Studies on the Role of Fibroblast Growth Factor Signaling in Neurogenesis Using Conjugated/Aged Animal Caps and Dorsal Ectoderm-Grafted Embryos (original) (raw)
Basic fibroblast growth factor in neuronal cultures of human fetal brain
Journal of Neuroscience Research, 1990
The presence of basic fibroblast growth factor (bFGF) was investigated in neuronal cells derived from 12 and 18 week-old human fetal brain cultures. To this purpose, the ability of bFGF to stimulate plasminogen activator (PA) production in fetal bovine aortic endothelial GM 7373 cells was used as an assay for this molecule in neuronal cell extracts. The identity of the PA-stimulating activity of neuronal cell extract with bFGF was confirmed by its high affinity for heparin and by its cross-reactivity with polyclonal antibodies to human placental bFGF. These antibodies recognized a Mr 18,000 cell-associated protein both in Western blot and in immuno-precipitation experiments. All the neurons showed bFGF immunoreactivity, as demonstrated hy immunocytochemical staining, while nonneuronal cells were unstained. The data demonstrate for the first time that cultured human fetal brain neurons contain and synthesize bFGF.
Fibroblast growth factors as regulators of central nervous system development and function
American journal of physiology. Regulatory, integrative and comparative physiology, 2003
Fibroblast growth factors (FGFs) are multifunctional signaling proteins that regulate developmental processes and adult physiology. Over the last few years, important progress has been made in understanding the function of FGFs in the embryonic and adult central nervous system. In this review, I will first discuss studies showing that FGF signaling is already required during formation of the neural plate. Next, I will describe how FGF signaling centers control growth and patterning of specific brain structures. Finally, I will focus on the function of FGF signaling in the adult brain and in regulating maintenance and repair of damaged neural tissues.
Experimental Neurology, 2000
The in vivo application of appropriate trophic factors may enhance regeneration of bulbospinal projections after spinal cord injury. Currently, little is known about the sensitivities of specific bulbospinal neuron populations to the many identified trophic factors. We devised novel in vitro assays to study trophic effects on the survival and neurite outgrowth of identified bulbospinal neurons. Carbocyanine dye crystals implanted into the cervical spinal cord of embryonic day (E)5 chick embryos retrogradely labeled developing bulbospinal neurons. On E8, dissociated cultures containing labeled bulbospinal neurons were prepared. Fibroblast growth factor (FGF)-2 (but not FGF-1) promoted the survival of bulbospinal neurons. FGF receptor expression was widespread in the E8 brainstem, but not detected in young bulbospinal neurons, suggesting that nonneuronal cells mediated the FGF-stimulated survival response. Astrocytes synthesize a variety of trophic factors, and astrocyte-conditioned medium (ACM) also promoted the survival of bulbospinal neurons. As might be expected, FGF-2 function blocking antibodies did not suppress ACM-promoted survival, nor did an ELISA detect FGF-2 in ACM. This suggests that nonneuronal cells synthesize other factors in response to exogenous FGF-2 which promote the survival of bulbospinal neurons. Focusing on vestibulospinal neurons, dissociated (survival assay) or explant (neurite outgrowth assay) cultures were prepared. FGF-2 promoted both survival and neurite outgrowth of identified vestibulospinal neurons. Interestingly, FGF-1 promoted neurite outgrowth but not survival; the converse was true of FGF-9. Thus, differential effects of specific growth factors on survival or neurite outgrowth of bulbospinal neurons were distinguished.
Annals of Neurology, 1998
The adult mammalian forebrain harbors neuronal precursor cells in the subependymal zone (SZ). Neuronal progenitors also persist in the adult human SZ and have been cultured from epileptic temporal lobe. In the present study, we sought to identify these neural progenitors in situ, and to direct their expansion and neuronal differentiation in vitro. We prepared explants of adult human SZ, obtained from temporal lobe resections of refractory epileptics. The resultant cultures were treated with fibroblast growth factor-2 (FGF-2) for a week, with concurrent exposure to [3H]thymidine, then switched to media containing brain-derived neurotrophic factor (BDNF) for up to 2 months. Sporadic neuronal outgrowth, verified antigenically and physiologically, was observed from SZ cultures regardless of FGF-2IBDNF treatment; however, only FGF-2/BDNF-treated cultures exhibited profuse outgrowth, and these displayed neuronal survival as long as 9 weeks in vitro. In addition, cortical cultures derived from two brains generated microtubule-associated protein-2+ neurons, which incorporated [3H]thymidine and exhibited significant calcium increments to depolarization. In histological sections of the subependyma, both uncommitted and restricted progenitors, defined respectively by musashi and Hu protein expression, were identified. Thus, the adult human subependyma harbors neural progenitors, which are able to give rise to neurons whose numbers can be supported for prolonged periods in vitro.
Fibroblast growth factor-9 inhibits astrocyte differentiation of adult mouse neural progenitor cells
Journal of Neuroscience Research, 2009
Fibroblast growth factor-9 (FGF9) is expressed in the CNS and is reported to be a mitogen for glial cells, to promote neuronal survival, and to retard oligodendrocyte differentiation. Here we examined the effects of FGF9 on the differentiation, survival, and proliferation of adult neural progenitor cells derived from the adult mouse subventricular zone. FGF9 by itself induced neurosphere proliferation, but its effects were modest compared with those of epidermal growth factor and FGF2. When neurospheres were dissociated and plated for differentiation, FGF9 increased total cell number over time in a dose-dependent manner. Ki67 immunostaining and bromodeoxyuridine incorporation indicated that this was at least partially due to the continued presence of proliferative nestin-positive neural progenitor cells and bIII tubulin-positive neuronal precursors. FGF9 also promoted cell survival as indicated by a decreased number of TUNEL-positive cells over time. Assessment of differentiation showed that FGF9 increased neuron generation that reflected the increase in total cell number; however, the percentage of progenitor cells differentiating into neurons was slightly decreased. FGF9 had a modest effect on oligodendrocyte generation, although it appeared to slow the maturation of oligodenrocytes at higher concentrations. The most marked effect on differentiation was an almost total lack of glial fibrillary acidic protein (GFAP)-positive astrocytes up to 7 days following FGF9 addition, indicating that astrocyte differentiation was strongly inhibited. Total inhibition required prolonged treatment, although a 1-hr pulse was sufficient for partial inhibition, and bone morphogenic protein-4 could partially overcome the FGF9 inhibition of astrocyte differentiation. FGF9 therefore has multiple effects on adult neural precursor cell function, enhancing neuronal precursor proliferation and specifically inhibiting GFAP expression. V V C 2009 Wiley-Liss, Inc.
In Vitro Cellular & Developmental Biology - Animal, 2009
Serum-free mouse embryo (SFME) cells are an epidermal growth factor (EGF)-dependent established line derived from brains of 16-d-old Balb/c mouse embryos. SFME cells grow indefinitely in serum-free medium without replicative senescence, chromosomal abnormalities, or malignant transformation. SFME cells express nestin, a neural stem cell marker, under serum-free conditions. Exposure to serum or transforming growth factor β (TGF-β) leads to a marked increase in differentiation toward the astrocytic lineage with expression of glial fibrillary acidic protein and other astrocyte markers. In this study, we show that treatment of SFME cells with bone morphogenetic protein-4 (BMP-4), another member of the TGF-β family, led to differentiation toward a neuronal lineage under conditions of low mitogenic stimulation (0.5 ng/mL) by EGF and fibroblast growth factor. Maximum mitogenic stimulation with 50 ng/mL EGF blocked the BMP-4 effect on neuronal differentiation, but did not block TGF-β-induced expression of markers of the astrocytic lineage. BMP-4 treatment also enhanced the activity of the neuron-specific enolase (NSE) promoter in SFME-NSE-lacZ cells that carry the gene for bacterial β-galactosidase under the control of the NSE promoter. Extended BMP-4 treatment caused SFME cells to express a neuronal phenotype synthesizing gamma-aminobutyric acid. These results indicate that SFME cells have the capacity to generate both neurons and astrocytes in vitro, which resemble the behavior of EGF-dependent multipotential stem cells in the central nervous system, and establish a relationship between effects of BMP-4 and degree of mitogenic stimulation by other peptide growth factors.
Expression of fibroblast growth factor in the nervous system of early avian embryos
Development
Basic fibroblast growth factor (bFGF) promotes the survival of a subpopulation of non-neuronal cells developing from trunk neural crest. It was therefore important to determine whether this factor is present in the nervous system at early developmental stages. Immunocytochemistry using specific polyclonal and monoclonal antibodies was combined with three highly sensitive assays: bFGF-induced proliferation of bovine adrenal cortex-derived capillary endothelial cells (ACE), a radioimmunoassay for bFGF (RIA) and Western blot analysis. bFGF immunoreactivity was localized to the cytoplasm of neuroepithelial cells derived from embryonic day 2 (E2) quail neural tubes and cultured for one day in a chemically defined medium. Specific staining was observed in young sensory neurons in cultures of neural crest clusters as well as in a subpopulation of non-neuronal cells. In cultured E7 dorsal root ganglia, immunostaining was confined to neuronal cell bodies and fibers. In situ, staining of spin...