Chronic sleep disruption, present in 25-60% of patients suffering from Alzheimer's Disease (AD), often precedes cardinal disease symptoms. While little is known about the mechanisms underlying chronic sleep disruption and the development of clinical pathology, both acute and chronic sleep deprivation have been found to increase biomarkers of AD including neuroinflammation and amyloid-beta accumulation. Additionally, in people without AD, sleep deprivation can result in a deterioration of working memory and attention. In this study, we examine the impact of chronic sleep disruption on cognition both at baseline and in the 5xFAD mouse model of AD. The 5xFAD mouse model is a transgenic mouse model of familial amyloidosis which expresses neurocognitive impairment as early as 2 months. To define the effect of chronic sleep disruption on cognition in the absence of AD pathology, 60 wild type mice were exposed to chronic sleep disruption or sham procedure for 8 weeks between 10 and 18 weeks of age. At 18 weeks of age, I evaluated the animals for changes in spatial memory (Barnes maze), short-term memory (Y-maze), locomotion and anxiety (open field test), and activities of daily living (burrowing trials). To test whether chronic sleep disruption specifically exacerbates AD-related neurocognitive decline, the same cognitive tests were performed on 60 5xFAD+ animals exposed to 8 weeks of sleep disruption or sham treatment. I then analyzed the collected data to isolate any trends of cognitive performance, finding that chronic sleep disruption impaired cognitive performance in 5xFAD+ and littermate controls, with a more significant impact on 5xFAD+ animals. These findings highlight the critical association between dysfunctional sleep and the development of cognitive impairment with AD disease progression which then guides us toward better preventative care and treatments.

The glymphatic system, which is primarily active during sleep, is a network of astroglial perivascular channels within the brain that allow for Cerebrospinal Fluid (CSF) influx and exchange. Glymphatics play a crucial role in the waste clearance of amyloid beta, a hallmark in the development of Alzheimer’s Disease and neurodegeneration. Recently, a bidirectional relationship between Alzheimer's Disease and sleep has also been suggested with the aggregation of amyloid beta 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 investigates whether chronic sleep disruption, similar to that experienced in humans, will impact downstream neuropathology. We hypothesize chronic sleep disruption will result in decreased glymphatic function and subsequently increased amyloid plaque burden. This experiment utilizes a chronic sleep fragmentation model in 120 5xFAD mice from 8 weeks to 16 weeks of age. In the Lafayette Sleep Fragmentation chambers, 60 animals are disturbed every two minutes during normal sleeping periods (daylight hours). 60 mice were placed in normal sleeping conditions. After eight weeks of sleep fragmentation or sham exposure, glymphatic function is assessed by in vivo near infrared imaging following stereotactic CSF tracer injection. Animals are perfusion fixed, cryosectioned, and glymphatic function is assessed by measurement of fluorescent cerebrospinal fluid tracers in brain tissue. Aquaporin-4 localization, amyloid plaque deposition, and markers of astroglial and microglial activation are assessed by immunofluorescence. In this project, I specifically work on cryosectioning the tissue, and understanding glymphatic function through the processes of immunofluorescence imaging and analysis. The collected data demonstrated that sleep disruption did increase 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.

Alzheimer’s Disease (AD) is a neurodegenerative disease that affects more than 5 million Americans. The glymphatic system (a network of perivascular spaces that facilitate fluid movement and solute clearance from the brain) and its dysfunction associated with aging has been implicated in the development of AD. The water channel aquaporin 4 (AQP4), located in astrocytic endfeet bordering the perivascular spaces, is crucial for the proper functioning of the glymphatic system. Data suggests that loss of AQP4 localization results in amyloid-ß deposition, a hallmark of AD pathology, and loss of AQP4 localization accompanies aging in rodents as well as AD in humans. In this study, we quantitatively analyze the expression of aquaporin- 4ex (AQP4ex)—a translational readthrough variant of AQP4 believed to play a role in its localization—to identify any correlation with aging and AD pathology. Selective deletion of AQP4ex results in the mislocalization of AQP4 all over the astrocytic membrane, indicating that AQP4ex is a crucial element in the localization of AQP4. We analyze young, old and AD groups in the murine (mouse) brain as well as AD versus control in a human case series. Currently, we see a trend towards decline in cortical perivascular AQP4ex in the AD group, with more analysis ongoing. This is the first characterization of AQP4ex expression in the murine brain and in a human case series, and these data will contribute to the small but growing body of research on AQP4ex and its relationship with AQP4 localization, creating opportunities to identify a new novel mechanism and novel target in AD pathology.

In the United States, 1.5 million individuals suffer a fracture due to bone disease each year. It is well documented that mechanical load affects bone development, but our understanding of the cellular mechanisms behind bone development under load is limited. Current human induced pluripotent stem cell (hiPSC) derived bone tissue models have more relevant human physiology compared to traditional animal models. However, there is a lack of dynamically loaded hiPSC bone tissue and diseased hiPSC bone tissue models in vitro. We propose a novel, three-dimensional bone tissue model as a platform for musculoskeletal disease modeling that allows for compressive loading that will enhance maturity as well as induce diseased bone phenotypes. We improved upon existing poly-L-lactide solvent cast scaffold techniques by incorporating a polyvinyl alcohol mold and an annealing step that increases the uniformity of the scaffolds and allows for higher throughput fabrication. Osteoblasts were derived from hiPSCs using established differentiation protocols and seeded into the 3D, porous, poly-L-lactide scaffold to generate in vitro bone tissue that generates significant extracellular calcium. We propose an arduino-powered, 3D-printed loading device that can apply physiologically relevant dynamic loads to the scaffold and hypothesize improved bone tissue maturity in comparison to 2D cultures and unloaded 3D scaffolds. By screening for markers of early bone development such as type I collagen, markers of later development such as osteocalcin, and assays for extracellular calcium, we can track the maturity and development of bone tissue. We expect that 3D bone growth with static loading will reveal diseased bone phenotypes such as decreased calcium deposition and immature bone, whereas dynamic loading will promote bone growth and lead to mature bone. Ultimately, this model will improve our ability to investigate the effects of mechanical loading in developing and diseased bone.

Induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) that have been engineered into three-dimensional heart tissues (EHTs) are valuable research tools for investigating debilitating genetic diseases that afflict the heart, such as Duchenne muscular dystrophy (DMD). Ensuring iPSC-CMs can be sufficiently matured to model such diseases remains a hurdle in current research, and maturational analysis techniques for iPSC-CMs are either qualitative, manual, or primarily based in two dimensions, leaving much to be desired. This poster details the creation of a suite of MATLAB image-processing scripts that can quantify the effect of three-dimensional culture and disease-causing DMD mutations on cardiomyocyte structure and maturation state. The iPSC-CMs were differentiated from stem cells, cast into EHTs, stained using immunofluorescence, and imaged using confocal microscopy. Using the scripts to analyze these 3D images of iPSC-CM stains, key maturational features of the cells can be quantified such as nuclei count; cardiomyocyte area; and sarcomere length, orientation, and z-disk width. Analyzing cardiomyocyte area can give key information on cardiomyocyte hypertrophy while examining sarcomere length, orientation, and Z-disk width can provide information on myofibril structure and organization. The suite allows analysis of these maturational features in both 2D and 3D cultures and offers a method for quantitatively assessing maturation in an automated manner. Measuring iPSC-CM maturation will also allow better comparison of existing maturational methods, such as mechanical loading, electrical stimulation, and small molecule treatment. The suite can also create graphical outputs to elegantly display data. Recent progress also includes a script that can count cell nuclei and quantify cell area. Overall, the suite will help improve maturational analysis of EHTs, and hopefully contribute to the discovery of new treatments for diseases that affect the heart. 

Adolescents with type 1 diabetes (T1D) are at risk for poor physical and psychosocial outcomes. Diabetes-related family conflict has previously been associated with youths' glycemic control (HbA1c). However, less is known about how family conflict associates with other health outcomes. This project aimed to explore correlations between adolescent and parent reported family conflict with diabetes-distress, depressive symptoms, resilience, and health-related quality of life (HRQOL) for both adolescent and parent. Eligible patients were enrolled in a two-site randomized controlled psychosocial intervention study. Participants were ages 13-18 with T1D for over a year and elevated levels of diabetes distress. At baseline, patients and their parent completed measures of: diabetes-specific family conflict (DFCS), HRQOL (T1DAL), diabetes distress (PAID-T), depressive symptoms (PHQ-8), and resilience (CD-RISC). HbA1c was pulled from participants medical charts. Descriptive statistics were used to summarize demographic variables, and bivariate correlation analyses were used to investigate the association between DFCS and the other psychosocial variables. Adolescents (N= 131; 53.4% female, 6.1% identified as another gender, 78.6% White, 9.9% Black, 2.3% Asian, 3.1% American Indian/Alaskan Native, and 80.9% Non-Hispanic, average age 15.38  1.5) DFCS scores correlated with more diabetes-distress (r=0.386, p<0.001), depressive symptoms (r=0.334, p<0.001), and less HQOL (r= -0.303, p<0.001). Parents’ (N=131; 79.4% White, 9.2% Latino/Mexican 6.1% Black, 2.3% Asian, 0.8% other, 79.4% private insurance) DFCS scores correlated with higher youth A1C (r=0.280, p<0.001), higher parent diabetes distress (r= 0.479, p <0.001), and lower parent resilience (r= - 0.200, p = 0.022) and HQOL (r = -0.369, p < 0.001). Both parent and patient reports appear to be an important area of further investigation for determining correlates of poor physical and psychosocial wellbeing in this high-risk group. While further investigation is needed, screening for family conflict may be important in clinical procedure, as it may become a future target for intervention.