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Overexpression of CD44 in neural precursor cells improves trans-endothelial migration and facilitates their invasion of perivascular tissues in vivo.

Deboux C, Ladraa S, Cazaubon S, Ghribi-Mallah S, Weiss N, Chaverot N, Couraud PO, Baron-Van Evercooren A - PLoS ONE (2013)

Bottom Line: In view of the role of CD44 in NPCs trans-endothelial migration in vitro, we questioned presently the benefit of CD44 overexpression by NPCs in vitro and in vivo, in EAE mice.Moreover, CD44 overexpression by NPCs improved significantly their elongation, spreading and number of filopodia over the extracellular matrix protein laminin in vitro.We then tested the effect of CD44 overexpression after i.v. delivery in the tail vein of EAE mice.

View Article: PubMed Central - PubMed

Affiliation: Université Pierre et Marie Curie-Paris 6, Centre de Recherche de l'Institut du Cerveau et de la Moelle Epinière, UMR-S975, Paris, France.

ABSTRACT
Neural precursor (NPC) based therapies are used to restore neurons or oligodendrocytes and/or provide neuroprotection in a large variety of neurological diseases. In multiple sclerosis models, intravenously (i.v) -delivered NPCs reduced clinical signs via immunomodulation. We demonstrated recently that NPCs were able to cross cerebral endothelial cells in vitro and that the multifunctional signalling molecule, CD44 involved in trans-endothelial migration of lymphocytes to sites of inflammation, plays a crucial role in extravasation of syngeneic NPCs. In view of the role of CD44 in NPCs trans-endothelial migration in vitro, we questioned presently the benefit of CD44 overexpression by NPCs in vitro and in vivo, in EAE mice. We show that overexpression of CD44 by NPCs enhanced over 2 folds their trans-endothelial migration in vitro, without impinging on the proliferation or differentiation potential of the transduced cells. Moreover, CD44 overexpression by NPCs improved significantly their elongation, spreading and number of filopodia over the extracellular matrix protein laminin in vitro. We then tested the effect of CD44 overexpression after i.v. delivery in the tail vein of EAE mice. CD44 overexpression was functional invivo as it accelerated trans-endothelial migration and facilitated invasion of HA expressing perivascular sites. These in vitro and in vivo data suggest that CD44 may be crucial not only for NPC crossing the endothelial layer but also for facilitating invasion of extravascular tissues.

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Time-course distribution of NPCs after i.v. delivery in the mouse tail.(A–F) Detection of GFP(green) to localize grafted cells and of von Willebrand factor (VWF, red) to identify endothelial cells, reveals that NPCs are localized in or around the vascular wall at 6 h (A, B) and 12–24 h (C, D) post injection, but are more distant from the vascular wall at 21 days (E, F). (A, C, E) actin eGFP control-NPCs and (B, D, F) CD44-NPCs. CD44 NPCs become more rapidly elongated and distant from the vascular wall. Moreover, in D and F, GFP+ CD44-NPCs form a double wave, which is not observed in control NPCs. Arrows point to regions enlarged in insets on top of each panel.
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pone-0057430-g004: Time-course distribution of NPCs after i.v. delivery in the mouse tail.(A–F) Detection of GFP(green) to localize grafted cells and of von Willebrand factor (VWF, red) to identify endothelial cells, reveals that NPCs are localized in or around the vascular wall at 6 h (A, B) and 12–24 h (C, D) post injection, but are more distant from the vascular wall at 21 days (E, F). (A, C, E) actin eGFP control-NPCs and (B, D, F) CD44-NPCs. CD44 NPCs become more rapidly elongated and distant from the vascular wall. Moreover, in D and F, GFP+ CD44-NPCs form a double wave, which is not observed in control NPCs. Arrows point to regions enlarged in insets on top of each panel.

Mentions: Having established that CD44 overexpression in NPCs improves their spreading and trans-endothelial migration in vitro, we tested the functionality of CD44 overexpression in vivo. Actin-GFP CD44-NPCs and control actin-GFP NPCs (1×106 cells per mouse) were i.v-delivered in the tail vein of C57/Bl6 mice affected with MOG (35–55)-induced chronic EAE, 4 days after disease onset as previously reported (6, 12, 13). To asses the distribution of i.v injected NPCs, a detailed histological analysis of brain, spinal cord, lung, spleen and liver was performed at 6 h, 24–48 h and 21 days after grafting. We did not find i.v. injected NPCs in the brain or spinal cord at any time point. By contrast the majority of injected NPCs were detected at the injection point in the tail (Fig. 4, Fig. 5, Fig. 6 and Fig. S1). In this site, immunohistochemistry for von Willebrand factor to detect endothelial cells (Fig. 4) and for laminin to detect basal lamina (Fig. S1), showed that the majority of control and CD44 overexpressing NPCs were localized in proximity of blood vessels. At early time-points (at 6 or 24–48 h after injection), cells were mainly round in shape with no obvious differences in size or shape between the two groups of cells. They had in great majority crossed blood vessel walls with few NPCs still localized within the blood vessel wall (Fig. 4A, Fig. 7). By 21 days post injection, control (Fig. 4E) and CD44-NPCs (Fig. 4F), were no longer in contact with blood vessel walls but were clearly localized further in the parenchyma. CD44 NPCs were obviously more elongated in shape than control NPCs.


Overexpression of CD44 in neural precursor cells improves trans-endothelial migration and facilitates their invasion of perivascular tissues in vivo.

Deboux C, Ladraa S, Cazaubon S, Ghribi-Mallah S, Weiss N, Chaverot N, Couraud PO, Baron-Van Evercooren A - PLoS ONE (2013)

Time-course distribution of NPCs after i.v. delivery in the mouse tail.(A–F) Detection of GFP(green) to localize grafted cells and of von Willebrand factor (VWF, red) to identify endothelial cells, reveals that NPCs are localized in or around the vascular wall at 6 h (A, B) and 12–24 h (C, D) post injection, but are more distant from the vascular wall at 21 days (E, F). (A, C, E) actin eGFP control-NPCs and (B, D, F) CD44-NPCs. CD44 NPCs become more rapidly elongated and distant from the vascular wall. Moreover, in D and F, GFP+ CD44-NPCs form a double wave, which is not observed in control NPCs. Arrows point to regions enlarged in insets on top of each panel.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585392&req=5

pone-0057430-g004: Time-course distribution of NPCs after i.v. delivery in the mouse tail.(A–F) Detection of GFP(green) to localize grafted cells and of von Willebrand factor (VWF, red) to identify endothelial cells, reveals that NPCs are localized in or around the vascular wall at 6 h (A, B) and 12–24 h (C, D) post injection, but are more distant from the vascular wall at 21 days (E, F). (A, C, E) actin eGFP control-NPCs and (B, D, F) CD44-NPCs. CD44 NPCs become more rapidly elongated and distant from the vascular wall. Moreover, in D and F, GFP+ CD44-NPCs form a double wave, which is not observed in control NPCs. Arrows point to regions enlarged in insets on top of each panel.
Mentions: Having established that CD44 overexpression in NPCs improves their spreading and trans-endothelial migration in vitro, we tested the functionality of CD44 overexpression in vivo. Actin-GFP CD44-NPCs and control actin-GFP NPCs (1×106 cells per mouse) were i.v-delivered in the tail vein of C57/Bl6 mice affected with MOG (35–55)-induced chronic EAE, 4 days after disease onset as previously reported (6, 12, 13). To asses the distribution of i.v injected NPCs, a detailed histological analysis of brain, spinal cord, lung, spleen and liver was performed at 6 h, 24–48 h and 21 days after grafting. We did not find i.v. injected NPCs in the brain or spinal cord at any time point. By contrast the majority of injected NPCs were detected at the injection point in the tail (Fig. 4, Fig. 5, Fig. 6 and Fig. S1). In this site, immunohistochemistry for von Willebrand factor to detect endothelial cells (Fig. 4) and for laminin to detect basal lamina (Fig. S1), showed that the majority of control and CD44 overexpressing NPCs were localized in proximity of blood vessels. At early time-points (at 6 or 24–48 h after injection), cells were mainly round in shape with no obvious differences in size or shape between the two groups of cells. They had in great majority crossed blood vessel walls with few NPCs still localized within the blood vessel wall (Fig. 4A, Fig. 7). By 21 days post injection, control (Fig. 4E) and CD44-NPCs (Fig. 4F), were no longer in contact with blood vessel walls but were clearly localized further in the parenchyma. CD44 NPCs were obviously more elongated in shape than control NPCs.

Bottom Line: In view of the role of CD44 in NPCs trans-endothelial migration in vitro, we questioned presently the benefit of CD44 overexpression by NPCs in vitro and in vivo, in EAE mice.Moreover, CD44 overexpression by NPCs improved significantly their elongation, spreading and number of filopodia over the extracellular matrix protein laminin in vitro.We then tested the effect of CD44 overexpression after i.v. delivery in the tail vein of EAE mice.

View Article: PubMed Central - PubMed

Affiliation: Université Pierre et Marie Curie-Paris 6, Centre de Recherche de l'Institut du Cerveau et de la Moelle Epinière, UMR-S975, Paris, France.

ABSTRACT
Neural precursor (NPC) based therapies are used to restore neurons or oligodendrocytes and/or provide neuroprotection in a large variety of neurological diseases. In multiple sclerosis models, intravenously (i.v) -delivered NPCs reduced clinical signs via immunomodulation. We demonstrated recently that NPCs were able to cross cerebral endothelial cells in vitro and that the multifunctional signalling molecule, CD44 involved in trans-endothelial migration of lymphocytes to sites of inflammation, plays a crucial role in extravasation of syngeneic NPCs. In view of the role of CD44 in NPCs trans-endothelial migration in vitro, we questioned presently the benefit of CD44 overexpression by NPCs in vitro and in vivo, in EAE mice. We show that overexpression of CD44 by NPCs enhanced over 2 folds their trans-endothelial migration in vitro, without impinging on the proliferation or differentiation potential of the transduced cells. Moreover, CD44 overexpression by NPCs improved significantly their elongation, spreading and number of filopodia over the extracellular matrix protein laminin in vitro. We then tested the effect of CD44 overexpression after i.v. delivery in the tail vein of EAE mice. CD44 overexpression was functional invivo as it accelerated trans-endothelial migration and facilitated invasion of HA expressing perivascular sites. These in vitro and in vivo data suggest that CD44 may be crucial not only for NPC crossing the endothelial layer but also for facilitating invasion of extravascular tissues.

Show MeSH
Related in: MedlinePlus