The retinal pigment epithelium (RPE) is a highly differentiated, post-mitotic cell layer that performs a host of functions critical to retinal homeostasis. To discharge its many functions, the RPE requires ample energy. Studies of cultured RPE cells show that they can derive energy from glucose by either aerobic glycolysis or oxidative phosphorylation (OXPHOS), depending upon culture conditions. However, the balance between these two primary modes of RPE glucose metabolism in vivo is unknown, and it is unclear whether alterations in this balance occur under normal and/or disease conditions. Abundant evidence links changes in cellular energy metabolism with alterations in cell phenotype in a variety of fields including cancer, development, stem cell differentiation and aging. In the outer retina, mutations in mitochondrial DNA that compromise OXPHOS cause macular retinopathy. Moreover, disproportionate damage to mitochondrial DNA has been documented in the RPE of individuals with age-related macular degeneration (AMD), suggesting a causal link. We previously showed that postnatal loss of mitochondrial DNA and OXPHOS capacity in the murine RPE in vivo has surprising effects on cell phenotype, causing activation of cell growth pathways, increased glycolytic flux, and loss of epithelial functions and integrity. Our findings demonstrate that enforced changes in cellular energy metabolism in vivo can drive dedifferentiation and transdifferentiation of the RPE, and support a causal connection between diminished RPE OXPHOS capacity and AMD. However, our results raise new questions about how particular aspects of altered cellular energy metabolism read out as changes in cell phenotype. Can increased aerobic glycolysis alone, in the presence of intact OXPHOS, activate cell growth pathways, cause dedifferentiation/transdifferentiation and loss of epithelial integrity? Are features of the RPE glycolytic phenotype reversible through rebalancing of metabolism? What aspects of the altered phenotype of OXPHOS- deficient RPE result from lack of ATP production via OXPHOS versus loss of electron transport to oxygen? Does diurnal variation in energy metabolism affect the capacity of the RPE to phagocytize outer segment tips? We propose an ensemble of experiments to address these questions. Specifically we will modulate RPE aerobic glycolysis in vivo in the context of intact OXPHOS, restore respiration without ATP generation to OXPHOS-deficient RPE in vivo, and probe the relationship between RPE energy metabolism and diurnal phagocytic capacity. Through detailed characterization and quantification of the alterations in RPE cell phenotype caused by these in vivo metabolic changes, we will uncover mechanistic connections between energy metabolism and RPE cell phenotype. Because we will work in vivo, the impact of RPE metabolic modulation on photoreceptors will be apparent. Thus, success of this project will not only provide foundational knowledge of in vivo RPE metabolism and its relationship to cell phenotype, it will also inform studies of abnormal RPE metabolism in human retinal disease.

Public Health Relevance

The nerves that sense light in the eye and the cells of the retinal pigment epithelium (RPE) form a critical partnership necessary for vision. We seek to understand how variation in the way the RPE obtains energy from food molecules affects the tissue's ability to function and support the nerves. Success of this project will provide insight into how aging and disease affect this partnership and suggest new avenues for development of therapies for disorders that originate with the RPE.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
1R01EY025790-01A1
Application #
9174545
Study Section
Biology of the Visual System Study Section (BVS)
Program Officer
Neuhold, Lisa
Project Start
2016-09-01
Project End
2021-08-31
Budget Start
2016-09-01
Budget End
2017-08-31
Support Year
1
Fiscal Year
2016
Total Cost
Indirect Cost
Name
Stanford University
Department
Genetics
Type
Schools of Medicine
DUNS #
009214214
City
Stanford
State
CA
Country
United States
Zip Code
94304
Benchorin, Gillie; Calton, Melissa A; Beaulieu, Marielle O et al. (2017) Assessment of Murine Retinal Function by Electroretinography. Bio Protoc 7: