Biology subjects

Szeto, G.

Publications and source records attributed to Szeto, G..

6 recordsLinked to original sources

bioRxiv2022

Brain injury accelerates the onset of a reversible age-related microglial phenotype associated with hyperphagocytosis and inflammatory neurodegeneration

Lipofuscin is an autofluorescent (AF) pigment formed by lipids and misfolded proteins that accumulates in post-mitotic cells with advanced age. Here we immunophenotyped microglia in the brain of old C57BL/6 mice (>18 months-old) and demonstrate that in comparison to young mice, one third of old microglia are AF, characterized by profound changes in lipid and iron content, phagocytic activity, and oxidative stress. Pharmacological depletion of microglia in old mice eliminated the AF microglia following repopulation and reversed microglial dysfunction. Age-related neurological deficits and neurodegeneration after traumatic brain injury (TBI) were attenuated in old mice lacking AF microglia. Furthermore, hyperphagocytic activity and lipid accumulation in microglia persisted for up to one year after TBI, were modified by Apoe4 genotype, and chronically driven by phagocyte-mediated oxidative stress. Thus, AF may reflect a pathological state in aging microglia associated with hyperphagocytosis and inflammatory neurodegeneration that can be further accelerated by TBI. TeaserTraumatic brain injury accelerates age-related pathological phagocytosis and lipofuscin formation in microglia.

neuroscience↗

bioRxiv2022

Persistent serum protein signatures define an inflammatory subset of long COVID

Long COVID or post-acute sequelae of SARS-CoV-2 (PASC) is a clinical syndrome featuring diverse symptoms that can persist for months after acute SARS-CoV-2 infection. The etiologies are unknown but may include persistent inflammation, unresolved tissue damage, or delayed clearance of viral protein or RNA. Attempts to classify subsets of PASC by symptoms alone have been unsuccessful. To molecularly define PASC, we evaluated the serum proteome in longitudinal samples from 55 PASC individuals with symptoms lasting [≥]60 days after onset of acute infection and compared this to symptomatically recovered SARS-CoV-2 infected and uninfected individuals. We identified subsets of PASC with distinct signatures of persistent inflammation. Type II interferon signaling and canonical NF-{kappa}B signaling (particularly associated with TNF), were the most differentially enriched pathways. These findings help to resolve the heterogeneity of PASC, identify patients with molecular evidence of persistent inflammation, and highlight dominant pathways that may have diagnostic or therapeutic relevance. One Sentence SummarySerum proteome profiling identifies subsets of long COVID patients with evidence of persistent inflammation including key immune signaling pathways that may be amenable to therapeutic intervention.

immunology↗

bioRxiv2019

Lipid-mediated insertion of Toll-like receptor (TLR) ligands for facile immune cell engineering

Cell-based immunotherapies have tremendous potential to treat many diseases, such as activating immunity in cancer or suppressing it in autoimmune diseases. Most cell-based cancer immunotherapies in the clinic provide adjuvant signals through genetic engineering to enhance T cell functions. However, genetically encoded signals have minimal control over dosing and persist for the life of a cell lineage. These properties make it difficult to balance increasing therapeutic efficacy with reducing toxicities. Here, we demonstrated the potential of phospholipid-coupled ligands as a non-genetic system for immune cell engineering. This system provides simple, controlled, non-genetic adjuvant delivery to immune cells via lipid-mediated insertion into plasma membranes. Lipid-mediated insertion (depoting) successfully delivered Toll-like receptor (TLR) ligands intracellularly and onto cell surfaces of diverse immune cells. These ligands depoted into immune cells in a dose-controlled fashion and did not compete during multiplex pairwise loading. Immune cell activation could be enhanced by autocrine and paracrine mechanisms depending on the biology of the TLR ligand tested. We determined that depoted ligands can functionally persist on plasma membranes for up to four days in naive and activated T cells, enhancing their activation, proliferation, and skewing cytokine secretion. Depoted ligands provide a persistent yet non-permanent adjuvant signal to immune cells that may minimize the intensity and duration of toxicities compared to permanent genetic delivery. Altogether, these findings demonstrate potential for lipid-mediated insertion (depoting) as a universal cell engineering approach with unique, complementary advantages to other cell engineering methods.

immunology↗

bioRxiv2019

Microglial depletion with CSF1R inhibitor during chronic phase of experimental traumatic brain injury reduces neurodegeneration and neurological deficits

Chronic neuroinflammation with sustained microglial activation occurs following moderate-to-severe traumatic brain injury (TBI) and is believed to contribute to subsequent neurodegeneration and neurological deficits. Microglia, the primary innate immune cells in brain, are dependent on colony stimulating factor 1 receptor (CSF1R) signaling for their survival. In this translational study, we examined the effects of delayed depletion and subsequent repopulation of microglia on chronic neurodegeneration and functional recovery up to three months posttrauma. A CSF1R inhibitor, PLX5622, was administered to injured adult male C57Bl/6 mice at one month after controlled cortical impact to remove chronically activated microglia, and the inihibitor was withdrawn 1 week later to allow microglial repopulation. Following TBI, the repopulated microglia displayed a ramified morphology, similar to that of sham control uninjured animals, whereas microglia in untreated injured animals showed the typical chronic posttraumatic hypertrophic morphology. PLX5622 treatment limited TBI-associated neuropathological changes at 3 months posttrauma; these included a smaller cortical lesion, reduced neuronal cell death in the injured cortex and ipsilateral hippocampus, and decreased NOX2-dependent reactive microgliosis. Furthermore, delayed depletion of microglia led to widespread changes in the cortical transcriptome, including alterations in gene pathways involved in neuroinflammation, oxidative stress, and neuroplasticity. PLX5622 treated animals showed significantly improved neurological recovery using a variety of complementary neurobehavioral evaluations. These included beam walk and rotorod tests for sensori-motor function, as well as Y-maze, novel object recognition, and Morris water maze tests to evaluate cognitive function. Together, our findings show that chronic phase removal of neurotoxic microglia using CSF1R inhibitors after experimental TBI can markedly reduce chronic neuroinflammation and neurodegeneration, as well as related long-term motor and cognitive deficits. Thus, CSF1R inhibition may provide a clinically feasible approach to limit posttraumatic neurodegeneration and neurological dysfunction following head injury.

neuroscience↗

bioRxiv2019

Collagen hydrogel confinement of amyloid-β accelerates aggregation and reduces cytotoxic effects

Alzheimers disease (AD) is the most common form of dementia and is associated with the accumulation of amyloid-{beta} (A{beta}), a peptide whose aggregation has been associated with neurotoxicity. Drugs targeting A{beta} have shown great promise in 2D in vitro models and mouse models, yet preclinical and clinical trials for AD have been highly disappointing. We propose that current in vitro culture systems for discovering and developing AD drugs have significant limitations; specifically, that A{beta} aggregation is vastly different in these 2D cultures carried out on flat plastic or glass substrates vs. in a 3D environment, such as brain tissue, where A{beta} confinement very likely alters aggregation kinetics and thermodynamics. In this work, we identified attenuation of A{beta} cytotoxicity in 3D hydrogel culture compared to 2D cell culture. We investigated A{beta} structure and aggregation in solution vs. hydrogel using Transmission Electron Microscopy (TEM), Fluorescence Correlation Spectroscopy (FCS), and Thioflavin T (ThT) assays. Our results reveal that the equilibrium is shifted to stable {beta}-sheet aggregates in hydrogels and away from the relatively unstable/unstructured presumed toxic oligomeric A{beta} species in solution. Volume exclusion imparted by hydrogel confinement stabilizes unfolded, presumably toxic species, promoting stable extended {beta}-sheet fibrils. These results, taken together with the many recent reports that 3D hydrogel cell cultures enable cell morphologies and epigenetic changes that are more similar to cells in vivo compared to 2D cultures, strongly suggest that AD drugs should be tested in 3D culture systems as a step along the development pathway towards new, more effective therapeutics.

bioengineering↗

bioRxiv2019

Impact of four common hydrogels on amyloid-β (Aβ) aggregation and cytotoxicity: Implications for 3D models of Alzheimer’s disease

The properties of a hydrogel utilized in 3D culture can influence cell phenotype and morphology, yielding striking similarities to cellular processes that occur in vivo. Indeed, research areas including regenerative medicine, tissue engineering, cancer models, and stem cell cultures have readily utilized 3D biomaterials to investigate cell biological questions. However, cells are only one component of this milieu. Macromolecules play roles as bioactive factors and physical structures. Yet, investigations of macromolecular biophysics largely focus on pure molecules in dilute solution. Biophysical processes such as protein aggregation underlie diseases including Alzheimers disease, which is hallmarked by accumulated neurotoxic amyloid-{beta} (A{beta}) aggregates. Previously, we demonstrated that A{beta} cytotoxicity is attenuated when cells are cultured within type I collagen hydrogels vs. on 2D substrates. Here, we investigated whether this phenomenon is conserved when A{beta} is confined within hydrogels of varying physiochemical properties, notably mesh size and bioactivity. We investigated A{beta} structure and aggregation kinetics in solution and in hydrogels (collagen, agarose, hyaluronic acid and polyethylene glycol) using fluorescence correlation spectroscopy and thioflavin T assays. Our results reveal that all hydrogels tested were associated with A{beta} cytotoxicity attenuation. We suggest that confinement itself imparts a profound effect, possibly by stabilizing A{beta} structures and shifting the aggregate equilibrium toward larger species. It is likely that the milieu that exist within cells and tissues also influences protein-protein interactions; thus, we suggest that it is critical to evaluate whether protein structure, function, and stability are altered in 3D systems vs. ideal solutions and 2D culture.

bioengineering↗