Division of Respiratory Diseases, Department of Internal Medicine, Jikei University School of Medicine, 3-25-8, Nishi-shimbashi, Minato-ku, Tokyo, 105-8461, Japan.
Am J Respir Cell Mol Biol. 2012 Mar;46(3):306-12. doi: 10.1165/rcmb.2011-0214OC. Epub 2011 Oct 6.
Cigarette smoke induces damage to proteins and organelles by oxidative stress, resulting in accelerated epithelial cell senescence in the lung, which is implicated in chronic obstructive pulmonary disease (COPD) pathogenesis. Although the detailed molecular mechanisms are not fully understood, cellular energy status is one of the most crucial determinants for cell senescence. Creatine kinase (CK) is a constitutive enzyme, playing regulatory roles in energy homeostasis of cells. Among two isozymes, brain-type CK (CKB) is the predominant CK in lung tissue. In this study, we investigated the role of CKB in cigarette smoke extract (CSE)-induced cellular senescence in human bronchial epithelial cells (HBECs). Primary HBECs and Beas2B cells were used. Protein carbonylation was evaluated as a marker of oxidative protein damage. Cellular senescence was evaluated by senescence-associated β-galactosidase staining. CKB inhibition was examined by small interfering RNA and cyclocreatine. Secretion of IL-8, a hallmark of senescence-associated secretary phenotype, was measured by ELISA. CKB expression levels were reduced in HBECs from patients with COPD compared with that of HBECs from nonsmokers. CSE induced carbonylation of CKB and subsequently decreased CKB protein levels, which was reversed by a proteasome inhibitor. CKB inhibition alone induced cell senescence, and further enhanced CSE-induced cell senescence and IL-8 secretion. CSE-induced oxidation of CKB is a trigger for proteasomal degradation. Concomitant loss of enzymatic activity regulating energy homeostasis may lead to the acceleration of bronchial epithelial cell senescence, which is implicated in the pathogenesis of COPD.
香烟烟雾通过氧化应激诱导蛋白质和细胞器损伤，导致肺部上皮细胞加速衰老，这与慢性阻塞性肺疾病（COPD）的发病机制有关。虽然详细的分子机制尚未完全了解，但细胞能量状态是细胞衰老的最重要决定因素之一。肌酸激酶（CK）是一种组成性酶，在细胞能量平衡中发挥调节作用。在两种同工酶中，脑型 CK（CKB）是肺组织中主要的 CK。在这项研究中，我们研究了 CKB 在香烟烟雾提取物（CSE）诱导的人支气管上皮细胞（HBEC）细胞衰老中的作用。使用了原代 HBEC 和 Beas2B 细胞。蛋白质羰基化被评估为氧化蛋白质损伤的标志物。通过衰老相关β-半乳糖苷酶染色评估细胞衰老。通过小干扰 RNA 和环磷酸肌酸检查 CKB 抑制作用。通过 ELISA 测量衰老相关分泌表型的标志性物质 IL-8 的分泌。与非吸烟者的 HBEC 相比，COPD 患者的 HBEC 中 CKB 的表达水平降低。CSE 诱导 CKB 的羰基化，随后降低 CKB 蛋白水平，这可以被蛋白酶体抑制剂逆转。CKB 抑制本身诱导细胞衰老，并且进一步增强 CSE 诱导的细胞衰老和 IL-8 分泌。CSE 诱导的 CKB 氧化是蛋白酶体降解的触发因素。同时丧失调节能量平衡的酶活性可能导致支气管上皮细胞衰老加速，这与 COPD 的发病机制有关。
