Development of a co-culture of keratinocytes and immune cells for in vitro investigation of cutaneous sulfur mustard toxicity
Development of a co-culture of keratinocytes and immune cells for in vitro investigation of cutaneous sulfur mustard toxicity
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DOI：
10.1016/j.cbi.2014.09.002
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发表时间：
2014-11-05
影响因子：
5.1
通讯作者：
Steinritz, Dirk
中科院分区：
文献类型：
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作者：
Balszuweit, Frank;Menacher, Georg;Steinritz, Dirk
芥子气（SM）是一种化学战剂，可导致皮肤起泡、溃疡和伤口愈合延迟。已知炎症和外源性凋亡在SM诱导的细胞毒性中具有重要作用。由于免疫细胞参与了这些过程，它们可能会显着调节SM毒性，但这些影响的程度尚不清楚。我们采用了永生化角质形成细胞（HaCaT）和免疫细胞（THP-1）的共培养模型，并将该模型暴露于SM。研究了坏死、凋亡和炎症的变化，这取决于SM攻击、THP-1细胞的存在或不存在以及数量。在SM暴露之前将THP-1共培养24小时，以模拟SM对皮肤中连续存在的免疫细胞的影响。我们的研究结果表明，THP-1的存在强烈增加坏死，凋亡和炎症。当THP-1和HaCaT细胞的比例与体内朗格汉斯免疫细胞和角质形成细胞的比例相似时，这种效果已经很显著。THP-1数量的任何进一步增加对SM诱导的细胞毒性仅有轻微的额外影响。为了评估免疫细胞迁移到SM损伤的皮肤区域的影响，我们将未暴露的THP-1添加到SM暴露的HaCaT中。这些THP-1对SM诱导的细胞毒作用只有轻微的影响。值得注意的是，在暴露于300 μ M SM的HaCaT中，通过添加完整的THP-1，坏死和炎症轻微减少。这种效应取决于免疫细胞的数量，随着未暴露的THP-1的加入而稳步增加。总之，我们已经证明了（a）所提出的共培养物是评估SM毒性的稳健模型，并且可以用于在体外测试潜在解毒剂的功效;（B）被SM强烈放大的细胞毒性损伤的免疫细胞，（c）相比之下，未暴露的THP-1（模拟免疫细胞在体内暴露后迁移到受影响区域）没有明显的不利影响，但是表现出一些保护作用。因此，保护免疫细胞免受SM毒性可能有助于减少整体损伤。(C)2014爱思唯尔爱尔兰有限公司版权所有。
Sulfur mustard (SM) is a chemical warfare agent causing skin blistering, ulceration and delayed wound healing. Inflammation and extrinsic apoptosis are known to have an important role in SM-induced cytotoxicity. As immune cells are involved in those processes, they may significantly modulate SM toxicity, but the extent of those effects is unknown. We adapted a co-culture model of immortalized keratinocytes (HaCaT) and immune cells (THP-1) and exposed this model to SM. Changes in necrosis, apoptosis and inflammation, depending on SM challenge, absence or presence and number of THP-1 cells were investigated. THP-1 were co-cultured for 24 h prior to SM exposure in order to model SM effects on immune cells continuously present in the skin. Our results indicate that the presence of THP-1 strongly increased necrosis, apoptosis and inflammation. This effect was already significant when the ratio of THP-1 and HaCaT cells was similar to the ratio of Langerhans immune cells and keratinocytes in vivo. Any further increases in the number of THP-1 had only slight additional effects on SM-induced cytotoxicity. In order to assess the effects of immune cells migrating into skin areas damaged by SM, we added non-exposed THP-1 to SM-exposed HaCaT. Those THP-1 had only slight effects on SM-induced cytotoxicity. Notably, in HaCaT exposed to 300 mu M SM, necrosis and inflammation were slightly reduced by adding intact THP-1. This effect was dependent on the number of immune cells, steadily increasing with the number of unexposed THP-1 added. In summary, we have demonstrated that (a) the presented co-culture is a robust model to assess SM toxicity and can be used to test the efficacy of potential antidotes in vitro; (b) immune cells, damaged by SM strongly amplified cytotoxicity, (c) in contrast, unexposed THP-1 (simulating migration of immune cells into affected areas after exposure in vivo) had no pronounced adverse, but exhibited some protective effects. Thus, protecting immune cells from SM toxicity may help to reduce overall injury. (C) 2014 Elsevier Ireland Ltd. All rights reserved.