James D. Akula

The Neurovascular Retina in Retinopathy of Prematurity

The continuing worldwide epidemic of retinopathy of prematurity (ROP), a leading cause of childhood visual impairment, strongly motivates further research into mechanisms of the disease. Although the hallmark of ROP is abnormal retinal vasculature, a growing body of evidence supports a critical role for the neural retina in the ROP disease process. The age of onset of ROP coincides with the rapid developmental increase in rod photoreceptor outer segment length and rhodopsin content of the retina with escalation of energy demands. Using a combination of non-invasive electroretinographic (ERG), psychophysical, and image analysis procedures, the neural retina and its vasculature have been studied in prematurely born human subjects, both with and without ROP, and in rats that model the key vascular and neural parameters found in human ROP subjects. These data are compared to comprehensive numeric summaries of the neural and vascular features in normally developing human and rat retina. In rats, biochemical, anatomical, and molecular biological investigations are paired with the non-invasive assessments. ROP, even if mild, primarily and persistently alters the structure and function of photoreceptors. Post-receptor neurons and retinal vasculature, which are intimately related, are also affected by ROP; conspicuous neurovascular abnormalities disappear, but subtle structural anomalies and functional deficits may persist years after clinical ROP resolves. The data from human subjects and rat models identify photoreceptor and post-receptor targets for interventions that promise improved outcomes for children at risk for ROP.

Retinal degenerative and hypoxic ischemic disease.

A broad spectrum of retinal diseases affects both the retinal vasculature and the neural retina, including photoreceptor and postreceptor layers. The accepted clinical hallmarks of acute retinopathy of prematurity (ROP) are dilation and tortuosity of the retinal vasculature. Additionally, significant early and persistent effects on photoreceptor and postreceptor neural structures and function are demonstrated in ROP. In this paper, we focus on the results of longitudinal studies of electroretinographic (ERG) and vascular features in rats with induced retinopathies that model the gamut of human ROP, mild to severe. Two potential targets for pharmaceutical interventions emerge from the observations. The first target is immature photoreceptors because the status of the photoreceptors at an early age predicts later vascular outcome; this approach is appealing as it holds promise to prevent ROP. The second target is the interplay of the neural and vascular retinal networks, which develop cooperatively. Beneficial pharmaceutical interventions may be measured in improved visual outcome as well as lessening of the vascular abnormalities.

Multimodal adaptive optics retinal imager: design and performance

Optical coherence tomography (OCT) and scanning laser ophthalmoscopy (SLO) are complementary imaging modalities, the combination of which can provide clinicians with a wealth of information to detect retinal diseases, monitor disease progression, or assess new therapies. Adaptive optics (AO) is a tool that enables correction of wavefront distortions from ocular aberrations. We have developed a multimodal adaptive optics system (MAOS) for high-resolution multifunctional use in a variety of research and clinical applications. The system integrates both OCT and SLO imaging channels into an AO beam path. The optics and hardware were designed with specific features for simultaneous SLO/OCT output, for high-fidelity AO correction, for use in humans, primates, and small animals, and for efficient location and orientation of retinal regions of interest. The MAOS system was tested on human subjects and rodents. The design, performance characterization, and initial representative results from the human and animal studies are presented and discussed.

Rod and rod-driven function in achromatopsia and blue cone monochromatism.

PURPOSE To evaluate rod photoreceptor and postreceptor retinal function in pediatric patients with achromatopsia (ACHR) and blue cone monochromatism (BCM) using contemporary electroretinographic (ERG) procedures. METHODS Fifteen patients (age range, 1-20 years) with ACHR and six patients (age range, 4-22 years) with BCM were studied. ERG responses to full-field stimuli were obtained in scotopic and photopic conditions. Rod photoreceptor (S(rod), R(rod)) and rod-driven postreceptor (log sigma, V(max)) response parameters were calculated from the a-wave and b-wave. ERG records were digitally filtered to demonstrate the oscillatory potentials (OPs); a sensitivity parameter, log SOPA(1/2), and an amplitude parameter, SOPA(max), were used to characterize the OP response. Response parameters were compared with those of 12 healthy control subjects. RESULTS As expected, photopic responses were nondetectable in patients with ACHR and BCM. In addition, mean scotopic photoreceptor (R(rod)) and postreceptor (V(max) and SOPA(max)) amplitude parameters were significantly reduced compared with those in healthy controls. The flash intensity required to evoke a half-maximum b-wave amplitude (log sigma) was significantly increased. CONCLUSIONS Results of this study provide evidence that deficits in rod and rod-mediated function occur in the primary cone dysfunction syndromes ACHR and BCM.

Omega-3 polyunsaturated fatty acids preserve retinal function in type 2 diabetic mice

Objective:Diabetic retinopathy (DR) is associated with hyperglycemia-driven microvascular pathology and neuronal compromise in the retina. However, DR is also linked to dyslipidemia. As omega-3 (ω-3) polyunsaturated fatty acids (PUFAs) are protective in proliferative retinopathy, we investigated the capacity of ω-3PUFAs to preserve retinal function in a mouse model of type 2 diabetes mellitus (T2DM).Design:Male leptin-receptor-deficient (db/db) mice were maintained for 22 weeks (4 weeks–26 weeks of life) on calorically and compositionally matched diets, except for 2% enrichment in either ω-3 or ω-6PUFAs. Visual function was assessed at 9, 14 and 26 weeks by electroretinography. Retinal capillary and neuronal integrity, as well as glucose challenge responses, were assessed on each diet.Results:The ω-3PUFA diet significantly preserved retinal function in the mouse model of T2DM to levels similar to those observed in nondiabetic control mice on normal chow. Conversely, retinal function gradually deteriorated in db/db mice on a ω-6PUFA-rich diet. There was also an enhanced ability of ω-3PUFA-fed mice to respond to glucose challenge. The protection of visual function appeared to be independent of cytoprotective or anti-inflammatory effects of ω-3PUFAs.Conclusion:This study identifies beneficial effects of dietary ω-3PUFAs on visual function in T2DM. The data are consistent with dyslipidemia negatively impacting retinal function. As ω-3PUFA lipid dietary interventions are readily available, safe and inexpensive, increasing ω-3PUFA intake in diabetic patients may slow the progression of vision loss in T2DM.

Broad spectrum antiangiogenic treatment for ocular neovascular diseases.

Pathological neovascularization is a hallmark of late stage neovascular (wet) age-related macular degeneration (AMD) and the leading cause of blindness in people over the age of 50 in the western world. The treatments focus on suppression of choroidal neovascularization (CNV), while current approved therapies are limited to inhibiting vascular endothelial growth factor (VEGF) exclusively. However, this treatment does not address the underlying cause of AMD, and the loss of VEGFs neuroprotective can be a potential side effect. Therapy which targets the key processes in AMD, the pathological neovascularization, vessel leakage and inflammation could bring a major shift in the approach to disease treatment and prevention. In this study we have demonstrated the efficacy of such broad spectrum antiangiogenic therapy on mouse model of AMD. Methods and Findings Lodamin, a polymeric formulation of TNP-470, is a potent broad-spectrum antiangiogenic drug. Lodamin significantly reduced key processes involved in AMD progression as demonstrated in mice and rats. Its suppressive effects on angiogenesis, vascular leakage and inflammation were studied in a wide array of assays including; a Matrigel, delayed-type hypersensitivity (DTH), Miles assay, laser-induced CNV and corneal micropocket assay. Lodamin significantly suppressed the secretion of various pro-inflammatory cytokines in the CNV lesion including monocyte chemotactic protein-1 (MCP-1/Ccl2). Importantly, Lodamin was found to regress established CNV lesions, unlike soluble fms-like tyrosine kinase-1 (sFlk-1). The drug was found to be safe in mice and have little toxicity as demonstrated by electroretinography (ERG) assessing retinal and by histology. Conclusions Lodamin, a polymer formulation of TNP-470, was identified as a first in its class, broad-spectrum antiangiogenic drug that can be administered orally or locally to treat corneal and retinal neovascularization. Several unique properties make Lodamin especially beneficial for ophthalmic use. Our results support the concept that broad spectrum antiangiogenic drugs are promising agents for AMD treatment and prevention.

The anatomy of the rat eye with oxygen-induced retinopathy.

Prior studies have documented the intertwined developmental courses of retinal blood vessel tortuosity (in fundus photographs) and retinal dysfunction (in electroretinographs) in Sprague–Dawley rat models of retinopathy of prematurity (ROP). Two such models, the “50/10 model” and the “75 model,” are named after the oxygen regimens used to induce retinopathy and are characterized by distinct neurovascular courses that span a range of disease severity. In this study of 50/10 and 75 model rats, retinal flatmounts were used to study the full vasculature at postnatal day (P) 15, P19 and P30. In addition, the layers of the neural retina were measured in toluidine blue-stained cross sections. Finally, gross anatomic features of the eye, including axial length, retinal surface area, and the ratio of anterior to posterior axial-lengths were evaluated. Both clock hours of neovascularization (NV) and percent avascular retina (AR) peaked at P19 and resolved by P30. Through P19, NV was found in every 50/10 model rat, but in only 60% of 75 model rats. AR was positively related to NV. All inner layers of the retina (outer plexiform layer through ganglion cell layer) were attenuated in 50/10 model rats but, in the 75 model, no layer differed significantly from that in controls. The eyes in both ROP models were smaller than those of age-matched controls. The ratio of anterior to posterior axial-lengths ranged from 0.45 in controls through 0.37 in the 75 model to 0.32 in the 50/10 model. Thus, eye growth is altered in these rat models of ROP.

Retinal degeneration in children: dark adapted visual threshold and arteriolar diameter.

To assess the condition of the retina in children with retinal degeneration due to Bardet-Biedl syndrome (BBS, n=41), Leber congenital amaurosis (LCA, n=31), or Usher syndrome (USH, n=13), the dark adapted visual threshold (DAT) and arteriolar diameters were measured. Compared to controls, the initial DATs of nearly all (83/85) were significantly elevated, and in 26/62 with serial DATs, significant progressive elevation occurred. Arteriolar diameters were significantly attenuated and narrowed with age in BBS and USH, but not LCA. Higher DATs were associated with narrower arterioles. Such non-invasive procedures can document the natural history of these retinal diseases and have the potential to assess response to future treatment.

Targeted and Reversible Blood-Retinal Barrier Disruption via Focused Ultrasound and Microbubbles

The blood-retinal barrier (BRB) prevents most systemically-administered drugs from reaching the retina. This study investigated whether burst ultrasound applied with a circulating microbubble agent can disrupt the BRB, providing a noninvasive method for the targeted delivery of systemically administered drugs to the retina. To demonstrate the efficacy and reversibility of such a procedure, five overlapping targets around the optic nerve head were sonicated through the cornea and lens in 20 healthy male Sprague-Dawley rats using a 690 kHz focused ultrasound transducer. For BRB disruption, 10 ms bursts were applied at 1 Hz for 60 s with different peak rarefactional pressure amplitudes (0.81, 0.88 and 1.1 MPa). Each sonication was combined with an IV injection of a microbubble ultrasound contrast agent (Definity). To evaluate BRB disruption, an MRI contrast agent (Magnevist) was injected IV immediately after the last sonication, and serial T1-weighted MR images were acquired up to 30 minutes. MRI contrast enhancement into the vitreous humor near targeted area was observed for all tested pressure amplitudes, with more signal enhancement evident at the highest pressure amplitude. At 0.81 MPa, BRB disruption was not detected 3 h post sonication, after an additional MRI contrast injection. A day after sonication, the eyes were processed for histology of the retina. At the two lower exposure levels (0.81 and 0.88 MPa), most of the sonicated regions were indistinguishable from the control eyes, although a few tiny clusters of extravasated erythrocytes (petechaie) were observed. More severe retinal damage was observed at 1.1 MPa. These results demonstrate that focused ultrasound and microbubbles can offer a noninvasive and targeted means to transiently disrupt the BRB for ocular drug delivery.

The neural retina in retinopathy of prematurity

ABSTRACT Retinopathy of prematurity (ROP) is a neurovascular disease that affects prematurely born infants and is known to have significant long term effects on vision. We conducted the studies described herein not only to learn more about vision but also about the pathogenesis of ROP. The coincidence of ROP onset and rapid developmental elongation of the rod photoreceptor outer segments motivated us to consider the role of the rods in this disease. We used noninvasive electroretinographic (ERG), psychophysical, and retinal imaging procedures to study the function and structure of the neurosensory retina. Rod photoreceptor and post‐receptor responses are significantly altered years after the preterm days during which ROP is an active disease. The alterations include persistent rod dysfunction, and evidence of compensatory remodeling of the post‐receptor retina is found in ERG responses to full‐field stimuli and in psychophysical thresholds that probe small retinal regions. In the central retina, both Mild and Severe ROP delay maturation of parafoveal scotopic thresholds and are associated with attenuation of cone mediated multifocal ERG responses, significant thickening of post‐receptor retinal laminae, and dysmorphic cone photoreceptors. These results have implications for vision and control of eye growth and refractive development and suggest future research directions. These results also lead to a proposal for noninvasive management using light that may add to the currently invasive therapeutic armamentarium against ROP. HIGHLIGHTSRetinopathy of prematurity (ROP) involves the neurosensory retina.The status of the immature rods may be a driver of ROP pathogenesis.Evidence of photoreceptor injury persists years after ROP is an active disease.Post‐receptor retina reorganizes to compensate for deficient photoreceptor inputs.The late maturing central retinal is especially vulnerable to the effects of ROP.