Spheroids from bladder smooth muscle cells for bladder tissue engineering
Tim Gerwinn, MD1, Souzan Salemi, PhD2, Lisa Krattiger, MSc2, Daniel Eberli, MD PhD2, Maya Horst, MD1.
1Universtiy Childrens Hospital Zurich, Zurich, Switzerland, 2Universtiy Hospital Zurich, Zurich, Switzerland.
BACKGROUND: Cell-based tissue engineering (TE) was proposed as an alternative to improve treatment outcomes in end stage bladder disease. However, TE approaches with 2D smooth muscle cell (SMC) culture lack success. We hypothesize that 3D SMC cultures (spheroids) with improved regenerative properties are superior building blocks for bladder TE.
METHODS: Bladder SMC spheroids were established. Culture conditions and development of contractile phenotype, as well as extracellular matrix (ECM) deposition were investigated using qRT-PCR, immunofluorescence, and immunoblotting and compared to 2D culture.
RESULTS: For the first time, we established and analysed primary bladder SMC spheroids. SMC spheroids were viable for up to 14 days. Unlike 2D culture, SMCs in spheroid culture did not appear to proliferate but mainly to differentiate. Spheroids predominantly expressed the late myogenic differentiation marker MyH11. On the contrary, 2D SMC expressed higher amounts of the general SMC differentiation marker α-SMA and lesser amounts of MyH11. Furthermore, the expression of bladder wall specific ECM proteins in SMC spheroids was markedly increased.
CONCLUSIONS: Primary bladder SMC spheroids offer several benefits over 2D SMCs; they show increase in late contractile protein markers, indicating the higher state of maturation and produce larger amounts of ECM proteins. We were able to confirm that SMC spheroids are promising building blocks to study detrusor regeneration in detail, and may provide improved function and regenerative potential, contributing to take bladder TE a significant step forward.
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