TBI causes loss of volume within the GCL of DG (29
). In this study, the volume decreased in both FGF-2–/–
mice, with FGF-2–/–
mice suffering a greater decrease by 35 days posttrauma (Figure , Table ). Furthermore, this volume loss corresponded to decreased numbers of granule cells, again with greater cell loss in FGF-2–/–
mice (Table ). We found that enhancing expression of FGF-2 preserved the integrity of GCL after TBI. There are at least two possible mechanisms: Production of new cells and reduction of cell loss over time (27
). Both may be operative in these experiments. We counted the number of cells in the GCL in FGF-2–/–
and WT mice (Table ). Thirty thousand fewer cells were lost in FGF-2+/+
animals compared with FGF-2–/–
mice. This would suggest that basal FGF-2 levels expressed in the WT can be neuroprotective. Interestingly, differences in TUNEL staining were not detected between groups 9 and 35 days after TBI, despite these group differences in cell loss. In addition, FGF-2 promoted neurogenesis in injured brain because 1100 more newly born neurons were found in FGF-2+/+
animals at 35 days after injury, whereas only 600 new neurons were detected in the KO. Thus, our data suggest that endogenously generated FGF-2 regulates cell populations through manipulating neurogenesis as well as attenuating neurodegeneration in the GCL after TBI.
The dual role of FGF-2 was also confirmed through gene transfer experiments. Increased levels of FGF-2 enhanced cell division in the SGZ by 2.2-fold compared with control vector (Figure ). At 5 weeks after TBI, FGF-2 vector yielded 2.5-fold more newborn granule neurons compared with the control vector (Table ), indicating that newly generated precursor cells proceed on to neuronal differentiation after FGF-2 gene transfer. Thus, neuronal production can be enhanced through upregulation of progenitor cell proliferation in the GCL after TBI by vector-mediated FGF-2 gene delivery. Moreover, posttraumatic overexpression of FGF-2 reduced the GCL volume loss by 10% (day 35) compared with animals injected with control vector (Figure ). These results suggest that acute treatment with HSV-1 amplicon vectors expressing FGF-2 after the onset of TBI can protect against the chronic, progressive volume loss in ipsilateral DG that occurs over several weeks after CCI. Furthermore, FGF-2 contributes to DG integrity after TBI by upregulating neurogenesis, based on the distribution pattern of newborn neurons labeled with BrdU. The total number of BrdU-labeled cells increased by gene transfer of FGF-2, and these cells migrated into the granule layer of DG and accumulated around the thinned part of the DG at day 35 (Figure ), presumably contributing to attenuation of volume loss of GCL at 35 days after TBI.
Although SGZ cell division was promoted after TBI compared with sham-operated mice (Figure b), FGF-2 levels in the ipsilateral hippocampus did not change after TBI. Normally, FGF-2 does not have a signal sequence for cell secretion through the endoplasmic reticulum/Golgi apparatus (47
), and it is probably released extracellularly only after cell damage. According to this hypothesis, it is speculated that FGF-2 plays a negligible role in adult neurogenesis in the normal state, and factors other than FGF-2 must contribute to regulation of proliferation and differentiation of neural progenitor cells in the DG. In fact, under basal conditions, there was no difference in dividing progenitor populations between FGF-2+/+
mice (Figure b), despite differences in FGF-2 levels in the hippocampus (15
). With respect to FGF-2 release, plasminogen activator-mediated proteolysis provides a mechanism for dissociation of biologically active FGF-2-heparan sulfate complexes from the extracellular matrix, rendering FGF-2 more biologically active (47
). Recently, TBI was shown to activate plasminogen activator (50
). We therefore speculate that TBI may promote FGF-2 dissociation from extracellular matrix and release from damaged cells. This phenomenon may account for the increase seen in SGZ cell division in FGF-2+/+
mice after TBI without measurable differences in FGF-2 levels from basal condition. Importantly, gene transfer of FGF-2
with an engineered secretion signal further enhanced the SGZ cell division in the injured DG with an increase in FGF-2 levels (Figure e). Thus, control of FGF-2 expression as well as its cellular release appears to be critically important in the regulation of progenitor cell proliferation after brain injury. In addition, injured tissue produces other factor(s) that convert latent FGF-2 to an active form (49
), consistent with a recent report suggesting that endogenous stimulatory signals are required for the action of growth factors potentiating ischemia-induced neurogenesis (31
In conclusion, we have shown that overexpression of FGF-2 increased neurogenesis in the adult hippocampus after TBI, whereas neurogenesis was reduced in FGF-2–deficient mice. The results obtained suggest that FGF-2 is, at least in part, responsible for regulating neuronal replacement, as well as attenuating neuronal loss after TBI. FGF-2 supplementation may provide a rational strategy to treat brain injury by simultaneously enhancing neurogenesis and reducing neurodegeneration. Insights into the functions of FGF-2, together with further understanding about other factors regulating regeneration and protection, should provide a strategy for repair of CNS injury after trauma, and for other CNS injuries and disorders, such as cerebral ischemia and neurodegenerative diseases.