Development of a 3D bone marrow adipose tissue model

Abstract

Over the past twenty years, evidence has accumulated that biochemically and spatially defined networks of extracellular matrix, cellular components, and interactions dictate cellular differentiation, proliferation, and function in a variety of tissue and diseases. Modeling in vivo systems in vitro has been undeniably necessary, but when simplified 2D conditions rather than 3D in vitro models are used, the reliability and usefulness of the data derived from these models decreases. Thus, there is a pressing need to develop and validate reliable in vitro models to reproduce specific tissue-like structures and mimic functions and responses of cells in a more realistic manner for both drug screening/disease modeling and tissue regeneration applications. In adipose biology and cancer research, these models serve as physiologically relevant 3D platforms to bridge the divide between 2D cultures and in vivo models, bringing about more reliable and translationally useful data to accelerate benchtop to bedside research. Currently, no model has been developed for bone marrow adipose tissue (BMAT), a novel adipose depot that has previously been overlooked as "filler tissue" but has more recently been recognized as endocrine-signaling and systemically relevant. Herein we describe the development of the first 3D, BMAT model derived from either human or mouse bone marrow (BM) mesenchymal stromal cells (MSCs). We found that BMAT models can be stably cultured for at least 3 months in vitro, and that myeloma cells (5TGM1, OPM2 and MM1S cells) can be cultured on these for at least 2 weeks. Upon tumor cell co-culture, delipidation occurred in BMAT adipocytes, suggesting a bidirectional relationship between these two important cell types in the malignant BM niche. Overall, our studies suggest that 3D BMAT represents a "healthier," more realistic tissue model that may be useful for elucidating the effects of MAT on tumor cells, and tumor cells on MAT, to identify novel therapeutic targets. In addition, proteomic characterization as well as microarray data (expression of >22,000 genes) coupled with KEGG pathway analysis and gene set expression analysis (GSEA) supported our development of less-inflammatory 3D BMAT compared to 2D culture. In sum, we developed the first 3D, tissue-engineered bone marrow adipose tissue model, which is a versatile, novel model that can be used to study numerous diseases and biological processes involved with the bone marrow.

Mesenchymal stem cells gene signature in high‐risk myeloma bone marrow linked to suppression of distinct IGFBP2‐expressing small adipocytes

“…Second, a subpopulation of distinct BM‐‘regulated’ small adipocytes that are metabolically active and systemically regulated in physiological conditions (Scheller et al , ) may be suppressed by MM. Third, as shown in a 3‐dimensional model of adipocyte‐MM cell interaction, MM cells may use fatty acids in adjacent large adipocytes, causing the large cells to shrink and eventually be eliminated in areas highly involved with MM (Fairfield et al , ), as documented in our work.…”

“…The effects of MM on BM adipocytes remain to be elucidated (Morris & Edwards, 2018). Although adipocytes are evident in interstitial BM and support MM growth in vitro (Trotter et al, 2016;Fairfield et al, 2018), adiponectin, an adipogenic factor, has been shown to induce anti-MM activity and is suppressed in MM (Fowler et al, 2011). Recent studies identified 'regulated' adipocytes that are smaller in size than constitutively present large adipocytes (Scheller et al, 2015), and that function as an endocrine tissue (Cawthorn et al, 2014), suggesting that BM adipocytes are heterogeneous.…”

Multiple Myeloma and Fatty Acid Metabolism

“…In vitro culture of BMAs isolated from MM patients has been shown to support myeloma growth and enhance chemoresistance by activating autophagy through leptin, leading to inhibition of caspase cleavage and apoptosis . We have seen similar results in our lab, and also observed that BMAs shrink when cocultured with MM cells, perhaps indicating lipolysis or some other form of delipidation . Furthermore, adipocytes have been shown to support ovarian cancer cells.…”

In Vitro 3D Cultures to Reproduce the Bone Marrow Niche

“…). As compared with 2D cultures, general metabolic, DNA replication, ribosome, and proliferation pathways were upregulated in BM adipocytes in this model . Implementation of this silk fibroin scaffold by Reagan and colleagues to study myeloma growth, osteogenesis inhibition, and MM–MSC interaction unveiled myeloma support of endothelial cell assembly into capillary‐like structures and cell adhesion‐mediated drug resistance …”

“…As compared with 2D cultures, general metabolic, DNA replication, ribosome, and proliferation pathways were upregulated in BM adipocytes in this model. (87) Implementation of this silk fibroin scaffold by Reagan and colleagues to study myeloma growth, osteogenesis inhibition, and MM-MSC interaction unveiled myeloma support of endothelial cell assembly into capillary-like structures and cell adhesion-mediated drug resistance. (59) Using a 3D Matrigel environment to recapitulate the BM niche, the tumoricidal potential of TEGs (αβT cells engineered to express a defined γδTCR), novel αβT cells engineered to express a defined γδTCR cell receptor (TCR), was examined and compared with a conventional 2D model.…”