Fluorophotometry is a sensitive method/technique to evaluate epithelial permeability in murine experimental dry eye

Purpose. Fluorophotometry is a well validated method for assessing corneal permeability in human subjects. However, with the growing importance of basic science animal research in ophthalmology, fluorophotometry’s use in animals must be further evaluated. The purpose of this study was to evaluate corneal epithelial permeability following desiccating stress using the modified Fluorotron Master™. ^ Methods. Corneal permeability was evaluated prior to and after subjecting 6-8 week old C57BL/6 mice to experimental dry eye (EDE) for 2 and 5 days (n=9/time point). Untreated mice served as controls. Ten microliters of 0.001% sodium fluorescein (NaF) were instilled topically into each mouse’s left eye to create an eye bath, and left to permeate for 3 minutes. The eye bath was followed by a generous wash with Buffered Saline Solution (BSS) and alignment with the Fluorotron Master™. Seven corneal scans using the Fluorotron Master were performed during 15 minutes (1 st post-wash scans), followed by a second wash using BSS and another set of five corneal scans (2nd post-wash scans) during the next 15 minutes. Corneal permeability was calculated using data calculated with the FM™ Mouse software. ^ Results. When comparing the difference between the Post wash #1 scans within the group and the Post wash #2 scans within the group using a repeated measurement design, there was a statistical difference in the corneal fluorescein permeability of the Post-wash #1 scans after 5 days (1160.21±108.26 vs. 1000.47±75.56 ng/mL, P<0.016 for UT-5 day comparison 8 [0.008]), but not after only 2 days of EDE compared to Untreated mice (1115.64±118.94 vs. 1000.47±75.56 ng/mL, P>0.016 for UT-2 day comparison [0.050]). There was no statistical difference between the 2 day and 5 day Post wash #1 scans (P=.299). The Post-wash #2 scans demonstrated that EDE caused a significant NaF retention at both 2 and 5 days of EDE compared to baseline, untreated controls (1017.92±116.25, 1015.40±120.68 vs. 528.22±127.85 ng/mL, P<0.05 [0.0001 for both]). There was no statistical difference between the 2 day and 5 day Post wash #2 scans (P=.503). The comparison between the Untreated post wash #1 with untreated post wash #2 scans using a Paired T-test showed a significant difference between the two sets of scans (P=0.000). There is also a significant difference between the 2 day comparison and the 5 day comparison (P values = 0.010 and 0.002, respectively). ^ Conclusion. Desiccating stress increases permeability of the corneal epithelium to NaF, and increases NaF retention in the corneal stroma. The Fluorotron Master is a useful and sensitive tool to evaluate corneal permeability in murine dry eye, and will be a useful tool to evaluate the effectiveness of dry eye treatments in animal-model drug trials.^

The function of math5 and myocilin in mammalian eye development and disease

Complex molecular events underlie vertebrate eye development and disease. The eye is composed of two major tissue types: the anterior and posterior segments. During development, the retinal progenitor cells differentiate into six neuronal and one non-neuronal cell types. These cell types later organize into the distinct laminar structure of the mature retina which occupies the posterior segment. In the developed anterior segment, both the ciliary body and trabecular meshwork regulate intraocular pressure created by the aqueous humor. The disruption in intraocular pressure can lead to a blinding condition called glaucoma. To characterize molecular mechanisms governing retinal development and glaucoma, two separate mouse knockout lines carrying mutations in math5 and myocilin were subjected to a series of in vivo analyses. ^ Math5 is a murine homologue of Drosophila atonal , a bHLH proneural gene essential for the formation of photoreceptor cells. The expression of math5 coincides with the onset of retinal ganglion cell differentiation. The targeted deletion of mouse math5 revealed that a null mutation inhibits the formation of a majority of the retinal ganglion cells. The mutation also interferes with the normal development of other retinal cell types such as amacrine, bipolar and photoreceptor cells. These results suggest that math5 is a proneural gene responsible for differentiation of retinal ganglion cells and may also have a role in normal development of other neuronal cell types within the retina. ^ Myocilin has two unique protein coding regions bearing homology to non-muscle myosin of Dictyostelium discoideum and to olfactomedin, an extracellular matrix molecule first described in the olfactory epithelium of the bullfrog. Recently, autosomal dominant forms of myocilin mutations have been found in individuals with primary open-angle glaucoma. The genetic linkage to glaucoma suggests a role of myocilin in normal intraocular pressure and ocular function. However, the analysis of mice heterozygous and homozygous for a targeted null mutation in myocilin indicates that it is dispensable for normal intraocular pressure or ocular function. Additionally, the lack of a discernable phenotype in both heterozygous and null mice suggests that haploinsufficiency is not a critical mechanism for MYOC-associated glaucoma in humans. Instead, disease-causing mutations likely act by gain of function. ^ In summary, these studies provide novel insights into the embryonic development of the vertebrate retina, and also begin to uncover the molecular mechanisms responsible for the pathogenesis of glaucoma. ^

Positional cloning and characterization of the human aniridia and murine small eye genes

Aniridia (AN) is a congenital, panocular disorder of the eye characterized by the complete or partial absence of the iris. The disease can occur in both the sporadic and familial forms which, in the latter case, is inherited as an autosomal dominant trait with high penetrance. The objective of this study was to isolate and characterize the genes involved in AN and Sey, and thereby to gain a better understanding of the molecular basis of the two disorders.^ Using a positional cloning strategy, I have approached and cloned from the AN locus in human chromosomal band 11p13 a cDNA that is deleted in two patients with AN. The deletions in these patients overlap by about 70 kb and encompass the 3$\sp\prime$ end of the cDNA. This cDNA detects a 2.7 kb mRNA encoded by a transcription unit estimated to span approximately 50 kb of genomic DNA. The message is specifically expressed in all tissues affected in all forms of AN, namely within the presumptive iris, lens, neuroretina, the superficial layers of the cornea, the olfactory bulbs, and the cerebellum. Sequence analysis of the AN cDNA revealed a number of motifs characteristic of certain transcription factors. Chief among these are the presence of the paired domain, the homeodomain, and a carboxy-terminal domain rich in serine, threonine and proline residues. The overall structure shows high homology to the Drosophila segmentation gene paired and members of the murine Pax family of developmental control genes.^ Utilizing a conserved human genomic DNA sequence as probe, I was able to isolate an embryonic murine cDNA which is over 92% homologous in nucleotide sequence and virtually identical at the amino acid level to the human AN cDNA. The expression pattern of the murine gene is the same as that in man, supporting the conclusion that it probably corresponds to the Sey gene. Its specific expression in the neuroectodermal component of the eye, in glioblastomas, but not in the neural crest-derived PC12 pheochromocytoma cell line, suggests that a defect in neuroectodermal rather mesodermal development might be the common etiological factor underlying AN and Sey. ^

The illness experience of persons with Parkinson's Disease

Background. Parkinson's disease is a chronic, progressive, age-related, neurodegenerative disorder with no known cause or promising cure. While substantial information is known about the pathophysiology of Parkinson's disease, little is known about the illness experience of persons living with the disease. The purpose of this study was to understand how persons with Parkinson's disease construct their illness experience and manage living with their illness on a daily basis. ^ Method. A qualitative study with an ethnographic approach employed the strategies of participant observations and fieldwork. Field data were generated from a two year exposure to two Parkinson's disease support groups in east Texas. Open-ended semi-structured interviews with seven men and seven women with Parkinson's disease were also conducted. These data were combined and analyzed using thematic analysis. ^ Findings. The illness experience is described through the metaphor "Sailing the Sea in The Eye of the Storm." This metaphor served as the overarching theme that covered the two interacting content themes of the voyage of Daily Negotiations in the Midst of Uncertainty and Reconstruction of the Self with Parkinson's Disease. Daily negotiations incorporated navigating daily activities with the uncertainty of both the progression and daily vicissitudes of the disease. Participants described their symptoms as progressive imprisonment that interfered with daily activities. The progressive nature of the disease required the participants to reconstruct their perceptions of themselves. Reconstructing the self involved the paradoxical balancing of preserving the self while simultaneously releasing aspects of the former self to reconstruct the self with Parkinson's disease. This process was reflected in four exemplars: I Know Me." "It's Still Me," "See Me." and "Remember Me." ^ Conclusions. This qualitative study illuminated the struggle of persons in dealing with the uncertainties and fluctuations of Parkinson's disease and the process of reconstructing their perceptions of themselves. The meaning and reconstruction of the illness experience expressed by participants will inform understanding beyond the disease itself to the illness experience that these participants must deal with on a daily basis. ^

Effects of treatment on orienting in Parkinson disease

Parkinson disease (PD) is a movement disorder affecting over one million Americans, and 1% of our population over 60 years of age. Currently, PD has an unknown cause, no predictive biomarker, and no cure, yet there are effective treatments (medicine and surgery) to chronically manage the motor symptoms. But, PD patients also develop cognitive symptoms (e.g., distractibility, executive dysfunction) that remain untreated or may decline as a result of treating the motor symptoms. To address this important issue, I measured covert orienting of attention and overt eye movements in PD patients to assess the patients' ability to automatically detect stimuli in their visual field, to predict and attend to where the stimuli would appear, and to volitionally look somewhere else. ^ PD patients completed the cognitive tasks under multiple treatment conditions, and their performance was compared to healthy adults. PD patients first completed the tasks after they had withdrawn from medication. Their unmedicated performance revealed exaggerated automatic orienting, poor predictability, and weak volitional orienting. PD patients then repeated the tasks while medication was giving its peak benefit. The medication returned automatic covert orienting toward normal but did not improve volitional covert orienting. Several PD patients completed the tasks a third time after receiving surgery (specifically, implantation of stimulating electrodes in a subcortical brain region to alleviate motor symptoms). The stimulation (without medication) returned automatic orienting toward normal, did not change predictability, and further impaired volitional orienting. Taken together, treatments prescribed to alleviate the motor symptoms (a patient's primary concern) only improve some cognitive functions. Future studies may establish criteria to predict which patients are more likely to have cognitive benefit from medication over surgery, or vice versa. ^ I have also hypothesized an anatomical model relating orienting circuitry to abnormal PD circuitry and the therapeutic targets. My results suggest medication is more effective restoring the orienting circuitry than stimulation. Further, automatic and volitional orienting abilities seem to be modulated independently, which differs from an earlier model proposing a dependent, inverse relationship. My results are further discussed in terms of response inhibition, response selection, and the location of the selection. ^

A case-control study of the association between corneal arcus and death by cardiovascular disease among cornea donors

The cornea of the human eye can develop deposits of lipids in the periphery known as corneal arcus. [2, 10] For over a century, these deposits have been of interest as possible indicators of the accumulation of lipids in arterial walls of the heart and body with implications for heart disease. [2, 10, 11, 29] Heart disease is currently the leading cause of death in this country. [5, 29] There have been several publications suggesting an association between the development of atherosclerotic lesions and corneal arcus. [2, 12, 29] Investigators have differed in their interpretation of the relevance of corneal arcus to coronary heart disease or cardiovascular disease. However, there is widespread consensus that the presence of corneal arcus in patients under the age of 50 should prompt physicians to further investigate for dyslipidemia or heart disease. [2, 3, 6, 8, 19] Earlier studies have often suffered from difficulty in determining the presence or severity of atherosclerosis and from inconsistencies in evaluating corneal arcus. This study involves the review of mortality data, medical and social history and standardized slit lamp examination of corneal tissue donors to evaluate the prevalence of corneal arcus in relation to death by CHD or CVD. The prevalence of arcus, odds ratio, and logistic regression was utilized for statistical analysis.^