Abstract

Circulatory System and Integrated Muscle Tissue for Drug and Toxicity Testing

George Truskey, R. Eugene and Susie E. Goodson Professor of Biomedical Engineering, Duke University

We have developed functional human tissue-engineered blood vessels (TEBVs) and skeletal muscle myobundles that model normal structure and function and represent disease states. We have fabricated specialized perfusion systems for the human TEBVs and skeletal muscle myobundles that enable in situ measurements of vasoactivity, dimensions, leukocyte adhesion, calcium transients, contractile force and oxygen uptake.  Offline measurements include glucose uptake, nitric oxide production, gene and protein expression, and metabolite levels. These microphysiological systems (MPS) have been prepared using primary human cells, induced pluripotent stem cells (iPSCs) from human fibroblasts, and reprogrammed endothelial cells.  Disease states that we examined including inflammation, Duchenne Muscular Dystrophy (DMD), metabolic storage disease, and Hutchison-Gilford Progeria Syndrome (HGPS). Since iPSC-derived cells exhibit reduced function relative to primary cells, addition of small molecules that activate specific transcription factors controlling differentiation improves MPS function.   We have generated an immortalized human primary DMD cell line to prepare functional myobundles. The bundles show some characteristics of a dystrophic muscle such as increased susceptibility to repeated eccentric contractions, evidence of myotube branching, and potential hypercontraction of myotubes. TEBVs prepared with cells from HGPS patients exhibit reduced numbers of smooth muscle cells, apoptosis and calcification, conditions found in vessels of adults with the disease.   Treatment of TEBVs with cells derived from HGPS patients with rapamycin appears to improve TEBV function. Cytokine addition does mimic inflammation in TEBVs and can be reversed following pretreatment with statins.  Cytokines reduce myobundle contractile force and exacerbate drug toxicity.  For myobundles, electrical stimulation frequencies above 10 Hz increased force and basal glucos