Downregulation of lysyl oxidase and upregulation of cellular thiols in rat fetal lung fibroblasts treated with cigarette smoke condensate
Downregulation of lysyl oxidase and upregulation of cellular thiols in rat fetal lung fibroblasts treated with cigarette smoke condensate
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DOI：
10.1093/toxsci/kfi019
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发表时间：
2005-02-01
影响因子：
3.8
通讯作者：
Li, WD
中科院分区：
文献类型：
--
作者：
Chen, LJ;Zhao, YZ;Li, WD
赖氨酰氧化酶（Lysyl oxidase，LO）是一种铜依赖性酶，通过催化弹性蛋白和胶原的交联，在细胞外基质（extracellular matrix，ECM）的形成和修复中起着重要作用。为了更好地理解香烟烟雾（CS）诱导肺气肿的机制，我们检查了LO及其底物的变化，即，弹性蛋白和1型胶原，细胞硫醇的主要成分，即，金属硫蛋白（MT）和谷胱甘肽（GSH），以及γ-谷氨酰半胱氨酸合成酶（γ-GCS），GSH生物合成的关键酶，在香烟烟雾冷凝物（CSC）处理的大鼠胎肺成纤维细胞（RFL 6）。RFL 6细胞暴露于CSC降低了LO催化活性、mRNA和蛋白质的水平，即，46 kDa前酶原、50 kDa酶原和32 kDa成熟酶以剂量依赖性方式。此外，CSC还抑制I型胶原和弹性蛋白的表达，LO的底物和肺ECM的重要成分。同时，细胞巯基包括MT和GSH以及γ-GCS在CSC处理的细胞中显著上调。为了评估细胞巯基对LO表达的调节，我们进一步研究了GSH水平增加对蛋白质和催化水平LO表达的影响。有趣的是，细胞暴露于谷胱甘肽单乙酯，GSH的传递系统，有效地提高细胞的GSH水平，并诱导剂量依赖性降低的蛋白质种类和LO的催化活性的水平。这些结果表明，上调CSC的细胞硫醇可能发挥重要作用，下调LO和随后不稳定的肺ECM在CS诱导的肺气肿。
Lysyl oxidase (LO), a copper-dependent enzyme, plays a critical role in the formation and repair of the extracellular matrix (ECM) by catalyzing the crosslinking of elastin and collagen. To better understand mechanisms of cigarette smoke (CS)-induced emphysema, we examined changes in LO and its substrates, i.e., elastin and collagen type 1, the major components of cellular thiols, i.e., metallothionein (MT) and glutathione (GSH), and gamma-glutamylcysteine synthetase (,y-GCS), a key enzyme for GSH biosynthesis, in cigarette smoke condensate (CSC)-treated rat fetal lung fibroblasts (RFL6). Exposure of RFL6 cells to CSC decreased levels of LO catalytic activity, mRNA, and protein, i.e., the 46 kDa preproenzyme, the 50 kDa proenzyme and the 32 kDa mature enzyme in a dose-dependent manner. In addition, CSC also inhibited the expression of collagen type I and elastin, substrates of LO and important components of the lung ECM. Meanwhile, cellular thiols including MT and GSH as well as gamma-GCS were markedly upregulated in CSC-treated cells. To evaluate modulation of LO expression by cellular thiols, we further examined the effect of increased levels of GSH on LO expression at protein and catalytic levels. Interestingly, exposure of cells to glutathione monoethyl ester, a GSH delivery system, effectively elevated cellular GSH levels and induced a dose-dependent decrease in levels of the protein species and catalytic activity of LO. These results suggest that upregulation by CSC of cellular thiols may play an important role in the downregulation of LO and subsequently destabilization of the lung ECM in CS-induced emphysema.