Acoustic emission(AE) is one of the most sensitive techniques to non invasively monitor deformation, fatigue, and fracture of many materials. The purpose of this study was to evaluate the potential to use AE to detect local failure events within porous ceramic materials. The primary material of interest was mineralized trabecular bone. A better understanding of the failure of trabecular bone is highly relevant to skeletal fragility diseases such as osteoporosis. This study sought to develop a post processing technique that could strengthen the relation between the events detected and the phenomena occurring as a specimen is loaded. The deficiency in other techniques is that they did not fully make a quantitative correlation between acoustic emission event characteristics and the physical occurrence of damage events. The study evaluated the use of seismic power laws because these laws were able to attach a quantitative model to an earthquake and its successive aftershocks. Earthquake transmission has similar propagation attributes when compared to acoustic emission; seismic waves radiate from the epicenter of an earthquake. Acoustic waves radiate from the source of energy release in an acoustic emission event. The study measured the acoustic emission response of trabecular bone and highly oriented ceramics. The bone and ceramics were extracted in two perpendicular directions so that the structural orientation was different. The study sought to evaluate if the power-laws could differentiate the acoustic emission response based on varying the material and varying the structural orientation. The samples were quasi-statically compressed; the mechanical and acoustic emission data were simultaneously recorded. The study found that using the seismic power-law did not statistically differentiate the directional orientation for trabecular bone or ceramic specimens. Acoustic emission did indicate that event detection was different for each type of the of material. Correlations were established with the acoustic emission response and the mechanical testing data. These relationships were explainable because of the mechanical properties of the material.

Thesis (doctoral)--Freie Universität Berlin, 1980.

In this study, trabecular architecture is considered as a cellular solid structure, including both intact and damaged bone models. ??Intact?? bone models were constructed based on ideal versions of 25, 60 and 80-year-old specimens with varying trabecular lengths and orientations to 5%, and 10% covariance of variation (COV). The models were also flipped between longer transverse and longer longitudinal trabeculae. With increasing COV of lengths and orientations of trabecular bone, the apparent modulus is linearly decreased, especially in the longer transverse trabeculae lengths. ??Damaged?? bone models were built from the 25 year old model at 5% COV of longer transverse trabeculae, and with removing trabeculae of 5% and 10% of trabecular volume in transverse and longitudinal directions, respectively, as well as in combination to total 10% and 15%. With increasing percent of trabeculae missing, the apparent modulus decreased, especially dramatically when removal was only in the transverse direction. The trabecular bone models were also connected to a cortical shell and it was found that the apparent modulus of an entire slice was increased in comparison to the modulus of trabecular bone alone. We concluded that the architecture of trabecular bone, especially both lengths and percent of trabecular missing in the longitudinal direction, significantly influences mechanical properties.

In this study, trabecular architecture is considered as a cellular solid structure, including both intact and damaged bone models. ??Intact?? bone models were constructed based on ideal versions of 25, 60 and 80-year-old specimens with varying trabecular lengths and orientations to 5%, and 10% covariance of variation (COV). The models were also flipped between longer transverse and longer longitudinal trabeculae. With increasing COV of lengths and orientations of trabecular bone, the apparent modulus is linearly decreased, especially in the longer transverse trabeculae lengths. ??Damaged?? bone models were built from the 25 year old model at 5% COV of longer transverse trabeculae, and with removing trabeculae of 5% and 10% of trabecular volume in transverse and longitudinal directions, respectively, as well as in combination to total 10% and 15%. With increasing percent of trabeculae missing, the apparent modulus decreased, especially dramatically when removal was only in the transverse direction. The trabecular bone models were also connected to a cortical shell and it was found that the apparent modulus of an entire slice was increased in comparison to the modulus of trabecular bone alone. We concluded that the architecture of trabecular bone, especially both lengths and percent of trabecular missing in the longitudinal direction, significantly influences mechanical properties.

No description available.

Epidemiological evidence indicates that selenium supplementation may increase risk for ocular hypertension and glaucoma. The purpose of this project was to determine the effects of selenium on the conventional "trabecular" aqueous outflow pathway, a likely site of pathology for glaucoma. Human trabecular meshwork (HTM) cells and human umbilical vein endothelial cells (HUVECs) were treated with selenium (MSeA) at or near physiologically relevant concentrations. Selenium uptake by cells was monitored using mass spectrometry. While detectible changes in intracellular selenium were observed after exposure to 1-10 uM MSeA for 24 hours, the majority remained in the conditioned medium. The high concentrations of extracellular selenium we observed raised the possibility that selenium has an extracellular target.To investigate the role of selenium in extracellular matrix turnover, I examined alterations in protein secretion and intracellular signaling. MSeA treatment (5-10 uM) led to a significant decrease in the secretion of matrix metalloproteinase -2 and its inhibitor after 6-24 hours and to a dose-dependent decrease in kinase signaling. Later, I investigated the possibility that integrins are an extracellular target of selenium by monitoring morphological changes in HTM cells and by treating them with divalent cations. MSeA stimulated morphological changes consistent with a decrease in integrin function. These occurred before (less than 3 hours) alterations in protein secretion and intracellular signaling (3-6 hours). Zinc treatment prevented MSeA-mediated alterations in protein secretion and changes in cell-matrix adhesion.Finally markers of HTM cell homeostasis were examined. MSeA treatment (5 uM) led to a 60% decrease in protein synthesis after 3 hours and a 60% reduction in protein secretion, without causing significant alterations in cell viability and total ATP. To assess the physiological relevance of my results, anterior segments were perfused with MSeA to determine its effects on aqueous outflow facility. Preliminary results suggest that MSeA leads to a decrease in outflow facility.The combination of MSeA-induced decreases in several indicators of HTM cell homeostasis (without adversely effects on cell viability at physiologically relevant doses) and decreases in outflow facility provide a possible mechanism for selenium-associated ocular hypertension.

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.

Previously, we showed that transcription factor activating protein 2-beta (AP-2β) deletion from the periocular mesenchyme (POM)-derived neural crest cells (NCCs) using Wnt1Cre (AP-2β NCC knockouts/AP-2β NCC KOs) resulted in anterior segment abnormalities and increased intraocular pressure (IOP). The present study investigated the role of AP-2β in development of structures of the conventional pathway including the trabecular meshwork and Schlemm’s canal, and the unconventional pathway including the ciliary muscle. Studies using NCC KOs revealed that the embryonic POM migrated appropriately, but a significant reduction in postnatal POM cell proliferation in the angle was observed, accompanied by reduced expression of trabecular meshwork and Schlemm's canal markers when compared to controls, which likely contributed to the elevated IOP in NCC KOs. However, since Wnt1Cre was expressed in multiple NCC derivatives, AP-2β was deleted specifically from the developing trabecular meshwork region (TMR) using Mgp-Cre knock-in (Mgp-Cre.KI) mice. Although migration of the POM giving rise to the trabecular meshwork was not affected, peripheral anterior synechia (PAS), a decrease in expression of trabecular meshwork and Schlemm’s canal markers, and significantly increased IOP was observed in TMR KOs compared to controls, paired with loss of retinal ganglion cells (RGCs), and reduced retinal thickness and function. However, treatment with latanoprost, a prostaglandin analog that increases outflow through the unconventional pathway, significantly reduced elevated IOP in TMR KOs. Overall, the results suggest that AP-2β plays a cell-autonomous role in trabecular meshwork development and a non-cell-autonomous role in Schlemm’s canal development, while also playing an indirect role in unconventional pathway function, and thus, is important for IOP homeostasis. Moreover, the AP-2β NCC KO and AP-2β TMR KO may serve as models of primary angle closure glaucoma that can be used to test IOP-lowering drugs, molecular targets and neuroprotective strategies to develop treatments for human glaucoma. / Thesis / Doctor of Philosophy (PhD) / Glaucoma is the leading cause of irreversible blindness worldwide. Primary angle closure glaucoma is one type of glaucoma resulting from abnormalities in structures that allow aqueous humour found in the front of the eye from exiting the eye through two major routes, including the conventional and unconventional pathways. Defects in these structures lead to increased intraocular pressure (IOP) that damages specialized cells important for vision. This project examines the role of transcription factor activating protein 2-beta (AP-2β) in development of structures responsible for IOP balance. Data from the current study showed that AP-2β is required for formation of the structures of the conventional pathway, but does not directly affect development of unconventional pathway structures. The two AP-2β deletion mutants used here can model human primary angle closure glaucoma to test the effect of various drugs and cell protection strategies aimed at treating glaucoma.

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No description available.