Section of Functional Proteomics, Department of Life Sciences, University of Siena, Via A. Moro 2, 53100 Siena, Italy.
Department of Pharmacological and Biomolecular Sciences "Rodolfo Paoletti", Università degli Studi di Milano, Via Balzaretti 9, 20133 Milan, Italy.
Int J Mol Sci. 2023 Mar 29;24(7):6431. doi: 10.3390/ijms24076431.
Cigarette smoke (CS) is a risk factor for inflammatory diseases, such as atherosclerosis. CS condensate (CSC) contains lipophilic components that may represent a systemic cardiac risk factor. To better understand CSC effects, we incubated mouse and human aortic smooth muscle cells (SMCs) with CSC. We evaluated specific markers for contractile [i.e., actin, aortic smooth muscle (), calponin-1 (), the Kruppel-like factor 4 (), and myocardin () genes] and inflammatory [i.e., IL-1β, and , , and galectin-3 () genes] phenotypes. CSC increased the expression of inflammatory markers and reduced the contractile ones in both cell types, with modulating the SMC phenotypic switch. Next, we performed a mass spectrometry-based differential proteomic approach on human SMCs and could show 11 proteins were significantly affected by exposition to CSC (FC ≥ 2.7, ≤ 0.05). These proteins are active in signaling pathways related to expression of pro-inflammatory cytokines and IFN, inflammasome assembly and activation, cytoskeleton regulation and SMC contraction, mitochondrial integrity and cellular response to oxidative stress, proteostasis control via ubiquitination, and cell proliferation and epithelial-to-mesenchymal transition. Through specific bioinformatics resources, we showed their tight functional correlation in a close interaction niche mainly orchestrated by the interferon-induced double-stranded RNA-activated protein kinase (alternative name: protein kinase RNA-activated; PKR) (EIF2AK2/PKR). Finally, by combining gene expression and protein abundance data we obtained a hybrid network showing reciprocal integration of the CSC-deregulated factors and indicating KLF4 and PKR as the most relevant factors.
香烟烟雾（CS）是炎症性疾病（如动脉粥样硬化）的一个风险因素。CS 冷凝物（CSC）含有亲脂性成分，可能代表一个系统性心脏危险因素。为了更好地了解 CSC 的影响，我们用 CSC 孵育了小鼠和人主动脉平滑肌细胞（SMCs）。我们评估了收缩性（如肌动蛋白、主动脉平滑肌（）、钙调蛋白-1（）、Kruppel 样因子 4（）和肌球蛋白（）基因）和炎症性（如 IL-1β 和 TNF-α、IL-6、MCP-1 和 galectin-3（）基因）表型的特定标志物。CSC 增加了两种细胞类型中炎症标志物的表达，并降低了收缩性标志物的表达，其中 调节 SMC 表型转换。接下来，我们对人 SMC 进行了基于质谱的差异蛋白质组学方法，结果表明有 11 种蛋白质的表达受到 CSC 暴露的显著影响（FC≥2.7， ≤ 0.05）。这些蛋白质在与促炎细胞因子和 IFN 表达、炎症小体组装和激活、细胞骨架调节和 SMC 收缩、线粒体完整性和细胞对氧化应激的反应、通过泛素化进行蛋白质稳态控制以及细胞增殖和上皮-间充质转化相关的信号通路中具有活性。通过特定的生物信息学资源，我们显示了它们在一个紧密的相互作用生态位中的紧密功能相关性，主要由干扰素诱导的双链 RNA 激活蛋白激酶（另一个名称：蛋白激酶 RNA 激活酶；PKR）（EIF2AK2/PKR）协调。最后，通过结合基因表达和蛋白质丰度数据，我们获得了一个混合网络，显示了 CSC 失调因子的相互整合，并表明 KLF4 和 PKR 是最相关的因子。
