3D bioprinting of skeletal muscle and neuromuscular junction tissue models

National Center For Advancing Translational Sciences

Investigators

Abstract

This project includes a collaboration between NCATS 3DTBP, the NCATS PaVe-GT program, and Dr Carsten Bonnemann and Dr. Michael Wards laboratories at NINDS to develop muscle and neuromuscular junction tissue models to test the effect of gene therapies being developed for the treatment of Congenital myasthenic syndromes. We selected a commercial platform by the company CurioBio that is a specialized instrument for measuring in vitro muscle contractility in a 24-well format. This muscle contraction assay platform contains two pillars hanging from the top in the individual wells. iMyoblasts embedded in bioink are bioprinted wrapping the pillars, and myotube bundles then mature in three weeks which contract upon electrical and chemical stimulation. One of the two pillars is elastic where a magnet was attached at the end and the plate reader detects the magnetic field changes representing displacement between the pillars. With known mechanical properties of the materials of the pillars, contracting forces are calculated in real time and simultaneously from the entire 24 wells. Our team has established a process to scale-up production of iMyoblast using automated tissue culture robots and a functional in vitro muscle model using bioprinting iPSC derived myoblasts, demonstrating muscle contracting response under electrical and acetylcholine stimulations. We have obtained CRISPR edited Congenital myasthenic syndromes imyoblasts and demosntrated that contractile force and relaxion of the disease mutant muscle bundles are different from healthy cells. We have also shown that the bioprinted muscles can be transduced with AVV-virus. We are now looking at disease correction with clinical AVV candidate for Congenital myasthenic syndromes. This year we spend most of the time troubleshooting the cells because after passage they lost the ability to contract in 3D. We have explored different expansion protocols and hope to be able to test AVV vectors by the end of 2025. We have also established a collaboration with Cure VCP and ADDSL1 foundations to model VCP and ADDSL1 muscle diseases with patient derived cells. Activities this year included obtained disease myoblasts and testind in the 3D emgineered musclet tissue. Troubleshooting is ongoing to purify myoblasts that contract in 3D.