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. 2006 May 31;26(22):5996-6003.
doi: 10.1523/JNEUROSCI.5380-05.2006.

Matrix metalloproteinase 2 (MMP2) and MMP9 secreted by erythropoietin-activated endothelial cells promote neural progenitor cell migration

Lei Wang  1 , Zheng Gang Zhang, Rui Lan Zhang, Sara R Gregg, Ann Hozeska-Solgot, Yvonne LeTourneau, Ying Wang, Michael Chopp
Affiliations

Matrix metalloproteinase 2 (MMP2) and MMP9 secreted by erythropoietin-activated endothelial cells promote neural progenitor cell migration

Lei Wang et al. J Neurosci. .

Abstract

We investigated the hypothesis that endothelial cells activated by erythropoietin (EPO) promote the migration of neuroblasts. This hypothesis is based on observations in vivo that treatment of focal cerebral ischemia with EPO enhances the migration of neuroblasts to the ischemic boundary, a site containing activated endothelial cells and angiogenic microvasculature. To model the microenvironment within the ischemic boundary zone, we used a coculture system of mouse brain endothelial cells (MBECs) and neural progenitor cells derived from the subventricular zone of the adult mouse. Treatment of MBECs with recombinant human EPO (rhEPO) significantly increased secretion of matrix metalloproteinase 2 (MMP2) and MMP9. rhEPO-treated MBEC supernatant as conditioned medium significantly increased the migration of neural progenitor cells. Application of an MMP inhibitor abolished the supernatant-enhanced migration. Incubation of neurospheres alone with rhEPO failed to increase progenitor cell migration. rhEPO activated phosphatidylinositol 3-kinase/Akt (PI3K/Akt) and extracellular signal-regulated kinase (ERK1/2) in MBECs. Selective inhibition of the PI3K/Akt and ERK1/2 pathways significantly attenuated the rhEPO-induced MMP2 and MMP9, which suppressed neural progenitor cell migration promoted by the rhEPO-activated MBECs. Collectively, our data show that rhEPO-activated endothelial cells enhance neural progenitor cell migration by secreting MMP2 and MMP9 via the PI3K/Akt and ERK1/2 signaling pathways. These data demonstrate that activated endothelial cells can promote neural progenitor cell migration, and provide insight into the molecular mechanisms underlying the attraction of newly generated neurons to injured areas in brain.

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Figures

Figure 1.
Figure 1.
Effect of rhEPO on angiogenesis and neurogenesis. A is a hematoxylin and eosin (H&E)-stained coronal section showing ischemic core outlined by a white line. Three-dimensional confocal images (B–G) were acquired from a boxed area of the ipsilateral hemisphere in A, as indicated by arrows, whereas images H–J were from the homologous area of the contralateral hemisphere. Confocal images show EBA-immunoreactive cerebral microvessels (B, E, H) and DCX-positive migrating neuroblasts (C, F, I) of representative rats treated with rhEPO (B–D) or saline (E–J). D, G, and J are merged images of EBA-positive vessels (green) and DCX-positive cells (red). LV, Lateral ventricle; Str, striatum; CC; corpus callosum. Scale bar, 100 μm.
Figure 2.
Figure 2.
Effect of rhEPO on MBECs coupled neurosphere migration. Coculture of MBECs (red) and neurospheres (green) with rhEPO (epoietin α) at doses of 0 (A), 1 (B), 5 (C), and 10 U/ml (D). E–H are bright-field images corresponding to A–D. I and J are quantitative data of average distance and farthest distance of neural progenitor cell migration. Error bars indicate SEM. ∗p < 0.05 versus the control. Scale bar, 100 μm.
Figure 3.
Figure 3.
Effect of conditioned medium on neurosphere migration. A–E are images of migration of neural progenitor cells out of neurospheres cultured with conditioned medium harvested from MBECs (A), MBECs and rhEPO (B), MBECs and rhEPO in the presence of OA Hy (C), LY294002 (D), or U0126 (E). F and G are quantitative data of average distance and farthest distance of neural progenitor cell migration. ∗p < 0.05 versus the control group and #p < 0.05 versus the rhEPO group. H shows quantitative data of fold changes of neural progenitor cell migration by blind-well chamber assay. ∗p < 0.01 and #p < 0.01 versus control and rhEPO groups, respectively. Error bars indicate SEM. I and J are images of double immunofluorescent staining showing TuJ1 (red) and GFAP (green)-positive cells in neural progenitor cells cocultured with endothelial cells treated with (I) and without rhEPO (J). OA, OA Hy; GM, GM6001; LY, LY294002; U, U0126. Scale bar, 100 μm.
Figure 4.
Figure 4.
Effect of rhEPO on expression of MMP2 and MMP9. Real-time RT-PCR analysis (A) shows MMP2 and MMP9 mRNA levels in MBECs treated with rhEPO at concentrations of 0, 1, 5, and 10 U/ml. B shows quantitative data of MMP2 and MMP9 mRNA levels. GAPDH was used as an internal control. Error bars indicate SEM. ∗p < 0.05 versus the control group.
Figure 5.
Figure 5.
Effect of rhEPO on MMP2 and MMP9 secretion. Gelatin zymography analysis (A) shows MMP2 and MMP9 activities in the conditioned medium harvested from control, rhEPO (EPO), and rhEPO with LY294002 (EPO+LY) or U0126 (EPO+U). B shows quantitative analysis of MMP2 and MMP9. ∗p < 0.05 and #p < 0.05 versus control and rhEPO groups, respectively. Error bars indicate SEM.
Figure 6.
Figure 6.
Effects of rhEPO on the PI3K/Akt and ERK1/2 signaling pathways. Western blot analysis shows phospho-Akt, total Akt (A), phospho-ERK1/2 and total ERK1/2 (C) in control, rhEPO (EPO), rhEPO with LY294002 (EPO+LY) or wortmannin (EPO+W), and rhEPO with U0126 (EPO+U) or PD9321 (EPO+PD). B and D show quantitative analysis of phospho-Akt and phospho-ERK1/2, respectively. ∗p < 0.05 and #p < 0.05 versus control and rhEPO groups, respectively. Error bars indicate SEM.
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References

    1. Anagnostou A, Liu Z, Steiner M, Chin K, Lee ES, Kessimian N, Noguchi CT (1994). Erythropoietin receptor mRNA expression in human endothelial cells. Proc Natl Acad Sci USA 91:3974–3978. - PMC - PubMed
    1. Arcasoy MO, Jiang X (2005). Co-operative signalling mechanisms required for erythroid precursor expansion in response to erythropoietin and stem cell factor. Br J Haematol 130:121–129. - PubMed
    1. Arkell J, Jackson CJ (2003). Constitutive secretion of MMP9 by early-passage cultured human endothelial cells. Cell Biochem Funct 21:381–386. - PubMed
    1. Arvidsson A, Collin T, Kirik D, Kokaia Z, Lindvall O (2002). Neuronal replacement from endogenous precursors in the adult brain after stroke. Nat Med 8:963–970. - PubMed
    1. Aye MM, Ma C, Lin H, Bower KA, Wiggins RC, Luo J (2004). Ethanol-induced in vitro invasion of breast cancer cells: the contribution of MMP-2 by fibroblasts. Int J Cancer 112:738–746. - PubMed

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