary astrocytic protein, mouse
keywords: 3D cell culture; HB-EGF; astrocyte culture; astrocytes; intermediate filament system