Extracellular signal-regulated kinase (ERK) inhibition attenuates cigarette smoke extract (CSE) induced-death inducing signaling complex (DISC) formation in human lung fibroblasts (MRC-5) cells.
Cigarette smoke (CS), a major risk factor in emphysema, causes cell death by incompletely understood mechanisms. Death-inducing signaling complex (DISC) formation is an initial event in Fas-mediated apoptosis. We demonstrated cigarette smoke extract (CSE) induced DISC formation in human lung fibroblasts (MRC-5). The aim of this study was to investigate the involvement of extracellular signal-regulated kinase (ERK) MAPK activation in CSE induced DISC formation. Immunoprecipitation (IP) for Fas and Western Immunoblot (IB) analysis for caspase 8 were then performed to show DISC. Lactate dehydrogenase (LDH) release was measured using a cytotoxicity detection kit. MTT assay was used as a measure of cell viability. We demonstrated that CSE induces DISC formation in MRC-5 using IP for Fas and IB for caspase 8. ERK was expressed in MRC-5 exposed to CSE. MEK-1 inhibitor (PD98059) decreased DISC formation in MRC-5 exposed to 20% CSE at 1 hr, and cell viability, as assessed by colorimetric MTT assay, was increased in MEK-1 inhibitor treated MRC-5 cells after 24 hr CSE exposure compared to the control. Inhibiting ERK significantly decreased the caspase-3,-8 activity in MEK-1 inhibitor treated MRC-5 cells compared to the control.The DISC formation, initial event of extrinsic apoptotic pathway, is a primary component of CSE- induced death in MRC-5, and ERK activation plays an active role in the DISC formation and downstream pathway. These results suggest that modulation of ERK may have therapeutic potential in the prevention of smoke-related lung injury.
Park
Jeong-Woong
JW
Department of Pulmonary and Critical Care Medicine, Gachon University, Korea. jwpark@gilhospital.com
Yoon
Jin Young
JY
Kim
Yu Jin
YJ
Kyung
Sun Young
SY
Lee
Sang Pyo
SP
Jeong
Sung Hwan
SH
Moon
Chanil
C
D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D005591:Chemical Fractionation; D053418:Death Domain Receptor Signaling Adaptor Proteins / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D006801:Humans; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008168:Lung; D012906:Smoke / Q000032:analysis*; D014026:Nicotiana / Q000737:chemistry / Q000633:toxicity*
D053418:Death Domain Receptor Signaling Adaptor Proteins; D004791:Enzyme Inhibitors; D012906:Smoke; D007770:L-Lactate Dehydrogenase; D048049:Extracellular Signal-Regulated MAP Kinases; D020169:Caspases