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Rob Pintwala, Cameron Postnikoff, Maud Gorbet; Co-culture with material-activated macrophages induce an inflammatory phenotype in lens epithelial cells. Invest. Ophthalmol. Vis. Sci. 2013;54(15):2951.
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© ARVO (1962-2015); The Authors (2016-present)
Posterior capsular opacification (PCO) is a frequent complication of cataract surgery and remains an important issue despite efforts to improve intraocular lens (IOL) materials and surgical techniques. Macrophages have been observed at the postoperative lens, although little is known about their interaction with both IOLs and lens cells. The aim of the current investigation is to develop an in vitro model of the interaction between human macrophages and lens epithelial cells to investigate the potential inflammatory mechanisms that occur in material-associated IOL complications.
The human acute monocytic leukemia cell line (THP-1) was cultured in RPMI 1640 cell culture medium supplemented with 10% fetal bovine serum (FBS). The monocytes were differentiated with phorbol 12-myristate 13-acetate into macrophage-like cells in 6-well tissue culture polystyrene plates. After 48 hours the media was replenished and cells were allowed to rest for a further 48 hours. Human lens epithelial cells (cell line: HLE B-3) were cultured in Dulbecco’s modified eagle medium (DMEM) supplemented with 20% FBS and were seeded into 12-well polyethylene terephthalate cell culture inserts. After 24 hours macrophages were transferred into the bottom of the 12-well polystyrene plates along with an IOL to physically separate it from the lens cells. The co-culture was incubated for 96 hours with media replenished every 48 hours. Lens epithelial cells were examined for changes in phenotype using immunocytochemistry to determine expression of fibronectin. The macrophages were analyzed via flow cytometry for changes in expression of CD36, CD14 and CD45.
Macrophages and lens cells grew well together. After 96 hours, increased expression of fibronectin was observed on lens epithelial cells co-cultured with macrophages interacting with PMMA and hydrophilic acrylic IOLs. Flow cytometry analysis of macrophages interacting with the IOL showed a down-regulation of CD14 and CD45 and an up-regulation of CD36 relative to the control (no IOL). No difference in macrophage activation was observed between the two IOL materials.
The novel co-culture model suggests that the presence of macrophages and their secretory products may induce an inflammatory phenotype in the human lens epithelium. Further work will include characterization of alpha-smooth muscle actin and E-cadherin expression.
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