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OBJECTIVE: The wound biofilm infections that develop tolerance to standard-of-care antimicrobial treatment has been increasing. The objective of this study was to demonstrate a proof-of-concept of mild magnetic nanoparticle (MNP)/alternating magnetic field (AMF) hyperthermia as an anti-biofilm therapy against multispecies biofilm infections. METHODS: Using both an in vitro cell culture and in vivo murine model of wound infection, we investigated whether MNP/AMF hyperthermia applied at a mild thermal dosage would be synergistically effective against dual species biofilm infection consisting of S. aureus and P. aeruginosa when combined with a broad-spectrum antibiotic, ciprofloxacin (CIP). RESULTS: The combined treatment of MNP/AMF hyperthermia and CIP to the wounds of diabetic mice (db/db mice) significantly reduced the CFU number of S. aureus and P. aeruginosa by 2-log and 3-log, respectively, compared to the untreated control group, whereas either mild MNP/AMF hyperthermia or CIP treatment alone had little effect on the eradication of both bacteria. Our gene microarray data obtained from the culture of S. aureus biofilm suggest that mild MNP/AMF could shift the expression of genes for cellular respiration from anaerobic fermentation to an aerobic glycolytic/tricarboxylic acid cycle (TCA) pathway, implicating that the beneficial effect of mild MNP/AMF hyperthermia on the increased susceptibility of biofilm bacteria to an antibiotic treatment is associated with an increased metabolic activity. CONCLUSION: Our results support the translational potential of mild MNP/AMF as an adjunctive therapy that can be combined with a broad-spectrum antibiotic treatment for the management of wound biofilm infections associated with multispecies bacteria.
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Diabetes Mellitus Experimental , Hipertermia Induzida , Staphylococcus aureus Resistente à Meticilina , Camundongos , Animais , Staphylococcus aureus , Pseudomonas aeruginosa , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Biofilmes , Ciprofloxacina/farmacologia , Ciprofloxacina/uso terapêutico , Fenômenos MagnéticosRESUMO
The formation of beta-amyloid (Aß) plaques is a classical hallmark of Alzheimer's disease (AD) that is associated with the promotion of neuroinflammation and subsequent neurotoxicity. Given the limited therapeutic options for targeting and clearing Aß plaques in AD, there is an urgent need to develop effective approaches to reduce plaque accumulation. The objective of this study was to validate mild magnetic nanoparticle (MNP) hyperthermia technology as a strategy to clear Aß deposits and determine the impact on microglia functionality. Our results demonstrated that the heating of MNPs localized to Aß aggregates upon exposure to high frequency alternating magnetic field (AMF) was sufficient to disrupt Aß plaques, resulting in its fragmentation. Importantly, this could facilitate the phagocytic clearance of Aß as well as attenuate pro-inflammatory responses by human microglial cells. Our results support the feasibility of mild MNP/AMF hyperthermia as a new strategy for reducing beta-amyloid burdens in Alzheimer's disease.
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Peptídeos beta-Amiloides/isolamento & purificação , Hipertermia Induzida/métodos , Magnetismo , Microglia/metabolismo , Nanopartículas , Placa Amiloide/metabolismo , Peptídeos beta-Amiloides/metabolismo , Linhagem Celular Transformada , Humanos , Microscopia Eletrônica de Varredura , FagocitoseRESUMO
Microglia are the primary resident immune cells of the central nervous system that are responsible for the maintenance of brain homeostasis. There is a plethora of evidence to suggest that microglia display distinct phenotypes that are associated with the alteration of cell morphology under varying environmental cues. However, it has not been fully explored how the varying states of microglial activation are linked to the alteration of microglia morphology, especially in the microdomain. The objective of this study was to quantitatively characterize the ultrastructural morphology of human microglia under neuroinflammatory cues. To address this, a human cell line of microglia was stimulated by antiinflammatory (IL-4), proinflammatory (TNF-α), and Alzheimer's disease (AD)-associated cues (Aß, Aß + TNF-α). The resulting effects on microglia morphology associated with changes in microdomain were analyzed using a high-resolution scanning electron microscopy. Our findings demonstrated that microglial activation under proinflammatory and AD-cues were closely linked to changes not only in cell shape but also in cell surface topography and higher-order branching of processes. Furthermore, our results revealed that microglia under proinflammatory cues exhibited unique morphological features involving cell-to-cell contact and the formation of vesicle-like structures. Our study provides insight into the fine details of microglia morphology associated with varying status of microglial activation.
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Doença de Alzheimer , Microglia , Doença de Alzheimer/metabolismo , Encéfalo/metabolismo , Sinais (Psicologia) , Humanos , Microglia/metabolismo , FenótipoRESUMO
Both neurons and glia in mammalian brains are highly ramified. Neurons form complex neural networks using axons and dendrites. Axons are long with few branches and form pre-synaptic boutons that connect to target neurons and effector tissues. Dendrites are shorter, highly branched, and form post-synaptic boutons. Astrocyte processes contact synapses and blood vessels in order to regulate neuronal activity and blood flow, respectively. Oligodendrocyte processes extend toward axons to make myelin sheaths. Microglia processes dynamically survey their environments. Here, we describe the local secretory system (ER and Golgi) in neuronal and glial processes. We focus on Golgi outpost functions in acentrosomal microtubule nucleation, cargo trafficking, and protein glycosylation. Thus, satellite ER and Golgi are critical for local structure and function in neurons and glia.
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Axônios , Dendritos , Animais , Axônios/metabolismo , Dendritos/metabolismo , Complexo de Golgi/metabolismo , Mamíferos , Neurônios , SinapsesRESUMO
Variants in the high confident autism spectrum disorder (ASD) gene ANK2 target both ubiquitously expressed 220 kDa ankyrin-B and neurospecific 440 kDa ankyrin-B (AnkB440) isoforms. Previous work showed that knock-in mice expressing an ASD-linked Ank2 variant yielding a truncated AnkB440 product exhibit ectopic brain connectivity and behavioral abnormalities. Expression of this variant or loss of AnkB440 caused axonal hyperbranching in vitro, which implicated AnkB440 microtubule bundling activity in suppressing collateral branch formation. Leveraging multiple mouse models, cellular assays, and live microscopy, we show that AnkB440 also modulates axon collateral branching stochastically by reducing the number of F-actin-rich branch initiation points. Additionally, we show that AnkB440 enables growth cone (GC) collapse in response to chemorepellent factor semaphorin 3 A (Sema 3 A) by stabilizing its receptor complex L1 cell adhesion molecule/neuropilin-1. ASD-linked ANK2 variants failed to rescue Sema 3A-induced GC collapse. We propose that impaired response to repellent cues due to AnkB440 deficits leads to axonal targeting and branch pruning defects and may contribute to the pathogenicity of ANK2 variants.
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Anquirinas/genética , Orientação de Axônios/genética , Axônios/fisiologia , Semaforina-3A/genética , Transdução de Sinais/genética , Animais , Anquirinas/metabolismo , Camundongos , Semaforina-3A/metabolismoRESUMO
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Melanocortin 4 receptor (MC4R) variants contribute to human obesity, and rats lacking functional MC4R (Mc4rK314X/K314X) are obese. We investigated the hypothesis that low energy expenditure (EE) and physical activity contribute to this obese phenotype in male rats, and determined whether lack of functional MC4R conferred protection from weight loss during 50% calorie restriction. Though Mc4rK314X/K314X rats showed low brown adipose Ucp1 expression and were less physically active than rats heterozygous for the mutation (Mc4r+/K314X) or wild-type (Mc4r+/+) rats, we found no evidence of lowered EE in Mc4rK314X/K314X rats once body weight was taken into account using covariance. Mc4rK314X/K314X rats had a significantly higher respiratory exchange ratio. Compared to Mc4r+/+ rats, Mc4rK314X/K314X and Mc4r+/K314X rats lost less lean mass during calorie restriction, and less body mass when baseline weight was accounted for. Limited regional overexpression of Mc3r was found in the hypothalamus. Although lower physical activity levels in rats with nonfunctional MC4R did not result in lower total EE during free-fed conditions, rats lacking one or two functional copies of Mc4r showed conservation of mass, particularly lean mass, during energy restriction. This suggests that variants affecting MC4R function may contribute to individual differences in the metabolic response to food restriction.