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Analysis of IL17RD effector molecules by mass spectrometry

ABSTRACT: Quantitative proteomics analysis of primary murine embryonic fibroblasts (MEFs) from Il17rd+/+ and Il17rd-/- mice stimulated with or without IL17A to identify novel targets of the IL17A/IL17RD signaling axis.

INSTRUMENT(S): TripleTOF 5600

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Embryo, Fibroblast, Embryonic Stem Cell

SUBMITTER: Carlos Gartner

LAB HEAD: Calvin Vary

PROVIDER: PXD048814 | Pride | 2024-02-09

REPOSITORIES: Pride

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			Action	DRS
	100VW25msSWATH-IL17andSEF-S1-2.wiff	Wiff
	100VW25msSWATH-IL17andSEF-S1-2.wiff.scan	Wiff
	100VW25msSWATH-IL17andSEF-S1-3.wiff	Wiff
	100VW25msSWATH-IL17andSEF-S1-3.wiff.scan	Wiff
	100VW25msSWATH-IL17andSEF-S1-4.wiff	Wiff

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Project description:The unique properties of the bone marrow allow for migration and proliferation of multipl myeloma (MM) cells, while also providing the perfect environment for development of quiescent, drug-resistant MM cell clones. Bone marrow adipocytes (BMAds), which have recently been identified as important contributors to systemic adipokine levels, bone strength, hematopoiesis, and progression of metastatic and primary bone marrow cancers, such as MM. Recent studies in myeloma suggest that BMAds can be reprogrammed by tumor cells to contribute to myeloma-induced bone disease, and reciprocally, BMAds support MM cells in vitro. Importantly, most data investigating BMAds have been generated using adipocytes derived by differentiating bone marrow-derived mesenchymal stromal cells (MSCs) into adipocytes in vitro using adipogenic media, due to the extreme technical challenges associated with isolating and culturing primary adipocytes. However, if studies could be performed with primary adipocytes, they likely will recapitulate in vivo biology better than MSC-derived adipocyte, as the differentiation process is artificial and differs from in vivo differentiation, and progenitor cell(s) of the primary BMAd may not be the same as the MSCs precursors used for adipogenic differentiation in vitro. Therefore, we developed and refined three methods for culturing primary BMAds (pBMAds): 2D coverslips, 2D transwells, and 3D silk scaffolds, all of which can be cultured alone or with MM cells to investigate bidirectional tumor-host signaling. To develop an in vitro model with a tissue-like structure to mimic the bone marrow microenvironment, we developed the first 3D, tissue engineered model utilizing pBMAds derived from human bone marrow. We found that pBMAds, which are extremely fragile, can be isolated and stably cultured in 2D for 10 days and in 3D for short term (~2 weeks) or long term (1 month) in vitro. To investigate the relationship between pBMAds and myeloma, MM cells can be added to investigate physical relationships through confocal imaging and soluble signaling molecules via mass spectrometry. In sum, we developed three in vitro cell culture systems to study primary bone marrow adipocytes and myeloma cells, which could be adapted to investigate many diseases and biological processes involving the bone marrow, including other bone-homing tumor types.

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Analysis of IL17RD effector molecules by mass spectrometry

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