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CD45RB is a novel molecular therapeutic target to inhibit Abeta peptide-induced microglial MAPK activation.

Zhu Y, Hou H, Nikolic WV, Ehrhart J, Rrapo E, Bickford P, Giunta B, Tan J - PLoS ONE (2008)

Bottom Line: Co-treatment of microglial cells with agonist CD45 antibodies results in significant inhibition of LPS-induced microglial TNF-alpha and IL-6 release through p44/42 and/or p38 pathways.Therefore, agonism of CD45RB PTP activity may be an effective therapeutic target for novel agents to treat AD due to its Abeta lowering, and inflammation reducing, properties that are particularly targeted at microglial cells.Such treatments may be more effective with less potential to produce systemic side-effects than therapeutics which induce non-specific, systemic down-regulation of inflammation.

View Article: PubMed Central - PubMed

Affiliation: Rashid Laboratory Developmental Neurobiology, Silver Child Development Center, Department of Psychiatry and Behavioral Medicine, University of South Florida College of Medicine, Tampa, Florida, United States of America.

ABSTRACT

Background: Microglial activation, characterized by p38 MAPK or p44/42 MAPK pathway signal transduction, occurs in Alzheimer's disease (AD). Our previous studies demonstrated CD45, a membrane-bound protein tyrosine phosphatase (PTP), opposed beta-amyloid (Abeta) peptide-induced microglial activation via inhibition of p44/42 MAPK. Additionally we have shown agonism of the RB isoform of CD45 (CD45RB) abrogates lipopolysaccharide (LPS)-induced microglial activation.

Methodology and results: In this study, CD45RB modulation of Abeta peptide or LPS-activated primary cultured microglial cells was further investigated. Microglial cells were co-treated with "aged" FITC-Abeta(1-42) and multiple CD45 isoform agonist antibodies. Data revealed cross-linking of CD45, particularly the CD45RB isoform, enhances microglial phagocytosis of Abeta(1-42) peptide and inhibits LPS-induced activation of p44/42 and p38 pathways. Co-treatment of microglial cells with agonist CD45 antibodies results in significant inhibition of LPS-induced microglial TNF-alpha and IL-6 release through p44/42 and/or p38 pathways. Moreover, inhibition of either of these pathways augmented CD45RB cross-linking induced microglial phagocytosis of Abeta(1-42) peptide. To investigate the mechanism(s) involved, microglial cells were co-treated with a PTP inhibitor (potassium bisperoxo [1,10-phenanthroline oxovanadate; Phen]) and Abeta(1-42) peptides. Data showed synergistic induction of microglial activation as evidenced by TNF-alpha and IL-6 release; both of which are demonstrated to be dependent on increased p44/42 and/or p38 activation. Finally, it was observed that cross-linking of CD45RB in the presence of Abeta(1-42) peptide, inhibits co-localization of microglial MHC class II and Abeta peptide; suggesting CD45 activation inhibits the antigen presenting phenotype of microglial cells.

Conclusion: In summary, p38 MAPK is another novel signaling pathway, besides p44/42, in which CD45RB cross-linking negatively regulates microglial Abeta phagocytosis while increasing potentially neurotoxic inflammation. Therefore, agonism of CD45RB PTP activity may be an effective therapeutic target for novel agents to treat AD due to its Abeta lowering, and inflammation reducing, properties that are particularly targeted at microglial cells. Such treatments may be more effective with less potential to produce systemic side-effects than therapeutics which induce non-specific, systemic down-regulation of inflammation.

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Cross-linking of CD45RB inhibits microglia MHC class II-Aβ co-localization.In order to examine microglia MHC II-Aβ peptide complex formation on the cell surface, microglia were treated with “aged” Cy3-Aβ1–42 peptide (300 nM) in the presence or absence of agonist CD45RB antibody for 48 h followed by staining with FITC-anti-mouse MHC class II antibody and Fluorescence microscopy. Note: Red indicates Aβ-positive; green indicates MHC class II-positive; yellow indicates the co-localization of MHC class II and Aβ, Ab indicates antibody. Blue indicates DAPI nuclear stain of the same fields. Original magnification = 40× for top and bottom panels.
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pone-0002135-g005: Cross-linking of CD45RB inhibits microglia MHC class II-Aβ co-localization.In order to examine microglia MHC II-Aβ peptide complex formation on the cell surface, microglia were treated with “aged” Cy3-Aβ1–42 peptide (300 nM) in the presence or absence of agonist CD45RB antibody for 48 h followed by staining with FITC-anti-mouse MHC class II antibody and Fluorescence microscopy. Note: Red indicates Aβ-positive; green indicates MHC class II-positive; yellow indicates the co-localization of MHC class II and Aβ, Ab indicates antibody. Blue indicates DAPI nuclear stain of the same fields. Original magnification = 40× for top and bottom panels.

Mentions: MHC class II plays an important role in loading and transporting extracellular pathogens and toxin to the APC surface, where they are recognized by CD+ T cells [35]. This cell surface protein is particularly interesting, as microglial cells express it in the frontal cortex and hippocampus of normally aging individuals and levels of expression are markedly increased in these brain regions in AD cases [36], [37]. The impairment of MHC II function results in a significant reduction of microglia-associated CNS inflammation [37], [38], suggesting the elevated level of MHC class II expression is strongly associated with a microglial immune response. Recently, we have shown that CD40 ligation increases MHC II-Aβ peptide complexes as examined by fluorescence microscopy [5]. In addition, we previously showed cross-linking of CD45 significantly opposes CD40-mediated microglial activation [14]. To examine whether cross-linking of CD45RB could inhibit formation of immunogenic MHC II-Aβ peptide complexes on the cell surface, we treated microglia with CD45RB antibody (2.5 µg/mL) in the presence or absence of “aged” Cy3-Aβ1–42 peptide (300 nM) for 48 h, followed by immunofluorescence staining with FITC-conjugated anti-mouse MHC class II antibody. Result show that cross-linking of CD45RB inhibits MHC class II-Aβ co-localization as detected by fluorescence microscopy (Fig. 5).


CD45RB is a novel molecular therapeutic target to inhibit Abeta peptide-induced microglial MAPK activation.

Zhu Y, Hou H, Nikolic WV, Ehrhart J, Rrapo E, Bickford P, Giunta B, Tan J - PLoS ONE (2008)

Cross-linking of CD45RB inhibits microglia MHC class II-Aβ co-localization.In order to examine microglia MHC II-Aβ peptide complex formation on the cell surface, microglia were treated with “aged” Cy3-Aβ1–42 peptide (300 nM) in the presence or absence of agonist CD45RB antibody for 48 h followed by staining with FITC-anti-mouse MHC class II antibody and Fluorescence microscopy. Note: Red indicates Aβ-positive; green indicates MHC class II-positive; yellow indicates the co-localization of MHC class II and Aβ, Ab indicates antibody. Blue indicates DAPI nuclear stain of the same fields. Original magnification = 40× for top and bottom panels.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2366070&req=5

pone-0002135-g005: Cross-linking of CD45RB inhibits microglia MHC class II-Aβ co-localization.In order to examine microglia MHC II-Aβ peptide complex formation on the cell surface, microglia were treated with “aged” Cy3-Aβ1–42 peptide (300 nM) in the presence or absence of agonist CD45RB antibody for 48 h followed by staining with FITC-anti-mouse MHC class II antibody and Fluorescence microscopy. Note: Red indicates Aβ-positive; green indicates MHC class II-positive; yellow indicates the co-localization of MHC class II and Aβ, Ab indicates antibody. Blue indicates DAPI nuclear stain of the same fields. Original magnification = 40× for top and bottom panels.
Mentions: MHC class II plays an important role in loading and transporting extracellular pathogens and toxin to the APC surface, where they are recognized by CD+ T cells [35]. This cell surface protein is particularly interesting, as microglial cells express it in the frontal cortex and hippocampus of normally aging individuals and levels of expression are markedly increased in these brain regions in AD cases [36], [37]. The impairment of MHC II function results in a significant reduction of microglia-associated CNS inflammation [37], [38], suggesting the elevated level of MHC class II expression is strongly associated with a microglial immune response. Recently, we have shown that CD40 ligation increases MHC II-Aβ peptide complexes as examined by fluorescence microscopy [5]. In addition, we previously showed cross-linking of CD45 significantly opposes CD40-mediated microglial activation [14]. To examine whether cross-linking of CD45RB could inhibit formation of immunogenic MHC II-Aβ peptide complexes on the cell surface, we treated microglia with CD45RB antibody (2.5 µg/mL) in the presence or absence of “aged” Cy3-Aβ1–42 peptide (300 nM) for 48 h, followed by immunofluorescence staining with FITC-conjugated anti-mouse MHC class II antibody. Result show that cross-linking of CD45RB inhibits MHC class II-Aβ co-localization as detected by fluorescence microscopy (Fig. 5).

Bottom Line: Co-treatment of microglial cells with agonist CD45 antibodies results in significant inhibition of LPS-induced microglial TNF-alpha and IL-6 release through p44/42 and/or p38 pathways.Therefore, agonism of CD45RB PTP activity may be an effective therapeutic target for novel agents to treat AD due to its Abeta lowering, and inflammation reducing, properties that are particularly targeted at microglial cells.Such treatments may be more effective with less potential to produce systemic side-effects than therapeutics which induce non-specific, systemic down-regulation of inflammation.

View Article: PubMed Central - PubMed

Affiliation: Rashid Laboratory Developmental Neurobiology, Silver Child Development Center, Department of Psychiatry and Behavioral Medicine, University of South Florida College of Medicine, Tampa, Florida, United States of America.

ABSTRACT

Background: Microglial activation, characterized by p38 MAPK or p44/42 MAPK pathway signal transduction, occurs in Alzheimer's disease (AD). Our previous studies demonstrated CD45, a membrane-bound protein tyrosine phosphatase (PTP), opposed beta-amyloid (Abeta) peptide-induced microglial activation via inhibition of p44/42 MAPK. Additionally we have shown agonism of the RB isoform of CD45 (CD45RB) abrogates lipopolysaccharide (LPS)-induced microglial activation.

Methodology and results: In this study, CD45RB modulation of Abeta peptide or LPS-activated primary cultured microglial cells was further investigated. Microglial cells were co-treated with "aged" FITC-Abeta(1-42) and multiple CD45 isoform agonist antibodies. Data revealed cross-linking of CD45, particularly the CD45RB isoform, enhances microglial phagocytosis of Abeta(1-42) peptide and inhibits LPS-induced activation of p44/42 and p38 pathways. Co-treatment of microglial cells with agonist CD45 antibodies results in significant inhibition of LPS-induced microglial TNF-alpha and IL-6 release through p44/42 and/or p38 pathways. Moreover, inhibition of either of these pathways augmented CD45RB cross-linking induced microglial phagocytosis of Abeta(1-42) peptide. To investigate the mechanism(s) involved, microglial cells were co-treated with a PTP inhibitor (potassium bisperoxo [1,10-phenanthroline oxovanadate; Phen]) and Abeta(1-42) peptides. Data showed synergistic induction of microglial activation as evidenced by TNF-alpha and IL-6 release; both of which are demonstrated to be dependent on increased p44/42 and/or p38 activation. Finally, it was observed that cross-linking of CD45RB in the presence of Abeta(1-42) peptide, inhibits co-localization of microglial MHC class II and Abeta peptide; suggesting CD45 activation inhibits the antigen presenting phenotype of microglial cells.

Conclusion: In summary, p38 MAPK is another novel signaling pathway, besides p44/42, in which CD45RB cross-linking negatively regulates microglial Abeta phagocytosis while increasing potentially neurotoxic inflammation. Therefore, agonism of CD45RB PTP activity may be an effective therapeutic target for novel agents to treat AD due to its Abeta lowering, and inflammation reducing, properties that are particularly targeted at microglial cells. Such treatments may be more effective with less potential to produce systemic side-effects than therapeutics which induce non-specific, systemic down-regulation of inflammation.

Show MeSH
Related in: MedlinePlus