Susceptibility of Skin Cells to UVA-induced Necrotic Cell Death Reflects the Intracellular Level of Labile Iron

Abstract

The mechanism of resistance of keratinocytes to ultraviolet A (UVA) (320-400 nm)-induced oxidative damage has not yet been elucidated. Here, we examined the possible link between the intracellular level of the labile iron pool (LIP) and the susceptibility to UVA-induced cell death using a series of human skin fibroblast and keratinocyte cell lines as a model. Resistance of keratinocytes to UVA-induced cell death was confirmed by flow cytometry and in fibroblasts necrosis was found to be the primary mode of cell death induced by UVA. The percentage of necrosis in fibroblasts also correlated with the extent of intracellular ATP depletion, a hallmark of necrotic cell death. The evaluation of the intracellular level of LIP by calcein assay revealed that both "basal" and "UVA-induced" levels of LIP in keratinocytes were several fold lower than in fibroblasts. Accordingly the dose to give an equivalent level of necrosis was several fold lower in fibroblasts than in keratinocytes. Furthermore, the modulation of "basal" or "UVA-induced" level of LIP by either Desferal and/or hemin treatment significantly affected the extent of UVA-induced necrotic cell death and ATP depletion in all the cell lines. Cellular susceptibility to UVA-induced necrotic cell death appears to reflect the intracellular level of LIP.

Nrf2-mediated protection against UVA radiation in human skin keratinocytes

“…Earlier work demonstrated that HaCaT keratinocytes are resistant to UVA-mediated membrane damage at lower doses of radiation (13,14). Study of cell morphology found that low and moderate doses (100 and 250 kJ•m −2 ) of UVA irradiation did not alter cell focal adhesions and overall cell morphology, which is consistent with results of an earlier study indicating that human primary keratinocytes are generally resistant to UVA-mediated damage as gauged by LDH and cell morphology (12).…”

“…As the top layer of skin, epidermal keratinocytes encounter more UV irradiation and are more resistant to UVA irradiation-mediated cell damage than fibroblasts (12,13). Nrf2 knockout mice have a prolonged inflammatory response after skin injury (8).…”

“…The current study investigated the cell response to different doses of UVA irradiation. Earlier studies found that a moderate dose (250 kJ•m −2 ) of UVA irradiation did not activate Nrf2 in HaCaT human skin keratinocytes while a high dose (> 500 kJ• m −2 ) of UVA irradiation caused significant damage to these cells (13). Therefore, a moderate to high dose (400 kJ•m −2 ) of UVA irradiation was used to examine Nrf2 accumulation following irradiation.…”

The importance of iron chelation and iron availability during PpIX-induced photodynamic therapy

“…Having free iron available may have therefore not been crucial for this purpose and it is also likely that iron-mediated Fenton reactions are only a small part of a much larger mechanism of damage mediated by PpIX-induced PDT which is known to be originally initiated by type-II singlet oxygen production [65]. Furthermore, although cellular iron levels are tightly regulated under normal circumstances, during oxidative stress iron homeostasis can be disrupted resulting in the release of labile iron [66][67][68]. This may also partially explain our findings, so that iron chelation initially reduces iron availability so that PpIX accumulation during PpIX-PDT is elevated and once a state of oxidative stress begins to occur on irradiation other transition metals and freshly released labile iron perpetuate the ROS cascades via Fenton reactions.…”

Enhancing Protoporphyrin IX induced Photodynamic Therapy with a Topical Iron Chelating Agent in a Normal Skin Model

“…A possible explanation for this may be that other transition metals (such as zinc or copper) may be able to mediate Fenton-type reactions instead of iron in the ROS cascades triggered by PpIX-PDT. Furthermore, although cellular iron levels are tightly regulated under normal circumstances, during oxidative stress iron homeostasis can be disrupted resulting in the release of labile iron [50][51][52]. This might mean that although iron chelation initially reduces iron availability, so that PpIX accumulation during PpIX-PDT is elevated, once a state of oxidative stress begins to occur on irradiation other transition metals and freshly released labile iron could perpetuate the ROS cascades via Fenton reactions in a timely and efficacious fashion.…”