Scientific and Regulatory Policy Committee Points to Consider: Fixation, Trimming, and Sectioning of Nonrodent Eyes and Ocular Tissues for Examination in Ocular and General Toxicity Studies.

A Working Group of the Society of Toxicologic Pathology's Scientific and Regulatory Policy Committee conducted a technical and scientific review of current practices relating to the fixation, trimming, and sectioning of the nonrodent eye to identify key points and species-specific anatomical landmarks to consider when preparing and evaluating eyes of rabbits, dogs, minipigs, and nonhuman primates from ocular and general toxicity studies. The topics addressed in this Points to Consider article include determination of situations when more comprehensive evaluation of the globe and/or associated extraocular tissues should be implemented (expanded ocular sampling), and what constitutes expanded ocular sampling. In addition, this manuscript highlights the practical aspects of fixing, trimming, and sectioning the eye to ensure adequate histopathological evaluation of all major ocular structures, including the cone-dense areas (visual streak/macula/fovea) of the retina for rabbits, dogs, minipigs, and nonhuman primates, which is a current regulatory expectation for ocular toxicity studies.[Box: see text].

Immunology and Pathology in Ocular Drug Development.

Clear vision is dependent on features that protect the anatomical integrity of the eye (cornea and sclera) and those that contribute to internal ocular homeostasis by conferring hemangiogenic (avascular tissues and antiangiogenic factors), lymphangiogenic (lack of draining lymphatics), and immunologic (tight junctions that form blood-ocular barriers, immunosuppressive cells, and modulators) privileges. The later examples are necessary components that enable the eye to maintain an immunosuppressive environment that responds to foreign invaders in a deviated manner, minimizing destructive inflammation that would impair vision. These conditions allowed for the observations made by Medawar, in 1948, of delayed rejection of allogenic tissue grafts in the anterior chamber of mouse eye and permit the sequestration of foreign invaders (eg, Toxoplasma gondii) within the retina of healthy individuals. Yet successful development of intraocular drugs (biologics and delivery devices) has been stymied by adverse ocular pathology, much of which is driven by immune pathways. The eye can be intolerant of foreign protein irrespective of delivery route, and endogenous ocular cells have remarkable plasticity when recruited to preserve visual function. This article provides a review of current understanding of ocular immunology and the potential role of immune mechanisms in pathology observed with intraocular drug delivery.

Zebrafish as an Animal Model for Ocular Toxicity Testing: A Review of Ocular Anatomy and Functional Assays.

Xenobiotics make their way into organisms from diverse sources including diet, medication, and pollution. Our understanding of ocular toxicities from xenobiotics in humans, livestock, and wildlife is growing thanks to laboratory animal models. Anatomy and physiology are conserved among vertebrate eyes, and studies with common mammalian preclinical species (rodent, dog) can predict human ocular toxicity. However, since the eye is susceptible to toxicities that may not involve a histological correlate, and these species rely heavily on smell and hearing to navigate their world, discovering visual deficits can be challenging with traditional animal models. Alternative models capable of identifying functional impacts on vision and requiring minimal amounts of chemical are valuable assets to toxicology. Human and zebrafish eyes are anatomically and functionally similar, and it has been reported that several common human ocular toxicants cause comparable toxicity in zebrafish. Vision develops rapidly in zebrafish; the tiny larvae rely on visual cues as early as 4 days, and behavioral responses to those cues can be monitored in high-throughput fashion. This article describes the comparative anatomy of the zebrafish eye, the notable differences from the mammalian eye, and presents practical applications of this underutilized model for assessment of ocular toxicity.

From the Cover: Inhibitors of Nicotinamide Phosphoribosyltransferase Cause Retinal Damage in Larval Zebrafish.

Nicotinamide phosphoribosyltransferase (NAMPT) has been investigated as a target for oncology because it catalyzes a rate-limiting step in cellular energy metabolism to produce nicotinamide adenine dinucleotide. Small molecule inhibitors of NAMPT have been promising drug candidates but preclinical development has been hindered due to associated retinal toxicity. Here we demonstrate that larval zebrafish can predict retinal toxicity associated with this mechanism revealing an attractive alternative method for identifying such toxicities. Zebrafish permit higher throughput testing while using far lower quantities of test article compared with mammalian systems. NAMPT inhibitor-associated toxicity manifested in zebrafish as a loss of response to visual cues compared with auditory cues. Zebrafish retinal damage associated with NAMPT inhibitor treatment was confirmed through histopathology. Ranking 6 NAMPT inhibitors according to their impact on zebrafish vision revealed a positive correlation with their in vitro potencies on human tumor cells. This correlation indicates translatable pharmacodynamics between zebrafish and human NAMPT and is consistent with on-target activity as the cause of retinal toxicity associated with NAMPT inhibition. Together, these data illustrate the utility of zebrafish for identifying compounds that may cause ocular toxicity in mammals, and, likewise, for accelerating development of compounds with improved safety margins.

Hormonal profiles of captive Galapagos tortoises (Chelonoidis nigra).

Monthly blood samples, daily mating observations from Galapagos tortoises (Chelonoidis nigra), and local rainfall and temperature were collected at the Honolulu Zoo as part of a fertility evaluation. Testosterone concentrations were measured for males (n = 6), two of which were seen copulating and were considered sexually active. Estrone sulfate and progesterone concentrations were measured for female tortoises (n = 9), two of which nested and only one had laid eggs. Testosterone profiles were similar for both sexually active and sexually inactive males, both of which were positively correlated with temperature but not rainfall. Peak testosterone concentrations (12.0 ± 1.4 ng/ml sexually active animals vs. 14.4 ± 2.4 ng/ml sexually inactive animals) occurred at the end of the nesting season, from April to July. Estrone sulfate concentrations were similar for nesting (n = 2) and non-nesting (n = 7) female tortoises, rising from non-detectable concentrations (September), and increasing to peak concentrations during the nesting season. Progesterone concentrations remained low and spiked (9.44 ng/ml) only for the female that nested and laid eggs. Testosterone was negatively correlated with mating behavior, and the male tortoises were likely capable of spermatogenesis even though only two of them engaged in mating behavior. The female tortoises were not senescent, as the estrone sulfate concentrations likely reflected waves of ovarian follicular activity. Endocrine parameters were not in synchrony with rainfall, and a disconnect between the timing of reproductive events and the environmental milieu may help to explain the poor fertility of these tortoises. Zoo Biol. 35:237-245, 2016. © 2016 Wiley Periodicals, Inc.