Traumatic brain injury (TBI) is a leading cause of death and disability worldwide and has been established as a risk factor for neurodegenerative diseases such as Alzheimer’s disease (AD). Neurofibrillary tangles (NFTs), aggregates of intracellular tau, are hallmarks of AD and are observed in the post-TBI brain; however, the mechanisms that contribute to tau aggregation and accumulation are not well understood. One key mechanism that may contribute to this tau aggregation is decreased clearance by the glymphatic system, a perivascular pathway that clears solutes, including tau, from the brain. PS19 mice with tau pathology were crossed with Aqp4-/- mice lacking the astroglial water channel aquaporin-4 (AQP4) or Snta1-/- mice lacking perivascular localization of AQP4. Behavioral tests were performed on the PS19:Aqp4 transgenic crosses at 4 or 6 months of age. Brain tissue was collected and stained for markers of phosphorylated-tau (p-tau) pathology. Sham or mild TBIs were performed on PS19:Snta1 transgenic crosses at 3 months of age. I performed behavioral testing at 4 months (1 month post-TBI) or 6 months (3 month post-TBI). Brain tissue was collected and stained for markers of p-tau pathology. I imaged this immunostained tissue and quantified the pathological tau burden. I observed that Aqp4 deletion was sufficient to exacerbate tau pathology in PS19 mice at 6 months old, in the absence of TBI, and more advanced tau pathology was observed in PS19+Snta1-/- mice at 6 months old (3 months post-TBI) compared to PS19+Snta1+/+ that also received a TBI. Loss of AQP4 or loss of perivascular AQP4 promotes tau pathology in a mouse model of tau pathology. These studies may provide a mechanistic basis for the vulnerability of the post-traumatic brain to tau aggregation and neurodegeneration and suggest that targeting glymphatic dysfunction may be useful in the prevention and treatment of neurodegeneration.

Activated neutrophils eject neutrophil extracellular traps (NETs) consisting of web-like structures of DNA, histones, and granular proteins. Neutrophils traffic into the brain in Alzheimer’s disease (AD) and along with NETs are associated with amyloid β (Aβ) plaques. However, whether NETs may impair glymphatic clearance of Aβ has never been examined. The glymphatic system is a network of perivascular pathways that provides a pathway for solutes to be cleared from the brain interstitium. Impairment of glymphatic function promotes Aβ plaque deposition. We proposed that NETs may decrease glymphatic exchange by impairing perivascular and interstitial fluid movement, resulting in impaired Aβ clearance. 5XFAD mice with amyloid pathology were crossed with Pad4^-/- mice lacking the ability to form NETs, generating 5XFAD mice lacking NET formation. I performed Barnes maze, spontaneous alternation Y maze, and open field behavioral tests on 5XFAD⁺Pad4^+/+ and 5XFAD⁺Pad4^-/- mice at 4 months old. The following week, tracer was intracisternally injected into the CSF and dynamic transcranial imaging was performed to measure tracer movement over the cortical surface. Brains were then collected after 45 minutes. I measured tracer influx in these brain slices and the tissue was stained for Aβ, GFAP, Iba1, and neutrophil markers. I imaged this stained tissue and analyzed the immunostaining. Overall, Pad4 gene deletion in a 5XFAD mouse model of Aβ pathology improved tracer influx, decreased Aβ burden, and improved cognitive function. These studies highlight the important role that neutrophils and NETs play in the pathology of AD and may be a novel therapeutic target.

Mild traumatic brain injury (mTBI) has emerged as a potential risk factor for development of neurodegenerative conditions such as Alzheimer’s disease (AD) and chronic traumatic encephalopathy (CTE). Blast mTBI, caused by exposure to a pressure wave from an explosion, is predominantly experienced by military personnel and has increased in prevalence and severity in recent decades. Yet the underlying pathology of blast mTBI and how it may differ from impact mTBI is largely unknown. We hypothesized that blast exposure impairs aquaporin-4 (AQP4)-dependent perivascular glymphatic exchange and is worsened in an Aqp4-/- mouse which exhibits glymphatic impairment. Tracer was intracisternally injected into the cerebrospinal fluid of wild type and Aqp4-/- mice following repetitive blast mTBI and dynamic transcranial imaging was performed to measure tracer movement over the cortical surface. Brains were then collected after 22 or 42 minutes. I imaged these brain slices and quantified tracer influx. I observed delayed impairment of glymphatic function in blast exposed mice and that glymphatic impairment and behavioral outcomes were worsened in a Aqp4-/- mouse model following blast injury. Given the role of glymphatic exchange in the clearance of interstitial solutes such as amyloid β and tau, these findings suggest that changes in AQP4 and glymphatic impairment following blast injury may render the post-traumatic brain vulnerable to neurodegeneration and suggest that pathophysiology and progression of blast injury may be fundamentally different from impact TBI.

Alzheimer’s Disease (AD) is a neurodegenerative disorder characterized by the aggregation of the amyloid-β (Aβ) peptide into fibrillar β-sheet plaques. While Aβ plaques have been the historical focus in the study of AD, it has recently been shown that Aβ also forms oligomeric intermediates of a novel α-sheet secondary structure along its aggregation pathway. Furthermore, the formation of these α-sheet oligomers is correlated with the neuronal death and subsequent cognitive impairment seen in AD, and it begins up to 10-20 years before plaque formation or symptom development. In this project, I utilize a novel diagnostic technique known as the soluble oligomer binding (SOBA) assay to characterize the in vitro aggregation of Aβ through α-sheet detection in various timepoints of incubated Aβ. Additionally, I use SOBA to characterize AD phenotypes in transgenic mouse models using α-sheet detection in brain homogenate samples. SOBA selectively detects α-sheet oligomers using plate surfaces functionalized with in-house α-sheet peptide designs, followed by the introduction of amyloid samples and then amyloid-specific primary and secondary antibodies for detection. This research allows for the further elucidation of Aβ's structure and aggregation pathway, paving the way for future treatment methods for AD. Finally, I aim to translate my work with AD to develop SOBA for the system of Type 2 Diabetes (T2D) to detect the presence of toxic islet amyloid polypeptide (IAPP), the amyloid peptide implicated in T2D. This will be achieved through the systematic tuning of antibody systems and other procedural parameters such as plate washing methods. Preliminary results have shown the ability of SOBA to detect α-sheet in synthetic IAPP samples with concentrations as low as 10 pM. This novel method for toxic IAPP detection has the potential for T2D diagnosis earlier in disease progression.

The glymphatic system, which is primarily active during sleep, is a network of astroglial perivascular channels within the brain that allows for cerebrospinal fluid (CSF) influx and exchange. Glymphatic exchange plays a crucial role in the clearance of amyloid, a hallmark in the development of Alzheimer’s. Recently, a bidirectional relationship between Alzheimer's disease and sleep has also been suggested with amyloid deposition associated with mid-life sleep disruption. However, the mechanistic link between sleep disruption, particularly over chronic time scales, and the development of Alzheimer’s pathology remains unclear. This study investigated whether chronic sleep disruption, similar to that experienced in aging population, impacts downstream Alzheimer’s-related neuropathology. We hypothesized chronic sleep disruption will result in decreased glymphatic function and increased amyloid plaque burden. This experiment utilized a chronic sleep disruption model using Lafayette Sleep Fragmentation chambers, where mice underwent either chronic sleep disruption every two minutes during normal sleeping periods (daylight hours) or normal sleeping conditions (sham) from 10 weeks to 18 weeks of age (n=120). After eight weeks of sleep disruption or sham exposure, glymphatic function was assessed by dynamic in vivo near infrared imaging following stereotactic CSF tracer injection. Animals were perfusion fixed, cryosectioned, and glymphatic function was further assessed by measurement of fluorescent cerebrospinal fluid tracers in brain tissue. Aquaporin-4 localization, amyloid plaque deposition, and markers of astroglial and microglial activation were assessed by immunofluorescence. The collected data demonstrated that sleep disruption significantly increased neuropathological outcomes. The measured impact of glymphatic function was also correlated with these downstream pathological effects. These findings could be an indicator of interactions between neurological disease progression and an inflammatory expression after sleep disruption. They can also shed more light on the complex relationship between Alzheimer’s disease progression, the glymphatic system, and chronic sleep disruption.