Overexpression of H- and L-ferritin subunits in lens epithelial cells: Fe metabolism and cellular response to UVB irradiation

Department of Anatomy, Physiology, and Radiology, College of Veterinary Medicine, North Carolina State University, 4700 Hillsborough Street, Raleigh, NC 27606, USA.
Investigative Ophthalmology & Visual Science (Impact Factor: 3.4). 08/2001; 42(8):1721-7.
Source: PubMed


To determine the effect of changes in ferritin subunit makeup on Fe metabolism and the resistance of lens epithelial cells (LEC) to photo-oxidative stress.
Cultured canine LEC were transiently transfected with pTargeT mammalian expression vector containing the whole coding sequence of H- or L-chain cDNA. The subunit composition of newly synthesized ferritin was analyzed by metabolic labeling and SDS-PAGE electrophoresis. Total ferritin concentration was measured by ELISA: Fe uptake and incorporation into ferritin was determined by incubating transfected cells with (59)Fe-labeled transferrin followed by native PAGE electrophoresis. The effect of UV irradiation was assessed by cell count after exposure of transfected cells to UVB radiation.
Transfected cells differentially expressed H- and L-ferritin chains from cDNA under the control of CMV promoter; overexpression of L-chain was much greater than that of H-chain. The expressed chains assembled into ferritin molecules under in vitro and in vivo condition. The ferritin of H-transfectants incorporated significantly more Fe than those of L-transfectants. The UVB irradiation reduced cell number of L-transfectants by half, whereas H-chain transfectants were protected.
Post-transfectional expression of ferritin H- and L-chains in LEC appears to be regulated differentially. Overexpression of L-chain ferritin did not have a major effect on cellular Fe distribution and did not protect LEC against UV irradiation, whereas overexpression of H-chain resulted in increased storage of Fe in ferritin and protected cells from UV damage.

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    • "Moreover, the free radical superoxide (•O2−), another product of IR, is relatively non-reactive towards DNA directly but highly reactive to [Fe-S] clusters in proteins, inactivating their function and indirectly oxidizing proximal DNA [14], [15]. Finally, a number of studies have demonstrated that increased ferritin expression can lead to protection against a variety of forms of oxidative stress [16], [17], [18], [19]. Hence, the production of indiscriminately damaging hydroxyl radicals by free iron, protein-bound iron's susceptibility to inactivation by peroxides in various enzymes, and the sensitivity of iron-sulfur clusters to superoxide suggests that limiting the role of iron within the cell via enhanced sequestration by ferritin proteins could lead to a survival advantage during exposure to IR. "
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