Monocyte-derived IL-6 programs microglia to rebuild damaged brain vasculature.

Cerebrovascular injury (CVI) is a common pathology caused by infections, injury, stroke, neurodegeneration and autoimmune disease. Rapid resolution of a CVI requires a coordinated innate immune response. In the present study, we sought mechanistic insights into how central nervous system-infiltrating monocytes program resident microglia to mediate angiogenesis and cerebrovascular repair after an intracerebral hemorrhage. In the penumbrae of human stroke brain lesions, we identified a subpopulation of microglia that express vascular endothelial growth factor A. These cells, termed 'repair-associated microglia' (RAMs), were also observed in a rodent model of CVI and coexpressed interleukin (IL)-6Ra. Cerebrovascular repair did not occur in IL-6 knockouts or in mice lacking microglial IL-6Ra expression and single-cell transcriptomic analyses revealed faulty RAM programming in the absence of IL-6 signaling. Infiltrating CCR2+ monocytes were the primary source of IL-6 after a CVI and were required to endow microglia with proliferative and proangiogenic properties. Faulty RAM programming in the absence of IL-6 or inflammatory monocytes resulted in poor cerebrovascular repair, neuronal destruction and sustained neurological deficits that were all restored via exogenous IL-6 administration. These data provide a molecular and cellular basis for how monocytes instruct microglia to repair damaged brain vasculature and promote functional recovery after injury.

GDE2-RECK controls ADAM10 α-secretase-mediated cleavage of amyloid precursor protein.

A disintegrin and metalloprotease 10 (ADAM10) is the α-secretase for amyloid precursor protein (APP). ADAM10 cleaves APP to generate neuroprotective soluble APPα (sAPPα), which precludes the generation of Aß, a defining feature of Alzheimer's disease (AD) pathophysiology. Reduced ADAM10 activity is implicated in AD, but the mechanisms mediating ADAM10 modulation are unclear. We find that the plasma membrane enzyme glycerophosphodiester phosphodiesterase 2 (GDE2) stimulates ADAM10 APP cleavage by shedding and inactivating reversion-inducing cysteine-rich protein with Kazal motifs (RECK), a glycosylphosphatidylinositol (GPI)-anchored inhibitor of ADAM10. In AD, membrane-tethered RECK is highly elevated and GDE2 is abnormally sequestered inside neurons. Genetic ablation of GDE2 phenocopies increased membrane RECK in AD, which is causal for reduced sAPPα, increased Aß, and synaptic protein loss. RECK reduction restores the balance of APP processing and rescues synaptic protein deficits. These studies identify GDE2 control of RECK surface activity as essential for ADAM10 α-secretase function and physiological APP processing. Moreover, our results suggest the involvement of the GDE2-RECK-ADAM10 pathway in AD pathophysiology and highlight RECK as a potential target for therapeutic development.

GDE2 expression in oligodendroglia regulates the pace of oligodendrocyte maturation.

BACKGROUND: Oligodendrocytes generate specialized lipid-rich sheaths called myelin that wrap axons and facilitate the rapid, saltatory transmission of action potentials. Extrinsic signals and surface-mediated pathways coordinate oligodendrocyte development to ensure appropriate axonal myelination, but the mechanisms involved are not fully understood. Glycerophosphodiester phosphodiesterase 2 (GDE2 or GDPD5) is a six-transmembrane enzyme that regulates the activity of surface glycosylphosphatidylinositol (GPI)-anchored proteins by cleavage of the GPI-anchor. GDE2 is expressed in neurons where it promotes oligodendrocyte maturation through the release of neuronally-derived soluble factors. GDE2 is also expressed in oligodendrocytes but the function of oligodendroglial GDE2 is not known. RESULTS: Using Cre-lox technology, we generated mice that lack GDE2 expression in oligodendrocytes (O-Gde2KO). O-Gde2KOs show normal production and proliferation of oligodendrocyte precursor cells. However, oligodendrocyte maturation is accelerated leading to the robust increase of myelin proteins and increased myelination during development. These in vivo observations are recapitulated in vitro using purified primary oligodendrocytes, supporting cell-autonomous functions for GDE2 in oligodendrocyte maturation. CONCLUSIONS: These studies reveal that oligodendroglial GDE2 expression is required for controlling the pace of oligodendrocyte maturation. Thus, the cell-type specific expression of GDE2 is important for the coordination of oligodendrocyte maturation and axonal myelination during neural development.

GDE2-Dependent Activation of Canonical Wnt Signaling in Neurons Regulates Oligodendrocyte Maturation.

Neurons and oligodendrocytes communicate to regulate oligodendrocyte development and ensure appropriate axonal myelination. Here, we show that Glycerophosphodiester phosphodiesterase 2 (GDE2) signaling underlies a neuronal pathway that promotes oligodendrocyte maturation through the release of soluble neuronally derived factors. Mice lacking global or neuronal GDE2 expression have reduced mature oligodendrocytes and myelin proteins but retain normal numbers of oligodendrocyte precursor cells (OPCs). Wild-type (WT) OPCs cultured in conditioned medium (CM) from Gde2-null (Gde2KO) neurons exhibit delayed maturation, recapitulating in vivo phenotypes. Gde2KO neurons show robust reduction in canonical Wnt signaling, and genetic activation of Wnt signaling in Gde2KO neurons rescues in vivo and in vitro oligodendrocyte maturation. Phosphacan, a known stimulant of oligodendrocyte maturation, is reduced in CM from Gde2KO neurons but is restored when Wnt signaling is activated. These studies identify GDE2 control of Wnt signaling as a neuronal pathway that signals to oligodendroglia to promote oligodendrocyte maturation.

GDE3 regulates oligodendrocyte precursor proliferation via release of soluble CNTFRα.

Oligodendrocyte development is tightly controlled by extrinsic signals; however, mechanisms that modulate cellular responses to these factors remain unclear. Six-transmembrane glycerophosphodiester phosphodiesterases (GDEs) are emerging as central regulators of cellular differentiation via their ability to shed glycosylphosphatidylinositol (GPI)-anchored proteins from the cell surface. We show here that GDE3 controls the pace of oligodendrocyte generation by negatively regulating oligodendrocyte precursor cell (OPC) proliferation. GDE3 inhibits OPC proliferation by stimulating ciliary neurotrophic factor (CNTF)-mediated signaling through release of CNTFRα, the ligand-binding component of the CNTF-receptor multiprotein complex, which can function as a soluble factor to activate CNTF signaling. GDE3 releases soluble CNTFRα by GPI-anchor cleavage from the plasma membrane and from extracellular vesicles (EVs) after co-recruitment of CNTFRα in EVs. These studies uncover new physiological roles for GDE3 in gliogenesis and identify GDE3 as a key regulator of CNTF-dependent regulation of OPC proliferation through release of CNTFRα.

Differentially expressed genes in iron-induced prion protein conversion.

The conversion of the cellular prion protein (PrPC) to the protease-resistant isoform is the key event in chronic neurodegenerative diseases, including transmissible spongiform encephalopathies (TSEs). Increased iron in prion-related disease has been observed due to the prion protein-ferritin complex. Additionally, the accumulation and conversion of recombinant PrP (rPrP) is specifically derived from Fe(III) but not Fe(II). Fe(III)-mediated PK-resistant PrP (PrPres) conversion occurs within a complex cellular environment rather than via direct contact between rPrP and Fe(III). In this study, differentially expressed genes correlated with prion degeneration by Fe(III) were identified using Affymetrix microarrays. Following Fe(III) treatment, 97 genes were differentially expressed, including 85 upregulated genes and 12 downregulated genes (≥1.5-fold change in expression). However, Fe(II) treatment produced moderate alterations in gene expression without inducing dramatic alterations in gene expression profiles. Moreover, functional grouping of identified genes indicated that the differentially regulated genes were highly associated with cell growth, cell maintenance, and intra- and extracellular transport. These findings showed that Fe(III) may influence the expression of genes involved in PrP folding by redox mechanisms. The identification of genes with altered expression patterns in neural cells may provide insights into PrP conversion mechanisms during the development and progression of prion-related diseases.

High fat diet produces brain insulin resistance, synaptodendritic abnormalities and altered behavior in mice.

Insulin resistance and other features of the metabolic syndrome are increasingly recognized for their effects on cognitive health. To ascertain mechanisms by which this occurs, we fed mice a very high fat diet (60% kcal by fat) for 17days or a moderate high fat diet (HFD, 45% kcal by fat) for 8weeks and examined changes in brain insulin signaling responses, hippocampal synaptodendritic protein expression, and spatial working memory. Compared to normal control diet mice, cerebral cortex tissues of HFD mice were insulin-resistant as evidenced by failed activation of Akt, S6 and GSK3ß with ex-vivo insulin stimulation. Importantly, we found that expression of brain IPMK, which is necessary for mTOR/Akt signaling, remained decreased in HFD mice upon activation of AMPK. HFD mouse hippocampus exhibited increased expression of serine-phosphorylated insulin receptor substrate 1 (IRS1-pS(616)), a marker of insulin resistance, as well as decreased expression of PSD-95, a scaffolding protein enriched in post-synaptic densities, and synaptopodin, an actin-associated protein enriched in spine apparatuses. Spatial working memory was impaired as assessed by decreased spontaneous alternation in a T-maze. These findings indicate that HFD is associated with telencephalic insulin resistance and deleterious effects on synaptic integrity and cognitive behaviors.

PKA modulates iron trafficking in the striatum via small GTPase, Rhes.

Ras homolog enriched in striatum (Rhes), is a highly conserved small guanosine-5'-triphosphate (GTP) binding protein belonging to the Ras superfamily. Rhes is involved in the dopamine receptor-mediated signaling and behavior though adenylyl cyclase. The striatum-specific GTPase share a close homology with Dexras1, which regulates iron trafficking in the neurons when activated though the post-translational modification called s-nitrosylation by nitric oxide (NO). We report that Rhes physiologically interacted with Peripheral benzodiazepine receptor-associated protein7 and participated in iron uptake via divalent metal transporter 1 similar to Dexras1. Interestingly, Rhes is not S-nitrosylated by NO-treatment, however phosphorylated by protein kinase A at the site of serine-239. Two Rhes mutants - the phosphomimetic form (serine 239 to aspartic acid) and constitutively active form (alanine 173 to valine) - displayed an increase in iron uptake compared to the wild-type Rhes. These findings suggest that Rhes may play a crucial role in striatal iron homeostasis.

Prion protein conversion induced by trivalent iron in vesicular trafficking.

Iron dyshomeostasis has been observed in prion diseases; however, little is known regarding the contribution of the oxidation state of iron to prion protein (PrP) conversion. In this study, PrP(C)-deficient HpL3-4 cells were exposed to divalent [Fe(II)] or trivalent [Fe(III)] iron, followed by exogenous recombinant PrP (rPrP) treatment. We then analyzed the accumulation of internalized rPrP and its biochemical properties, including its resistance to both proteinase K (PK) digestion and detergent solubility. Fe(III), but not Fe(II), induced the accumulation of internalized rPrP, which was partially converted to detergent-insoluble and PK-resistant PrP (PrP(res)). The Fe(III)-induced PrP(res) generation required an intact cell structure, and it was hindered by U18666A, an inhibitor of vesicular trafficking, but not by NH4Cl, an inhibitor of endolysosomal acidification. These observations implicated that the Fe(III)-mediated PrP(res) conversion likely occurs during endosomal vesicular trafficking rather than in the acidic environment of lysosomes.