Precision Cut Lung Slices (PCLS) as ex vivo Human Lung Tissue Model of Cellular Senescence Assessed by Multi-Omic Modalities v1

This protocol outlines the experimental workflow for evaluating the efficacy of senescence inducers and senotherapeutic / senolytic compounds in modulating the senescence-associated secretory phenotype (SASP) using precision-cut lung slices (PCLS) as an ex vivo model system. PCLS retain the native tissue architecture and cell-type complexity of the lung, making them a relevant multi-cellular platform to study spatial and functional responses of lung tissue to pharmacological agents. Version 1 of the protocol outlines: Ex vivo induction of senescence by irradiation of PCLS using defined doses (e.g., 5–10 Gy) to model DNA damage-induced senescence and inflammatory activation. Application of senolytic drug treatment (e.g., Dasatinib, Quercetin and Fisetin, or other candidate compounds) administered after recovery from irradiation. Collection of PCLS supernatants for secretome profiling via Luminex multiplex cytokine immunoassays, focused on senescence-associated secretory phenotype (SASP) factors. Recovery and preservation of PCLS tissue following senescence induction / inhibition for 1) RNA extraction and NanoString nCounter transcriptomic profiling, targeting genes involved in senescence, DNA damage response, and immune modulation, 2) collection of supernatants for secretome profiling via Luminex multiplex cytokine immunoassays, and 3) fixation, embedding and sectioning for histochemistry, mutiplexed immunofluorescence (MxIF) and/or spatial transcriptomics.