A Novel Choroidal Endothelial Cell Line Has a Decreased Affinity for the Age-Related Macular Degeneration–Associated Complement Factor H Variant 402H

Abstract

PURPOSE. Choroidal endothelial cells play a central role in the pathogenesis of age-related macular degeneration (AMD). Protocols for isolating primary choroidal endothelial cells have been described but require access to human donor eyes, which is a limiting factor. Therefore, a conditionally immortalized choroidal endothelial cell (ciChEnC) line has been established. METHODS.Choroidal endothelial cells were selected by magnetic-activated cell sorting and conditionally immortalized using temperature-sensitive simian virus 40 large T antigen and human telomerase. The cell line obtained was characterized based on expression of endothelial marker proteins and endothelial cell-specific responses to various stimuli. Binding of AMDassociated and non-AMD variants of complement factor H in the context of a recombinant CCP6-8 (complement control protein domains 6-8) construct was determined using ELISA.RESULTS. ciChEnCs maintained morphology and von Willebrand factor and vascular endothelial cadherin expression for up to 27 passages. The cells internalized acetylated low-density lipoprotein, formed tubes on Matrigel, and increased intercellular adhesion molecule-1 expression in response to tumor necrosis factor-a. Cells grew into dense monolayers with barrier function and showed characteristics of choriocapillary cells, such as expression of plasmalemma vesicle-associated protein, human leukocyte antigen ABC, carbonic anhydrase IV, and membrane indentations reflecting fenestrations. ciChEnCs synthesized glycosaminoglycans chondroitin sulfate and the complement factor H ligand heparan sulfate. Interestingly, binding of the AMD-associated 402H variant of factor H to ciChEnC was significantly decreased compared to the 402Y variant. CONCLUSIONS.A novel ciChEnC cell line with choriocapillary characteristics has been established and should greatly facilitate investigation of the pathogenesis of AMD in the context of the choriocapillary microenvironment.

RF/6A Chorioretinal Cells Do Not Display Key Endothelial Phenotypes

“… 22 Primary and immortalized human choroidal endothelial cells are also reported to be VWF positive. 23 , 24 Thus, bona fide choroidal and retinal endothelial cells would be expected to contain WPB.…”

“…Another long established functional behavior of endothelial cells, including an immortalized human choroidal endothelial cell line, 24 is uptake and metabolism of acetylated low density lipoprotein (ac-LDL). 29 Primary HUVEC and HREC exhibited robust ac-LDL uptake.…”

Mechanisms of FH Protection Against Neovascular AMD

“…Furthermore, Toomey et al hypothesize that FH and RPE lipoproteins (with or without oxidative modifications) compete for binding to the GAGs ≪zip codes≫ observed on the Bruch's membrane leading to increased lipoprotein accumulation and drusen formation with the presence of FH 402H (36). Moreover, mixed data have been observed regarding the altered affinity of the FH 402H variant on RPE cells (10,16) or on a choroidal endothelial cell line (37). It remains undetermined whether the FH 402H polymorphism directly affects choriocapillaris and how it participates to pathogenesis of AMD, including CNV.…”

Red risks for a journey to the red planet: The highest priority human health risks for a mission to Mars

“…Primarily, these include edema (swelling) of the optic disc and retinal nerve fiber layer (RNFL), chorioretinal folds (wrinkles in the retina), globe flattening, and refractive error shifts 47 . Flight duration is thought to play a role in the pathogenesis of SANS, as nearly all cases have been diagnosed during or immediately after long-duration spaceflight (i.e., missions of 30 days duration or longer), although signs have been discovered as early as mission day 10 48 . Because of SANS, ocular data are nominally collected during ISS missions.…”

“…OCT has been utilized onboard the ISS since late 2013, and it has revolutionized NASA’s ability to objectively detect and monitor SANS and build a high-resolution database of retinal and optic nerve head images. Through this technology, it has been recently discovered that that a majority of long-duration astronauts (including females) present with some level of ODE and engorgement of the choroidal vasculature 48 , 55 . The trends and patterns of these ocular anatomical changes may hold the key to deciphering the pathophysiology of SANS 48 , 55 .…”