Toward discovering a novel family of peptides targeting neuroinflammatory states of brain microglia and astrocytes.

Microglia are immune-derived cells critical to the development and healthy function of the brain and spinal cord, yet are implicated in the active pathology of many neuropsychiatric disorders. A range of functional phenotypes associated with the healthy brain or disease states has been suggested from in vivo work and were modeled in vitro as surveying, reactive, and primed sub-types of primary rat microglia and mixed microglia/astrocytes. It was hypothesized that the biomolecular profile of these cells undergoes a phenotypical change as well, and these functional phenotypes were explored for potential novel peptide binders using a custom 7 amino acid-presenting M13 phage library (SX7) to identify unique peptides that bind differentially to these respective cell types. Surveying glia were untreated, reactive were induced with a lipopolysaccharide treatment, recovery was modeled with a potent anti-inflammatory treatment dexamethasone, and priming was determined by subsequently challenging the cells with interferon gamma. Microglial function was profiled by determining the secretion of cytokines and nitric oxide, and expression of inducible nitric oxide synthase. After incubation with the SX7 phage library, populations of SX7-positive microglia and/or astrocytes were collected using fluorescence-activated cell sorting, SX7 phage was amplified in Escherichia coli culture, and phage DNA was sequenced via next-generation sequencing. Binding validation was done with synthesized peptides via in-cell westerns. Fifty-eight unique peptides were discovered, and their potential functions were assessed using a basic local alignment search tool. Peptides potentially originated from proteins ranging in function from a variety of supportive glial roles, including synapse support and pruning, to inflammatory incitement including cytokine and interleukin activation, and potential regulation in neurodegenerative and neuropsychiatric disorders.

Supporting microglial niches for therapeutic benefit in psychiatric disorders.

Inflammation is an essential tissue response to injury, stress, or infection resulting in debris and/or pathogen clearance intended to promote healing and recovery. Due to the status as an immune 'privileged' tissue, microglia serve as endogenous regulators of inflammation in the central nervous system, but maintain communication with peripheral immune system to enable recruitment of peripheral immune cells in case of injury or infection. While microglia retain the functional capacity for a full range of inflammatory functions - microglia express a range of pattern-recognition receptors and function as innate immune cells, carry out phagocytosis of pathogens, and act as antigen presenting cells - in the healthy central nervous system (CNS) these functions are rarely engaged. Subsequently microglia are being recognized to occupy an increasing number of homeostatic niches, and in many cases have adopted immune or inflammatory mechanisms to carry out these niche functions absent immune activation. These sterile inflammatory functions are challenging long-held views of the role of inflammation in the central nervous system while simultaneously expanding the potential for the development of truly novel therapeutic interventions for a range of neuroinflammatory, neurodegenerative, and neuropsychiatric disorders. In the present review we discuss recent preclinical evidence for conserved niche functions for microglia whose disruption may causally contribute to various psychiatric disorders, and prospective targets for restoring disrupted niches.

Biomaterials and glia: Progress on designs to modulate neuroinflammation.

Microglia are multi-functional cells that play a vital role in establishing and maintaining the function of the nervous system and determining the fate of neurons following injury or neuropathology. The roles of microglia are diverse and essential to the capacity of the nervous system to recover from injury, however sustained inflammation can limit recovery and drive chronic disease processes such as neurodegenerative disorders. When assessing implantable therapeutic devices in the central nervous system, an improved lifetime of the implant is considered achievable through the attenuation of microglial inflammation. Consequently, there is a tremendous underexplored potential in biomaterial and engineered design to modulate neuroinflammation for therapeutic benefit. Several strategies for improving device compatibility reviewed here include: biocompatible coatings, improved designs in finer and flexible shapes to reduce tissue shear-related scarring, and loading of anti-inflammatory drugs. Studies about microglial cell cultures in 3D hydrogels and nanoscaffolds to assess various injuries and disorders are also discussed. A variety of other microglia-targeting treatments are also reviewed, including nanoparticulate systems, cellular backpacks, and gold plinths, with the intention of delivering anti-inflammatory drugs by targeting the phagocytic nature of microglia. Overall, this review highlights recent advances in biomaterials targeting microglia and inflammatory function with the potential for improving implant rejection and biocompatibility studies. STATEMENT OF SIGNIFICANCE: Microglia are the resident immune cells of the central nervous system, and thus play a central role in the neuroinflammatory response against conditions than span acute injuries, neuropsychiatric disorders, and neurodegenerative disorders. This review article presents a summary of biomaterials research that target microglia and other glial cells in order to attenuate neuroinflammation, including but not limited to: design of mechanically compliant and biocompatible stimulation electrodes, hydrogels for high-throughput 3D modelling of nervous tissue, and uptake of nanoparticle drug delivery systems. The goal of this paper is to identify strengths and gaps in the relevant literature, and to promote further consideration of microglia behaviour and neuroinflammation in biomaterial design.

Brain biocompatibility and microglia response towards engineered self-assembling (RADA)4 nanoscaffolds.

(RADA)4-based nanoscaffolds have many inherent properties making them amenable to tissue engineering applications: ease of synthesis, ease of customization with bioactive moieties, and amenable for in situ nanoscaffold formation. There is a dearth in the literature on their biocompatibility in brain tissues; where the glia response is key to regulating the local host response. Herein, nanoscaffolds composed of (RADA)4 and (RADA)4-IKVAV mixtures were evaluated in terms of their effect on primary microglia in culture and general tissue (in vivo) biocompatibility (astrocyte and migroglia). Laminin-derived IKVAV peptide was chosen to promote beneficial cell interaction and attenuate deleterious glial responses. Microglia remained ramified when cultured with these nanoscaffolds, as observed using TNF-α and IL-1ß, NO, and proliferation assays. Evidence suggests that cultured microglia phagocytise the matrix whilst remaining ramified and viable, as shown visually and metabolically (MTT). Nanoscaffold intracerebral injection did not lead to microglia migration or proliferation, nor were glial scarring and axonal injury observed over the course of this study. IKVAV had no affect on microglia activation and astrogliosis. (RADA)4 should be advantageous for localized injection as a tuneable-platform device, which may be readily cleared without deleterious effects on tissue-resident microglia. STATEMENT OF SIGNIFICANCE: Self-assembling nanoscaffolds have many inherent properties making them amenable to tissue engineering applications: ease of synthesis, ease of customization with bioactive moieties, and amenable for in situ nanoscaffold formation. A dearth of literature exists on their biocompatibility in brain tissues; where the glia response is key to regulating the local host response. Herein, nanoscaffolds composed of the peptides (RADA)4 and (RADA)4-IKVAV mixtures were evaluated in terms of their effect on microglia cells in culture and general tissue (in vivo) biocompatibility (astrocyte and migroglia). Laminin-derived IKVAV peptide was chosen to promote beneficial cell interaction and attenuate deleterious glial responses. (RADA)4 nanoscaffolds showed no adverse effect from these cell types and should be advantageous for localized injection as a tuneable-platform device.

Fluoxetine and citalopram decrease microglial release of glutamate and D-serine to promote cortical neuronal viability following ischemic insult.

Depression is one of the most common disorders appearing following a stroke, and is also a major factor limiting recovery and rehabilitation in stroke patients. Antidepressants are the most common prescribed treatment for depression and have shown to have anti-inflammatory properties within the central nervous system (CNS). The major source of pro-inflammatory factors within the CNS is from activated microglia, the innate immune cells of the CNS. Antidepressants have been shown to promote midbrain and hippocampal neuronal survival following an ischemic insult and this survival is mediated through the anti-inflammatory effects on microglia, but the effects on cortical neuronal survival after this insult have yet to be investigated. The present study aimed to test and compare antidepressants from three distinct classes (tricyclics, monoamine oxidase inhibitors, and selective serotonin-reuptake inhibitors [SSRIs]) on the release of inflammatory factors and amino acids from activated microglia and whether altering this release could affect cortical neuronal viability after an ischemic insult. Primary microglia were treated with 1 µg/ml LPS and/or 10 µM antidepressants, and the various factors released into medium were assayed. Co-cultures consisting of microglia and primary cortical neurons were used to assess the effects of antidepressant-treated activated microglia on the viability of ischemic injured neurons. Of the antidepressants tested, most decreased the release of the proinflammatory factors nitric oxide, tumor necrosis factor-alpha, and interleukin 1-beta from activated microglia. Fluoxetine and citalopram, the SSRIs, decreased the release of the amino acids glutamate and d-serine from LPS-activated microglia. oxygen-glucose deprived (OGD) cortical neurons cocultured with LPS-activated microglia pre-treated with fluoxetine and citalopram showed greater survival compared to injured neurons co-cultured with untreated activated microglia. Microglial release of glutamate and d-serine was shown to be the most important factor mediating neuronal survival following antagonism studies. To our knowledge, our results demonstrate for the first time that fluoxetine and citalopram decrease the release of glutamate and d-serine from LPS-activated microglia and this causes an increase in the survival of OGD-injured cortical neurons after co-culture.