Micro-Fat Tissue on Chip

Obesity has been defined as abnormal or excessive fat accumulation. The prevalence of obesity in the Western world has reached more than 35 % of the population, and thus has grown to epidemic proportions. The most effective treatment for obesity is dietary and lifestyle changes; however, these approaches are difficult to maintain over the long term. Therefore, existing pharmacological and surgical interventions largely act by either suppressing caloric intake or reducing stomach size, respectively. Despite all efforts, obesity and related diseases remain major health risks for patients and are associated with high healthcare costs in most countries. An attractive alternative for treating obesity is to reduce the lipid storage function of the adipose tissue by reprogramming its cell type composition. The human fat is mainly composed of white adipose tissue, which exhibits low capacities to burn fat on its own. Nevertheless, a small fraction of fat cells, named brown and beige fat cells, is able to burn fat and produce heat in order to maintain the body temperature. The cells are present primarily in newborns before reducing in number upon aging. Knowledge of the developmental factors, which govern fat cell type specification can potentially be used to increase the brown-like properties of white fat depots in obese patients. Fatal but also postnatal development information of fat cells is hardly existent due to either inaccessibility of primary tissue or the relatively long processes of the cell type evolvement. Thus, the overall goal of MicroAdiPSChip is to engineer miniaturized human adipose tissue on a chip in order to understand how white and brown fat cells develop from a pluripotent stem cell. Upon integration of the pluripotent stem cell on chip, we aim to investigate systematically how changing chemical, tissue architectural, and mechanical microenvironments shape the colour of the fat cells.

To address the question which in vivo factors influence fat cell type development, we started the of the MicroAdiPSChip project by engineering and integrating an in vitro three-dimensional (3D) tissue model for the induced pluripotent stem cells on new microfluidic chip platforms. In general, microfluidic chip technologies are used (1) to gain control over the chemical microenvironment of in vitro cell cultures over longer time scales and (2) upon miniaturization, allow for multiplexed workflow for tens of cell cultures. Most chip platforms, however, are designed for holding two-dimensional (2D) immortalized laboratory cell lines rather than fragile and sensitive stem cells in a 3D environment. Therefore, to enable the 3D stem cell approach on a chip, we had to scale microfluidic designs and production technologies towards holding cultures with dimension of hundreds rather than tens of microns. Using two independent approaches, we implemented new design rules and additive production technologies for microfluidics: while the first approach focused on new fluidic structures needed for generating 3D cultures on chip from human stem cells, the second approach used 3D printing of defined microfluidic channel structures to enable fluidic control mechanisms for long-term culturing. Combination of both processes allowed generating a new platform to automate the simultaneous differentiation of 96 human pluripotent stem cell cultures into white adipocytes under defined temporal chemical programs on a single chip. Currently, we are performing deep cellular characterization of these generated adipose cell types. In the second phase of the project, the research advances will allow us in to generate “browning” of the stem cell-derived white adipocytes.
To investigate tissue architectural influences on fat cell development, we proposed to generate an assembling method for complex micro-fat tissue on a chip. Here, our working hypothesis was that paracrine communication between stem cells and cells of the perivascular microenvironment define the developmental fat cell trajectory. Therefore, we induced the development of adipocytes, endothelial and pericytes from the same human pluripotent stem cells. From the latter two cell types, we were able to generate functional vessels in vitro. In addition, single cell transcriptomic analysis of the vessel formation process in a chip-compatible hydrogel revealed the major cell-cell interaction proteins required for establishing these vessels. In combination with our new open microfluidic chip platform, we will exploit the molecular knowledge of vessel formation to generate vascularized stem cell-derived white adipose tissue on a chip. Through changes in the cell type composition and spatial organization of the vascularized adipose tissue, we will investigate if we can induce “browning” of the tissue solely by modifications of the its architecture.

The MicroAdiPSChip project led to the following novel technologies and scientific advances beyond the state of the art:

1) The generation of two new microfluidic chip platforms for long-term culturing and processing 3D of human stem cell cultures
2) A new additive manufacturing technology for microfluidic chip production
3) A differentiation platform to derive white adipocytes from human induced pluripotent stem cell in high yield
4) The engineering of vascularized human adipose tissue on chip
5) A single cell study of in vitro blood vessel formation from stem cell-derived endothelial cells within chip compatible hydrogel

With these advances, we set the foundation to provide much needed insights into adipocyte cell type development and therewith create the basis for new strategies for obesity therapies as proposed.

Periodic Reporting for period 3 - MicroAdiPSChip (Micro-Fat Tissue on Chip)

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