Project description:While aging is well established as an important risk factor for the development and progression of atherosclerosis, the underlying molecular mechanisms of this relationship remain poorly defined, and its role in atherosclerosis regression is unknown. We investigated the response of bone marrow-derived macrophages isolated from old and young mice to atherogenic aggregated LDL.
Project description:Bone marrow macrophages were isolated from CD40-TRAF2/3/5 -/- and CD40-TRAF6 -/- mice, followed by induction of differentiation and maturation. Apoptotic cardiomyocytes were then added to trigger efferocytosis for 3 hours. Subsequently, total proteins were extracted for proteomic sequencing.
Project description:Bone marrow cells (BM) were isolated and primed with M-CSF (M BMDM) or GM-CSF (GM BMDM) for 7 days. M-BMDMs were treated with IL6 (20 ng/ml) for 3 h or 24 h while GM-BMDMs were treated for 3 h. The difference between BM, M-BMDM, and GM-BMDM were analyzed to describe the phenotye and function of each population. Moreover, by RNA-seq analysis, the influence of IL6 treatment on GM-BMDMs and M-BMDMs were analyzed.
Project description:LRRK2 is commonly mutated in Parkinson’s disease and has cell type-specific mechanisms of activation and function. In macrophages, LRRK2 is associated with lysosomes and is activated following lysosomal damage. However, the functional outcomes of clinically relevant pathogenic mutations in macrophages are unknown. Here, using primary mouse and patient-derived iPSC-derived macrophage (iPSDM) models of LRRK2-G2019S, we defined the substrates of LRRK2 after lysosomal damage. Using phosphoproteomics we found that LRRK2-G2019S and wild-type macrophages showed similar levels of Rab GTPases phosphorylation after lysosomal damage. However, the levels of pRab12 increased whereas the levels of pRab35 decreased in LRRK2-G2019S macrophages. Functionally, LRRK2-G2019S macrophages showed a deficit in lysosomal membrane repair that resulted in more cell death and increased apoptosis. Importantly, we recapitulated this phenotype in iPSDM from patients carrying the G2019S mutation, but not in isogenic control iPSDM. Altogether, we define here the signaling downstream of G2019S in macrophages and identify susceptibility to cell death after lysosomal damage as an important phenotype of this mutation.