Involvement of mitochondrial dysfunction during transforming growth factor β1-induced senescence-like growth arrest of MvlLu cells

PURPOSE: Normal cells enter a nondividing state after a finite number of population doubling, which is termed replicative senescence. Although the free radical theory has long been implicated in the senescence, recent studies have emphasized mitochondrial DNA hypothesis because mitochondria is the major ROS generator in addition to its high vulnerability to oxidative damage. Transforming growth factor β1 (TGF β1) has well been known to suppress epithelial cell growth and recently been described to be involved in cellular senescence. The purpose of this study is to investigate how TGF β1 could induce senescence in Mv1Lu mink lung epithelial cells, whether and how mitochondrial defects are involved in the senescence. MATERIALS AND METHODS: Mink lung epithelial Mv1Lu cells were treated with TGF β1. Cell cycle, ROS generation, and mitochondrial membrane potential were analyzed by flow cytometric analysis. The activities of mitochondrial respiratory complex were analyzed by spectrophotometric analysis and cellular respiration. Expression pattern of proteins were analyzed by immunocytochemistry and immunoblotting. Mitochondrial morphology was examined by electron microscope. RESULTS: TGF β1 arrested mink lung epithelial Mv1Lu cells at G1 phase of the cell cycle with acquisition of senescent phenotypes in the presence of 10 % serum, whereas it gradually induced apoptosis with lower concentrations of serum. The senescent arrest was accompanied by prolonged generation of reactive oxygen species (ROS) and persistent disruption of mitochondrial membrane potential (ΔΨm). Therefore, we investigated the relationship between mitochondrial defect and the ROS production. Pretreatment of Mv1Lu cells with antioxidants effectively removed the generated ROS without preventing the ΔΨm disruption, and complete inhibition of respiratory chain activities with KCN or oligomycin totally blocked the ROS generation. In addition, the mitochondrial respiration rate gradually diminished upon treatment with TGF β1 and the decrease was accompanied by reduced complex IV activity. These data suggested that the mitochondrial respiratory defect via decreased complex IV activity was the cause of the prolonged ROS generation. The respiratory defect was also followed by mitochondrial oxidative stress, evidenced by oxidation of Prx III, and morphological change of mitochondria to highly elongated form. Finally, we demonstrated that hepatocyte growth factor released Mv1Lu cells from the arrest by protecting decrease of mitochondrial respiration, thereby preventing both the ΔΨm disruption and the ROS generation. CONCLUSIONS: Our results describe that TGF β1 induces senescence-like growth arrest in Mv1Lu cells in the presence of serum (higher than 5 %). During the arrest, ROS is persistently produced from the defective respiration, which is caused by decrease of mitochondrial complex IV activity. Moreover, HGF releases the arrest by preventing mitochondrial dysfunction. These results demonstrate that the mitochondrial defects are involved in the senescence induced by TGF β1, a single cytokine, supporting the importance of ‘mitochondrial theory of aging’ and propose a potential intimate coupling between TGF β1-induced signaling and modulation of mitochondrial metabolism.

Author dc:creator

• 윤, 영실

Contributors dc:contributor

• 윤, 계순
• 대학원 의학과
• 200224342

• Transforming Growth Factor beta1 (TGF β1)
• senescence-like growth arrest
• mitochondrial dysfunction
• Reactive Oxygen Species (ROS)
• cellular respiration
• Hepatocyte Growth Factor (HGF)

Language dc:language

en

Identifier

http://dcoll.ajou.ac.kr:9080/dcollection/jsp/common/DcLoOrgPer.jsp?sItemId=000000000229
000000000229

OAI identifier oai:identifier

oai:repository.ajou.ac.kr:201003/2293