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Understanding The Crosstalk Between Adipose-derived Stem Cells And Chondrocytes Using Co-culture Systems
Mary E. Ziegler, PhD, Alexandria M. Sorensen, Derek A. Banyard, MD, MBA, MS, Philipp Neßbach, MSc, Gregory RD Evans, MD, Alan D. Widgerow, MBBCh, MMed.
University of California, Irvine, Orange, CA, USA.

Purpose: Microtia, a congenital condition that leads to external ear deformities of varying degrees, is currently treated using surgical reconstruction techniques, which all require a multi-stage approach and have other inherent disadvantages. Cartilage engineering serves as an alternative treatment approach in the field of external ear reconstruction. Typically, this technique involves isolating and culturing chondrocytes that are derived from the remnants of the patient's auricular cartilage, with the aim of creating bioengineered cartilage matrices. However, cartilage engineering is inundated with a variety of challenges, including the difficulty in culturing sufficient chondrocytes and the production of cartilage that does not possess the proper mechanical properties. To overcome these difficulties, we propose a novel cartilage engineering model that involves co-culturing chondrocytes with adipose-derived stem cells (ADSCs). However, to establish this model, a better understanding of the crosstalk between these cells in co-culture is necessary. We hypothesize that ADSCs have a trophic effect on chondrocytes, making them more chondrogenic compared to chondrocytes that are cultured alone.
Methods: Auricular chondrocytes (ACs) were isolated from porcine ear and ADSCs were isolated from human lipoaspirate. ACs and ADSCs were co-cultured either directly or via a trans-well system at defined ratios. In addition, the cells were cultured alone. Cell proliferation was assessed using a CCK-8 assay for the transwell co-culture and using flow cytometry for the direct co-culture. In addition, the supernatants from these co-cultures were collected, and a secretome analysis was performed to assess the differential secretion of proteins under these conditions.
Results: When ADSCs were co-cultured with ACs, proliferation was reduced compared to ACs that were cultured alone. The secretome analysis revealed that the cells in the co-culture conditions produced more chondrogenic factors compared to the chondrocytes alone. In addition, we validated that the factors important for chondro-induction were secreted by the ADSCs.
Conclusion: These data revealed that the ADSCs provided paracrine support to the ACs to induce chondrogenesis, which supports cartilage engineering when the number of ACs available is limited. Furthermore, revealing the mechanism of chondrogenesis in this setting contributed to refining cartilage engineering in order to ultimately utilize a patient's own chondrocytes and ADSCs for the creation of a customized ear framework that could be used for surgical reconstruction.


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