The Role of SPARC in Aqueous Humor Outflow and TGFß2-mediated Ocular Hypertension in a Murine Model

Glaucoma is the leading cause of irreversible blindness worldwide, and is a major cause of blindness in the United States. It affects approximately 5% of Caucasians and 10% of African- Americans over the age of 60 years. Elevated intraocular pressure (IOP) is currently the only modifiable risk factor for glaucoma. Impaired outflow of aqueous humor from the eye is thought to be the cause of pathologically elevated IOP. However, the etiology of outflow impairment is unknown. Anatomically, the aqueous humor drains into the iridocorneal angle of the eye, where the iris inserts at the transition between the cornea and sclera. In humans, approximately 80-90% of the aqueous traverses through the trabecular meshwork (TM), juxtacanalicular connective tissue (JCT), Schlemm’s canal, collector channels and empties into episcleral veins. Abnormalities at these sites are thought to cause impaired outflow. Abnormal accumulation of extracellular matrix (ECM) in the TM or JCT, abnormal endothelial function in Schlemm’s canal, or a combination of these components have been strongly implicated. Our laboratory has focused on the role of Secreted Protein Acidic and Rich in Cysteine (SPARC) in regulating outflow. SPARC is the prototypical matricellular protein that mediates ECM organization and turnover in numerous human tissues. Our lab was first to demonstrate that SPARC is highly expressed in the TM and JCT regions of the eye, and that the SPARC knockout (KO) mouse has a significant decrease in IOP of 15-20%. SPARC may affect the degree of segmental flow, a theory that states that variable aqueous outflow occurs around the circumference of the eye; only certain portions of the TM are thought to display active outflow at any particular moment. The cytokine transforming growth factor-ß2 (TGFß2) has been shown to modulate multiple ECM proteins, including SPARC. TGFß2 is significantly upregulated by 2 to 3-fold in the aqueous humor of glaucoma patients compared to controls. In addition, when TGFß2 is overexpressed in rodent eyes, increased ECM deposition is observed within the trabecular meshwork leading to IOP elevation. SPARC is one of the most highly upregulated proteins by TGFß2, and is downstream of TGFß2. We hypothesized that wild-type (WT) mice would demonstrate segmental flow, while SPARC KO mice would display a more continuous pattern of outflow around the eye. We also believed that IOP would be inversely correlated with outflow area. We also hypothesized that SPARC is essential to the process of TGFß2-mediated ocular hypertension, and that the lack of SPARC would impair IOP elevation. We conducted a tracer study utilizing fluorescent microbeads to determine the location of outflow circumferentially around the mouse TM. Microbeads were injected intracamerally into the eyes of WT and KO mice. After a 45-minute incubation period, the mice were euthanized and eyes were processed for confocal, light, and electron microscopy. During the second group of experiments, empty or TGFß2-containing adenovirus was injected intravitreally into WT and SPARC KO mice and IOP was measured for 2 weeks. Immunohistochemistry was completed on all tissues to assess for changes in major ECM proteins. Percentage effective filtration length (PEFL), or area of the TM labeled by tracer, was significantly increased in SPARC KO mice (70.61% ± 11.36%, p<0.005; N=11) compared to WT mice (54.68% ± 9.95%; N=11). In addition, the pressures between the two sets of eyes were significantly different with mean pressures of 16.3 mm Hg in WT mice and 12.6 mm Hg in KO mice (p<0.005, N=11 pairs). In addition, PEFL and IOP were inversely correlated with R2 = 0.72 (N=10 pairs); in eyes with higher IOP, PEFL was reduced. Electron microscopy demonstrated that high-tracer TM areas had a greater separation between trabecular beams. Collagen fibril diameter was found to be smaller in the KO (28.272 nm) compared to WT (34.961 nm; p<0.0005, N=3 pairs). These data provided structural correlations to the functional data regarding segmental flow. In the second set of experiments, IOP was found to be significantly elevated in TGFß2- injected WT mice compared to empty vector-injected WT mice during days 4-11 (p<0.05, N=8). However, IOP was not significantly elevated in TGFß2-injected KO mice compared to controls. Immunohistochemistry demonstrated that TGFß2 increased expression of collagen IV, fibronectin, plasminogen activator inhibitor-1 (PAI-1), connective tissue growth factor (CTGF), and SPARC within the TM of WT mice, but only PAI-1 and CTGF in KO mice (p<0.05, N=3 pairs). These data support our hypotheses, indicating that SPARC plays an integral role in the modulation of aqueous humor outflow. In addition, it appears as though SPARC is essential to the regulation of TGFß2-mediated ocular hypertension. Aside from providing further evidence of the importance of ECM in IOP regulation, our work presents the novel discovery of segmental flow in the mouse. Given the potential role of SPARC in TGFß2-mediated ocular hypertension, SPARC may not only play an integral role in ECM homeostasis within the trabecular meshwork, but may be a valuable target for pharmacologic therapy in treating primary open-angle glaucoma

ETMS: Efficient Traffic Management System for Congestion Detection and Alert using HAAR Cascade

Rapid social development has resulted in the emergence of a new major societal issue: urban traffic congestion, which many cities must address. In addition to making&nbsp; it more difficult for people to get around town, traffic jams are a major source of the city's pollution crisis. In order to address the problems of automobile exhaust pollution and congestion, this paper uses the system dynamics approach to develop a model to study the urban traffic congestion system from the perspectives of trucks,private cars, bikes&nbsp;and public transportation. This project proposes a system for detecting vehicles and sending alerts when traffic levels rise to dangerous levels using Haar Cascade and Fuzzy Cognitive Maps (FCP). The proposed system uses Haar Cascade to detect moving vehicles, which are then classified using FCP. The system can make decisions based on partial or ambiguous information by utilising FCP, a soft computing technique, which allows it to learn from past actions. An algorithm for estimating traffic density is also used by the system to pinpoint active areas. In congested areas, the system will alert the driver if it anticipates a collision with another vehicle and also Experiments show that the proposed system is able to accurately detect vehicles and provide timely alerts to the driver, drastically lowering the probability of accidents occurring in heavily travelled areas. The importance of introducing such a system cannot be overstated in today's transportation system. It's a big deal for the future of intelligent urban planning and traffic control. Congestion relief, cleaner air, and increased security are just some of the long-term benefits that justify the high initial investment. To add, this system is adaptable to suburban and rural areas, which can also experience traffic congestion issues

An Exploration of the Virtual Digital Twin Capture for Spatial Tasks and its Applications

Our generation is currently at the juncture of the fourth industrial revolution - Industry 4.0. Emergent technology such as Augmented Reality (AR), Internet of Things (IoT), Artificial Intelligence (AI), cloud computing, big data, and more are at the center of this. Amidst all these, the concept of Digital Twinning is a promising technology for realizing Industry 4.0. Simply put, a Digital Twin is a virtual representation of a real task, action, or object. This thesis explores the parameters and details required to generate a Digital Twin. Using these insights, we propose two applications that utilize digital twinning - EditAR and AnnotateXR. EditAR is an AR workflow for authoring kinesthetic instructions for spatial tasks. AnnotateXR is an Extended Reality (XR) workflow for automating data annotation to support multiple Computer Vision (CV) applications. We evaluate these systems through user studies and report the results on the usability and viability of these workflows. From an evaluation study, EditAR received an average system usability score (SUS) of 82.0. Over the course of a user study, using AnnotateXR, users were able to generate a total of 112,737 semantically segmented images and 144 videos annotated for action segmentation in 66.55 minutes. AnnotateXR received an average SUS score of 91.0

Gene networks and systems biology in Alzheimer's disease: Insights from multi‐omics approaches

Despite numerous studies in the field of dementia and Alzheimer's disease (AD), a comprehensive understanding of this devastating disease remains elusive. Bulk transcriptomics have provided insights into the underlying genetic factors at a high level. Subsequent technological advancements have focused on single-cell omics, encompassing techniques such as single-cell RNA sequencing and epigenomics, enabling the capture of RNA transcripts and chromatin states at a single cell or nucleus resolution. Furthermore, the emergence of spatial omics has allowed the study of gene responses in the vicinity of amyloid beta plaques or across various brain regions. With the vast amount of data generated, utilizing gene regulatory networks to comprehensively study this disease has become essential. This review delves into some techniques employed in the field of AD, explores the discoveries made using these techniques, and provides insights into the future of the field

Bioinspired ZnS:Gd Nanoparticles Synthesized from an Endophytic Fungi <i>Aspergillus flavus</i> for Fluorescence-Based Metal Detection

Recently, several nonconventional sources have emerged as strong hotspots for the biosynthesis of chalcogenide quantum dots. However, studies that have ascertained the biomimetic methodologies that initiate biosynthesis are rather limited. The present investigation portrays a few perspectives of rare-earth(Gd)-doped ZnS biosynthesis using the endophytic fungi Aspergillus flavus for sensing metals based on their fluorescence. Analysis of ZnS:Gd nanoparticles was performed by elemental analysis, energy-dispersive X-ray spectroscopy (EDS), atomic force microscopy (AFM), X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), photoluminescence (PL), and transmission electron microscopy (TEM). The results of TEM demonstrated that the particles were polycrystalline in nature, with a mean size of 10&#8315;18 nm. The fluorescence amenability of the biogenic ZnS nanoparticles was further used for the development of a simple and efficient sensing array. The results showed sensitive and detectable quenching/enhancement in the fluorescence of biogenic colloidal ZnS nanoparticles, in the presence of Pb (II), Cd (II), Hg (II), Cu (II) and Ni (II), respectively. The fluorescence intensity of the biogenic ZnS:Gd nanoparticles was found to increase compared to that of the ZnS nanoparticles that capacitate these systems as a reliable fluorescence sensing platform with selective environmental applications

Towards a Comprehensive and Robust Micromanipulation System with Force-Sensing and VR Capabilities

In this modern world, with the increase of complexity of many technologies, especially in the micro and nanoscale, the field of robotic manipulation has tremendously grown. Microrobots and other complex microscale systems are often to laborious to fabricate using standard microfabrication techniques, therefore there is a trend towards fabricating them in parts then assembling them together, mainly using micromanipulation tools. Here, a comprehensive and robust micromanipulation platform is presented, in which four micromanipulators can be used simultaneously to perform complex tasks, providing the user with an intuitive environment. The system utilizes a vision-based force sensor to aid with manipulation tasks and it provides a safe environment for biomanipulation. Lastly, virtual reality (VR) was incorporated into the system, allowing the user to control the probes from a more intuitive standpoint and providing an immersive platform for the future of micromanipulation

The Circadian Clock That Controls Gene Expression in Arabidopsis Is Tissue Specific

The expression of CHALCONE SYNTHASE (CHS) expression is an important control step in the biosynthesis of flavonoids, which are major photoprotectants in plants. CHS transcription is regulated by endogenous programs and in response to environmental signals. Luciferase reporter gene fusions showed that the CHS promoter is controlled by the circadian clock both in roots and in aerial organs of transgenic Arabidopsis plants. The period of rhythmic CHS expression differs from the previously described rhythm of chlorophyll a/b-binding protein (CAB) gene expression, indicating that CHS is controlled by a distinct circadian clock. The difference in period is maintained in the wild-type Arabidopsis accessions tested and in the de-etiolated 1 and timing of CAB expression 1 mutants. These clock-affecting mutations alter the rhythms of both CAB and CHS markers, indicating that a similar (if not identical) circadian clock mechanism controls these rhythms. The distinct tissue distribution of CAB and CHS expression suggests that the properties of the circadian clock differ among plant tissues. Several animal organs also exhibit heterogeneous circadian properties in culture but are believed to be synchronized in vivo. The fact that differing periods are manifest in intact plants supports our proposal that spatially separated copies of the plant circadian clock are at most weakly coupled, if not functionally independent. This autonomy has apparently permitted tissue-specific specialization of circadian timing