In a recent study published in International Journal of Molecular Sciencesresearchers have found that loss of pigment epithelium-derived factor (PEDF) can cause eye aging.
The retina is the light-sensitive tissue located at the back of the eye. Additionally, age-related retinal disorders such as age-related macular degeneration (AMD) can cause blindness.
The retinoprotective protein PEDF is produced by the retinal pigment epithelium (RPE)-expressed serine protease inhibitor family 1 gene (Serpinf1) and decreases with retinal degeneration, cellular senescence, and aging. PEDF, a member of the serpin superfamily, has been compared to the guardian of the eye because it protects retinal photoreceptors, neurons, and the RPE from pathological damage and prevents choroidal and retinal neovascularization.
The authors of the current study previously reported that knockdown of patatin-like phospholipase domain-containing 2 (Pnpla2) slows the digestion of lipids and rhodopsin across RPE-passing phagocytosis. This conclusion suggests that the PEDF receptor (PEDF-R) is essential for RPE phagocytosis and possibly links PEDF to the process. However, the involvement of PEDF in RPE phagocytosis and the activities directed by PEDF-R are not specified.
This work used a Serpinf1 null mouse model to determine whether PEDF can help prevent aging in the RPE.
The authors evaluated the expression of genes associated with aging across the RPE of three-month-old Serpinf1 null mice. They performed quantitative polymerase chain reaction (qPCR) using complementary deoxyribonucleic acid (cDNA) made from ribonucleic acid (RNA) collected from freshly dissected Serpinf1+/-, Serpinf1+/+ and Serpinf1-/- choroid/RPE.
5-Diaminofluorescein (5-DAF) fluorescence intensity per RPE cell was assessed in both genotypes and plotted to quantitatively investigate the impact of PEDF ablation on senescence-associated galactosidase (SA-β-gal) function. Furthermore, because senescent cells experience changes in nuclear size, the researchers used fluorescence confocal imaging to analyze the subcellular morphology of the RPE.
The team examined the expression of the Pnpla2 gene, which was necessary for the RPE to degrade photoreceptor outer segments, as this function of the RPE declines with aging. In addition, they measured the amount of phagocytosed rhodopsin and lipids across the RPE of Serpinf1-deficient mice. For each genotype, the number of particles per RPE cell was calculated and plotted to quantitatively assess the impact of PEDF deficiency on rhodopsin turnover. Boron-dipyrromethane (BODIPY) intensity per region of interest (ROI) was calculated for each genotype and plotted for quantitative analysis of the effect of PEDF deficiency on RPE lipid accumulation.
The team found that Serpinf1 deletion triggered the cyclin-dependent kinase inhibitor 1A (Cdkn1a) for the p21 protein, the histone family member H2A X (H2ax) for the histone H2AX protein, and the galactosidase beta 1 (Glb1) gene for β-galactosidase. The results of the study showed that the RPE undergoes senescent changes after the loss of PEDF. This conclusion was supported by the identified 1) nuclear enlargement, 2) increased SA-β-gal function, 3) activation of senescence-associated genes Glb1, Cdkn1a, and H2ax, and 4) disorganized F-actin distribution pattern and similar change phenotypes across the RPE of Serpinf1- /- (Serpinf1 null) versus wild-type RPE.
The researchers found that ablation of Serpinf1 increased the volume of RPE cell nuclei and the number of nucleoli, indicating chromatin rearrangement. They observed that after deletion of the Serpinf1 gene, the Pnpla2 gene and the related PEDF-R protein decreased.
In addition, rhodopsin and lipids were deposited in the RPE of Serpinf1-deficient animals compared with littermate controls. The observation that Serpinf1/RPE accumulates rhodopsin and lipids and downregulates the phagocytosis-related gene Pnpla2 indicates an impairment of RPE phagocytosis caused by PEDF ablation.
The study findings support earlier findings that aging causes a decrease in the phagocytic activity of the RPE. Together, they demonstrate how PEDF helps prevent aging and promotes phagocytosis, demonstrating its dual function in promoting RPE activity and its effects.
The researchers noted that this study was the first to use a Serpinf1-deficient mouse to examine RPE aging and phagocytosis. The study findings highlight PEDF as a retinoprotective and regulatory protein of age-like changes associated with defective degradation of the photoreceptor outer segment across the RPE, demonstrating the lack of PEDF as a contributor to senescence-like modifications in the RPE.
Taken together, the current research demonstrates the importance of PEDF signaling in the development of age-related eye diseases. The team reported that using PEDF-null mouse models may help define new signaling pathways involving phagocytosis that affect lipid and visual pigment recycling across eye diseases and offer mechanistic insight into the investigation of aging and senescence.
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