(J-385) A Multicellular Approach to Control Adipocyte Characteristics in Tissue-Engineered Skeletal Muscle

Introduction:: White adipose tissue is known for its ability to store energy, and it also plays a role in changes in hormone status at the organismal level. Brown adipose tissue is identified by an increased energy expenditure and is being considered as a potential remedy for obesity. Less is understood about brown adipose tissue; however, exciting research has identified brown adipose tissue as a potential treatment for obesity, at least in part, due to its ability to increase energy expenditure. To this end, the ability for white adipocytes to transition into brown adipocytes (beiging) is of great interest. The vast majority of the research underlying white or brown adipose tissue dynamics has been directed towards the understanding of white or brown adipocytes in adipose tissue depots. Conversely, efforts to understand the role of white and brown adiopocytes, and the interconversion between the two in skeletal muscle are less common. The goal of the present work is to develop an in vitro model of skeletal muscle (tissue engineered skeletal muscle (TE-SkM)) in order to investigate phenomena related to adipocyte dynamics in skeletal muscle. In these preliminary experiments the objective was to determine if muscle precursor cells (MPCs) and fibroadipogenic progenitor cells (FAPs) derived from skeletal muscle could be used to create TE-SkM containing both muscle cells and adipocytes, and whether or not the adipocyte type could be controlled.

Materials and Methods:: MPCs and FAPs were obtained from skeletal muscles of male Lewis rat using a prelate technique combined with magnetic-activated cell sorting (MACS). To produce tissue-engineered skeletal muscle (TE-SkM) constructs containing different types of adipocytes, MPCs and FAPs were placed in a fibrin hydrogel contained in a custom polydimethylsiloxane mold, and the constructs were treated with brown adipogenic media (BAM) or white adipogenic media (WAM). After a period of 21 days, samples were stained with F-Actin, DAPI, and BODIPY, to identify myotubes, nuclei, and adipocytes, respectively. Confocal images of the TE-SkM constructs were analyzed to determine if the media type (WAM or BAM) had an effect on adipocyte size. RT-PCR was used to identify key genetic markers of brown adipogenesis, including UCP1 and CIDEA.

Results, Conclusions, and Discussions:: Results and Discussion: TE-SkM constructs treated with WAM had an ~ 3-fold increase in adipocyte size when compared to those treated with BAM. There was an ~3-fold higher expression of UCP1 in constructs treated with BAM as compared to those treated with MM, while those treated with WAM had a decrease in UCP1 expression when compared to MM. In constructs treated with either WAM or BAM there was an ~4-5-fold increase in CIDEA expression as compared to those treated with MM.

Conclusions: These initial findings provide support for the development of this approach to control adipocyte characteristics in TE-SkM in order to facilitate investigations into mechanisms and applications of these phenomena. A particularly critical piece of evidence to support this contention is the increased expression of UCP1 which is a key marker of brown adipogenic differentiation. These studies also support the idea that MPCs and FAPs are appropriate cells to serve as the foundation for TE-SkM development in studies to understand adipocyte phenomena in skeletal muscle. Future research efforts will examine the effects of diabetes on white and brown adipogenesis using this TE-SkM model, as well as determining methods to gain an even greater control over adipocyte content and type.


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References (Optional): : 1. Kim, S. H., & Plutzky, J. (2016). Diabetes & Metabolism Journal, 40(1), 12-21. 2. Cypess, A. M., & Kahn, C. R. (2010). Current opinion in endocrinology, diabetes, and obesity, 17(2), 143.