Radiation Inducing Pulmonary Cellular Senescence through the LPA-LPA1-Akt Pathway

Abstract

Delivering ionizing radiation to thoracic region in NSCLC patients faces a major limitation known as the radiation-induced lung injuries (RILIs). During this process, DNA damage to exposed cells could lead to cellular senescence. Accumulating evidence suggests that cellular senescence may play an important role in the pathogenesis of inflammatory and fibrotic lung diseases. Our group has previously established that, through the lysophosphatidic acid (LPA)-LPA receptor 1 (LPA1) signaling pathway, high levels of LPA in irradiated lungs play an important role in the development of RILIs. This study is designed to discover whether selectively blocking the LPA-LPA1 signaling in the early phase can effectively ameliorate the cellular senescence caused by IR treatment. Firstly, SPF C57BL/6J mice were exposed to X-rays (16 Gy/f) on the whole thorax. Irradiated mouse lungs were analyzed at different points of time to understand the development of cellular senescence. Then AM966 (30 mg/kg, qod, intraperitoneal injection), a selective antagonist to LPA1, was prescribed to irradiated mice from the 1st day to the 7th day and from the 1st day to the 30th day after irradiation. Finally, to explore the relations between cellular senescence and LPA- LPA1 signaling, HBE and HUVEC cells were exposed to the combinatory treatment of irradiation (16 Gy/f) and LPA (10 μM) to simulate the conditions in vivo. The effects of AM966 (40 μΜ), Akt inhibitor MK 2006 (5 μM), and Akt stimulator SC79 (5 μM) on cells were studied 48 hours after irradiation. The irradiated lung tissue had been substantially expressing senescence-associated β-galactosidase (SA-β-gal) and p21 from the 14th day after irradiation, both of which culminated on the 60th day with extensive pulmonary fibrosis. AM966 prescription effectively reduced SA-β-gal activity on the 7th and 30th day as well as the levels of senescence-associated secretory phenotype (SASP) factors and p21. The expression of Ki-67 was well-maintained in the AM966 group. At the 48-hour time point after irradiation, more than 80 percent of the HBE and HUVEC cells were arrested in the G2/M phase, and exhibited high activity of SA-β-gal, significantly increased γH2AX foci and ROS production. All above were ameliorated by AM966 or MK2206 pretreatment, while SC79 could counteract the effect of AM966. LPA in high concentrations participates in the radiation-induced pulmonary cellular senescence through the LPA-LPA1-Akt signaling. Selectively blocking LPA1 by AM966 in the early stage could significantly ameliorate pulmonary cellular senescence and injuries.