Sofie Van Kelst

Activation of p38 MAPK is required for Bax translocation to mitochondria, cytochrome c release and apoptosis induced by UVB irradiation in human keratinocytes

This study establishes that activation of p38 MAPK by UVB represents a crucial signal required for the conformational change and translocation of Bax to the mitochondria in human keratinocytes. UVB‐induced Bax translocation and mitochondrial cytochrome c release, which precede caspase activation and other endpoints of the apoptotic program such as chromatin fragmentation and loss of mitochondrial transmembrane potential, are blocked by genetic or pharmacological inhibition of the p38α MAPK. Inhibition of p38 MAPK strongly reduces the UVB‐induced formation of sunburn cells and blocks Bax conformational change both in cultured human keratinocytes and in human skin, providing clear evidence for the physiological role of the p38 MAPK‐Bax pathway in the removal of precancerous, UVB‐damaged keratinocytes. Furthermore, we show that Bcl‐2 overexpression, but not the pan‐caspase inhibitor zVAD‐fmk, blocks Bax conformational change and its subsequent translocation downstream of p38 MAPK. These data indicate that the activation of p38 MAPK by UVB engages a caspase‐independent death signal leading to mitochondrial membrane permeabilization and apoptosis in human keratinocytes and suggest that p38 MAPK might have a preventive role in the process of photocarcinogenesis.

Concomitant inhibition of AKT and autophagy is required for efficient cisplatin-induced apoptosis of metastatic skin carcinoma

Cutaneous squamous cell carcinoma (cSCC) is one of the most common cancers in the Caucasian population. Although early stages of skin cancer have a high curability and excellent prognosis, advanced cSCC shows resistance to chemotherapy, including cisplatin. The PI3‐K/AKT pathway is known to have a role in both skin cancer development and resistance to therapeutic drugs. In this study, we used isogenic cell lines representing different stages of malignant transformation of the keratinocytes that were derived from dysplastic forehead skin (PM1), primary cutaneous SCC (MET1) and its lymph node metastasis (MET4) of an immunosuppressed patient. We show that skin tumor progression parallels enhanced AKT activation and increased resistance to cisplatin‐induced apoptosis. Pharmacological AKT inhibition, or specific AKT1 knock down, sensitizes the apoptosis‐resistant MET1 and, to a lesser extent, MET4 cells to cisplatin‐mediated cell death. Concomitantly autophagy induction was observed in MET4, as demonstrated by accumulation of the autophagic protein marker LC3‐II, by analysis of full autophagosome maturation process using tandem mRFP‐GFP fluorescence microscopy and by electron microscopy. Counteracting the autophagic process by 3‐methyladenine or specific ATG5 knock down enhanced cytotoxicity of cisplatin combined with AKT inhibitor, thus revealing a key role for autophagy in chemoresistance. Taken together, these results indicate that concomitant inhibition of autophagy is required to increase the therapeutic benefit of AKT inhibition for combination therapy with the standard chemotherapeutic agent cisplatin in advanced skin carcinoma.

The Flavonoid Luteolin Increases the Resistance of Normal, but Not Malignant Keratinocytes, Against UVB-Induced Apoptosis

Adequate protection of skin against the carcinogenic effects of UVB irradiation is essential. Flavonoids may have a conspicuous role in cancer prevention because of their antioxidant, anti-inflammatory, and growth-inhibitory effects. Therefore, we tested the effects of the flavone luteolin (LUT) on selected parameters of the sunburn response in normal human keratinocytes, exposed to physiological doses of UVB. LUT attenuated UVB-induced cell death through delay and inhibition of intrinsic apoptotic signaling. Moreover, LUT not only predominantly affected the mitochondrial apoptosis pathway through its antioxidant capacity, but also changed the balance of Bcl2 (B-cell leukemia/lymphoma 2)-family members. Furthermore, LUT had inhibitory effects on the UVB-induced release of the inflammatory mediators, IL-1alpha and prostaglandin-E(2). Using different cell lines derived from squamous cell carcinomas, we showed that LUT did not increase the resistance of malignant keratinocytes to UVB. Our data suggest that LUT inhibits different aspects of the sunburn response, which results ultimately in an increased survival of normal keratinocytes, whereas the sensitivity of malignant cells to UVB remain unchanged. Hence, LUT might have value in new photoprotective applications or improve existing ones.