Möller Winfried, Brown David M, Kreyling Wolfgang G, Stone Vicki
GSF National Research Center for Environment and Health, Robert Koch Allee 29, D-82131 Munich-Gauting, Germany.
Part Fibre Toxicol. 2005 Oct 4;2:7. doi: 10.1186/1743-8977-2-7.
Particulate air pollution is reported to cause adverse health effects in susceptible individuals. Since most of these particles are derived form combustion processes, the primary composition product is carbon with a very small diameter (ultrafine, less than 100 nm in diameter). Besides the induction of reactive oxygen species and inflammation, ultrafine particles (UFP) can cause intracellular calcium transients and suppression of defense mechanisms of alveolar macrophages, such as impaired migration or phagocytosis.
In this study the role of intracellular calcium transients caused by UFP was studied on cytoskeleton related functions in J774A.1 macrophages. Different types of fine and ultrafine carbon black particles (CB and ufCB, respectively), such as elemental carbon (EC90), commercial carbon (Printex 90), diesel particulate matter (DEP) and urban dust (UD), were investigated. Phagosome transport mechanisms and mechanical cytoskeletal integrity were studied by cytomagnetometry and cell viability was studied by fluorescence microscopy. Macrophages were exposed in vitro with 100 and 320 microg UFP/ml/million cells for 4 hours in serum free medium. Calcium antagonists Verapamil, BAPTA-AM and W-7 were used to block calcium channels in the membrane, to chelate intracellular calcium or to inhibit the calmodulin signaling pathways, respectively.
Impaired phagosome transport and increased cytoskeletal stiffness occurred at EC90 and P90 concentrations of 100 microg/ml/million cells and above, but not with DEP or UD. Verapamil and W-7, but not BAPTA-AM inhibited the cytoskeletal dysfunctions caused by EC90 or P90. Additionally the presence of 5% serum or 1% bovine serum albumin (BSA) suppressed the cytoskeletal dysfunctions. Cell viability showed similar results, where co-culture of ufCB together with Verapamil, W-7, FCS or BSA produced less cell dead compared to the particles only.
据报道，颗粒物空气污染会对易感个体造成不良健康影响。由于这些颗粒大多源自燃烧过程，其主要组成产物是直径非常小的碳（超细颗粒，直径小于100纳米）。除了诱导活性氧生成和炎症反应外，超细颗粒（UFP）还可引起细胞内钙瞬变，并抑制肺泡巨噬细胞的防御机制，如迁移或吞噬功能受损。
在本研究中，研究了UFP引起的细胞内钙瞬变对J774A.1巨噬细胞中细胞骨架相关功能的作用。研究了不同类型的细颗粒和超细颗粒炭黑（分别为CB和ufCB），如元素碳（EC90）、商用碳（Printex 90）、柴油颗粒物（DEP）和城市灰尘（UD）。通过细胞磁强计研究吞噬体运输机制和机械细胞骨架完整性，通过荧光显微镜研究细胞活力。巨噬细胞在无血清培养基中以100和320微克UFP/毫升/百万细胞的浓度体外暴露4小时。分别使用钙拮抗剂维拉帕米、BAPTA-AM和W-7阻断细胞膜上的钙通道、螯合细胞内钙或抑制钙调蛋白信号通路。
在EC90和P90浓度为100微克/毫升/百万细胞及以上时，吞噬体运输受损和细胞骨架硬度增加，但DEP或UD不会出现这种情况。维拉帕米和W-7可抑制EC90或P90引起的细胞骨架功能障碍，而BAPTA-AM则不能。此外，5%血清或1%牛血清白蛋白（BSA）的存在可抑制细胞骨架功能障碍。细胞活力也有类似结果，与仅使用颗粒相比，ufCB与维拉帕米、W-7、胎牛血清（FCS）或BSA共同培养时细胞死亡较少。
