School of Medical Sciences, RMIT University, PO Box 71, Bundoora, VIC, 3083, Australia,
Arch Dermatol Res. 2013 Nov;305(9):817-33. doi: 10.1007/s00403-013-1412-z. Epub 2013 Sep 26.
Ultraviolet (UV) radiation can activate the p38 mitogen-activated protein kinase (MAPK), Jun N-terminal kinase (JNK) and nuclear factor-κB (NFκB) pathways in skin cells. HaCaT cells are widely used as a primary keratinocyte substitute to study these pathways. However, like most squamous cell carcinomas (SCCs), it contains a dysfunctional p53. It is unclear if HaCaT cells activate these signalling pathways similarly to SCC cells (Colo16) or to primary human epidermal keratinocytes (HEK). In this study, the UV activation (UVA, UVB, UVA+B, UVB+A) of p38 MAPK, JNK and NFκB pathways, and TNFα secretion by HEK, HaCaT and Colo16 cells were investigated. The signalling pathway activation was UV-type and dose-dependent with UVB+A radiation inducing a high p38 and JNK activation. HaCaT cells exhibited 2- to 4-fold higher activity of the p38 (771% at 60 min) and JNK (794% at 30 min) pathways following UVB+A radiation than did HEK cells (p38: 367% at 15 min and JNK: 184% at 30 min). While both HaCaT and Colo16 cells did not activate the NFκB pathway, Colo16 cells had a lower p38 and higher JNK activity than HaCaT cells. Irradiated HaCaT cells produced less TNFα (UVB: 3.5 pg/ml), while HEK cells produced the most (UVB: 1,296 pg/ml). When co-exposed to IL1α, irradiated HaCaT had the greatest fold of TNFα release (UVB: 16.2-fold, UVA+B: 8.9-fold and UVB+A: 6.1-fold). The pattern of activation and TNFα secretion of HaCaT cells mirrored that of Colo16 cells. It is likely that the presence of molecular alterations in HaCaT cells may be responsible for its different responses to that seen for HEK cells. The results of this study suggest caution in using HaCaT cells as a substitute for normal keratinocytes in investigating UV-induced cells signalling pathways.
紫外线（UV）辐射可以激活皮肤细胞中的 p38 丝裂原活化蛋白激酶（MAPK）、Jun N 末端激酶（JNK）和核因子-κB（NFκB）途径。HaCaT 细胞被广泛用作研究这些途径的主要角质形成细胞替代物。然而，与大多数鳞状细胞癌（SCC）一样，它含有功能失调的 p53。目前尚不清楚 HaCaT 细胞是否与 SCC 细胞（Colo16）或原代人表皮角质形成细胞（HEK）相似地激活这些信号通路。在这项研究中，研究了 HEK、HaCaT 和 Colo16 细胞中 p38 MAPK、JNK 和 NFκB 途径的 UV 激活（UVA、UVB、UVA+B、UVB+A）以及 TNFα 的分泌。信号通路的激活是 UV 类型和剂量依赖性的，其中 UVB+A 辐射诱导 p38 和 JNK 的高激活。与 HEK 细胞（p38：15 分钟时为 367%，JNK：30 分钟时为 184%）相比，HaCaT 细胞在接受 UVB+A 辐射后，p38（60 分钟时为 771%）和 JNK（30 分钟时为 794%）途径的活性高 2-4 倍。虽然 HaCaT 和 Colo16 细胞均未激活 NFκB 途径，但 Colo16 细胞的 p38 活性较低，JNK 活性较高。辐照的 HaCaT 细胞产生的 TNFα 较少（UVB：3.5 pg/ml），而 HEK 细胞产生的 TNFα 最多（UVB：1,296 pg/ml）。当共同暴露于 IL1α 时，辐照的 HaCaT 细胞具有最大的 TNFα 释放倍数（UVB：16.2 倍，UVA+B：8.9 倍，UVB+A：6.1 倍）。HaCaT 细胞的激活和 TNFα 分泌模式与 Colo16 细胞相似。HaCaT 细胞中存在分子改变可能是其对 HEK 细胞反应不同的原因。本研究结果表明，在研究 UV 诱导的细胞信号通路时，应谨慎使用 HaCaT 细胞作为正常角质形成细胞的替代物。
