Slc6a3-dependent expression of a CAPS-associated Nlrp3 allele results in progressive behavioral abnormalities and neuroinflammation in aging mice.

BACKGROUND: An association between neuroinflammation and age-related neurologic disorders has been established but the molecular mechanisms and cell types involved have not been thoroughly characterized. Activity of the proinflammatory NLRP3 inflammasome is implicated in Alzheimer's and Parkinson's disease and our recent studies in patients suggest that dopaminergic neurons within the degenerating mesencephalon express NLRP3 throughout the progression of PD. Here, we directly test the impact of enhanced inflammasome activity in mesencephalic neurons by characterizing motor function, tissue integrity, and neuroinflammation in aging mice harboring hyperactivating mutations within the endogenous murine Nlrp3 locus, enabled only in cells expressing the dopaminergic neuron-specific Slc6a3 promoter. METHODS: We compared mice harboring inducible alleles encoding the cryopyrin-associated periodic syndrome activating mutations Nlrp3A350V and Nlrp3L351P inserted into the endogenous mouse Nlrp3 locus. Tissue specific expression was driven by breeding these animals with mice expressing Cre recombinase under the control of the dopaminergic neuron-specific Slc6a3 promoter. The experimental mice, designed to express hyperactive NLRP3 only when the endogenous mouse Nlrp3 promotor is active in dopaminergic neurons, were analyzed throughout 18 months of aging using longitudinal motor function assessments. Biochemical and histologic analyses of mesencephalic tissues were conducted in 1- and 18-month-old animals. RESULTS: We observed progressive and significant deficits in motor function in animals expressing Nlrp3L351P, compared with animals expressing Nlrp3WT and Nlrp3A350V. Age-dependent neuroinflammatory changes in the mesencephalon were noted in all animals. Analysis of GFAP-immunoreactive astrocytes in the substantia nigra revealed a significant increase in astrocyte number in animals expressing Nlrp3L351P compared with Nlrp3WT and Nlrp3A350V. Further analysis of Nlrp3L351P striatal tissues indicated genotype specific gliosis, elevated Il1b expression, and both morphologic and gene expression indicators of proinflammatory A1 astrocytes. CONCLUSIONS: Dopaminergic neurons have the potential to accumulate NLRP3 inflammasome activators with age, including reactive oxygen species, dopamine metabolites, and misfolded proteins. Results indicate the Nlrp3 locus is active in dopaminergic neurons in aging mice, and that the hyperactive Nlrp3L351P allele can drive neuroinflammatory changes in association with progressive behavioral deficits. Findings suggest neuronal NLRP3 inflammasome activity may contribute to neuroinflammation observed during normal aging and the progression of neurologic disorders.

Efficacy of Oral Cryotherapy During Oxaliplatin Infusion in Preventing Oral Thermal Hyperalgesia: A Randomized Trial.

BACKGROUND: Chemotherapy-induced oral thermal hyperalgesia (OTH) is a common and debilitating side effect of platinum-based anticancer agents. This study evaluated the efficacy of oral cryotherapy in preventing OTH during oxaliplatin chemotherapy infusion. METHODS: Patients with gastrointestinal cancer treated with biweekly oxaliplatin (85 mg/m2 over 120 minutes) at Abramson Cancer Center at the University of Pennsylvania were randomized to receive oral cryotherapy (ice chips) during oxaliplatin infusion or standard-of-care treatment. All patients completed baseline questionnaires regarding oral and peripheral symptoms and on-treatment questionnaires on day 1 of each subsequent chemotherapy cycle. Those in the treatment arm were asked to document how long they kept the ice chips in their mouths (0, <30, 30, 60, 90, or 120 minutes) and to report their discomfort associated with oral cryotherapy. Evaluable patients were those who had completed at least 2 cycles of oxaliplatin therapy. RESULTS: Of 62 randomized patients with a variety of gastrointestinal malignancies, 50 (25 per treatment arm) were evaluable for efficacy. The rate of patients with oral symptoms after the first treatment cycle was significantly lower in the intervention arm (n=8; 32%) than in the control arm (n=18; 72%), meeting the primary study objective (P=.01). The magnitude of difference in symptom scores before versus after the first treatment cycle was significantly less in the intervention versus control arm (P=.001). No difference in oral symptoms over time was seen between the intervention and control groups (P=.20), although a high attrition rate was noted. Duration of ice chip exposure was associated with improved oral symptoms over time (P=.02). CONCLUSIONS: Oral cryotherapy is a tolerable and cost-effective method of diminishing OTH in patients receiving oxaliplatin chemotherapy, and seems to be most effective in the early stages of treatment.

Fate of Triclosan in Irrigated Soil: Degradation in Soil and Translocation into Onion and Tomato.

This study determined the fate of triclosan, a prevalent wastewater contaminant in recycled waters and surface streams, when soil and crop plants were irrigated at environmentally relevant concentrations. Soil triclosan concentrations were monitored in an 8-wk and in a 16-wk study without plants to determine triclosan degradation. Onion ( O. Fedtsch.) and tomato ( L.) were assessed for growth and triclosan accumulation at four levels of triclosan exposure (0, 0.015, 0.15, and 1.5 µg L) in irrigation waters within ranges of those found in recycled waters and associated receiving streams. Onions were grown for 8 wk and tomatoes were grown for 8 wk (short-term study) and 12 wk (long-term study) in potting soil. Soil triclosan concentrations increased (5-fold) with triclosan levels applied to soils alone. With repeated application, the half-life of triclosan was 18 d, with low-level accumulation in soil. Bioaccumulation of triclosan was observed in all edible portions of onions (115-435 ng g), primarily in bulbs, with no discernible impact on biomass. In both short- and long-term tomato studies, triclosan translocated to shoots and fruits (approaching a translocation factor of 1) at the highest level examined. Even at low triclosan concentrations typically found in recycled waters and receiving streams, agricultural irrigation presents an additional exposure route for organic contaminants to humans via commercial crops. Our study indicates that bulb crops, in particular, would likely accumulate high levels of triclosan. However, concentrations detected in both onions and tomato fruits determined here are below current human exposure limits.

NLRP3 expression in mesencephalic neurons and characterization of a rare NLRP3 polymorphism associated with decreased risk of Parkinson's disease.

Neuroinflammation is a well-characterized pathophysiology occurring in association with the progression of Parkinson's disease. Characterizing the cellular and molecular basis of neuroinflammation is critical to understanding its impact on the incidence and progression of PD and other neurologic disorders. Inflammasomes are intracellular pro-inflammatory pattern-recognition receptors capable of initiating and propagating inflammation. These cellular complexes are well characterized in the innate immune system and activity of the NLRP3 inflammasome has been reported in microglia. NLRP3 inflammasome activity has been associated with Alzheimer's disease, and recent reports, from our laboratory and others, indicate that Nlrp3 is required for neuroinflammation and nigral cell loss in animal models of PD. NLRP3 has not yet been characterized in PD patients. Here we characterize NLRP3 in PD using immunohistologic and genetic approaches. Histologic studies revealed elevated NLRP3 expression in mesencephalic neurons of PD patients. Analysis of exome sequencing data for genetic variation of NLRP3 identified multiple single-nucleotide polymorphisms (SNPs) including rs7525979 that was associated with a significantly reduced risk of developing PD. Mechanistic studies conducted in HEK293 cells indicated that the synonymous SNP, NLRP3 rs7525979, alters the efficiency of NLRP3 translation impacting NLRP3 protein stability, ubiquitination state, and solubility. These data provide evidence that dopaminergic neurons are a cell-of-origin for inflammasome activity in PD and are consistent with recent animal studies, suggesting that inflammasome activity may impact the progression of PD.

Editor's Highlight: Nlrp3 Is Required for Inflammatory Changes and Nigral Cell Loss Resulting From Chronic Intragastric Rotenone Exposure in Mice.

Complex interactions between genetic and environmental factors are widely believed to underlie the incidence and progression of Parkinson's disease (PD). Rotenone is a naturally occurring metabolic toxin employed as an insecticide and piscicide identified as a risk factor for the development of PD in agricultural workers. The Nlrp3 inflammasome is an intracellular mediator that can initiate an inflammatory cascade in response to cellular stress. Reports by others indicating that NLRP3 expression was detectable in tissues obtained from Alzheimer's disease patients and that the PD-associated protein α-synuclein could activate inflammasomes in cultured glial cells, prompted us to test the prediction that Nlrp3 was required for the development of Parkinson's-like changes resulting from rotenone exposure in mice. We exposed wild type and Nlrp3-/- mice to chronic low doses of intragastric rotenone and conducted longitudinal behavioral and serum cytokine analysis followed by evaluation of neuroinflammatory and neurodegenerative endpoints in brain tissues. We observed progressive rotenone-dependent changes in serum cytokine levels and circulating leukocytes in wild type mice not observed in Nlrp3-/- mice. Analysis of brain tissues revealed Nlrp3-dependent neuroinflammation and nigral cell loss in mice exposed to rotenone as compared with mice exposed to vehicle alone. Together, our findings provide compelling evidence of a role for Nlrp3 in nigral degeneration and neuroinflammation resulting from systemic rotenone exposure and suggest that the suppression of NLRP3 activity may be a rational neuroprotective strategy for toxin-associated PD.