Optic Nerve Head Tissue Research Articles

A Method to Estimate Biomechanics and Mechanical Properties of Optic Nerve Head Tissues From Parameters Measurable Using Optical Coherence Tomography

Optic nerve head (ONH) tissue properties and biomechanics remain mostly unmeasurable in the experiment. We hypothesized that these can be estimated numerically from ocular parameters measurable in vivo with optical coherence tomography (OCT). Using parametric models representing human ONHs we simulated acute intraocular pressure (IOP) increases (10 mmHg). Statistical models were fit to predict, from OCT-measurable parameters, 15 outputs, including ONH tissue properties, stresses, and deformations. The calculations were repeated adding parameters that have recently been proposed as potentially measurable with OCT. We evaluated the sensitivity of the predictions to variations in the experimental parameters. Excellent fits were obtained to predict all outputs from the experimental parameters, with cross-validated R2s between 0.957 and 0.998. Incorporating the potentially measurable parameters improved fits significantly. Predictions of tissue stiffness were accurate to within 0.66 MPa for the sclera and 0.24 MPa for the lamina cribrosa. Predictions of strains and stresses were accurate to within 0.62% and 4.9 kPa, respectively. Estimates of ONH biomechanics and tissue properties can be obtained quickly from OCT measurements using an applet that we make freely available. These estimates may improve understanding of the eye sensitivity to IOP and assessment of patient risk for development or progression of glaucoma.

Peripapillary ring: histology and correlations

To assess the histologic correlate of the peripapillary ring (PPR) as the optic disc boundary, to measure it histomorphometrically and to assess its associations with glaucoma and axial length. Using a light microscope, we measured the width of the PPR and optic nerve pia mater. We additionally had measured the axial length, length of parapapillary beta and gamma zones, and thickness of the sclera and choroid. The study included 83 human globes with a mean axial length of 26.1±3.3mm (range: 20.0-35.0mm). In the non-highly myopic (axial length <26.5mm) non-glaucomatous eyes, mean PPR width was 55±14μm (range: 26-88μm) and mean thickness of the optic nerve pia mater was 52±22μm (range: 20-84μm). PPR width and pia mater thickness were significantly associated with each other (p=0.001; correlation coefficient r=0.37). PPR width increased significantly with longer axial length (p=0.001; r=0.35) and with the length of the peripapillary scleral flange (p=0.02; r=0.34). Pia mater thickness increased significantly with longer axial length (p=0.03; r=0.24) and with presence of glaucoma (p=0.03). In eyes without parapapillary gamma zone, PPR was covered or was at least touched by the end of Bruch's membrane and separated the peripapillary choroid from the prelaminar optic nerve head tissue. In eyes with parapapillary gamma zone, PPR was the continuation of the optic nerve pia mater and reached the undersurface of the peripapillary retinal nerve fibre layer. In these eyes, Bruch's membrane started in a distance of up to 5mm from PPR, and the thin space of peripapillary choroid ended by an attachment of Bruch's membrane to the thinned sclera. Peripapillary ring width (mean: 55±14μm) increases slightly with longer axial length and is independent of glaucoma. It is strongly correlated with the pia mater thickness and is the continuation of the pia mater to the ocular surface.

In vivo optic nerve head biomechanics: performance testing of a three-dimensional tracking algorithm

Measurement of optic nerve head (ONH) deformations could be useful in the clinical management of glaucoma. Here, we propose a novel three-dimensional tissue-tracking algorithm designed to be used in vivo. We carry out preliminary verification of the algorithm by testing its accuracy and its robustness. An algorithm based on digital volume correlation was developed to extract ONH tissue displacements from two optical coherence tomography (OCT) volumes of the ONH (undeformed and deformed). The algorithm was tested by applying artificial deformations to a baseline OCT scan while manipulating speckle noise, illumination and contrast enhancement. Tissue deformations determined by our algorithm were compared with the known (imposed) values. Errors in displacement magnitude, orientation and strain decreased with signal averaging and were 0.15 µm, 0.15° and 0.0019, respectively (for optimized algorithm parameters). Previous computational work suggests that these errors are acceptable to provide in vivo characterization of ONH biomechanics. Our algorithm is robust to OCT speckle noise as well as to changes in illumination conditions, and increasing signal averaging can produce better results. This algorithm has potential be used to quantify ONH three-dimensional strains in vivo, of benefit in the diagnosis and identification of risk factors in glaucoma.

Changes in Optic Nerve Head Circulation in Response to Vasoactive Agents: Intereye Comparison in Monkeys with Experimental Unilateral Glaucoma

To investigate circulatory changes in the optic nerve head (ONH) in response to vasoactive agents including calcium antagonists, a substrate of nitric oxide (NO), and an inhibitor of NO synthase (NOS) in monkeys with unilateral experimental glaucoma. Argon laser cautery to the trabecular meshwork was used to create experimental unilateral glaucoma in nine monkeys. The effects of systemic lomerizine or nilvadipine (calcium-antagonists), L-arginine (a substrate of NO), and NG-nitro-L-arginine methyl ester (L-NAME, a NOS inhibitor) on the ONH tissue blood velocity (NB(ONH)) was studied by the laser speckle method. Lomerizine and nilvadipine significantly increased NB(ONH) in the untreated normal eyes (P = 0.039 and 0.008, respectively), while significant, less increases were found in the laser-treated experimental glaucomatous eyes with significant intereye differences (P = 0.036 and 0.011, respectively). L-arginine significantly increased NB(ONH) in both eyes without intereye difference (P = 0.71). L-NAME had no significant effects on NB(ONH) in the experimental glaucoma eyes; however, it produced a significant decrease in the nonlaser treated eyes (P = 0.036). In experimental glaucomatous eyes, the reactivity of ONH vessels to calcium antagonists was preserved, but was significantly reduced. The response to a NOS inhibitor was lost; however, reactivity to a substrate of NO was normal. These data indicate that in experimental glaucoma, vasodilator reactivity in the peripheral vasculature of the ONH is preserved, but functional alterations are likely to affect reactivity to the NO system.

Shadow Removal and Contrast Enhancement in Optical Coherence Tomography Images of the Human Optic Nerve Head

To improve the quality of optical coherence tomography (OCT) images of the optic nerve head (ONH). Two algorithms were developed, one to compensate for light attenuation and the other to enhance contrast in OCT images. The former was borrowed from developments in ultrasound imaging and was proven suitable with either time- or spectral-domain OCT. The latter was based on direct application of pixel intensity exponentiation. The performances of these two algorithms were tested on spectral-domain OCT images of four adult ONHs. Application of the compensation algorithm significantly reduced the intralayer contrast (from 0.74 ± 0.16 to 0.17 ± 0.12; P < 0.001), indicating successful blood vessel shadow removal. Furthermore, compensation dramatically improved the visibility of deeper ONH tissues, such as the peripapillary sclera and lamina cribrosa. Application of the contrast-enhancement algorithm significantly increased the interlayer contrast (from 0.48 ± 0.22 to a maximum of 0.89 ± 0.05; P < 0.001) and thus allowed a better differentiation of tissue boundaries. Contrast enhancement was robust only when compensation was considered. The proposed algorithms are simple and can significantly improve the quality of ONH images clinically captured with OCT. This study has important implications, as it will help improve our ability to perform automated segmentation of the ONH; quantify the morphometry and biomechanics of ONH tissues in vivo; and identify potential risk indicators for glaucoma.

Visualization of Fundus Vessel Pulsation Using Principal Component Analysis

Spontaneous venous pulsation is one of the clinical signs with which to rule out elevated intracranial pressure and papilledema. More subtle pulsatile retinal movements are difficult to observe because of eye movements. Recording a fundus movie and aligning (registering) the images helps, but the images still contain distracting microsaccadic distortions and noise. The authors hypothesized that addressing these latter points should allow observation of minute pulsating features in fundus movies. Principal component analysis (PCA), a basic form of blind source analysis, is applied to recorded fundus image sequences. The authors demonstrate this method in 5-second image sequences acquired with a near-infrared SLO (HRA+OCT Spectralis). The images are first registered to correct for eye drift, then microsaccade-distorted images are rejected, and the remaining image sequence is decomposed into principal components. Finally, a movie is constructed using the first five principal components (these had pulsatile features). Each of the processing steps (registration, cleaning, PCA filtering) improves the detection of pulsatile features, ultimately allowing clear visualization of spontaneous venous pulsation. Depending on the subject, additional features can be observed: pulsation amplitude of the arterial tree of approximately 10 μm, pulsation of arterioles down to 70-μm diameter, complete venous collapse, overall optic nerve head tissue pulsation, and mechanical links between veins and arteries. By disentangling pulsatile motion from other dynamic components of retinal images, unprecedented resolution in physiologic motion of retinal vessel structure is achievable.

Neuroprotective effect on retinal ganglion cells by transpupillary laser irradiation of the optic nerve head

This study demonstrates that subthreshold transpupillary thermotherapy (TTT) laser irradiation on optic nerve head protects retinal ganglion cells (RGCs) in an optic nerve crush (ONC) model. TTT was performed in right eyes with an 810-nm diode laser aimed at the center of the optic nerve head, using the following protocol: power 60 mW, duration 60 s, spot size 500 μm. Fluoro-Gold was injected into bilateral superior colliculi 5 days before sacrifice and fluorescent gold labeled RGCs were counted under fluorescence microscopy. In the ONC group, a progressive loss of RGCs was observed; however, in comparison with the ONC group, RGCs density was significantly higher ( P = 0.001, independent samples t-test) at day 7 postoperative and only borderline significances were obtained at days 14 and 28 postoperative ( P = 0.044 and P = 0.045, respectively, independent samples t-test) in ONC + TTT group, which implies the potential neuroprotective role of TTT. This protective effect seems to be heat shock proteins (HSPs) related, because intraperitoneal Quercetin (an inhibitor of HSPs, 4 mg/kg/day for 7 days) could completely abolish this protective effect at days 7, 14 and 28 postoperative ( P = 0.012, P = 0.002, and P = 0.000, respectively, independent samples t-test). Minimal collateral damage of TTT on optic nerve head tissue, peripapillary RGCs and the myelin sheath of the optic nerve were observed under transmission electron microscopy. These findings suggested that subthreshold TTT might be a safe and practical approach to protect RGCs. The underlying mechanisms may involve TTT-induced HSPs in RGCs.

Effects of Scleral Stiffness Properties on Optic Nerve Head Biomechanics

The biomechanical environment within the optic nerve head, important in glaucoma, depends strongly on scleral biomechanical properties. Here we use a range of measured nonlinear scleral stress-strain relationships in a finite element (FE) model of the eye to compute the biomechanical environment in the optic nerve head at three levels of intraocular pressure (IOP). Three stress-strain relationships consistent with the 5th, 50th and 95th percentiles of measured human scleral stiffness were selected from a pool of 30 scleral samples taken from 10 eyes and implemented in a generic FE model of the eye using a hyperelastic five-parameter Mooney-Rivlin material model. Computed strains within optic nerve head tissues depended strongly on scleral properties, with most of this difference occurring between the compliant and median scenarios. Also, the magnitudes of strains were found to be substantial even at normal IOP (up to 5.25% in the lamina cribrosa at 15 mmHg), being larger than previously reported values even at normal levels of IOP. We conclude that scleras that are "weak", but still within the physiologic range, will result in appreciably increased optic nerve head strains and could represent a risk factor for glaucomatous optic neuropathy. Estimations of the deformation at the optic nerve head region, particularly at elevated IOP, should take into account the nonlinear nature of scleral stiffness.

Subtilisin-like Proprotein Convertase Expression, Localization, and Activity in the Human Retina and Optic Nerve Head

Subtilisin-like proprotein convertases (SPCs) are a family of calcium-dependent cleavage enzymes that act on dibasic sites of various peptide/protein substrates. The purpose of this study was to investigate the expression, localization, and activity of SPCs in the human retina and optic nerve head. mRNA expression of the SPC family in the human retina and optic nerve head tissues was evaluated by quantitative reverse transcription polymerase chain reaction (QRT-PCR). Double immunofluorescence staining was performed on paraffin-embedded human posterior sections to localize SPC family members. Western blot analysis was used to identify PACE4 isoform expression within the optic nerve head and retina. In addition, a fluorogenic SPC substrate-based assay was used to elucidate SPC enzyme activity within human retina and optic nerve head (ONH) tissues. QPCR results indicated that PC1 and PC2 were expressed 4.1- and 5.7-fold higher in retina compared to optic nerve head, whereas PACE4 was expressed 4.1-fold higher in the ONH. PC1 and PC2 were localized primarily in neuronal cells, whereas PACE4 and PC5 were limited to the glia of the retina and optic nerve head. SPC activity in ONH lysate was significantly higher than that of retinal lysate; however, when an SPC inhibitor was added, activity in ONH decreased more than that in retina. These results indicate that the SPCs are expressed in distinct patterns throughout the human retina and ONH. PC1 and PC2 were primarily expressed in neurons, whereas PACE4 appeared to be largely restricted to glia. Thus, elevated PACE4 may modulate the bioactivity of proteins secreted in the ONH and retina.

Bone morphogenetic protein 4 inhibits TGF‐β2 stimulation of extracellular matrix proteins in optic nerve head cells: Role of gremlin in ECM modulation

The characteristic cupping of the optic nerve head (ONH) in glaucoma is associated with elevated TGF-beta2 and increased synthesis and deposition of extracellular matrix (ECM) proteins. In addition to TGF-beta2, the human ONH also expresses bone morphogenetic proteins (BMPs) and BMP receptors, which are members of the TGF-beta superfamily. We examined the potential effects of BMP4 and the BMP antagonist gremlin on TGF-beta2 induction of ECM proteins in ONH cells. BMP-4 dose dependently inhibited TGF-beta2-induced fibronectin (FN) and PAI-1 expression in ONH astrocytes and lamina cribrosa (LC) cells and also reduced TGF-beta2 stimulation of collagen I, collagen VI, and elastin. Addition of gremlin blocked this BMP-4 response, increasing cellular and secreted FN as well as PAI-1 levels in both cell types. Gremlin was expressed in ONH tissues and ONH cells, and gremlin protein levels were significantly increased in the LC region of human glaucomatous ONH tissues. Interestingly, recombinant gremlin dose dependently increased ECM protein expression in cultured ONH astrocytes and LC cells. Gremlin stimulation of ECM required activation of TGF-beta receptor and R-Smad3. TGF-beta2 increased gremlin mRNA expression and protein levels in ONH cells. Inhibition of either the type I TGF-beta receptor or Smad3 phosphorylation blocked TGF-beta2-induced gremlin expression. In conclusion, BMP4 blocked the TGF-beta2 induction of ECM proteins in ONH cells. The BMP antagonist gremlin reversed this inhibition, allowing TGF-beta2 stimulation of ECM synthesis. Increased expression of gremlin in the glaucomatous ONH may further exacerbate TGF-beta2 effects on ONH ECM metabolism by inhibiting BMP-4 antagonism of TGF-beta2 signaling. Modulation of the ECM via gremlin provides a novel therapeutic target for glaucoma.

Intraocular Pressure Spikes in Keratectasia, Axial Myopia, and Glaucoma

To review a range of activities associated with intraocular pressure (IOP) spikes. To examine the possible significance of IOP spikes in conditions such as keratectasia, axial myopia, and glaucoma. Hypotheses concerning mechanisms for adverse responses to IOP spikes were examined. Apart from the possibility that IOP spikes might cause susceptible corneal, posterior scleral, or optic nerve head tissue to yield to associated distending forces, there is the possibility that these tissues will be also be damaged by increased hydrostatic pressure. In-office tonometry does not indicate the degree to which ocular tissues are exposed to IOP spikes. For eyes that are exposed to IOP spikes of longer duration, that occur frequently and which result in a larger IOP increment, the risk of an adverse response may be greater. Changes in ocular tissues because of increased hydrostatic pressure may include morphological cellular changes and alterations to enzyme function. Eye rubbing may be the most significant mechanism for creating IOP spikes because of the large IOP increments that may be involved, as well as the possibility that abnormal rubbing can become a chronic habit. As appears to be the case in keratoconus, asymmetric exposure to IOP spikes may help to explain some asymmetric presentations of post-laser-assisted in situ keratomileusis, glaucoma, or myopia. Ideally methods for the objective assessment of patient risk for adverse responses to IOP spikes will continue to be developed. A self-administered questionnaire may help identify patients who are significantly exposed to IOP spikes. Family history may indicate an increased risk of diseases for which IOP spikes may have significant implications. Patient counseling regarding the possibility that IOP spiking activities may contribute to the development and/or progression of conditions such as keratectasia, axial myopia, and glaucoma may be indicated.

Neuroprotective effect of transpupillary thermotherapy in the optic nerve crush model of the rat

Transpupillary thermotherapy (TTT) has been shown to induce heat shock protein (Hsp) 72 in optic nerve head tissue. The neuroprotective effect of TTT was investigated in an optic nerve crush rat model. TTT was performed onto the optic nerve head in the right eye of subject rats. After 24 h, an optic nerve crush injury using an aneurysm clip was performed at 2 mm from the optic nerve head for 60 s. At 7 and 14 days later, retrograde labelling of retinal ganglion cells (RGCs) with DTMR crystal was carried out and the density of the surviving RGCs was evaluated. Immunohistochemical staining was performed to confirm the expression of Hsp72. At 7 days after optic nerve crush injury, the mean density of surviving RGCs was higher in TTT group (372.7+/-149.8 per mm(2)) than in optic nerve crush group (252.9+/-96.7 per mm(2)) with borderline significance. In the retinal areas at 1 mm from the optic nerve head, a significant increase in surviving RGCs from TTT treated eyes was observed at both 7 and 14 days after optic nerve crush injury. However, no significant differences in surviving RGCs were demonstrated 2 and 3 mm from the optic nerve head. These results demonstrate that TTT aimed onto the optic nerve head showed a neuroprotective effect.

Modeling individual-specific human optic nerve head biomechanics. Part I: IOP-induced deformations and influence of geometry

Glaucoma, the second most common cause of blindness worldwide, is an ocular disease characterized by progressive loss of retinal ganglion cell (RGC) axons. Biomechanical factors are thought to play a central role in RGC loss, but the specific mechanism underlying this disease remains unknown. Our goal was to characterize the biomechanical environment in the optic nerve head (ONH)--the region where RGC damage occurs--in human eyes. Post mortem human eyes were imaged, fixed at either 5 or 50 mmHg pressure and processed histologically to acquire serial sections through the ONH. Three-dimensional models of the ONH region were reconstructed from these sections and embedded in a generic scleral shell to create a model of an entire eye. We used finite element simulations to quantify the effects of an acute change in intraocular pressure from 5 to 50 mmHg on the ONH biomechanical environment. Computed strains varied substantially within the ONH, with the pre-laminar neural tissue and the lamina cribrosa showing the greatest strains. The mode of strain having the largest magnitude was third principal strain (compression), reaching 12-15% in both the lamina cribrosa and the pre-laminar neural tissue. Shear strains were also substantial. The distribution of strains in all ONH tissues was remarkably similar between eyes. Inter-individual variations in ONH geometry (anatomy) have only modest effects on ONH biomechanics, and may not explain inter-individual susceptibility to elevated intraocular pressure. Consistent with previous results using generic ONH models, the displacements of the vitreo-retinal interface and the anterior surface of the lamina cribrosa can differ substantially, suggesting that currently available optical imaging methods do not provide information of the acute deformations within ONH tissues. Predicted strains within ONH tissues are potentially biologically significant and support the hypothesis that biomechanical factors contribute to the initial insult that leads to RGC loss in glaucoma.

Modeling individual-specific human optic nerve head biomechanics. Part II: influence of material properties

Biomechanical factors acting within the optic nerve head (ONH) likely play a role in the loss of vision that occurs in glaucoma. In a companion paper (Sigal et al. 2008), we quantified the biomechanical environment within individual-specific ONH models reconstructed from human post mortem eyes. Our goal in this manuscript was to use finite element modeling to investigate the influence of tissue material properties on ONH biomechanics in these same individual-specific models. A sensitivity analysis was carried out by simulating the effects of changing intraocular pressure on ONH biomechanics as tissue mechanical properties were systematically varied over ranges reported in the literature. This procedure was repeated for each individual-specific model described in the companion paper (Sigal et al. 2008). The outcome measures of the analysis were first and third principal strains, as well as the derived quantity of maximum shear strain, in ONH tissues. Scleral stiffness had by far the largest influence in ONH biomechanics, and this result was remarkably consistent across ONH models. The stiffnesses of the lamina cribrosa and pia mater were also influential. These results are consistent with those obtained using generic ONH models. The compressibility of the pre-laminar neural tissue influenced compressive and shearing strains. Overall, tissue material properties had a much greater influence on ONH biomechanics than did tissue geometry, as assessed by comparing results between our individual-specific models. Material properties of ONH tissues, particularly of the peripapillary sclera, play a dominant role in the mechanical response of an ONH to acute changes in IOP and may be important in the pathogenesis of glaucoma. We need to better understand inter-individual differences in scleral biomechanical properties and whether they are clinically important.

Activation of the BMP Canonical Signaling Pathway in Human Optic Nerve Head Tissue and Isolated Optic Nerve Head Astrocytes and Lamina Cribrosa Cells

Bone morphogenetic proteins (BMPs) are members of the transforming growth factor (TGF)-beta superfamily that controls multiple functions in a variety of cells. We have previously shown that human optic nerve head (ONH) astrocytes and lamina cribrosa (LC) cells express BMP and BMP receptor mRNA and proteins. The purpose of the present study was to determine whether human ONH tissues express the canonical BMP signaling pathway and whether ONH cells secrete BMP-4 and respond to exogenous BMP-4 through this pathway. Well-characterized human ONH astrocytes (N = 2) and LC cells (N = 3) were treated with exogenous BMP-4 (20 ng/mL) for various times. Western immunoblot analysis was used to detect secreted BMP-4 in serum-free conditioned media of ONH cells and in human ONH tissues (N = 4) and Smad proteins in total cell lysate of ONH cells and tissues. Intracellular colocalization of p-R-Smad1 with Co-Smad4 and localization of inhibitory Smads (e.g., I-Smad6 and I-Smad7) were studied through immunocytochemistry. In addition, coimmunoprecipitation was used to verify the interaction of p-R-Smad1 with Co-Smad4. ONH astrocytes and LC cells secrete BMP-4 and synthesize R-Smad1, R-Smad5, I-Smad6, I-Smad7, and Co-Smad4 proteins. Exposure to BMP-4 for either 10 or 60 minutes resulted in increased p-R-Smad1 and p-R-Smad1/5/8 protein levels that declined after 12 hours of treatment. Immunocytochemistry and coimmunoprecipitation studies revealed that p-R-Smad1/5/8 and Co-Smad4 interact and colocalize in the nucleus. BMP-4 treatment resulted in increased coprecipitation of p-R-Smad1/5/ 8 and Co-Smad4. I-Smad6 and I-Smad7 are localized in the nucleus and cytoplasm of ONH astrocytes and LC cells. Proteins for BMP-4, p-R-Smad1/5/8, R-Smad1, R-Smad5, R-Smad8, and Co-Smad4 are present in human ONH tissues. In addition, phosphorylated Smad1 and Smad5 colocalize with Smad4 in the nuclei of ONH tissues. These results indicate that BMP-4 and Smad signaling proteins are present in human ONH tissues, isolated ONH astrocytes, and LC cells. In addition, exogenous BMP-4 treatment of ONH astrocytes and LC cells results in downstream signaling through the canonical Smad pathway. Thus, cells within the human ONH may respond to locally released BMP through paracrine or autocrine mechanisms.

Population differences in elastin maturation in optic nerve head tissue and astrocytes.

Glaucomatous optic neuropathy is characterized by remodeling of the extracellular matrix with disorganization of elastic fibers in the optic nerve head (ONH). There are significant differences in prevalence of glaucomatous optic neuropathy between African Americans (AAs) and Caucasian Americans (CAs). The goal of this study was to evaluate differences in elastin synthesis and maturation in ONH tissue and cells of AA and CA donors with no eye disease, to provide a basis for underlying racial differences in susceptibility to elevated intraocular pressure. The amount of mature elastin in ONHs from each group of donors was evaluated by desmosine radioimmunoassay. The distribution of elastic fibers in ONH tissue was investigated by immunofluorescent staining. Elastin and lysyl oxidase mRNA levels and alternative splicing of elastin in ONH astrocytes were investigated by quantitative PCR. Tropoelastin protein expression was assessed by immunoblot analysis. ONHs from AA donors had significantly reduced levels of desmosine compared with those of CAs. In contrast, elastin mRNA and tropoelastin synthesis were elevated in ONH astrocytes from AA individuals. The inclusion of exon 23 in elastin mRNA and lysyl oxidase-like 2 mRNA levels was significantly reduced in astrocytes from AA compared with CA donors. A reduced number of cross-linking domains in elastin and decreased lysyl oxidase-like 2 expression leads to decreased amount of mature elastin in ONHs from healthy AA individuals compared with CA donors. These results suggest ELN and LOXL2 as candidate susceptibility genes for population-specific genetic risk of primary open-angle glaucoma (POAG).

Predicted extension, compression and shearing of optic nerve head tissues

Glaucomatous optic neuropathy may be in part due to an altered biomechanical environment within the optic nerve head (ONH) produced by an elevated intraocular pressure (IOP). Previous work has characterized the magnitude of the IOP-induced deformation of ONH tissues but has not focused specifically on the mode of deformation (strain), i.e. whether the ONH tissues and cells are stretched, compressed or sheared. Circumstantial evidence indicates that the mode of deformation has biological consequences. Here we use computational models to study the different modes of deformation that occur in an ONH as a result of an increase in IOP. One generic and three individual-specific 3D models of the human ONH were reconstructed as previously described. Each model consisted of five tissue regions: pre and post-laminar neural tissue, lamina cribrosa, sclera and pia mater. Finite element methods were then used to predict the biomechanical response to changes in IOP. For each model we computed six local measures of strain, including the magnitude and direction of maximum stretching, maximum compression and maximum shearing strain. We compared the spatial and population distributions of the various measures of strain by using semi-quantitative (contour plots) and quantitative (histograms) methods. For all models, as IOP increased, the tissues of the ONH were subjected simultaneously to various modes of strain, including compression, extension and shearing. The highest magnitudes of all modes of strain occurred within the neural tissue regions. There were substantial differences in the magnitudes of the various modes of strain, with the largest strains being in compression, followed by shearing and finally by extension. The biomechanical response of an individual-specific ONH to changes in IOP is complex and cannot be fully captured by one measure of deformation. We predict that cells within the ONH are subjected to very different modes of deformation as IOP increases. The largest deformations are compressive, followed by shearing and stretching. Models of IOP-induced RGC damage need to be further refined by characterizing the cellular response to these different modes of strain.

Oxygen Saturation in Optic Nerve Head Structures by Hyperspectral Image Analysis

Purpose: A method is presented for the calculation and visualization of percent blood oxygen saturation from specific tissue structures in hyperspectral images of the optic nerve head (ONH). Methods: Trans-pupillary images of the primate optic nerve head and overlying retinal blood vessels were obtained with a hyperspectral imaging (HSI) system attached to a fundus camera. Images were recorded during normal blood flow and after partially interrupting flow to the ONH and retinal circulation by elevation of the intraocular pressure (IOP) from 10 mmHg to 55 mmHg in steps. Percent oxygen saturation was calculated from groups of pixels associated with separate tissue structures, using a linear least-squares curve fit of the recorded hemoglobin spectrum to reference spectra obtained from fully oxygenated and deoxygenated red cell suspensions. Color maps of saturation were obtained from a new algorithm that enables comparison of oxygen saturation from large vessels and tissue areas in hyperspectral images. Results: Percent saturation in retinal vessels and from the average over ONH structures (IOP = 10 mmHg) was (mean ± SE): artery 81.8 ± 0.4%, vein 42.6 ± 0.9%, average ONH 68.3 ± 0.4%. Raising IOP from 10 mmHg to 55 mmHg for 5 min caused blood oxygen saturation to decrease (mean ± SE): artery 46.1 ± 6.2%, vein 36.1 ± 1.6%, average ONH 41.9 ± 1.6%. The temporal cup showed the highest saturation at low and high IOP (77.3 ± 1.0% and 60.1 ± 4.0%) and the least reduction in saturation at high IOP (22.3%) compared with that of the average ONH (38.6%). A linear relationship was found between saturation indices obtained from the algorithm and percent saturation values obtained by spectral curve fits to calibrated red cell samples. Conclusions: Percent oxygen saturation was determined from hyperspectral images of the ONH tissue and retinal vessels overlying the ONH at normal and elevated IOP. Pressure elevation was shown to reduce blood oxygen saturation in vessels and ONH structures, with the smallest reduction in the ONH observed in the temporal cup. IOP-induced saturation changes were visualized in color maps using an algorithm that follows saturation-dependent changes in the blood spectrum and blood volume differences across tissue. Reduced arterial saturation at high IOP may have resulted from a flow-dependent mechanism.

Factors Influencing Optic Nerve Head Biomechanics

The biomechanical environment within the optic nerve head (ONH) may play a role in retinal ganglion cell loss in glaucomatous optic neuropathy. This was a systematic analysis in which finite element methods were used to determine which anatomic and biomechanical factors most influenced the biomechanical response of the ONH to acute changes in IOP. Based on a previously described computational model of the eye, each of 21 input factors, representing the biomechanical properties of relevant ocular tissues, the IOP, and 14 geometric factors were independently varied. The biomechanical response of the ONH tissues was quantified through a set of 29 outcome measures, including peak and mean stress and strain within each tissue, and measures of geometric changes in ONH tissues. Input factors were ranked according to their aggregated influence on groups of outcome measures. The five input factors that had the largest influence across all outcome measures were, in ranked order: stiffness of the sclera, radius of the eye, stiffness of the lamina cribrosa, IOP, and thickness of the scleral shell. The five least influential factors were, in reverse ranked order: retinal thickness, peripapillary rim height, cup depth, cup-to-disc ratio, and pial thickness. Factor ranks were similar for various outcome measure groups and factor ranges. The model predicts that ONH biomechanics are strongly dependent on scleral biomechanical properties. Acute deformations of ONH tissues, and the consequent high levels of neural tissue strain, were less strongly dependent on the action of IOP directly on the internal surface of the ONH than on the indirect effects of IOP on the sclera. This suggests that interindividual variations in scleral properties could be a risk factor for the development of glaucoma. Eye size and lamina cribrosa biomechanical properties also have a strong influence on ONH biomechanics.

Autoantibodies in patients with glaucoma: a comparison of IgG serum antibodies against retinal, optic nerve, and optic nerve head antigens

The aim of this study was to analyze and compare the entire IgG autoantibody patterns against different ocular antigens (retina, optic nerve, and optic nerve head) in sera of glaucoma patients and healthy subjects. Sixty-six patients were included in this study: healthy volunteers without any ocular disorders (CO, n=30), patients with primary open-angle glaucoma (POAG, n=19), and patients with normal-tension glaucoma (NTG, n=17). The sera were tested for antibodies against retinal, optic nerve, and optic nerve head tissues. Immunodetection was performed using 4-chloro-1-naphthol staining. The autoantibody patterns were digitized and subsequently analyzed by multivariate statistical techniques. All patients showed a complex repertoire of IgG antibodies against retinal, optic nerve, and optic nerve head antigens. The analysis of discriminance revealed a statistically significant differences between the patterns of all three groups. Our multivariate approach could quantify the differences in immunoreactivities of patient sera against the three antigens. The POAG group had the most significant difference against retinal antigens (P=0.0021) compared with the other antigens. In the NTG group the highest reactivity appeared against optic nerve head (P=0.00053) and optic nerve (P=0.0025). All groups showed different and complex antibody patterns against the three ocular tissues. These autoantibodies are highly specific for each patient group. The analysis of these patterns could provide further information about possible autoimmune mechanisms in the pathogenesis of glaucoma.