(B-76) Fabricating 3-dimensional human brown adipose microtissues for transplantation studies

Introduction:: Transplanting cell cultured brown adipocytes (BAs) represents a promising approach to prevent and treat obesity (OB) and its associated metabolic disorders, including type 2 diabetes mellitus (T2DM). However, transplanted BAs have a very low survival rate in vivo. The enzymatic dissociation during the harvest of fully differentiated BAs also loses significant cells. There is a critical need for novel methods that can avoid cell death during cell preparation, transplantation, and in vivo. Here, we reported that preparing BAs as injectable microtissues could overcome the problem.

Materials and Methods:: Human BA microtissues were fabricated using a 3D suspension culture system with a PEG hydrogel. We investigated the effects of 3D culture on BA differentiation and UCP-1 expression and identified the optimal initial cell aggregate size. The microtissues were then cryopreserved and transplanted into the high-fat diet (HFD)HF-induced obesity and T2DM mice. We evaluated the survival, safety, and efficacy of the microtissues by monitoring body weight, fat gain, glucose tolerance, insulin sensitivity, and the impact on multiple organs and the secretion of protein factors and endogenous adipokines.

Results, Conclusions, and Discussions:: We found that 3D culture promoted BA differentiation and UCP-1 expression, and the optimal initial cell aggregate size was 100 μm. The microtissues could be produced at large scales via 3D suspension assisted with a PEG hydrogel and could be cryopreserved. Fabricated microtissues could survive in vivo for the long term. They alleviated body weight and fat gain and improved glucose tolerance and insulin sensitivity in HFD-induced OB and T2DM mice. Transplanted microtissues impacted multiple organs, secreted protein factors, and influenced the secretion of endogenous adipokines. To our best knowledge, this is the first report on fabricating human BA microtissues and showing their safety and efficacy in T2DM mice. The proposal of transplanting fabricated BA microtissues, the microtissue fabrication method, and the demonstration of efficacy in T2DM mice are all new. Our results show that engineered 3D human BA microtissues have considerable advantages in product scalability, storage, purity, safety, dosage, survival, and efficacy.

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