ABS 004: Absolute Quantification of Amino Acids within Male and Female Ocular Tissues

Cloe Ratliff ¹ ², Mark Eminhizer ¹ ², Tuan Ngo ¹ ², Jianhai Du ¹ ²

¹ Department of Ophthalmology & Visual Sciences, West Virginia University
² Department of Biochemistry & Molecular Medicine, West Virginia University

Van Wickle (2025) Volume 1, ABS004

Introduction: The eye is one of the most metabolically active organs within the human body, largely due to the high energy demands of the photoreceptors’ renewal and visual transduction. The photoreceptors are specialized neurons that play a crucial role in detecting light through specialized light-sensing proteins located in their outer segment (OS) discs. These discs are regularly shed and renewed in a daily cycle, helping to maintain the photoreceptors’ optimal length while also eliminating toxic photo-oxidative byproducts that accumulate during the visual transduction process. The disc-shedding process peaks in the morning right after the eyes are opened and decreases throughout the day. As the energy expenditure varies throughout the day the concentration and requirement of metabolites will also vary. The metabolic processes within the eye are influenced by several factors, including genetic expression, aging, diet, and environmental conditions, all of which can impact the overall health of the eye. Metabolic dysregulation can lead to diseases such as diabetic retinopathy and age-related macular degeneration (AMD), the leading cause of blindness among the elderly. Sex significantly impacts eye metabolism and susceptibility to ocular diseases, with women being more prone to conditions such as AMD, cataracts, and glaucoma. While morphological differences between male and female ocular tissues are known, the metabolic basis for these sex differences remains unknown and could be vital to the growth of how we treat these diseases.

Methods: We collected four different tissues, retina, retinal pigment epithelium (RPE), cornea, and lens of 8-week old wild type (WT) C57BL/6J mice; male (6) and female (6), at either 10 am or 2 pm. Amino acids were extracted using an 80% cold methanol method with internal standard. We were able to differentiate the metabolic usages between sex and different times of the day. The mix was centrifuged and supernatant was used to analyze the targeted metabolites present using Agilent 7890B gas chromatography-mass spectrometry (GC-MS). Protein concentrations from extraction pellets were measured for data normalization using MetaboAnalyst 5.0.

Results: Multivariate analysis helped identify key metabolic changes that are sex and tissue specific. There was no difference in male retina or RPE based on time. The female displayed differences in isoleucine, leucine, and lysine in the retina, as well as metabolites in the TCA and glycolysis cycles in RPE. In cornea there is downregulation of taurine in both sexes at 2 pm. Females show a decrease in 21 metabolites whereas males display only 4. Remarkably, metabolites associated with nucleotide and glutathione pathways are upregulated in male lens tissue only, the opposite trend for the other tissues.

Discussion: Time plays an important, yet differing, role in male versus female mice. In the retina, cornea, and RPE female amino acid abundance is dependent on time but male amino acid abundance is not. Upregulation of BCAAs in the female retina later in the day suggests greater muscle usage. Downregulation of amino acids involved in the TCA cycle in female RPE suggests less energy production and usage later in the day. The opposite trend is true in the lens in which male amino acid abundance is dependent on time and female amino acid abundance is less affected by time.