Phenotypic Analysis of Human ES Cell-Derived Muscle Stem Cells

The overall goal of this project was to use hESCs to define the cellular and functional phenotypes of human muscle stem cells as they differentiate along the muscle lineage, and specifically evaluate their ability to augment tissue and function of dystrophic muscle. Toward this goal, we have evaluated one muscle-specific reporter hESC line, generated a second line using another muscle-specific promoter, developed three alternative methods for directing myoblast differentiation from hESCs in culture, and piloted injections of hESC-derived myogenic precursor cells into hind limb muscle of immunodeficient mice.

Muscle cells derived from human embryonic stem cell (hESC) can be potential source for cell therapy to regenerate muscular diseases. The focus of this grant has been to develop efficient methods for isolating muscle stem cells from hESCs that avoid animal products so that we can use these to both understand how muscle cells form, and determine which of these may be best for treating muscular dystrophy.

Our progress over the past year has significantly advanced the aims of this work: 1) Determine the cellular phenotypes of human muscle stem cells as they differentiate into myoblasts, and 2) Determine the ability of human muscle stem cells at different stages of development to engraft, proliferate and differentiate into muscle in a mouse model of muscular dystrophy, and determine their functional and myo-mechanical effects on dystrophic muscle. We now have a working system to derive early progenitor muscle cells from human embryonic stem cells. The differentiation protocol has been developed sufficiently such that skeletal muscle cells can be generated from human ES cells. We have identified points along the differentiation process at which muscle cells that are less mature and possibly more stem-like are prevalent. The data suggests that based on the genes the cells express at early stages, isolation and transplantation of cells at that stage but not further along will be most beneficial for transplantation and clinical application. This brings us a step closer to obtaining useful muscle cells that can be transplanted to treat muscle disorders. The current plan is to test these cells in muscle injury preclinical models to evaluate their capacity to regenerate injured muscle.

Our progress over the past year has advanced the aims of this work: 1) Determine the cellular phenotypes of human muscle stem cells as they differentiate into myoblasts, and 2) Determine the ability of human muscle stem cells at different stages of development to engraft, proliferate and differentiate into muscle in a mouse model of muscular dystrophy, and determine their functional and myo-mechanical effects on dystrophic muscle. We have derived early progenitor muscle cells from human embryonic stem cells. We have used a surface marker profile that we have characterized on endogenous human muscle stem cells to identify and induced human muscle precursors in our differentiation assays. We have tested various differentiation assays by transplanting human cells into mouse muscle. We are using the information gained from characterizing induced muscle cells to compare with endogenous muscle cells that naturally exist in order to learn how to efficiently derive human muscle stem cells that can be used to treat muscle diseases.