Replicating heterochronic parabiosis on a 3D microphysiological circuit to study the systemic regulation of aging muscle
Authors: Jang Young Charles , Lee Yunki , Choi Jennifer , Ahn Song ih , Kim YongTae 
Published: June 2021

Skeletal muscle possesses a robust ability to regenerate following injury due to the presence of muscle satellite cells (MuSCs) but this capacity declines with aging. Recent in vivo studies using heterochronic parabiosis, in which two animals are surgically attached to share blood circulation, demonstrated that systemic factors in the blood of young animals can rejuvenate the regenerative capacity of aged muscle. The exact mechanisms by which circulating youthful factors mediate the rejuvenating abilities of muscle stem cells, however, have yet to be elucidated. Due to the complexity of animal parabiosis and the dynamic nature of blood‐borne factors, reliable identification of these humoral factors remains a major hurdle. To address this challenge, we leverage microengineering technologies to build 3D co‐culture system of cells and tissues exposed to controlled biochemical cues with pathophysiological relevance. We engineered a 3D microvascularized MuSC niche on a chip (VMoC) designed to harness key characteristics of the native muscle microenvironment where MuSCs are cultured on a myofiber‐laden 3D hydrogel covered by vascular endothelial cells (ECs). We then employ our VMoC devices integrated to mimic heterochronic parabiosis to identify potential rejuvenating factors that enhance myogenesis of MuSCs. Our VMoC was engineered using a double‐layered microfluidic device that has 1) a top channel for the luminal side over a porous membrane, and 2) a bottom channel with a series of micropillars designed to confine the hydrogel. The primary MuSCs harvested from hindlimb muscles of GFP‐mice established multi‐nucleated myotubes on the myofiber surface in the bottom channel, while mouse aortic ECs created a monolayer in the top channel. We first found that the MuSCs co‐cultured with ECs significantly enhanced myogenic proliferation and differentiation as compared to the monoculture, while exhibiting similar differentiation to vascular endothelial growth factor (VEGF) treatment to the monoculture. This result demonstrated that a biological paracrine effect of vascular ECs can be investigated in our chip 3D co‐culture platform. More interestingly, we observed that the administration of serum respectively isolated from young, old, and Sod1−/− (oxidative stress‐accelerated model) mice showed distinct myogenic activities of MuSCs; those treated with old and Sod1−/− serum showed significantly lower myogenic differentiation. This result confirmed the ability of our VMoC platform to study systemic circulating factors in blood serum and to test the rejuvenating effects in our integrated chip‐to‐chip platform that can replicate heterochronic blood exchange. Furthermore, we can expand this technology to study heterochronic parabiosis between young‐ and old‐VMoCs. With precise control of systemic flow, our platform would address the critical barriers of in vivo parabiosis study, and serve as an experimental model to investigate the systemic regulation of aging and facilitate the identification of putative rejuvenation factors.Support or Funding InformationNIH R21AR072287 (YJ) S&R Foundation Ryuji Ueno Award (YJ)NIH NIH DP2 HL142050 (YK)Schematic diagram of vascularized muscle on a chip (Figure 1) and representative images of cells in 3D co‐culture (Figure 2).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.