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Mesenchymal stem cells stabilize the blood-brain barrier through regulation of astrocytes.

Park HJ, Shin JY, Kim HN, Oh SH, Song SK, Lee PH - Stem Cell Res Ther (2015)

Bottom Line: Consequently, MSC treatment reduced neutrophil infiltration and enhanced survival of midbrain dopaminergic neurons in LPS-treated animals.In cellular system, MSC treatment led to a significant reversion of VEGF-A-induced eNOS and tight junction protein expression in endothelial cells, which led to increased EBA expressing cells.Additionally, MSC treatment significantly attenuated LPS-induced increased expressions of IL-1β in microglia and VEGF-A in astrocytes with an increase in IL-10 levels.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology and Brain Research Institute, Yonsei University College of Medicine, 250 Seongsanno, Seodaemun-gu, Seoul, 120-752, South Korea. pphj0105@yuhs.ac.

ABSTRACT

Introduction: The blood-brain barrier (BBB) protects the brain against potentially neurotoxic molecules in the circulation, and loss of its integrity may contribute to disease progression in neurodegenerative conditions. Recently, the active role of reactive astrocytes in BBB disruption has become evident in the inflamed brain. In the present study, we investigated whether mesenchymal stem cell (MSC) treatment might modulate reactive astrocytes and thus stabilize BBB integrity through vascular endothelial growth factor A (VEGF-A) signaling in inflammatory conditions.

Methods: For the inflamed brain, we injected LPS using a stereotaxic apparatus and MSCs were injected into the tail vein. At 6 hours and 7 days after LPS injection, we analyzed modulatory effects of MSCs on the change of BBB permeability through VEGF-A signaling using immunochemistry and western blot. To determine the effects of MSCs on VEGF-A-related signaling in cellular system, we had used endothelial cells treated with VEGF-A and co-cultured astrocyte and BV 2 cells treated with lipopolysaccharide (LPS) and then these cells were co-cultured with MSCs.

Results: In LPS-treated rats, MSCs restored Evans blue infiltration and the number of endothelial-barrier antigen (EBA) and P-glycoprotein (p-gp)-expressing cells, which were significantly altered in LPS-treated animals. Additionally, MSC administration following LPS treatment markedly increased the density of astrocytic filaments around vessels and reversed LPS-induced elevations in VEGF-A levels as well as endothelial nitric oxide synthase (eNOS)-dependent downregulation of tight junction proteins in the endothelium. Consequently, MSC treatment reduced neutrophil infiltration and enhanced survival of midbrain dopaminergic neurons in LPS-treated animals. In cellular system, MSC treatment led to a significant reversion of VEGF-A-induced eNOS and tight junction protein expression in endothelial cells, which led to increased EBA expressing cells. Additionally, MSC treatment significantly attenuated LPS-induced increased expressions of IL-1β in microglia and VEGF-A in astrocytes with an increase in IL-10 levels.

Conclusion: The present study indicated that MSCs may stabilize BBB permeability by modulating astrocytic endfeet and VEGF-A signaling, which may be relevant to the treatment of Parkinsonian diseases as a candidate for disease modifying therapeutics.

No MeSH data available.


Related in: MedlinePlus

MSCs modulate VEGF-A-regulated tight junction protein expression in bEnd.3 cells. Western blots revealed that VEGF-A treatment significantly increased eNOS expression and decreased CLN-5 expression in bEnd.3 cells; however, the expressions of these proteins were reverted to control levels after MSC treatment a (n = 3). *p <0.05, **p <0.01. Immunocytochemical analysis showed that MSC treatment attenuated VEGF-A-induced increases in eNOS immunoreactivity and downregulation of CLN-5 immunoreactivity b, c. Scale bar: 20 μm. In addition, the number of detected EBA-expressing cells was significantly increased in MSC-treated cells relative to those treated only with VEGF-A d, e (n = 3). ***p <0.001. Scale bar: 100 μm. CLN-5 claudin-5, DAPI 4',6-diamidino-2-phenylindole, EBA endothelial-barrier antigen, eNOS endothelial nitric oxide synthase, LPS lipopolysaccharide, MSC mesenchymal stem cell, VEGF vascular endothelial growth factor
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Fig5: MSCs modulate VEGF-A-regulated tight junction protein expression in bEnd.3 cells. Western blots revealed that VEGF-A treatment significantly increased eNOS expression and decreased CLN-5 expression in bEnd.3 cells; however, the expressions of these proteins were reverted to control levels after MSC treatment a (n = 3). *p <0.05, **p <0.01. Immunocytochemical analysis showed that MSC treatment attenuated VEGF-A-induced increases in eNOS immunoreactivity and downregulation of CLN-5 immunoreactivity b, c. Scale bar: 20 μm. In addition, the number of detected EBA-expressing cells was significantly increased in MSC-treated cells relative to those treated only with VEGF-A d, e (n = 3). ***p <0.001. Scale bar: 100 μm. CLN-5 claudin-5, DAPI 4',6-diamidino-2-phenylindole, EBA endothelial-barrier antigen, eNOS endothelial nitric oxide synthase, LPS lipopolysaccharide, MSC mesenchymal stem cell, VEGF vascular endothelial growth factor

Mentions: To determine the effects of MSCs on VEGF-A-related signaling in a cellular system, endothelial cells were treated with VEGF-A and then co-cultured with MSCs. Western blots revealed that VEGF-A treatment significantly increased eNOS expression and decreased CLN-5 expression in bEnd.3 cells; however, the expressions of these proteins were reverted to control levels after MSC treatment (Fig. 5a). This result was further confirmed by immunocytochemical analysis, which showed that MSC treatment attenuated VEGF-A-induced increases in eNOS immunoreactivity and downregulation of CLN-5 immunoreactivity (Fig. 5b, c). Additionally, we assessed survival of endothelial cells after VEGF-A treatment using EBA immunocytochemistry. Whereas VEGF-A treatment decreased EBA immunoreactivity, MSC treatment in these cells markedly increased EBA-positive cells (Fig. 5d). Stereological analysis revealed that the number of detected EBA-positive cells was significantly increased in MSC-treated cells relative to those treated only with VEGF-A (p <0.001; Fig. 5d).Fig. 5


Mesenchymal stem cells stabilize the blood-brain barrier through regulation of astrocytes.

Park HJ, Shin JY, Kim HN, Oh SH, Song SK, Lee PH - Stem Cell Res Ther (2015)

MSCs modulate VEGF-A-regulated tight junction protein expression in bEnd.3 cells. Western blots revealed that VEGF-A treatment significantly increased eNOS expression and decreased CLN-5 expression in bEnd.3 cells; however, the expressions of these proteins were reverted to control levels after MSC treatment a (n = 3). *p <0.05, **p <0.01. Immunocytochemical analysis showed that MSC treatment attenuated VEGF-A-induced increases in eNOS immunoreactivity and downregulation of CLN-5 immunoreactivity b, c. Scale bar: 20 μm. In addition, the number of detected EBA-expressing cells was significantly increased in MSC-treated cells relative to those treated only with VEGF-A d, e (n = 3). ***p <0.001. Scale bar: 100 μm. CLN-5 claudin-5, DAPI 4',6-diamidino-2-phenylindole, EBA endothelial-barrier antigen, eNOS endothelial nitric oxide synthase, LPS lipopolysaccharide, MSC mesenchymal stem cell, VEGF vascular endothelial growth factor
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4588687&req=5

Fig5: MSCs modulate VEGF-A-regulated tight junction protein expression in bEnd.3 cells. Western blots revealed that VEGF-A treatment significantly increased eNOS expression and decreased CLN-5 expression in bEnd.3 cells; however, the expressions of these proteins were reverted to control levels after MSC treatment a (n = 3). *p <0.05, **p <0.01. Immunocytochemical analysis showed that MSC treatment attenuated VEGF-A-induced increases in eNOS immunoreactivity and downregulation of CLN-5 immunoreactivity b, c. Scale bar: 20 μm. In addition, the number of detected EBA-expressing cells was significantly increased in MSC-treated cells relative to those treated only with VEGF-A d, e (n = 3). ***p <0.001. Scale bar: 100 μm. CLN-5 claudin-5, DAPI 4',6-diamidino-2-phenylindole, EBA endothelial-barrier antigen, eNOS endothelial nitric oxide synthase, LPS lipopolysaccharide, MSC mesenchymal stem cell, VEGF vascular endothelial growth factor
Mentions: To determine the effects of MSCs on VEGF-A-related signaling in a cellular system, endothelial cells were treated with VEGF-A and then co-cultured with MSCs. Western blots revealed that VEGF-A treatment significantly increased eNOS expression and decreased CLN-5 expression in bEnd.3 cells; however, the expressions of these proteins were reverted to control levels after MSC treatment (Fig. 5a). This result was further confirmed by immunocytochemical analysis, which showed that MSC treatment attenuated VEGF-A-induced increases in eNOS immunoreactivity and downregulation of CLN-5 immunoreactivity (Fig. 5b, c). Additionally, we assessed survival of endothelial cells after VEGF-A treatment using EBA immunocytochemistry. Whereas VEGF-A treatment decreased EBA immunoreactivity, MSC treatment in these cells markedly increased EBA-positive cells (Fig. 5d). Stereological analysis revealed that the number of detected EBA-positive cells was significantly increased in MSC-treated cells relative to those treated only with VEGF-A (p <0.001; Fig. 5d).Fig. 5

Bottom Line: Consequently, MSC treatment reduced neutrophil infiltration and enhanced survival of midbrain dopaminergic neurons in LPS-treated animals.In cellular system, MSC treatment led to a significant reversion of VEGF-A-induced eNOS and tight junction protein expression in endothelial cells, which led to increased EBA expressing cells.Additionally, MSC treatment significantly attenuated LPS-induced increased expressions of IL-1β in microglia and VEGF-A in astrocytes with an increase in IL-10 levels.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology and Brain Research Institute, Yonsei University College of Medicine, 250 Seongsanno, Seodaemun-gu, Seoul, 120-752, South Korea. pphj0105@yuhs.ac.

ABSTRACT

Introduction: The blood-brain barrier (BBB) protects the brain against potentially neurotoxic molecules in the circulation, and loss of its integrity may contribute to disease progression in neurodegenerative conditions. Recently, the active role of reactive astrocytes in BBB disruption has become evident in the inflamed brain. In the present study, we investigated whether mesenchymal stem cell (MSC) treatment might modulate reactive astrocytes and thus stabilize BBB integrity through vascular endothelial growth factor A (VEGF-A) signaling in inflammatory conditions.

Methods: For the inflamed brain, we injected LPS using a stereotaxic apparatus and MSCs were injected into the tail vein. At 6 hours and 7 days after LPS injection, we analyzed modulatory effects of MSCs on the change of BBB permeability through VEGF-A signaling using immunochemistry and western blot. To determine the effects of MSCs on VEGF-A-related signaling in cellular system, we had used endothelial cells treated with VEGF-A and co-cultured astrocyte and BV 2 cells treated with lipopolysaccharide (LPS) and then these cells were co-cultured with MSCs.

Results: In LPS-treated rats, MSCs restored Evans blue infiltration and the number of endothelial-barrier antigen (EBA) and P-glycoprotein (p-gp)-expressing cells, which were significantly altered in LPS-treated animals. Additionally, MSC administration following LPS treatment markedly increased the density of astrocytic filaments around vessels and reversed LPS-induced elevations in VEGF-A levels as well as endothelial nitric oxide synthase (eNOS)-dependent downregulation of tight junction proteins in the endothelium. Consequently, MSC treatment reduced neutrophil infiltration and enhanced survival of midbrain dopaminergic neurons in LPS-treated animals. In cellular system, MSC treatment led to a significant reversion of VEGF-A-induced eNOS and tight junction protein expression in endothelial cells, which led to increased EBA expressing cells. Additionally, MSC treatment significantly attenuated LPS-induced increased expressions of IL-1β in microglia and VEGF-A in astrocytes with an increase in IL-10 levels.

Conclusion: The present study indicated that MSCs may stabilize BBB permeability by modulating astrocytic endfeet and VEGF-A signaling, which may be relevant to the treatment of Parkinsonian diseases as a candidate for disease modifying therapeutics.

No MeSH data available.


Related in: MedlinePlus