Defined adipocyte differentiation and long term stability through a cellulose-based culture matrix

White adipose tissue (WAT) comprises about one fourth of the human body, interacts with many other organs via paracrine and endocrine signals and sequestrates lipophilic substances. Thereby it impacts distribution and levels of drugs [1]. Additionally WAT is the origin of different diseases and thus target of several drugs addressing those [2]. Therefore an adipose tissue testsystem is highly needed to analyze fundamental biological issues in physiological or diseased state, screen for potential drugs or create affiliated safety profiles. Additionally adipose tissue substitutes are highly desired to treat lost, deformed or burned subcutaneous fatty tissue [3]. In several promising attempts substantial progress was achieved e.g. by the encapsulation of adipocytes in a 3D environment with native matrix components like gelatine [4] or the optimization of media composition [5]. However, regardless of its potential in vivo or in vitro application, an adipose tissue model has to fulfill two main requirements which could not be implemented so far. First, the model has to show in vitro stability for a period of several weeks to enable its maturation and time-dependent investigations. Second, culture conditions have to be free of animal derived components or preferably completely defined to exclude possible impairments through unknown constituents and allow for GMP-compliant production. In this approach a novel cellulose based matrix and a specific media supplementation are combined to achieve defined adipocyte differentiation and long term maintenance. Human primary adipose-derived stem cells (ASCs) were isolated, characterized and expanded xenofree. ASCs were next to tissue culture polystyrol applied to a collagen coated surface and the cellulose-based matrix. Adipogenic differentiation of ASCs was induced by addition of a composed defined differentiation medium and continued for 14 days. On day 14 medium was switched to a composed defined adipocyte maintenance medium. Obtained adipocytes were subsequently cultured for additional 28 days. Preservation of adipocyte characteristics were evaluated by immunofluorescence staining of specific markers like perilipin A and the quantitative analysis of stored lipids. Retained adipocyte metabolism was determined by released leptin, lipolysis by released glycerol levels. Our results classify the cellulose-based matrix as a useful tool to realize and optimize defined adipogenic differentiation of ASCs and the subsequent adipocyte maintenance. The matrix constitution enables strong cell matrix interaction and thereby strengthens cell adherence which is often diminished under defined culture conditions. Due to matrix-induced cell quiescence adipocytes' long-term stability was enhanced. Based on matrix-derived support successful differentiation and maintenance were confirmed via the composed media under completely defined conditions. Therefore the cellulose-based matrix is a promising biomaterial for stable long-term culture of quiescent cells with preservation of cell specific functions and characteristics which could as well be applied in other setups. Consecutive defined adipose tissue models could be further expanded e.g. to co-culture models and used as testsystems or tissue substitutes.