TREM2 is an innate immune receptor expressed in microglia and macrophages. ApoE isoforms (E2/E3/E4) are ligands of TREM2. TREM2 variants and ApoE4 are top risk factors of Alzheimer’s disease. TREM2 can be cleaved from cell membrane as soluble TREM2 (sTREM2), the level of which fluctuates during Alzheimer’s progression. However, the mechanisms that sTREM2 and the interactions between TREM2 and ApoE may contribute to Alzheimer’s neuroinflammation are largely uncharacterized. The project objectives were to investigate whether sTREM2 and ApoE isoforms can affect cytokine expression profiles in myeloid-derived cell models. My results show that sTREM2 can stimulate inflammatory cytokine expression at early time-point but anti-inflammatory cytokine expression at later time-point mainly via MAPK-JNK signaling pathway. sTREM2 has differential effects on cytokine expression in M0, M1, and M2 macrophages. ApoE isoforms also differentially induce cytokine expression and regulate TREM2 expression in M0, M1, and M2 macrophages. My study reveals a complex interplay of sTREM2, TREM2 and ApoE isoforms and differential effects of those in the models.

Alzheimer’s disease (AD) is a widespread neurodegenerative disorder that affects tens of millions of patients worldwide. Throughout the last two decades an incredible amount of time and resources have been funneled into hopefully finding medications that would provide a cure. Unfortunately, no such compound has been identified and instead the only FDA approved medications for AD to date target symptomatic management and may not even be effective for longer than a couple of years. To this end, this paper sets out to identify modifiable risk factors for AD as well as provide recommendations for clinicians on how best to utilize the tools currently available to them to treat AD. Additionally this paper addresses common flaws in AD clinical trial study designs and provides future research directions to expand outside of the popular amyloid hypothesis and instead potentially focus on a multi-pathway mechanism of the disease. The following thesis will outline several potential mechanisms that can lead to the hallmark pathologies seen in AD, primarily amyloid deposition and neurofibrillary tangles as well as neuronal death. The majority of commercial and research interest into AD has been focused on the amyloid hypothesis and the notion that stopping the formation of amyloid plaques would stop the disease course. However, in recent years other mechanisms and neurotoxic pathways such as inhibition of tricarboxylic acid (TCA) cycle enzymes, neuroinflammation, and tauopathy have been shown to contribute both to the formation of amyloid plaques as well as contributing to AD pathology in their own right. The modifiable risk factors explored in this paper include the effects of triglycerides as well as intake of antioxidant vitamins and omega-3-fatty acids, both of which are beneficial for brain health. This paper will also highlight some of the extensive research on the Apolipoprotein E gene and the effects the various alleles have on AD risk. These being the putative protective effect of the APOE2 allele, “neutral” effect of the most commonly found APOE3 allele, and finally the deleterious effects of the APOE4 allele, believed to be the strongest genetic risk factor for late-onset AD.

Microglia are a unique type of immune cell found within the brain, spinal cord and retina. In the healthy brain, their job is to support neurons, defend against infectious microbes, clear extracellular debris and remove dead or dying cells through phagocytosis. This diverse array of functions presents the possibility of unique subsets of microglia existing in the healthy brain, yet none have been described thus far. By utilizing cellular autofluorescence as a discriminating characteristic, we identified two novel subsets of microglia present in the healthy brains of mice and non-human primates. Approximately 70% of microglia displayed autofluorescence (AF+) while the remaining 30% did not (AF–). While the proportion of AF+ and AF– microglia remained constant throughout most of adult life, the autofluorescence intensity increased exclusively in the AF+ subset at an almost linear rate with age. This gain in autofluorescence correlated with equivalent increases in the size and complexity of storage bodies, as detected by transmission electron microscopy and increases in LAMP1 levels, a key component of the lysosomal compartment. As the brain ages, lysosomal storage material builds up inside AF+ microglia, further increasing the accumulation of autofluorescence as a result. The analysis of protein content in autofluorescent subsets revealed that AF+ microglia produced more proteins and enzymes involved in the storage and degradation of waste material, as well as more proteins involved in the regulation of mTOR, a key cellular pathway governing nutrient availability and energy production. Interestingly, the disruption of lysosomal function in microglia through genetic mutations accelerated the accumulation of storage material in AF+ cells, which led to impaired microglia physiology and increased cell death, mimicking the effects observed during advanced aging. Increasing evidence suggests that the accumulation of waste materials inside the brain contributes to diseases of aging and these data are suggestive of a mechanistic convergence between aging and lysosomal storage disorders.

Alzheimer's disease (AD) is defined as a progressive neurodegenerative disorder and is characterized by a devastating mental decline. There are three pathological hallmarks of the disease necessary for its diagnosis, these are extracellular amyloid plaques comprised of the beta-amyloid (Aβ) protein, intracellular neurofibrillary tangles comprised of hyperphosphorylated tau protein, and marked neuronal loss. Active immunization against Aβ1-42 or passive immunization with monoclonal anti-Aβ antibodies has been shown to reduce amyloid deposition and improve cognition in transgenic mouse models of AD, aged beagles, and nonhuman primates. Unfortunately, due to cerebrovascular adverse events, both active and passive immunization strategies targeting Aβ have failed in clinical trials. It is, therefore, necessary to identify novel amyloid-clearing therapeutics that do not induce cerebrovascular adverse events. We hypothesized that neuroinflammatory modulation could be a potential novel target. Triggering receptor expressed on myeloid cells-2 (TREM2) is a lipid and lipoprotein binding receptor expressed exclusively in the brain by microglia. Homozygous TREM2 loss of function mutations cause early-onset progressive presenile dementia while heterozygous, function-reducing point mutations triple the risk of sporadic, late-onset AD. Heterozygous TREM2 point mutations, which reduce either ligand binding or cell surface expression, are associated with a reduction in the number of microglia surrounding amyloid plaques, microglial inability to phagocytose compact Aβ deposits and form a barrier between plaques and neurons, an increase in the number of phospho- tau-positive dystrophic neurites and increased tau in the cerebrospinal fluid. Heterozygous mutations also double the rate of brain atrophy and decrease the age of AD onset by 3-6 years. Although human genetics supports the notion that loss of TREM2 function exacerbates neurodegeneration, it is unclear whether activation of TREM2 in a disease state is beneficial. The work we present here characterizes a TREM2 agonizing antibody as a potential therapeutic for amyloid reduction. We found that its administration results in immune modulation, recruitment of microglia to the site of amyloid plaques, reduced amyloid deposition and improvement in spatial learning and novel object recognition memory in the 5xFAD model of AD. More specifically, we show that intracranial injection of TREM2 agonizing antibodies into the frontal cortex and hippocampus of 5xFAD mice leads to clearance of diffuse and compact amyloid. We also show that systemic injection of TREM2 agonizing antibodies weekly over a period of 14 weeks results in clearance of diffuse and compact amyloid as well as elevated plasma concentrations of Aβ1-40 and Aβ1-42. Furthermore, systemic administration of these antibodies led to immune modulation and enhanced cognitive performance on radial arm water maze and novel object recognition tests. Importantly, we show the TREM2 agonizing antibody does not induce the adverse cerebrovascular events known to accompany amyloid modifying therapies. Though systemic administration of both TREM2 agonizing and anti-Abantibodies does not further enhance amyloid clearance or cognitive performance, co-administration mitigates the adverse cerebrovascular events associated with anti-Aβ antibodies. Collectively, these data indicate TREM2 activators may be an effective therapeutic target for the treatment of AD.

Indiana University-Purdue University Indianapolis (IUPUI) / Triggering Receptor Expressed on Myeloid Cells 2 (TREM2), a receptor expressed in myeloid cells including microglia in brain and osteoclasts in bone has been proposed as a link between brain and bone disease. Previous studies identified an AD-associated mutation (R47H) which is known to confer an increased risk for developing AD. In these studies, we used a heterozygous model of the TREM2 R47H variant (TREM2R47H/+), which does not exhibit cognitive defects, as a translational model of genetic risk factors that contribute to AD, and investigated whether alterations to TREM2 signaling could also contribute to bone and skeletal muscle loss, independently of central nervous system defects. Our study found that female TREM2R47H/+ animals experience bone loss in the femoral mid-diaphysis between 4 and 13 months of age as measured by microCT, which stalls out by 20 months of age. Female TREM2R47H/+ animals also experience significant decreases in the mechanical and material properties of the femur measured by three-point bending at 13 months of age, but not at 4 or 20 months. Interestingly, male TREM2R47H/+ animals do not demonstrate any discernable differences in bone geometry or strength until 20 months of age, where we observed slight changes in the bone volume and material properties of male TREM2R47H/+ bones. Ex vivo osteoclast differentiation assays demonstrate that only male TREM2R47H/+ osteoclasts differentiate more after 7 days with osteoclast differentiation factors compared to WT, but qPCR follow-up showed sexdependent differences in intracellular signaling. However, bone is not the only musculoskeletal tissue affected by the TREM2 R47H variant. Skeletal muscle strength measured by both in vivo plantar flexion and ex vivo contractility of the soleus is increased and body composition is altered in female TREM2R47H/+ mice compared to WT, and this is not likely due to bone-muscle crosstalk. These studies suggests that TREM2 R47H expression in the bone and skeletal muscle are likely impacting each tissue independently. These data demonstrate that AD-associated variants in TREM2 can alter bone and skeletal muscle strength in a sex-dimorphic manner independent of the presence of central neuropathology.

No description available.