Background: Air pollution is linked to central nervous system disease, but the mechanisms responsible are poorly understood.Objectives: Here, we sought to address the brain-region–specific effects of diesel exhaust (DE) and key cellular mechanisms underlying DE-induced microglia activation, neuroinflammation, and dopaminergic (DA) neurotoxicity.Methods: Rats were exposed to DE (2.0, 0.5, and 0 mg/m3) by inhalation over 4 weeks or as a single intratracheal administration of DE particles (DEP; 20 mg/kg). Primary neuron–glia cultures and the HAPI (highly aggressively proliferating immortalized) microglial cell line were used to explore cellular mechanisms.Results: Rats exposed to DE by inhalation demonstrated elevated levels of whole-brain IL-6 (interleukin-6) protein, nitrated proteins, and IBA-1 (ionized calcium-binding adaptor molecule 1) protein (microglial marker), indicating generalized neuroinflammation. Analysis by brain region revealed that DE increased TNFα (tumor necrosis factor-α), IL-1β, IL-6, MIP-1α (macrophage inflammatory protein-1α) RAGE (receptor for advanced glycation end products), fractalkine, and the IBA-1 microglial marker in most regions tested, with the midbrain showing the greatest DE response. Intratracheal administration of DEP increased microglial IBA-1 staining in the substantia nigra and elevated both serum and whole-brain TNFα at 6 hr posttreatment. Although DEP alone failed to cause the production of cytokines and chemokines, DEP (5 μg/mL) pretreatment followed by lipopolysaccharide (2.5 ng/mL) in vitro synergistically amplified nitric oxide production, TNFα release, and DA neurotoxicity. Pretreatment with fractalkine (50 pg/mL) in vitro ameliorated DEP (50 μg/mL)-induced microglial hydrogen peroxide production and DA neurotoxicity.Conclusions: Together, these findings reveal complex, interacting mechanisms responsible for how air pollution may cause neuroinflammation and DA neurotoxicity.

Redox-signaling is implicated in deleterious microglial activation underlying CNS disease, but how ROS program aberrant microglial function is unknown. Here, the oxidation of NF-κB p50 to a free radical intermediate is identified as a marker of dysfunctional M1 (pro-inflammatory) polarization in microglia. Microglia exposed to steady fluxes of H2O2 showed altered NF-κB p50 protein-protein interactions, decreased NF-κB p50 DNA binding, and augmented late-stage TNFα expression, indicating that H2O2 impairs NF-κB p50 function and prolongs amplified M1 activation. NF-κB p50−/− mice and cultures exhibited a disrupted M2 (alternative) response and impaired resolution of the M1 response. Persistent neuroinflammation continued 1 week after LPS (1mg/kg, IP) administration in the NF-κB p50−/− mice. However, peripheral inflammation had already resolved in both strains of mice. Treatment with the spin-trap DMPO mildly reduced LPS-induced 22 h TNFα in the brain in NF-κB p50+/+ mice. Interestingly, DMPO failed to reduce and strongly augmented brain TNFα production in NF-κB p50−/− mice, implicating a fundamental role for NF-κB p50 in the regulation of chronic neuroinflammation by free radicals. These data identify NF-κB p50 as a key redox-signaling mechanism regulating the M1/M2 balance in microglia, where loss of function leads to a CNS-specific vulnerability to chronic inflammation.

Increasing reports support that air pollution causes neuroinflammation and is linked to central nervous system (CNS) disease/damage. Diesel exhaust particles (DEP) are a major component of urban air pollution, which has been linked to microglial activation and Parkinson’s disease-like pathology. To begin to address how DEP may exert CNS effects, microglia and neuron-glia cultures were treated with either nanometer-sized DEP (<0.22 µM; 50µg/mL), ultrafine carbon black (ufCB, 50µg/ml), or DEP extracts (eDEP; from 50 µg/ml DEP) and the effect of microglial activation and dopaminergic (DA) neuron function was assessed. All three treatments showed enhanced amoeboid microglia morphology, increased H2O2 production, and decreased DA uptake. Mechanistic inquiry revealed that the scavenger receptor inhibitor fucoidan blocked DEP internalization in microglia, but failed to alter DEP-induced H2O2 production in microglia. However, pretreatment with the MAC1/CD11b inhibitor antibody blocked microglial H2O2 production in response to DEP. MAC1−/− mesencephalic neuron-glia cultures were protected from DEP-induced loss of DA neuron function, as measured by DA uptake. These findings support that DEP may activate microglia through multiple mechanisms, where scavenger receptors regulate internalization of DEP and the MAC1 receptor is mandatory for both DEP-induced microglial H2O2 production and loss of DA neuron function.

The identity of muscle secreted factors critical for the development and maintenance of neuromuscular junctions (NMJs) remains largely unknown. Here, we show that muscle fibers secrete and concentrate the fibroblast growth factor binding protein 1 (FGFBP1) at NMJs. Although FGFBP1 expression increases during development, its expression decreases before NMJ degeneration during aging and in SOD1 G93A mice, a mouse model for amyotrophic lateral sclerosis (ALS). Based on these findings, we examined the impact of deleting FGFBP1 on NMJs. In the absence of FGFBP1, NMJs exhibit structural abnormalities in developing and middle age mice. Deletion of FGFBP1 from SOD1 G93A mice also accelerates NMJ degeneration and death. Based on these findings, we sought to identify the mechanism responsible for decreased FGFBP1 in stressed skeletal muscles. We show that FGFBP1 expression is inhibited by increased accumulation of the transforming growth factor-␤1 (TGF-␤1) in skeletal muscles and at their NMJs. These findings suggest that targeting the FGFBP1 and TGF-␤1 signaling axis holds promise for slowing age-and disease-related degeneration of NMJs.

Perisynaptic Schwann cells (PSCs) are specialized, non-myelinating, synaptic glia of the neuromuscular junction (NMJ), that participate in synapse development, function, maintenance, and repair. The study of PSCs has relied on an anatomy-based approach, as the identities of cell-specific PSC molecular markers have remained elusive. This limited approach has precluded our ability to isolate and genetically manipulate PSCs in a cell specific manner. We have identified neuron-glia antigen 2 (NG2) as a unique molecular marker of S100β+ PSCs in skeletal muscle. NG2 is expressed in Schwann cells already associated with the NMJ, indicating that it is a marker of differentiated PSCs. Using a newly generated transgenic mouse in which PSCs are specifically labeled, we show that PSCs have a unique molecular signature that includes genes known to play critical roles in PSCs and synapses. These findings will serve as a springboard for revealing drivers of PSC differentiation and function.