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Role of P2X7 and P2Y2 receptors on α-secretase-dependent APP processing: Control of amyloid plaques formation "in vivo" by P2X7 receptor.

Miras-Portugal MT, Diaz-Hernandez JI, Gomez-Villafuertes R, Diaz-Hernandez M, Artalejo AR, Gualix J - Comput Struct Biotechnol J (2015)

Bottom Line: In Neuro-2a, a neuroblastoma cell line that constitutively expresses APP, activation of the P2X7 receptor leads to reduction of α-secretase activity, the opposite effect being obtained by P2Y2 receptor activation.In vivo inhibition of the P2X7 receptor induced a significant decrease in the number and size of hippocampal amyloid plaques.This reduction is mediated by an increase in the proteolytic processing of APP through α-secretase activity, which correlates with an increase in the phosphorylated form of GSK-3, a less active form of this enzyme.

View Article: PubMed Central - PubMed

Affiliation: Biochemistry Department, School of Veterinary Sciences, Complutense University of Madrid, Madrid, Spain, Institute of Neurochemistry (IUIN), Complutense University of Madrid, Madrid, Spain.

ABSTRACT
Amyloid precursor protein (APP) is expressed in a large variety of neural and non-neural cells. The balance between non-pathogenic and pathologic forms of APP processing, mediated by α-secretase and β-secretase respectively, remains a crucial step to understand β-amyloid, Aβ42 peptide, formation and aggregation that are at the origin of the senile plaques in the brain, a characteristic hallmark of Alzheimer's disease (AD). In Neuro-2a, a neuroblastoma cell line that constitutively expresses APP, activation of the P2X7 receptor leads to reduction of α-secretase activity, the opposite effect being obtained by P2Y2 receptor activation. The in vivo approach was made possible by the use of J20 mice, a transgenic mouse model of familial Alzheimer's disease (FAD) expressing human APP mutant protein. This animal exhibits prominent amyloid plaques by six months of age. In vivo inhibition of the P2X7 receptor induced a significant decrease in the number and size of hippocampal amyloid plaques. This reduction is mediated by an increase in the proteolytic processing of APP through α-secretase activity, which correlates with an increase in the phosphorylated form of GSK-3, a less active form of this enzyme. The in vivo findings corroborate the therapeutic potential of P2X7 antagonists in the treatment of FAD.

No MeSH data available.


Related in: MedlinePlus

Brilliant Blue-G (BBG) treatment reduces the number of amyloid plaques and microglia in the J20 mouse hippocampus. (A) Immunostaining of WO2 in hippocampal slices from 6 to 8 months-old J20 mice injected intraperitoneal with vehicle or BBG. Scale bar, 500 μm. (B) Histogram represents the mean ± SEM of Aβ-amyloid plaques per slice in the hippocampus of J20 mice treated with vehicle or BBG, being 16 slices per mouse (n = 7 mice per condition). ***p < 0.005, unpaired Student's t test. (C) Immunostaining of microglial marker Iba-1 in hippocampal slices from 6 to 8 months-old J20 mice treated with vehicle or BBG. Scale bar, 500 μm. (D) Quantification of microglial cells in the hippocampus of J20 mice treated with vehicle or BBG. Histograms represent the mean ± SEM of microglial cells per hippocampal area of 0.1 mm2 being 16 slices per mouse (n = 7 mice per treatment). For methods see Ref. [49].
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f0020: Brilliant Blue-G (BBG) treatment reduces the number of amyloid plaques and microglia in the J20 mouse hippocampus. (A) Immunostaining of WO2 in hippocampal slices from 6 to 8 months-old J20 mice injected intraperitoneal with vehicle or BBG. Scale bar, 500 μm. (B) Histogram represents the mean ± SEM of Aβ-amyloid plaques per slice in the hippocampus of J20 mice treated with vehicle or BBG, being 16 slices per mouse (n = 7 mice per condition). ***p < 0.005, unpaired Student's t test. (C) Immunostaining of microglial marker Iba-1 in hippocampal slices from 6 to 8 months-old J20 mice treated with vehicle or BBG. Scale bar, 500 μm. (D) Quantification of microglial cells in the hippocampus of J20 mice treated with vehicle or BBG. Histograms represent the mean ± SEM of microglial cells per hippocampal area of 0.1 mm2 being 16 slices per mouse (n = 7 mice per treatment). For methods see Ref. [49].

Mentions: After BBG treatment the number and size of amyloid plaques at the hippocampal structures of J20 mice were significantly reduced compared to their littermates treated with vehicle, as shown in Fig. 4A and B. In addition, the treatment with BBG did not significantly modify either the P2X7 receptor or murine APP and human APP mRNA expression. The levels of these proteins and their distribution patterns in the hippocampus were also not modified by the treatment with BBG, as demonstrated by western blot and immunohistochemical studies. However, a dramatic change was observed concerning the pattern of the C83 and C99 peptides, which correspond to the carboxyterminal fragments generated by APP cleavage by α-secretase and β-secretase, respectively. C99 fragment was under the limits of detection in wild type mouse brain, but was very abundant in the brain of J20 mice, which exhibit a much lower concentration of the α-secretase generated C83 fragment. However, BBG reversed the situation and a significant increase in the C83 fragment was achieved in the brain of BBG-treated J20 mice [48]. This situation mimics the APP processing pattern observed in N2a cells treated with P2X7 receptor inhibitors. On the other hand, an increase in the phosphorylated form of GSK-3 was observed in the hippocampus of BBG-treated J20 mice, when compared with vehicle-treated animals. Thus, BBG was able to reduce hippocampal GSK-3 activity in FAD animal models in the same way as in N2a cells.


Role of P2X7 and P2Y2 receptors on α-secretase-dependent APP processing: Control of amyloid plaques formation "in vivo" by P2X7 receptor.

Miras-Portugal MT, Diaz-Hernandez JI, Gomez-Villafuertes R, Diaz-Hernandez M, Artalejo AR, Gualix J - Comput Struct Biotechnol J (2015)

Brilliant Blue-G (BBG) treatment reduces the number of amyloid plaques and microglia in the J20 mouse hippocampus. (A) Immunostaining of WO2 in hippocampal slices from 6 to 8 months-old J20 mice injected intraperitoneal with vehicle or BBG. Scale bar, 500 μm. (B) Histogram represents the mean ± SEM of Aβ-amyloid plaques per slice in the hippocampus of J20 mice treated with vehicle or BBG, being 16 slices per mouse (n = 7 mice per condition). ***p < 0.005, unpaired Student's t test. (C) Immunostaining of microglial marker Iba-1 in hippocampal slices from 6 to 8 months-old J20 mice treated with vehicle or BBG. Scale bar, 500 μm. (D) Quantification of microglial cells in the hippocampus of J20 mice treated with vehicle or BBG. Histograms represent the mean ± SEM of microglial cells per hippocampal area of 0.1 mm2 being 16 slices per mouse (n = 7 mice per treatment). For methods see Ref. [49].
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4372621&req=5

f0020: Brilliant Blue-G (BBG) treatment reduces the number of amyloid plaques and microglia in the J20 mouse hippocampus. (A) Immunostaining of WO2 in hippocampal slices from 6 to 8 months-old J20 mice injected intraperitoneal with vehicle or BBG. Scale bar, 500 μm. (B) Histogram represents the mean ± SEM of Aβ-amyloid plaques per slice in the hippocampus of J20 mice treated with vehicle or BBG, being 16 slices per mouse (n = 7 mice per condition). ***p < 0.005, unpaired Student's t test. (C) Immunostaining of microglial marker Iba-1 in hippocampal slices from 6 to 8 months-old J20 mice treated with vehicle or BBG. Scale bar, 500 μm. (D) Quantification of microglial cells in the hippocampus of J20 mice treated with vehicle or BBG. Histograms represent the mean ± SEM of microglial cells per hippocampal area of 0.1 mm2 being 16 slices per mouse (n = 7 mice per treatment). For methods see Ref. [49].
Mentions: After BBG treatment the number and size of amyloid plaques at the hippocampal structures of J20 mice were significantly reduced compared to their littermates treated with vehicle, as shown in Fig. 4A and B. In addition, the treatment with BBG did not significantly modify either the P2X7 receptor or murine APP and human APP mRNA expression. The levels of these proteins and their distribution patterns in the hippocampus were also not modified by the treatment with BBG, as demonstrated by western blot and immunohistochemical studies. However, a dramatic change was observed concerning the pattern of the C83 and C99 peptides, which correspond to the carboxyterminal fragments generated by APP cleavage by α-secretase and β-secretase, respectively. C99 fragment was under the limits of detection in wild type mouse brain, but was very abundant in the brain of J20 mice, which exhibit a much lower concentration of the α-secretase generated C83 fragment. However, BBG reversed the situation and a significant increase in the C83 fragment was achieved in the brain of BBG-treated J20 mice [48]. This situation mimics the APP processing pattern observed in N2a cells treated with P2X7 receptor inhibitors. On the other hand, an increase in the phosphorylated form of GSK-3 was observed in the hippocampus of BBG-treated J20 mice, when compared with vehicle-treated animals. Thus, BBG was able to reduce hippocampal GSK-3 activity in FAD animal models in the same way as in N2a cells.

Bottom Line: In Neuro-2a, a neuroblastoma cell line that constitutively expresses APP, activation of the P2X7 receptor leads to reduction of α-secretase activity, the opposite effect being obtained by P2Y2 receptor activation.In vivo inhibition of the P2X7 receptor induced a significant decrease in the number and size of hippocampal amyloid plaques.This reduction is mediated by an increase in the proteolytic processing of APP through α-secretase activity, which correlates with an increase in the phosphorylated form of GSK-3, a less active form of this enzyme.

View Article: PubMed Central - PubMed

Affiliation: Biochemistry Department, School of Veterinary Sciences, Complutense University of Madrid, Madrid, Spain, Institute of Neurochemistry (IUIN), Complutense University of Madrid, Madrid, Spain.

ABSTRACT
Amyloid precursor protein (APP) is expressed in a large variety of neural and non-neural cells. The balance between non-pathogenic and pathologic forms of APP processing, mediated by α-secretase and β-secretase respectively, remains a crucial step to understand β-amyloid, Aβ42 peptide, formation and aggregation that are at the origin of the senile plaques in the brain, a characteristic hallmark of Alzheimer's disease (AD). In Neuro-2a, a neuroblastoma cell line that constitutively expresses APP, activation of the P2X7 receptor leads to reduction of α-secretase activity, the opposite effect being obtained by P2Y2 receptor activation. The in vivo approach was made possible by the use of J20 mice, a transgenic mouse model of familial Alzheimer's disease (FAD) expressing human APP mutant protein. This animal exhibits prominent amyloid plaques by six months of age. In vivo inhibition of the P2X7 receptor induced a significant decrease in the number and size of hippocampal amyloid plaques. This reduction is mediated by an increase in the proteolytic processing of APP through α-secretase activity, which correlates with an increase in the phosphorylated form of GSK-3, a less active form of this enzyme. The in vivo findings corroborate the therapeutic potential of P2X7 antagonists in the treatment of FAD.

No MeSH data available.


Related in: MedlinePlus