Preferential and Increased Uptake of Hydroxyl-Terminated PAMAM Dendrimers by Activated Microglia in Rabbit Brain Mixed Glial Culture.

Polyamidoamine (PAMAM) dendrimers are multifunctional nanoparticles with tunable physicochemical features, making them promising candidates for targeted drug delivery in the central nervous system (CNS). Systemically administered dendrimers have been shown to localize in activated glial cells, which mediate neuroinflammation in the CNS. These dendrimers delivered drugs specifically to activated microglia, producing significant neurological improvements in multiple brain injury models, including in a neonatal rabbit model of cerebral palsy. To gain further insight into the mechanism of dendrimer cell uptake, we utilized an in vitro model of primary glial cells isolated from newborn rabbits to assess the differences in hydroxyl-terminated generation 4 PAMAM dendrimer (D4-OH) uptake by activated and non-activated glial cells. We used fluorescently-labelled D4-OH (D-Cy5) as a tool for investigating the mechanism of dendrimer uptake. D4-OH PAMAM dendrimer uptake was determined by fluorescence quantification using confocal microscopy and flow cytometry. Our results indicate that although microglial cells in the mixed cell population demonstrate early uptake of dendrimers in this in vitro system, activated microglia take up more dendrimer compared to resting microglia. Astrocytes showed delayed and limited uptake. We also illustrated the differences in mechanism of uptake between resting and activated microglia using different pathway inhibitors. Both resting and activated microglia primarily employed endocytotic pathways, which are enhanced in activated microglial cells. Additionally, we demonstrated that hydroxyl terminated dendrimers are taken up by primary microglia using other mechanisms including pinocytosis, caveolae, and aquaporin channels for dendrimer uptake.

In vitro and in situ visualization of cytoskeletal deformation under load: traumatic axonal injury.

It is difficult to obtain insight into the mechanisms occurring within live cells during mechanical loading, because this complex environment is dynamic and evolving. This is a particular challenge from a subcellular mechanics perspective, where temporal and spatial information on the evolving cytoskeletal structures is required under loading. Using fluorescently labeled proteins, we visualize 3-dimensional live subcellular cytoskeletal populations under mechanical loading using a high-resolution confocal microscope. The mechanical forces are determined using a computational (finite element) model that is validated by integrating instrumentation into the testing platform. Transfected microtubules and neurofilaments of E17 rat neuronal axons are imaged before, during, and after loading. Comparisons between unloaded and loaded live cells demonstrate both spatial and temporal changes for cytoskeletal populations within the imaged volume. NF signal decreases by 24%, yet the microtubule signal exhibits no significant change 20-35 s after loading. Transmission electron microscopy assesses cytoskeletal structure spatial distribution for undeformed and deformed axons. While cytoskeletal degeneration occurs at prolonged time intervals following loads, our data provides insights into real time cytoskeletal evolution occurring in situ. Our findings suggest that, for axons undergoing traumatic injury in response to applied mechanical loads, changes at the substructural level of neurofilaments may precede microtubule rupture and degeneration.

Toll-like receptor 4 deficiency impairs microglial phagocytosis of degenerating axons.

Microglia are rapidly activated in the central nervous system (CNS) in response to a variety of injuries, including inflammation, trauma, and stroke. In addition to modulation of the innate immune response, a key function of microglia is the phagocytosis of dying cells and cellular debris, which can facilitate recovery. Despite emerging evidence that axonal debris can pose a barrier to regeneration of new axons in the CNS, little is known of the cellular and molecular mechanisms that underlie clearance of degenerating CNS axons. We utilize a custom micropatterned microfluidic system that enables robust microglial-axon co-culture to explore the role of Toll-like receptors (TLRs) in microglial phagocytosis of degenerating axons. We find that pharmacologic and genetic disruption of TLR4 blocks induction of the Type-1 interferon response and inhibits phagocytosis of axon debris in vitro. Moreover, TLR4-dependent microglial clearance of unmyelinated axon debris facilitates axon outgrowth. In vivo, microglial phagocytosis of CNS axons undergoing Wallerian degeneration in a dorsal root axotomy model is impaired in adult mice in which TLR4 has been deleted. Since purinergic receptors can influence TLR4-mediated signaling, we also explored a role for the microglia P2 receptors and found that the P2X7R contributes to microglial clearance of degenerating axons. Overall, we identify TLR4 as a key player in axonal debris clearance by microglia, thus creating a more permissive environment for axonal outgrowth. Our findings have significant implications for the development of protective and regenerative strategies for the many inflammatory, traumatic, and neurodegenerative conditions characterized by CNS axon degeneration.

Behavioural and social drivers of COVID-19 vaccination in Vietnam: a scoping review.

BACKGROUND: Understanding of the behavioural and social drivers (BeSD) of vaccination is key to addressing vaccine hesitancy and accessibility issues. Vietnam's national COVID-19 vaccination programme resulted in high uptake of primary doses among adults, but lower booster doses for adults and primary doses for 5-11 years. This scoping review assessed BeSD influencing COVID-19 vaccine uptake in Vietnam to design interventions on reaching the national vaccination targets. METHOD: We conducted a scoping review by searching PubMed, MedRxiv, LitCOVID, COVID-19 LOVE platform, WHO's COVID-19 research database and seven dominant Vietnamese language medical journals published in English or Vietnamese between 28 December 2019 and 28 November 2022. Data were narratively synthesised and summarised according to the four components of the WHO BeSD framework. The drivers were then mapped along the timeline of COVID-19 vaccine deployment and the evolution of the pandemic in Vietnam. RESULTS: We identified 680 records, of which 39 met the inclusion criteria comprising 224 204 participants. Adults' intention to receive COVID-19 vaccines for themselves (23 studies) ranged from 58.0% to 98.1%. Parental intention to vaccinate their under 11-year-old children (six studies) ranged from 32.8% to 79.6%. Key drivers of vaccination uptake were perceived susceptibility and severity of disease, perceived vaccine benefits and safety, healthcare worker recommendation, and positive societal perception. Commonly reported COVID-19 vaccines' information sources (six studies) were social and mainstream media (82%-67%), television (72.7%-51.6%) and healthcare workers (47.5%-17.5%). Key drivers of COVID-19 uptake remained consistent for both adults and children despite changes in community transmission and vaccine deployment. CONCLUSION: Key enablers of vaccine uptake for adults and children included perceived disease severity, perceived vaccine benefits and safety and healthcare worker recommendations. Future studies should assess vaccine communication targeted to these drivers, national policies and political determinants to optimise vaccine uptake.

Use of telehealth services among Nepali living overseas during Covid-19 pandemic: The opportunities, limitations, lessons learned and recommendations.

There are estimated over 8 million Nepali migrants spread across various countries around the globe. Though the majority of them enjoy good health in general, a large proportion of them suffer from non-communicable diseases, mental health issues and communicable diseases. Telemedicine services, which are organized by Non-Resident Nepali Association (NRNA), have been proven to be effective in addressing some of the health and medical needs of the migrant Nepali workers. The purpose of this study is to assess the use of tele-health services among Nepali migrant population and examine the limitations. During the pandemic period from March 2020 through August 2021, Nepali in different countries utilized telehealth services. Mental health issues, chronic diseases, skin diseases were the most common ailments people sought telehealth services for. Many of them sought for urgent medical consultations related to Covid-19 symptoms and ailments. Digital gap, lack of cross-border regulations and unwillingness to utilize telemedicine were the challenges the service faced in the optimal utilization of such services. Training and education, use of easy Apps and subsidies from the government would help in the long-term use and sustainability of telehealth services amongst the Nepali migrants.

Differential regulation of brain-derived neurotrophic factor (BDNF) expression in sensory neuron axons by miRNA-206.

Distinct subcellular localization and subsequent translational control of 3' UTR variants of mRNA encoding brain-derived neurotrophic factor (BDNF) are critical for the development and plasticity of neurons. Although the processes that lead to preferential localization of BDNF have been well studied, it is still not clear how neurons ensure differential BDNF production in a spatial-specific manner. Here, we identified that microRNA (miRNA)-206 has the potential to specifically regulate BDNF with a long 3' UTR without affecting its short 3' UTR counterpart. Overexpression of miRNA-206 in sensory neurons resulted in a 30% and 45% reduction of BDNF protein expression in the cell bodies and axons, respectively. The work described in the present study indicates that miRNAs can differentially and specifically regulate the expression of transcript variants with different localization patterns.

Changes in Neurofilament and Microtubule Distribution following Focal Axon Compression.

Although a number of cytoskeletal derangements have been described in the setting of traumatic axonal injury (TAI), little is known of early structural changes that may serve to initiate a cascade of further axonal degeneration. Recent work by the authors has examined conformational changes in cytoskeletal constituents of neuronal axons undergoing traumatic axonal injury (TAI) following focal compression through confocal imaging data taken in vitro and in situ. The present study uses electron microscopy to understand and quantify in vitro alterations in the ultrastructural composition of microtubules and neurofilaments within neuronal axons of rats following focal compression. Standard transmission electron microscopy processing methods are used to identify microtubules, while neurofilament identification is performed using antibody labeling through gold nanoparticles. The number, density, and spacing of microtubules and neurofilaments are quantified for specimens in sham Control and Crushed groups with fixation at <1 min following load. Our results indicate that the axon caliber dependency known to exist for microtubule and neurofilament metrics extends to axons undergoing TAI, with the exception of neurofilament spacing, which appears to remain constant across all Crushed axon diameters. Confidence interval comparisons between Control and Crushed cytoskeletal measures suggests early changes in the neurofilament spatial distributions within axons undergoing TAI may precede microtubule changes in response to applied loads. This may serve as a trigger for further secondary damage to the axon, representing a key insight into the temporal aspects of cytoskeletal degeneration at the component level, and suggests the rapid removal of neurofilament sidearms as one possible mechanism.

Curcumin protects axons from degeneration in the setting of local neuroinflammation.

Axon degeneration is a hallmark of several central nervous system (CNS) disorders, including multiple sclerosis (MS), Alzheimer's disease (AD) and Parkinson's disease (PD). Previous neuroprotective approaches have mainly focused on reversal or prevention of neuronal cell body degeneration or death. However, experimental evidence suggests that mechanisms of axon degeneration may differ from cell death mechanisms, and that therapeutic agents that protect cell bodies may not protect axons. Moreover, axon degeneration underlies neurologic disability and may, in some cases, represent an important initial step that leads to neuronal death. Here, we develop a novel quantitative microfluidic-based methodology to assess mechanisms of axon degeneration caused by local neuroinflammation. We find that LPS-stimulated microglia release soluble factors that, when applied locally to axons, result in axon degeneration. This local axon degeneration is mediated by microglial MyD88/p38 MAPK signaling and concomitant production of nitric oxide (NO). Intra-axonal mechanisms of degeneration involve JNK phosphorylation. Curcumin, a compound with both anti-oxidant and JNK inhibitory properties, specifically protects axons, but not neuronal cell bodies, from NO-mediated degeneration. Overall, our platform provides mechanistic insights into local axon degeneration, identifies curcumin as a novel axon protectant in the setting of neuroinflammation, and allows for ready screening of axon protective drugs.

The interacting effect of cognitive and motor task demands on performance of gait, balance and cognition in young adults.

Mobility limitations and cognitive impairments, each common with aging, reduce levels of physical and mental activity, are prognostic of future adverse health events, and are associated with an increased fall risk. The purpose of this study was to examine whether divided attention during walking at a constant speed would decrease locomotor rhythm, stability, and cognitive performance. Young healthy participants (n=20) performed a visuo-spatial cognitive task in sitting and while treadmill walking at 2 speeds (0.7 and 1.0 m/s).Treadmill speed had a significant effect on temporal gait variables and ML-COP excursion. Cognitive load did not have a significant effect on average temporal gait variables or COP excursion, but variation of gait variables increased during dual-task walking. ML and AP trunk motion was found to decrease during dual-task walking. There was a significant decrease in cognitive performance (success rate, response time and movement time) while walking, but no effect due to treadmill speed. In conclusion walking speed is an important variable to be controlled in studies that are designed to examine effects of concurrent cognitive tasks on locomotor rhythm, pacing and stability. Divided attention during walking at a constant speed did result in decreased performance of a visuo-spatial cognitive task and an increased variability in locomotor rhythm.