ATP-Sensitive Potassium Channel: A Novel Target for Protection Against UV-Induced Human Skin Cell Damage CONG CAO, 1 SARAH HEALEY, 1 ASHLEY AMARAL, 1 AVERY LEE-COUTURE, 1 SHU WAN, 1 NICOLA KOUTTAB, 2 WENMING CHU, 3 AND YINSHENG WAN 1 * 1 Department of Biology, Providence College, Providence, Rhode Island 2 Department of Pathology, Roger Williams Medical Center, Boston University, Providence, Rhode Island 3 Department of Molecular Microbiology and Immunology, Brown University, Providence, Rhode Island Ultraviolet radiation (UV) induces cell damages leading to skin photoaging and skin cancer. ATP-sensitive potassium (K ATP ) channel openers (KCOs) have been shown to exert significant myocardial preservation and neuroprotection in vitro and in vivo, and yet the potential role of those KCOs in protection against UV-induced skin cell damage is unknown. We investigated the effects of pinacidil and diazoxide, two classical KCOs, on UV-induced cell death using cultured human keratinocytes (HaCat cells). Here, we demonstrated for the first time that Kir 6.1, Kir 6.2 and SUR2 subunits of K ATP channels are functionally expressed in HaCaT cells and both non- selective K ATP channel opener pinacidil and mitoK ATP (mitochondrial K ATP ) channel opener diazoxide attenuated UV-induced keratinocytes cell death. The protective effects were abolished by both non-selective K ATP channel blocker glibenclamide and selective mitoK ATP channel blocker 5-hydroxydecanoate (5-HD). Also, activation of K ATP channel with pinacidil or diazoxide resulted in suppressive effects on UV-induced MAPK activation and reactive oxygen species (ROS) production. Unexpectedly, we found that the level of intracellular ROS was slightly elevated in HaCaT cells when treated with pinacidil or diazoxide alone. Furthermore, UV-induced mitochondrial membrane potential loss, cytochrome c release and ultimately apoptotic cell death were also inhibited by preconditioning with pinacidil and diazoxide, and their effects were reversed by glibenclamide and 5-HD. Taken together, we contend that mitoK ATP is likely to contribute the protection against UV-induced keratinocytes cell damage. Our findings suggest that K ATP openers such as pinacidil and diazoxide may be utilized to prevent from UV-induced skin aging. J. Cell. Physiol. 212: 252–263, 2007. ß 2007 Wiley-Liss, Inc. ATP-sensitive K þ (K ATP ) channels, originally discovered in the heart, are widely distributed in many tissues and cell types, including pancreatic b-cell, brain, skeletal and smooth muscles, and kidney (Ishibashi et al., 1998; Garlid et al., 2003). K ATP channels are located in various parts of the cell, including plasmalemmal membrane (sK ATP ) and inner mitochondrial membrane (mitoK ATP ) (Ishibashi et al., 1998; Garlid et al., 2003). It is generally believed that opening of sK ATP channels results in an enhancement of membrane repolarization and thereby shortening of action potential duration, leading to reduction in calcium entry into cells through voltage-gated calcium channels (Auchampach and Gross, 1993). Such reduction in calcium entry may decrease cardiac work and prevent calcium- dependent pathological events. While in pancreatic b-cells, increased ATP concentration due to increased glucose metabolism closes K ATP channels allows calcium influx, the resultant rise in intracellular calcium concentration ([Ca 2þ ] i ) in the b-cells triggers insulin secretion (Ashcroft et al., 1984; Cook and Hales, 1984). One exciting new direction has been the selective activation of mitoK ATP channels as a means of protecting cells against lethal ischemia or chemical stress. Activation of mitoK ATP channels protects mitochondria and the cells primarily by decreasing mitochondrial transmembrane potential and inhibiting or delaying massive calcium influx into mitochondria induced by ischemia or chemical stress, preserving intermembrane space or matrix volume, ultimately reducing ROS production and release of cytochrome c as well as apoptotic cell death (Garlid et al., 2003; Busija et al., 2004). Numerous recent studies have shown that opening of mitoK ATP channels with drugs or by physiological stimuli protects the heart against subsequent ischemic stress (Hu et al., 1999; Sato et al., 2000). Also, recent but limited studies, including our previously works (Hu et al., 2005; Wang et al., 2005b, 2006; Liu et al., 2006; Yang et al., 2006), provide evidence that activation of mitoK ATP channels protects the brain against ischemic or chemical challenge (Ishibashi et al., 1998; Busija et al., 2004). The role of K ATP channel and K ATP channel openers (KCOs) in protection against ultraviolet radiation (UV)-induced cell death have not been studied thus far. One of the major biological features of UV is the induction of apoptotic cell death of keratinocytes, which in vivo appear as sunburn cells (SC) within the epidermis. And the major action spectrum inducing the formation of SC is UVB and A2. Previous works from human skin keratinocytes exposed to UVB radiation have indicated that various genes might be involved in concomitant apoptosis Contract grant sponsor: NIH (from INBRE Program of the National Center for Research Resources); Contract grant number: P20 RR016457. *Correspondence to: Yinsheng Wan, Department of Biology, Providence College, 549 River Ave. Providence, RI 02918-0001. E-mail: yswan@providence.edu Received 27 September 2006; Accepted 19 December 2006 DOI: 10.1002/jcp.21026 ORIGINAL ARTICLE 252 Journal of Journal of Cellular Physiology Cellular Physiology ß 2007 WILEY-LISS, INC.