Dynamic culture induces a cell type-dependent response impacting on the thickness of engineered connective tissues

Abstract

Mesenchymal cells are central to connective tissue homeostasis and are widely used for tissue-engineering applications. Dermal fibroblasts and adipose-derived stromal cells (ASCs) allow successful tissue reconstruction by the self-assembly approach of tissue engineering. This method leads to the production of multilayered tissues, devoid of exogenous biomaterials, that can be used as stromal compartments for skin or vesical reconstruction. These tissues are formed by combining cell sheets, generated through cell stimulation with ascorbic acid, which favours the cell-derived production/organization of matrix components. Since media motion can impact on cell behaviour, we investigated the effect of dynamic culture on mesenchymal cells during tissue reconstruction, using the self-assembly method. Tissues produced using ASCs in the presence of a wave-like movement were nearly twice thicker than under standard conditions, while no difference was observed for tissues produced from dermal fibroblasts. The increased matrix deposition was not correlated with an increased proliferation of ASCs, or by higher transcript levels of fibronectin or collagens I and III. A 30% increase of type V collagen mRNA was observed. Interestingly, tissues engineered from dermal fibroblasts featured a four-fold higher level of MMP-1 transcripts under dynamic conditions. Mechanical properties were similar for tissues reconstructed using dynamic or static conditions. Finally, cell sheets produced using ASCs under dynamic conditions could readily be manipulated, resulting in a 2 week reduction of the production time (from 5 to 3 weeks). Our results describe a distinctive property of ASCs' response to media motion, indicating that their culture under dynamic conditions leads to optimized tissue engineering.

Methylcellulose Based Thermally Reversible Hydrogel System for Tissue Engineering Applications

“…The single or multilayer cell sheets developed in the present study may be used in the applications to tissue reconstructions [45,46,47,48,49,50,51,52,53,54]. Single cell sheets can be transplanted directly to host in the case of skin [18,52], corneal epithelium [47,49], and periodontal ligament [48].…”

Human adipose-derived stromal cells for the production of completely autologous self-assembled tissue-engineered vascular substitutes

“…However, complete and stable differentiation of ASC in vitro is hard to achieve and, in opposition to our goal, it would increase the production time of the substitutes. However, undifferentiated ASC share many features with fibroblasts [10,12,34], the main cellular component of the adventitia. In this study, we demonstrated the potential of undifferentiated ASC to be used, as an alternative to DF and SMC, to engineer in vitro autologous and completely biological self-assembled vascular substitutes.…”

“…However, since the TEVS-14R did not fuse as well as the TEVS-7R, intermediate thicknesses should be assessed. Moreover, ASC have been shown to deposit more ECM and form thicker cell sheets when cultured under dynamic conditions [34]. Taking advantage of this characteristic could allow for the production of thicker ASC-TEVS without increasing the number of revolutions or culture time.…”

Magnetic Resonance Imaging of Human Tissue-Engineered Adipose Substitutes

“…32 Tissue reconstruction proceeded as a multistep protocol based on adaptations of previously published methods from our group. 32,33,35 The adipose substitutes were produced according to two main models based on initial cell seeding into six-well plates for series 1 and 2 (9.6 cm 2 ), while series 3 consisted of a model produced from initially larger culture vessels (57-87 cm 2 ). For each model, cell sheets were gradually formed into the culture plates and lifted with forceps after 31-43 days.…”

“…The resulting connective cell sheets can then be superposed to produce thicker connective tissues. 32,33 If an adipogenic induction step is performed in parallel to matrix stimulation, then the resulting tissue sheets feature functional adipocytes filled with lipids. 32,34 These reconstructed tissues do not contain synthetic or exogenous biomaterials, and their production can be customized for shape and thickness.…”