Introduction
Neurogenesis occurs in the adult mammalian brain in two specialized geminal zones: the subventricular zone (produces new neurons destinated for the olfactory bulb) and the granular cell layer of the dentate gyrus of the hippocampus. Stroke leads to a marked increase of cell proliferation in the subventricular zone 1 . Moreover, the fate of BrdU positive cells is influenced by growth factors like epidermal growth factor (EGF), directing cells into the glial lineage and fibroblast growth factor (FGF), directing cells into the neural lineage 2 . However it is unclear whether this proliferation can be induced by growth factors in a model of mild ischemia and whether these factors can promote the neural lineage in this model. Therefore this study aimed at verifying the occurrence of post-stroke nerve cell division and influence of growth factors in a transient, endothelin-1- induced middle cerebral artery occlusion model (e-MCAO), and the number of cells, directing to the neural lineage in different areas of the post ischemic brain.
Materials
Male Sprague Dawley rats were subjected unilaterally to eMCAO by application of endothelin-1. After ischaemia a miniosmotic pump was implanted and the thymidine analogon bromodeoxyuridine (BrdU; 1 mg/ml), a marker of DNA synthesis that labels dividing cells and their terminal progeny, and BrdU + EGF/FGF was infused directly into the ventricle. After 12 days pumps were removed and after 14 days brains were perfusion-fixed and sections were processed for histochemistry for evaluation of the infarct area or immunofluorescence. Primary antibodies used were the specific markers anti-BrdU for newly generated cells and doublecortin for migrating cells. For detection and quantification we used confocal laser scanning microscopy.
Results
2 weeks after eMCAO we found BrdU-positive cells in the striatum, ventricle wall and the dentate gyrus of the hippocampus. EGF/FGF-2 had a promotional effect on the number of BrdU-positive and DCX-positive cells in the lateral ventricle, whereas ischemia itself had no effects on the BrdU-positive cells in the lateral ventricle, but in the striatum. Partly these cells are doublelabeled. Interestingly, the growth factor effect was inversed in the dentate gyrus.
Conclusions
Our results suggests: that EGF/FGF post-stroke treatment has a promotional effect on the number of BrdU-positive and DCX-positive cells in the ventricle wall and the striatum, whereas doblelabelling was only altered in the ipsilateral striatum. In the non-damaged hippocampus, the effect of growth factors attenuated cell proliferation and doublelabelling.