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Pentraxin 3 mediates neurogenesis and angiogenesis after cerebral ischaemia.

Rodriguez-Grande B, Varghese L, Molina-Holgado F, Rajkovic O, Garlanda C, Denes A, Pinteaux E - J Neuroinflammation (2015)

Bottom Line: In vitro, recombinant PTX3 induced marked endothelial cellular proliferation and promoted formation of tube-like structures of endothelial cell line bEnd.5.Finally, a lack of PTX3 potentiated motor deficits 14 days after MCAo.These results indicate that PTX3 mediates neurogenesis and angiogenesis and contributes to functional recovery after stroke, highlighting a key role of PTX3 as a mediator of brain repair and suggesting that PTX3 could be used as a new target for stroke therapy.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Life Sciences, A,V, Hill Building, University of Manchester, Oxford Road, Manchester M13 9PT, UK. emmanuel.pinteaux@manchester.ac.uk.

ABSTRACT

Background: The acute phase protein pentraxin 3 (PTX3) is a new biomarker of stroke severity and is a key regulator of oedema resolution and glial responses after cerebral ischaemia, emerging as a possible target for brain repair after stroke. Neurogenesis and angiogenesis are essential events in post-stroke recovery. Here, we investigated for the first time the role of PTX3 in neurogenesis and angiogenesis after stroke.

Methods: PTX3 knockout (KO) or wild-type (WT) mice were subjected to experimental cerebral ischaemia (induced by middle cerebral artery occlusion (MCAo)). Poststroke neurogenesis was assessed by nestin, doublecortin (DCX) and bromodeoxyuridine (BrdU) immunostaining, whereas angiogenesis was assessed by BrdU, vascular endothelial growth factor receptor 2 (VEGFR2) and PECAM-1 immunostaining. In vitro neurogenesis and angiogenesis assays were carried out on neurospheres derived from WT or interleukin-1β (IL-1β) KO mice, and mouse endothelial cell line bEnd.5 respectively. Behavioural function was assessed in WT and PTX3 KO mice using open-field, motor and Y-maze tests.

Results: Neurogenesis was significantly reduced in the dentate gyrus (DG) of the hippocampus of PTX3 KO mice, compared to WT mice, 6 days after MCAo. In addition, recombinant PTX3 was neurogenic in vitro when added to neurospheres, which was mediated by IL-1β. In vivo poststroke angiogenesis was significantly reduced in PTX3 KO mice compared to WT mice 14 days after MCAo, as revealed by reduced vascular density, less newly formed blood vessels and decreased expression of VEGFR2. In vitro, recombinant PTX3 induced marked endothelial cellular proliferation and promoted formation of tube-like structures of endothelial cell line bEnd.5. Finally, a lack of PTX3 potentiated motor deficits 14 days after MCAo.

Conclusions: These results indicate that PTX3 mediates neurogenesis and angiogenesis and contributes to functional recovery after stroke, highlighting a key role of PTX3 as a mediator of brain repair and suggesting that PTX3 could be used as a new target for stroke therapy.

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Lack of PTX3 impairs angiogenesis 14 days after middle cerebral artery occlusion(MCAo). Proliferating vessels (indicated by white arrows in (A) and (D)) are seen in the striatum 6 days (A) and 14 days (D) after MCAo. Angiogenesis is similar for both genotypes 6 days after MCAo (B), but PTX3 knockout (KO) have less striatal proliferating vessels than wild-type (WT) mice 14 days after MCAo (E), even after normalisation with the total amount of PECAM-1 (E) and (F). Scale bars = 50 μm. Students t-test (n = 4) was used. *P <0.05. Error bars show SEM.
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Fig3: Lack of PTX3 impairs angiogenesis 14 days after middle cerebral artery occlusion(MCAo). Proliferating vessels (indicated by white arrows in (A) and (D)) are seen in the striatum 6 days (A) and 14 days (D) after MCAo. Angiogenesis is similar for both genotypes 6 days after MCAo (B), but PTX3 knockout (KO) have less striatal proliferating vessels than wild-type (WT) mice 14 days after MCAo (E), even after normalisation with the total amount of PECAM-1 (E) and (F). Scale bars = 50 μm. Students t-test (n = 4) was used. *P <0.05. Error bars show SEM.

Mentions: Since neurogenesis is coupled with angiogenesis in the injured brain [15], we tested whether PTX3 also promotes angiogenesis after experimental stroke. Lack of PTX3 did not significantly affect the levels of PECAM-1 staining 48 h (Figure 2A) or 6 days (Figure 2B) after MCAo, although there was a trend towards decreased PECAM-1 staining in PTX3 KO mice. In contrast, PECAM-1 staining was significantly reduced in PTX3 KO mice 14 days after MCAo, compared to WT mice (Figure 2C). To confirm that this variation in PECAM-1 staining was due to angiogenesis, we quantified the amount of BrdU incorporated within the vascular endothelium. After 6 days (Figure 3A,B and C) and 14 days (Figure 3D,E and F) post-MCAo, several blood vessels had incorporated BrdU. Lack of PTX3 significantly affected the amount of proliferating vessels 14 days (Figure 3E) but not 6 days (Figure 3B) after MCAo. To determine if the reduced amount of vascular BrdU was only due to the fact that PTX3 KO mice had fewer blood vessels, we normalised the amount of BrdU to the amount of PECAM-1 staining (Figures 3C and F). This also indicated a significant deficit in angiogenesis in PTX3 KO mice 14 days after MCAo. In addition to fewer proliferating vessels, PTX3 KO mice had less vascular VEGFR2 staining 14 days after MCAo (Figure 4A), both as percentage of the total area (Figure 4B) and when normalised and expressed as percentage of the PECAM-1-positive area (Figure 4C). VEGF exerts a potent pro-angiogenic effect through the activation of VEGFR2 [29], whose levels increase after ischaemia [30].Figure 2


Pentraxin 3 mediates neurogenesis and angiogenesis after cerebral ischaemia.

Rodriguez-Grande B, Varghese L, Molina-Holgado F, Rajkovic O, Garlanda C, Denes A, Pinteaux E - J Neuroinflammation (2015)

Lack of PTX3 impairs angiogenesis 14 days after middle cerebral artery occlusion(MCAo). Proliferating vessels (indicated by white arrows in (A) and (D)) are seen in the striatum 6 days (A) and 14 days (D) after MCAo. Angiogenesis is similar for both genotypes 6 days after MCAo (B), but PTX3 knockout (KO) have less striatal proliferating vessels than wild-type (WT) mice 14 days after MCAo (E), even after normalisation with the total amount of PECAM-1 (E) and (F). Scale bars = 50 μm. Students t-test (n = 4) was used. *P <0.05. Error bars show SEM.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4308938&req=5

Fig3: Lack of PTX3 impairs angiogenesis 14 days after middle cerebral artery occlusion(MCAo). Proliferating vessels (indicated by white arrows in (A) and (D)) are seen in the striatum 6 days (A) and 14 days (D) after MCAo. Angiogenesis is similar for both genotypes 6 days after MCAo (B), but PTX3 knockout (KO) have less striatal proliferating vessels than wild-type (WT) mice 14 days after MCAo (E), even after normalisation with the total amount of PECAM-1 (E) and (F). Scale bars = 50 μm. Students t-test (n = 4) was used. *P <0.05. Error bars show SEM.
Mentions: Since neurogenesis is coupled with angiogenesis in the injured brain [15], we tested whether PTX3 also promotes angiogenesis after experimental stroke. Lack of PTX3 did not significantly affect the levels of PECAM-1 staining 48 h (Figure 2A) or 6 days (Figure 2B) after MCAo, although there was a trend towards decreased PECAM-1 staining in PTX3 KO mice. In contrast, PECAM-1 staining was significantly reduced in PTX3 KO mice 14 days after MCAo, compared to WT mice (Figure 2C). To confirm that this variation in PECAM-1 staining was due to angiogenesis, we quantified the amount of BrdU incorporated within the vascular endothelium. After 6 days (Figure 3A,B and C) and 14 days (Figure 3D,E and F) post-MCAo, several blood vessels had incorporated BrdU. Lack of PTX3 significantly affected the amount of proliferating vessels 14 days (Figure 3E) but not 6 days (Figure 3B) after MCAo. To determine if the reduced amount of vascular BrdU was only due to the fact that PTX3 KO mice had fewer blood vessels, we normalised the amount of BrdU to the amount of PECAM-1 staining (Figures 3C and F). This also indicated a significant deficit in angiogenesis in PTX3 KO mice 14 days after MCAo. In addition to fewer proliferating vessels, PTX3 KO mice had less vascular VEGFR2 staining 14 days after MCAo (Figure 4A), both as percentage of the total area (Figure 4B) and when normalised and expressed as percentage of the PECAM-1-positive area (Figure 4C). VEGF exerts a potent pro-angiogenic effect through the activation of VEGFR2 [29], whose levels increase after ischaemia [30].Figure 2

Bottom Line: In vitro, recombinant PTX3 induced marked endothelial cellular proliferation and promoted formation of tube-like structures of endothelial cell line bEnd.5.Finally, a lack of PTX3 potentiated motor deficits 14 days after MCAo.These results indicate that PTX3 mediates neurogenesis and angiogenesis and contributes to functional recovery after stroke, highlighting a key role of PTX3 as a mediator of brain repair and suggesting that PTX3 could be used as a new target for stroke therapy.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Life Sciences, A,V, Hill Building, University of Manchester, Oxford Road, Manchester M13 9PT, UK. emmanuel.pinteaux@manchester.ac.uk.

ABSTRACT

Background: The acute phase protein pentraxin 3 (PTX3) is a new biomarker of stroke severity and is a key regulator of oedema resolution and glial responses after cerebral ischaemia, emerging as a possible target for brain repair after stroke. Neurogenesis and angiogenesis are essential events in post-stroke recovery. Here, we investigated for the first time the role of PTX3 in neurogenesis and angiogenesis after stroke.

Methods: PTX3 knockout (KO) or wild-type (WT) mice were subjected to experimental cerebral ischaemia (induced by middle cerebral artery occlusion (MCAo)). Poststroke neurogenesis was assessed by nestin, doublecortin (DCX) and bromodeoxyuridine (BrdU) immunostaining, whereas angiogenesis was assessed by BrdU, vascular endothelial growth factor receptor 2 (VEGFR2) and PECAM-1 immunostaining. In vitro neurogenesis and angiogenesis assays were carried out on neurospheres derived from WT or interleukin-1β (IL-1β) KO mice, and mouse endothelial cell line bEnd.5 respectively. Behavioural function was assessed in WT and PTX3 KO mice using open-field, motor and Y-maze tests.

Results: Neurogenesis was significantly reduced in the dentate gyrus (DG) of the hippocampus of PTX3 KO mice, compared to WT mice, 6 days after MCAo. In addition, recombinant PTX3 was neurogenic in vitro when added to neurospheres, which was mediated by IL-1β. In vivo poststroke angiogenesis was significantly reduced in PTX3 KO mice compared to WT mice 14 days after MCAo, as revealed by reduced vascular density, less newly formed blood vessels and decreased expression of VEGFR2. In vitro, recombinant PTX3 induced marked endothelial cellular proliferation and promoted formation of tube-like structures of endothelial cell line bEnd.5. Finally, a lack of PTX3 potentiated motor deficits 14 days after MCAo.

Conclusions: These results indicate that PTX3 mediates neurogenesis and angiogenesis and contributes to functional recovery after stroke, highlighting a key role of PTX3 as a mediator of brain repair and suggesting that PTX3 could be used as a new target for stroke therapy.

Show MeSH
Related in: MedlinePlus