Astrocyte activation and reactive gliosis : A new target in stroke?

Stroke is an acute insult to the central nervous system (CNS) that triggers a sequence of responses in the acute, subacute as well as later stages, with prominent involvement of astrocytes. Astrocyte activation and reactive gliosis in the acute stage of stroke limit the tissue damage and contribute to the restoration of homeostasis. Astrocytes also control many aspects of neural plasticity that is the basis for functional recovery. Here, we discuss the concept of intermediate filaments (nanofilaments) and the complement system as two handles on the astrocyte responses to injury that both present attractive opportunities for novel treatment strategies modulating astrocyte functions and reactive gliosis.Peer reviewe

The role of the intermediate filament (nanofilament) protein nestin in neural progenitor cell and astrocyte differentiation

Nestin, a class VI intermediate filament (nanofilament) protein, is commonly used as a marker for neural stem/progenitor cells (NSPCs), but its role in neurogenesis remains elusive. Nestin is also expressed in immature and reactive astrocytes. The up-regulation of intermediate filament proteins glial fibrillary acidic protein (GFAP), vimentin and nestin is a characteristic feature of reactive astrocytes, accompanied by alterations in the expression of many other genes. We found that transgenic mice deficient for nestin have increased number of newly born hippocampal neurons 6 weeks after BrdU in vivo labeling, suggesting increased generation and/or survival of newly born neurons. We also showed that in vitro nestin deficient astrocytes provide a more pro-neurogenic environment that results in a 2-fold increase in neuronal differentiation of NSPCs. Astrocytes are highly heterogeneous cells and fulfill a variety of important functions in healthy as well as diseased brain. In addition, astrocytes can exhibit features characteristic of NSPCs and modulate neurogenesis by inhibiting neuronal differentiation of NSPCs through Jagged1-mediated Notch signaling. Given the vast array of astrocyte functions, classification of astrocyte subpopulations on a molecular level is highly desirable. We used single cell quantitative real-time PCR to investigate the heterogeneity of astrocytes with respect to their Notch signaling competence. Our results show that the Notch signal sending but not Notch signal receiving competence of astrocytes depends on GFAP and vimentin. Further, we showed that nestin and heparin binding EGF-like growth factor (HB-EGF) may serve as classifiers of astrocyte subpopulations in vitro. Utilizing our newly developed Bioactive3D and standard 2D cell culture systems, we showed that HB-EGF alters astrocyte morphology towards a more radial glia-like phenotype. HB-EGF affects proliferation, differentiation and expression of Notch signal pathway related genes and leads to the up regulation of nestin expression. HB-EGF in cell culture media results in partial de-differentiation of astrocytes and therefore should be used with caution

HB-EGF affects astrocyte morphology, proliferation, differentiation, and the expression of intermediate filament proteins

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.HB-EGF (heparin-binding EGF-like growth factor) was previously suggested to replace serum, a common and undefined component in primary astrocyte cultures. We show that both in standard 2D and in our newly developed Bioactive3D culture system, HB-EGF affects astrocyte morphology, proliferation, gene/protein expression and leads to partial de-differentiation of astrocytes. Thus, HB-EGF should only be used with caution as a serum replacement in astrocyte cultures

HB-EGF affects astrocyte morphology, proliferation, differentiation, and the expression of intermediate filament proteins

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.HB-EGF (heparin-binding EGF-like growth factor) was previously suggested to replace serum, a common and undefined component in primary astrocyte cultures. We show that both in standard 2D and in our newly developed Bioactive3D culture system, HB-EGF affects astrocyte morphology, proliferation, gene/protein expression and leads to partial de-differentiation of astrocytes. Thus, HB-EGF should only be used with caution as a serum replacement in astrocyte cultures