Suppression of Inflammation with Conditional Deletion of the Prostaglandin E₂EP2 Receptor in Macrophages and Brain Microglia

Abstract

Prostaglandin E 2 (PGE 2 ), a potent lipid signaling molecule, modulates inflammatory responses through activation of downstream G-protein coupled EP 1-4 receptors. Here, we investigated the cell-specific in vivo function of PGE 2 signaling through its E-prostanoid 2 (EP2) receptor in murine innate immune responses systemically and in the CNS. In vivo, systemic administration of lipopolysaccharide (LPS) resulted in a broad induction of cytokines and chemokines in plasma that was significantly attenuated in EP2-deficient mice. Ex vivo stimulation of peritoneal macrophages with LPS elicited proinflammatory responses that were dependent on EP2 signaling and that overlapped with in vivo plasma findings, suggesting that myeloid-lineage EP2 signaling is a major effector of innate immune responses. Conditional deletion of the EP2 receptor in myeloid lineage cells in Cd11bCre;EP2 lox/lox mice attenuated plasma inflammatory responses and transmission of systemic inflammation to the brain was inhibited, with decreased hippocampal inflammatory gene expression and cerebral cortical levels of IL-6. Conditional deletion of EP2 significantly blunted microglial and astrocytic inflammatory responses to the neurotoxin MPTP and reduced striatal dopamine turnover. Suppression of microglial EP2 signaling also increased numbers of dopaminergic (DA) neurons in the substantia nigra independent of MPTP treatment, suggesting that microglial EP2 may influence development or survival of DA neurons. Unbiased microarray analysis of microglia isolated from adult Cd11bCre;EP2 lox/lox and control mice demonstrated a broad downregulation of inflammatory pathways with ablation of microglial EP2 receptor. Together, these data identify a cell-specific proinflammatory role for macrophage/microglial EP2 signaling in innate immune responses systemically and in brain.

Prostaglandin signaling suppresses beneficial microglial function in Alzheimer’s disease models

“…This conclusion was based on the selectivity of the Cd11b promoter-driven expression of Cre recombinase in brain to microglia, as demonstrated by Boillee et al: crosses of Cd11b-Cre mice to Rosa26-lacZ mice showed Cre recombination in the CNS that was restricted to microglia (31). In a previous study using this Cre recombinase line, we observed approximately 50% reduction of floxed Ep2 sequences in microglia and in peritoneal macrophages (26). Peripherally, CD11b protein is normally expressed in monocytes and tissuespecific macrophages, and to a lesser extent in subpopulations of granulocytes, mature B lymphocytes, and CD4 + T lymphocytes (59,60).…”

“…Consistently, studies in AD model mice demonstrate reduced amyloid pathology with global deletion of individual PGE 2 G protein-coupled receptors (21)(22)(23), and additional studies have shown a suppressive signaling effect of the PGE 2 receptor EP2 on Aβ 42 phagocytosis (24,25). These studies, along with the recent demonstration of a broad regulatory function of EP2 signaling on cell cycle, cytoskeletal, and immune genes in quiescent microglia (26), suggest that microglial EP2 signaling may be a general suppressor of immune and nonimmune processes that protect against onset and progression of AD pathology. To investigate this hypothesis, we used in vitro and in vivo mouse models that recapitulate acute and chronic aspects of microglial responses to Aβ peptides.…”

“…APP-PS1 mice (APPSwe-PS1ΔE9; ref. 63 (26,29,31). C57BL/6 Ep2 fl/+ mice have been previously described (26).…”

“…We next used a micro glial conditional knockout strategy to directly examine microglial EP2 function in chemotaxis and clearance of Aβ peptide in vivo. The Cd11b-Cre recombinase line leads to excision of floxed sequences in cells of the myeloid lineage, including monocytes, macrophages, and microglia, and has been successfully used to examine neuroinflammatory responses in brain (26,29,31 Figure 3A). The remaining FITC + staining area and intensity, quantified in a blinded fashion (see Methods), indicated that clearance of Aβ 42 peptides was significantly higher in Cd11b-Cre Ep2 fl/fl mice than in Cd11b-Cre controls ( Figure 3B).…”

“…Thus, the suppression of MIP-1α expression coupled to the induction of proinflammatory responses in the context of Aβ stimulation and EP2 signaling is unexpected, but likely suggests a complex transcriptional regulation of MIP-1α and cytokine release. Nonetheless, that the Cd11b-Cre recombinase line is approximately 50% efficient in excising floxed Ep2 sequences (26). Incomplete excision of floxed sequences is common in many Cre-mediated systems, in which recombinase activity frequently results in cell-specific knockdown of gene expression.…”