Exploring senescence-driven miscommunication in the lung microenvironment

Lung cells, being the interface with the external environment, rely on stress response pathways throughout the lifetime. Among these responses, senescence might have detrimental effects, as the inhibition of cell cycle hampers tissue regeneration and the Senescent Associated Secretory Phenotype (SASP) promotes inflammation and parenchymal destruction. As a paradigm of senescence-driven lung impairment, we developed an in vitro model of Lymphangioleiomyomatosis (LAM), a pulmonary low-grade, destructive, metastasizing rare neoplasm. Distinctive of LAM cells are the metastatic capability, the dysregulated mechanistic Target Of Rapamycin (mTOR) and the active secretion of factors that remodel the lung parenchyma to establish a LAM-favouring microenvironment without forming of solid masses. While there is no cure for LAM, the only approved pharmacological treatment is rapamycin, which selectively inhibits the mTOR Complex 1 (mTORC1). In senescence, mTORC1 induces SASP, through which cells reinforce their senescent phenotype and promote senescence in neighbouring cells. We demonstrated that primary LAM/TSC cells are senescent depending on mTOR hyperactivation and induce senescence in non-LAM pulmonary lung fibroblasts (PLFs) through their conditioned medium (CM). This sustains the hypothesis that LAM cells have a SASP capable to modify LAM microenvironment. In this context, we demonstrated the role of Interleukin-8 (IL-8), highly secreted by LAM/TSC cells, in sustaining LAM/TSC cells senescence and in inducing senescence on PLFs. In our model, the inhibition of one IL-8 receptor, CXCR2, by the small molecule SB225002 impairs autocrine senescence in LAM/TSC cells. Since this molecule only partially counteracts IL-8-induced senescence in PLFs, we investigated the role of Extracellular Vesicles (EVs) among SASP factors, demonstrating a significantly higher release of EVs by LAM/TSC cells than PLFs. Notably, the positivity to the senescence markers β-Galactosidase, p16 and p21 is increased in PLFs cultured with LAM/TSC EVs, indicating that they contribute to senescence spreading. Finally, we demonstrated that IL-8 is specifically present in LAM/TSC EVs but not in PLFs EVs and we observed that mTORC1 inhibition reduces EV release by LAM/TSC cells. Thus, our results indicate that modulating senescence through molecules targeting SASP may be a promising strategy to interfere with the pathological communication in lung microenvironment, with potential therapeutic applications