Human ear, nasal, and rib chondrocytes were compared with respect to their suitability to generate autologous cartilage grafts for nonarticular reconstructive surgery. Cells were expanded for two passages in medium containing 10% fetal bovine serum without (control) or with transforming growth factor beta(1) (TGF-beta(1)), fibroblast growth factor 2 (FGF-2), and platelet-derived growth factor bb (PDGF-bb) (TFP). Expanded cells were cultured as three-dimensional pellets in chondrogenic serum-free medium containing insulin, dexamethasone, and TGF-beta(1). Chondrocytes from all three sources were successfully isolated, increased their proliferation rate in response to TFP, and dedifferentiated during passaging. Redifferentiation by ear and nasal, but not rib, chondrocytes was enhanced after TFP expansion, as assessed by the significant increase in glycosaminoglycan (GAG)/DNA content and collagen type II mRNA expression in the resulting pellets. TFP-expanded ear and nasal chondrocytes generated pellets of better quality than rib chondrocytes, as assessed by the significantly higher GAG/DNA content and collagen type II mRNA expression, and by the relative stain intensities for GAG and collagen types I and II. In conclusion, postexpansion cell yields suggest that all three sources investigated could be used to generate autologous grafts of clinically relevant size. However, ear and nasal chondrocytes, if expanded with TFP, display superior postexpansion chondrogenic potential and may be a preferred cell source for cartilage tissue engineering.

NCT01138891 (http://www.clinicaltrials.gov).

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The fate of human adipose tissue stem cells (ASCs) is largely determined by biochemical and mechanical cues from the extracellular matrix (ECM), which are sensed and transmitted by integrins. It is well known that specific ECM constituents influence ASC proliferation and differentiation. Nevertheless, knowledge on how individual integrins regulate distinct processes is still limited. We performed gene profiling of 18 alpha integrins in sorted ASCs and adipocytes, identifying downregulations of RGD-motif binding integrins integrin-alpha-V (ITGAV) and integrin-alpha-5 (ITGA5), upregulation of laminin binding and leukocyte-specific integrins and individual regulations of collagen and LDV-receptors in differentiated adipocytes in-vivo. Gene function analyses in in-vitro cultured ASCs unraveled differential functions of ITGA5 and ITGAV. Knockdown of ITGAV, but not ITGA5 reduced proliferation, caused p21Cip1 induction, repression of survivin and specific regulation of Hippo pathway mediator TAZ. Gene knockdown of both integrins promoted adipogenic differentiation, while transgenic expression impaired adipogenesis. Inhibition of ITGAV using cilengitide resulted in a similar phenotype, mimicking loss of pan-ITGAV expression using RNAi. Herein we show ASC specific integrin expression patterns and demonstrate distinct regulating roles of both integrins in human ASCs and adipocyte physiology suggesting a negative impact of RDG-motif signaling on adipogenic differentiation of ASCs via ITGA5 and ITGAV.