Shelley Boyd

Immunopathology of the Noninfectious Posterior and Intermediate Uveitides

The posterior and intermediate uveitides share an underlying immune etiology; however, they can be clinically and immunopathologically distinguished. Although the initiating stimuli for posterior and intermediate uveities are not known, it is believed that an exogenous agent (such as a bacterium or a virus) or an endogenous molecule may induce disease. In either case, T-helper lymphocytes in conjunction with human leukocyte antigens are likely to be involved. This review examines the epidemiology, histology, immunopathology, and theories of pathogenesis of several posterior and intermediate uveitides, including sympathetic ophthalmia, Vogt-Koyanagi-Harada syndrome, Behçets disease, sarcoidosis, intermediate uveitis, white dot syndromes, and birdshot retinochoroidopathy.

The development of a patchy organization of the rat striatum

The rat striatum can be divided into patch and matrix compartments. Patches, as marked by high opiate receptor binding, first emerge perinatally from a dense, diffuse field of striatal opiate binding. Our quantitative analysis revealed that the patch compartment formed its peak proportion of the total striatal area at postnatal day 7. After this time, patches occupied a smaller proportion of the striatum, reflecting the fact that the number of patches and mean area per patch reached near adult levels during the first postnatal week, yet the volume of the striatum as a whole continued to increase for several weeks postnatally. Results from transplant and early postnatal lesion experiments suggested that connections between the striatum and other brain areas are important for the formation and/or maintenance of the patch and matrix compartments. Transplants of embryonic striatum to cavities in the cortex of young adult hosts developed diffuse opiate receptor binding but not dopamine receptor binding. Significantly, the opiate receptor binding seen in the transplants was never organized into the dense patches normally seen in the adult striatum. In a few transplants areas of relatively higher opiate receptor binding occurred in areas of relatively low neuronal cell density, as is seen in early normal development, but the dense adult patches never developed. Coronal diencephalic hemisections, but not decortications, in the early postnatal period produced drastic shrinkage of the striatum and, more importantly, a large decrease in opiate receptor patches when expressed as a proportion of total striatal area. Neuronal connections with more caudal brain structures may play a role in the final differentiation and maintenance of the striatal compartments.

High-speed, high-resolution Fourier-domain optical coherence tomography system for retinal imaging in the 1060 nm wavelength region

A high-speed (47,000 A-scans/s), ultrahigh axial resolution Fourier domain optical coherence tomography (OCT) system for retinal imaging at approximately 1060 nm, based on a 1024 pixel linear array, 47 kHz readout rate InGaAs camera is presented. When interfaced with a custom superluminescent diode (lambda(c) = 1020 nm, Deltalambda = 108 nm, Pout = 9 mW), the system provides 3.3 microm axial OCT resolution at the surface of biological tissue, approximately 4.5 microm in vivo in rat retina, approximately 5.7 microm in vivo in human retina, and 110 dB sensitivity for 870 microW incident power and 21 mus integration time. Retinal tomograms acquired in vivo from a human volunteer and a rat animal model show clear visualization of all intraretinal layer and increased penetration into the choroid.

Maternal anti-platelet β3 integrins impair angiogenesis and cause intracranial hemorrhage

Fetal and neonatal alloimmune thrombocytopenia (FNAIT) is a life-threatening disease in which intracranial hemorrhage (ICH) is the major risk. Although thrombocytopenia, which is caused by maternal antibodies against β3 integrin and occasionally by maternal antibodies against other platelet antigens, such as glycoprotein GPIbα, has long been assumed to be the cause of bleeding, the mechanism of ICH has not been adequately explored. Utilizing murine models of FNAIT and a high-frequency ultrasound imaging system, we found that ICH only occurred in fetuses and neonates with anti-β3 integrin-mediated, but not anti-GPIbα-mediated, FNAIT, despite similar thrombocytopenia in both groups. Only anti-β3 integrin-mediated FNAIT reduced brain and retina vessel density, impaired angiogenic signaling, and increased endothelial cell apoptosis, all of which were abrogated by maternal administration of intravenous immunoglobulin (IVIG). ICH and impairment of retinal angiogenesis were further reproduced in neonates by injection of anti-β3 integrin, but not anti-GPIbα antisera. Utilizing cultured human endothelial cells, we found that cell proliferation, network formation, and AKT phosphorylation were inhibited only by murine anti-β3 integrin antisera and human anti-HPA-1a IgG purified from mothers with FNAIT children. Our data suggest that fetal hemostasis is distinct and that impairment of angiogenesis rather than thrombocytopenia likely causes FNAIT-associated ICH. Additionally, our results indicate that maternal IVIG therapy can effectively prevent this devastating disorder.

Limiting factors to the OCT axial resolution for in-vivo imaging of human and rodent retina in the 1060nm wavelength range

A computational model was developed to evaluate the limitations to the highest axial resolution, achievable with ultrahigh resolution optical coherence tomography (UHROCT) in the 1060 nm wavelength region for in-vivo imaging of the human and rodent retina. The model considers parameters such as the wavelength dependent water absorption, the average length of the human and rodent eyes, and the power limitations for the imaging beam as defined in the ANSI standard. A custom-built light source with re-shaped spectrum was used to verify experimentally the results from the computational model. Axial OCT resolution of 4.2 microm and 7.7 microm was measured from a mirror reflection with the custom light source by propagating the imaging beam through water cells with 5 mm and 25 mm thickness, corresponding to the average axial length of the rodent and human eye respectively. Assuming an average refractive index of 1.38 for retinal tissue, the expected axial OCT resolution in the rodent and human retina is 3 microm and 5.7 microm respectively. Retinal tomograms acquired in-vivo from the rat eye with the modified light source show clear visualization of all intraretinal layers, as well as a network of capillaries (approximately 10 microm in diameter) in the inner- and outer plexiform layers of the retina.

The PI3K/Akt/mTOR pathway mediates retinal progenitor cell survival under hypoxic and superoxide stress

Oxygen (O₂) tension has emerged as a major regulator of stem cell (SC) biology. Low O₂ concentrations that are toxic to mature cells can confer advantage to stem and early progenitors, while superoxide stress remains a constant threat in aerobic biology and may be partially avoided through sequestration of SCs in the relatively hypoxic stem or regenerative niche. Using primary retina-derived retinal progenitor cells (RPCs) and the R28 progenitor cell line in vitro, we show that RPCs are sensitive to hydrogen peroxide (H₂O₂) induced damage and resistant to moderate levels of low oxygen stress (1% O₂). Under hypoxic conditions, multipotent RPCs upregulate Epo receptors, and Epo, along with insulin, protects against both superoxide- and severe hypoxia- (0.25% O₂) induced apoptosis through activation of the canonical PI3K/Akt/mTOR pathway. This survival advantage is sensitive to inhibitors of PI3K and mTOR. We further demonstrate phosphorylation of the p70S6 ribosomal kinase, a downstream mediator of PI3K/Akt/mTOR and translational activator. Overall, these data confirm that RPCs are sensitive to superoxide stress and resistant to hypoxia and that this resistance is mediated in part by Epo. They further suggest that manipulation of RPCs ex vivo prior to ocular delivery, or the in vivo delivery of exogenous survival factors at the time of cell implantation, could enhance the success of regenerative therapies aimed to restore sight.

Uveitis in the elderly—is it easy to identify the masquerade?

Uveitis tends to occur in working age adults and is a major cause of visual loss in this age group.1 It does however occur both in childhood and in the elderly, and considerably less information is available regarding these two groups, particularly the latter. Regardless, in all age groups, it is necessary to accurately diagnose the form of uveitis, and to detect any underlying or associated systemic problems. Management is guided by the significant features of a patient’s history, the signs present within the eye, and the results of relevant investigations.2 While the causes of uveitis are legion, most are immune mediated, requiring the patient’s immune system to mount and perpetuate an immune response, irrespective of the initiating stimulus.3 It is known that the aging immune system is less able to respond to foreign antigens and that autoimmune antibody production rises. Overall, however, it is believed that the incidence of true immune mediated uveitis declines in the elderly. Infectious endophthalmitis, particularly that arising after surgery, and lymphoma both occur at higher frequency in the elderly. These constitute the masquerade syndromes. In this review, information available on the types of uveitis found in the elderly is presented, along with data on the aging immune system. Specific information is given on the causes of the masquerade syndromes, and an approach to clinical management is suggested. In most published series, there is a decline in the incidence of uveitis presenting in patients over the age of 65 years. Uveitis starting in the younger age groups may still be active in the older patient, although in many the disease has become quiescent. Very few studies address the epidemiology of uveitis in the elderly per se, and still fewer provide useful data regarding patterns of presentation and the likely diagnoses in this …

Origin-specific epigenetic program correlates with vascular bed-specific differences in Rgs5 expression

Cells from multiple origins contribute to vascular smooth muscle cell (VSMC) development. Phenotypic heterogeneity of VSMCs is associated with their point of developmental origin; however, the mechanisms driving such differences are unknown. We here examined the mechanisms controlling vascular bed‐specific differences in Rgs5 expression during development. Rgs5 levels were similar across different regions of the vasculature in neonatal animals but were >15‐fold higher in descending aortas compared with carotid arteries of adult mice. Thus, vessel bed‐specific changes in regulation of Rgs5 expression occurred during vessel maturation. Examination of adult Rgs5‐LacZ reporter mice revealed lower Rgs5 expression in VSMCs originating from the third (carotid artery) branchial arch compared with those originating in the fourth and sixth (aortic B segment, right subclavian, and ductus arteriosus) branchial arches. Indeed, a mosaic Rgs5 expression pattern, with discreet LacZ boundaries between VSMCs derived from different developmental origins, was observed. Furthermore, Rgs5‐LacZ expression was correlated with the site of VSMC origin (splanchic mesoderm ≈ local mesenchyme > somites > proepicardium > mesothelium). Surprisingly, Rgs5 reporter activity in cultured carotid artery‐ and descending aorta‐derived cells did not recapitulate the differences observed in vivo. Consistent with a developmental origin‐specific epigenetic mechanism driving the observed expression differences in vivo, the Rgs5 promoter showed increased methylation on CpG dinucleotides in carotid arteries compared with that in descending aortas in adult but not in neonatal mice. In vitro methylation of the Rgs5 promoter confirmed that its activity is sensitive to transcriptional down‐regulation by CpG methylation. These data suggest that an origin‐dependent epigenetic program regulates vascular bed‐ and maturation state‐dependent regulation of VSMC‐specific gene transcription.—Zhang, H., Gu, S., Al‐Sabeq, B., Wang, S., He, J., Tam, A., Cifelli, C., Mathalone, N., Tirgari, S., Boyd, S., Heximer, S. P. Origin‐specific epigenetic program correlates with vascular bedspecific differences in Rgs5 expression. FASEB J. 26, 181–191 (2012). www.fasebj.org

Intact endothelial autophagy is required to maintain vascular lipid homeostasis

The physiological role of autophagic flux within the vascular endothelial layer remains poorly understood. Here, we show that in primary endothelial cells, oxidized and native LDL stimulates autophagosome formation. Moreover, by both confocal and electron microscopy, excess native or modified LDL appears to be engulfed within autophagic structures. Transient knockdown of the essential autophagy gene ATG7 resulted in higher levels of intracellular 125I‐LDL and oxidized LDL (OxLDL) accumulation, suggesting that in endothelial cells, autophagy may represent an important mechanism to regulate excess, exogenous lipids. The physiological importance of these observations was assessed using mice containing a conditional deletion of ATG7 within the endothelium. Following acute intravenous infusion of fluorescently labeled OxLDL, mice lacking endothelial expression of ATG7 demonstrated prolonged retention of OxLDL within the retinal pigment epithelium (RPE) and choroidal endothelium of the eye. In a chronic model of lipid excess, we analyzed atherosclerotic burden in ApoE−/−mice with or without endothelial autophagic flux. The absence of endothelial autophagy markedly increased atherosclerotic burden. Thus, in both an acute and chronic in vivo model, endothelial autophagy appears critically important in limiting lipid accumulation within the vessel wall. As such, strategies that stimulate autophagy, or prevent the age‐dependent decline in autophagic flux, might be particularly beneficial in treating atherosclerotic vascular disease.

Compartmentalization of the embryonic striatum after intraocular transplantation

In order to assess the intrinsic potential of the isolated embryonic striatum to develop its adult patch and matrix compartments, embryonic day 16 striata were transplanted into the anterior eye chamber of adult host rats. After 2-12 weeks of survival the transplants showed heterogeneous and in the majority of cases complementary distributions of opiate receptor binding and acetylcholinesterase staining, which mark the patch and matrix compartments of the adult striatum, respectively. The complementarity of patch and matrix markers in the transplants shows that the transplants do compartmentalize. However, the density of the markers in the transplants did not reach the levels seen in the adult striatum. The results suggest that the commitment of cells to a striatal compartment is a very early event in the embryonic development of the forebrain.