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tPA Deficiency in Mice Leads to Rearrangement in the Cerebrovascular Tree and Cerebroventricular Malformations.

Stefanitsch C, Lawrence AL, Olverling A, Nilsson I, Fredriksson L - Front Cell Neurosci (2015)

Bottom Line: Our analysis demonstrates that life-long deficiency of tPA is associated with rearrangements in the cerebrovascular tree, including a reduction in the number of vascular smooth-muscle cell covered, large diameter, vessels and a decrease in vessel-associated PDGFRα expression as compared to wild-type (WT) littermate controls.In addition, we found that ablation of tPA results in an increased number of ERG-positive endothelial cells and increased junctional localization of the tight junction protein ZO1.In addition, we found that tPA (-/-) mice displayed mild cerebral ventricular malformations, a feature previously associated with ablation of PDGF-C, thereby providing an in vivo link between tPA and PDGF signaling in central nervous system (CNS) development.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Biochemistry and Biophysics, Division of Vascular Biology, Karolinska Institutet Stockholm, Sweden.

ABSTRACT
The serine protease tissue-type plasminogen activator (tPA) is used as a thrombolytic agent in the management of ischemic stroke, but concerns for hemorrhagic conversion greatly limits the number of patients that receive this treatment. It has been suggested that the bleeding complications associated with thrombolytic tPA may be due to unanticipated roles of tPA in the brain. Recent work has suggested tPA regulation of neurovascular barrier integrity, mediated via platelet derived growth factor (PDGF)-C/PDGF receptor-α (PDGFRα) signaling, as a possible molecular mechanism affecting the outcome of stroke. To better understand the role of tPA in neurovascular regulation we conducted a detailed analysis of the cerebrovasculature in brains from adult tPA deficient (tPA(-/-) ) mice. Our analysis demonstrates that life-long deficiency of tPA is associated with rearrangements in the cerebrovascular tree, including a reduction in the number of vascular smooth-muscle cell covered, large diameter, vessels and a decrease in vessel-associated PDGFRα expression as compared to wild-type (WT) littermate controls. In addition, we found that ablation of tPA results in an increased number of ERG-positive endothelial cells and increased junctional localization of the tight junction protein ZO1. This is intriguing since ERG is an endothelial transcription factor implicated in regulation of vascular integrity. Based on these results, we propose that the protection of barrier properties seen utilizing these tPA (-/-) mice might be due, at least in part, to these cerebrovascular rearrangements. In addition, we found that tPA (-/-) mice displayed mild cerebral ventricular malformations, a feature previously associated with ablation of PDGF-C, thereby providing an in vivo link between tPA and PDGF signaling in central nervous system (CNS) development. Taken together, the data presented here will advance our understanding of the role of tPA within the CNS and in regulation of cerebrovascular permeability.

No MeSH data available.


Related in: MedlinePlus

Normal basement membrane and astrocyte distribution in tPA−/− mice. Immunofluorescent staining of murine brain sections from tPA−/− mice (n = 5) and WT littermate controls (n = 5) was conducted with antibodies directed against (A) collagen IV, a marker of the basement membrane, (B) glial fibrillary acidic protein (GFAP), a marker for astrocytes, and (C) aquaporin 4 (AQP4), a marker of perivascular astrocytic endfeet. These analyses revealed normal distribution of basement membrane (A) and perivascular astrocytes (arrows, B,C) around similar sized vessels in tPA−/− mice and WT littermates. In addition, normal distribution of parenchymal astrocyte staining (arrowheads, B) was detected. The pictures are representative maximum intensity projections images generated from confocal Z-stacks from stainings on brain sections from WT and tPA−/− mice and the stainings have been repeated at least two independent times. Cell nuclei were visualized with DAPI and vessels with podocalyxin (Podo) or CD31. Scale bars 20 μm.
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Figure 4: Normal basement membrane and astrocyte distribution in tPA−/− mice. Immunofluorescent staining of murine brain sections from tPA−/− mice (n = 5) and WT littermate controls (n = 5) was conducted with antibodies directed against (A) collagen IV, a marker of the basement membrane, (B) glial fibrillary acidic protein (GFAP), a marker for astrocytes, and (C) aquaporin 4 (AQP4), a marker of perivascular astrocytic endfeet. These analyses revealed normal distribution of basement membrane (A) and perivascular astrocytes (arrows, B,C) around similar sized vessels in tPA−/− mice and WT littermates. In addition, normal distribution of parenchymal astrocyte staining (arrowheads, B) was detected. The pictures are representative maximum intensity projections images generated from confocal Z-stacks from stainings on brain sections from WT and tPA−/− mice and the stainings have been repeated at least two independent times. Cell nuclei were visualized with DAPI and vessels with podocalyxin (Podo) or CD31. Scale bars 20 μm.

Mentions: To determine whether tPA deficiency affects the basement membrane and/or perivascular astrocyte distribution we analyzed sections from tPA−/− and WT littermate control brains stained with antibodies directed against collagen IV, a marker of the basement membrane; GFAP, a marker for astrocytes; and aquaporin 4 (AQP4), a marker of perivascular astrocytic endfeet. These analyses revealed what appeared as normal distribution of basement membrane around similar sized vessels in tPA−/− mice (Figure 4A). Further, we detected similar distribution of GFAP+ astrocytes in the parenchyma (arrowheads) as well as GFAP+ perivascular astrocytes associated with comparable sized vessels (arrows) in the brains of tPA−/− mice relative to WT littermate controls (Figure 4B). This was supported by stainings of astrocytic endfeet with AQP4 showing apparently normal perivascular astrocyte coverage in vessels of comparable size (Figure 4C).


tPA Deficiency in Mice Leads to Rearrangement in the Cerebrovascular Tree and Cerebroventricular Malformations.

Stefanitsch C, Lawrence AL, Olverling A, Nilsson I, Fredriksson L - Front Cell Neurosci (2015)

Normal basement membrane and astrocyte distribution in tPA−/− mice. Immunofluorescent staining of murine brain sections from tPA−/− mice (n = 5) and WT littermate controls (n = 5) was conducted with antibodies directed against (A) collagen IV, a marker of the basement membrane, (B) glial fibrillary acidic protein (GFAP), a marker for astrocytes, and (C) aquaporin 4 (AQP4), a marker of perivascular astrocytic endfeet. These analyses revealed normal distribution of basement membrane (A) and perivascular astrocytes (arrows, B,C) around similar sized vessels in tPA−/− mice and WT littermates. In addition, normal distribution of parenchymal astrocyte staining (arrowheads, B) was detected. The pictures are representative maximum intensity projections images generated from confocal Z-stacks from stainings on brain sections from WT and tPA−/− mice and the stainings have been repeated at least two independent times. Cell nuclei were visualized with DAPI and vessels with podocalyxin (Podo) or CD31. Scale bars 20 μm.
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Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4663266&req=5

Figure 4: Normal basement membrane and astrocyte distribution in tPA−/− mice. Immunofluorescent staining of murine brain sections from tPA−/− mice (n = 5) and WT littermate controls (n = 5) was conducted with antibodies directed against (A) collagen IV, a marker of the basement membrane, (B) glial fibrillary acidic protein (GFAP), a marker for astrocytes, and (C) aquaporin 4 (AQP4), a marker of perivascular astrocytic endfeet. These analyses revealed normal distribution of basement membrane (A) and perivascular astrocytes (arrows, B,C) around similar sized vessels in tPA−/− mice and WT littermates. In addition, normal distribution of parenchymal astrocyte staining (arrowheads, B) was detected. The pictures are representative maximum intensity projections images generated from confocal Z-stacks from stainings on brain sections from WT and tPA−/− mice and the stainings have been repeated at least two independent times. Cell nuclei were visualized with DAPI and vessels with podocalyxin (Podo) or CD31. Scale bars 20 μm.
Mentions: To determine whether tPA deficiency affects the basement membrane and/or perivascular astrocyte distribution we analyzed sections from tPA−/− and WT littermate control brains stained with antibodies directed against collagen IV, a marker of the basement membrane; GFAP, a marker for astrocytes; and aquaporin 4 (AQP4), a marker of perivascular astrocytic endfeet. These analyses revealed what appeared as normal distribution of basement membrane around similar sized vessels in tPA−/− mice (Figure 4A). Further, we detected similar distribution of GFAP+ astrocytes in the parenchyma (arrowheads) as well as GFAP+ perivascular astrocytes associated with comparable sized vessels (arrows) in the brains of tPA−/− mice relative to WT littermate controls (Figure 4B). This was supported by stainings of astrocytic endfeet with AQP4 showing apparently normal perivascular astrocyte coverage in vessels of comparable size (Figure 4C).

Bottom Line: Our analysis demonstrates that life-long deficiency of tPA is associated with rearrangements in the cerebrovascular tree, including a reduction in the number of vascular smooth-muscle cell covered, large diameter, vessels and a decrease in vessel-associated PDGFRα expression as compared to wild-type (WT) littermate controls.In addition, we found that ablation of tPA results in an increased number of ERG-positive endothelial cells and increased junctional localization of the tight junction protein ZO1.In addition, we found that tPA (-/-) mice displayed mild cerebral ventricular malformations, a feature previously associated with ablation of PDGF-C, thereby providing an in vivo link between tPA and PDGF signaling in central nervous system (CNS) development.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Biochemistry and Biophysics, Division of Vascular Biology, Karolinska Institutet Stockholm, Sweden.

ABSTRACT
The serine protease tissue-type plasminogen activator (tPA) is used as a thrombolytic agent in the management of ischemic stroke, but concerns for hemorrhagic conversion greatly limits the number of patients that receive this treatment. It has been suggested that the bleeding complications associated with thrombolytic tPA may be due to unanticipated roles of tPA in the brain. Recent work has suggested tPA regulation of neurovascular barrier integrity, mediated via platelet derived growth factor (PDGF)-C/PDGF receptor-α (PDGFRα) signaling, as a possible molecular mechanism affecting the outcome of stroke. To better understand the role of tPA in neurovascular regulation we conducted a detailed analysis of the cerebrovasculature in brains from adult tPA deficient (tPA(-/-) ) mice. Our analysis demonstrates that life-long deficiency of tPA is associated with rearrangements in the cerebrovascular tree, including a reduction in the number of vascular smooth-muscle cell covered, large diameter, vessels and a decrease in vessel-associated PDGFRα expression as compared to wild-type (WT) littermate controls. In addition, we found that ablation of tPA results in an increased number of ERG-positive endothelial cells and increased junctional localization of the tight junction protein ZO1. This is intriguing since ERG is an endothelial transcription factor implicated in regulation of vascular integrity. Based on these results, we propose that the protection of barrier properties seen utilizing these tPA (-/-) mice might be due, at least in part, to these cerebrovascular rearrangements. In addition, we found that tPA (-/-) mice displayed mild cerebral ventricular malformations, a feature previously associated with ablation of PDGF-C, thereby providing an in vivo link between tPA and PDGF signaling in central nervous system (CNS) development. Taken together, the data presented here will advance our understanding of the role of tPA within the CNS and in regulation of cerebrovascular permeability.

No MeSH data available.


Related in: MedlinePlus