Son E S, Kyung S Y, Lee S P, Jeong S H, Shin J Y, Ohba M, Yeo E J, Park J W
Department of Pulmonary, Allergy and Critical Care Medicine, Gachon University, Gil Medical Center, Namdong-Gu, Incheon, Republic of Korea.
Institute of Molecular Oncology, Showa University, Hatanodai, Shinagawa-ku, Tokyo, Japan.
Hum Exp Toxicol. 2015 Sep;34(9):869-77. doi: 10.1177/0960327114561343. Epub 2014 Dec 9.
Cigarette smoke (CS) is a major risk factor for emphysema, which causes cell death in structural cells of the lung by mechanisms that are still not completely understood. We demonstrated previously that CS extract (CSE) induces caspase activation in MRC-5 human lung fibroblasts, activated protein kinase C-η (PKC-η), and translocated PKC-η from the cytosol to the membrane. The objective of this study was to investigate the involvement of PKC-η activation in a CSE-induced extrinsic apoptotic pathway. We determined that CSE increases expression of caspase 3 and 8 cleavage in MRC-5 cells and overexpression of PKC-η significantly increased expression of caspase 3 and 8 cleavage compared with control LacZ-infected cells. In contrast, dominant negative (dn) PKC-η inhibited apoptosis in MRC-5 cells exposed to CSE and decreased expression of caspase 3 and 8 compared with control cells. Exposure to 10% CSE for >8 h significantly increased lactate dehydrogenase release in PKC-η-infected cells compared with LacZ-infected cells. Additionally, PKC-η-infected cells had an increased number of Hoechst 33342 stained nuclei compared with LacZ-infected cells, while dn PKC-η-infected cells exhibited fewer morphological changes than LacZ-infected cells under phase-contrast microscopy. In conclusion, PKC-η activation plays a pro-apoptotic role in CSE-induced extrinsic apoptotic pathway in MRC-5 cells. These results suggest that modulation of PKC-η may be a useful tool for regulating the extrinsic apoptosis of MRC-5 cells by CSE and may have therapeutic potential in the treatment of CS-induced lung injury.
香烟烟雾（CS）是肺气肿的主要危险因素，其通过尚未完全明确的机制导致肺结构细胞死亡。我们先前已证明，香烟烟雾提取物（CSE）可诱导MRC-5人肺成纤维细胞中的半胱天冬酶激活、激活蛋白激酶C-η（PKC-η），并使PKC-η从胞质溶胶转位至细胞膜。本研究的目的是探讨PKC-η激活在CSE诱导的外源性凋亡途径中的作用。我们确定，CSE可增加MRC-5细胞中半胱天冬酶3和8的裂解表达，与对照LacZ感染细胞相比，PKC-η的过表达显著增加了半胱天冬酶3和8的裂解表达。相反，显性负性（dn）PKC-η抑制暴露于CSE的MRC-5细胞中的凋亡，并与对照细胞相比降低了半胱天冬酶3和8的表达。与LacZ感染细胞相比，暴露于10% CSE超过8小时显著增加了PKC-η感染细胞中乳酸脱氢酶的释放。此外，与LacZ感染细胞相比，PKC-η感染细胞中经Hoechst 33342染色的细胞核数量增加，而在相差显微镜下，dn PKC-η感染细胞比LacZ感染细胞表现出更少的形态学变化。总之，PKC-η激活在CSE诱导的MRC-5细胞外源性凋亡途径中起促凋亡作用。这些结果表明，调节PKC-η可能是调节CSE诱导的MRC-5细胞外源性凋亡的有用工具，并且可能在治疗CS诱导的肺损伤方面具有治疗潜力。
