Project description:Satellite cells, the stem cells of skeletal muscle tissue, hold a remarkable regeneration capacity and therapeutic potential in regenerative medicine. However, low satellite cell yield from autologous or donor-derived muscles hinders the adoption of satellite cell transplantation for the treatment of muscle diseases, including Duchenne muscular dystrophy (DMD). To address this limitation, here we investigated whether satellite cells can be derived in allogeneic or xenogeneic animal hosts. First, injection of CRISPR/Cas9-corrected mouse DMD-induced pluripotent stem cells (iPSCs) into mouse blastocysts carrying an ablation system of host satellite cells gave rise to intraspecies chimeras exclusively carrying iPSC-derived satellite cells. Furthermore, injection of genetically corrected DMD-iPSCs into rat blastocysts resulted in the formation of interspecies rat-mouse chimeras harboring mouse satellite cells. Remarkably, iPSC-derived satellite cells or derivative myoblasts produced in intraspecies or interspecies chimeras restored dystrophin expression in DMD mice following intramuscular transplantation, and contributed to the satellite cell pool. Collectively, this study demonstrates the feasibility of producing therapeutically competent stem cells across divergent animal species, raising the possibility of generating human muscle stem cells in large animals for regenerative medicine purposes.
Project description:Satellite cells, the stem cells of skeletal muscle tissue, hold a remarkable regeneration capacity and therapeutic potential in regenerative medicine. However, low satellite cell yield from autologous or donor-derived muscles hinders the adoption of satellite cell transplantation for the treatment of muscle diseases, including Duchenne muscular dystrophy (DMD). To address this limitation, here we investigated whether satellite cells can be derived in allogeneic or xenogeneic animal hosts. First, injection of CRISPR/Cas9-corrected mouse DMD-induced pluripotent stem cells (iPSCs) into mouse blastocysts carrying an ablation system of host satellite cells gave rise to intraspecies chimeras exclusively carrying iPSC-derived satellite cells. Furthermore, injection of genetically corrected DMD-iPSCs into rat blastocysts resulted in the formation of interspecies rat-mouse chimeras harboring mouse satellite cells. Remarkably, iPSC-derived satellite cells or derivative myoblasts produced in intraspecies or interspecies chimeras restored dystrophin expression in DMD mice following intramuscular transplantation, and contributed to the satellite cell pool. Collectively, this study demonstrates the feasibility of producing therapeutically competent stem cells across divergent animal species, raising the possibility of generating human muscle stem cells in large animals for regenerative medicine purposes.
Project description:Immune rejection has long hindered allogenic cell transplantation therapy. Current genetic modification approaches, including direct targeting of major histocompatibility complex or constitutive expression of immune inhibitory molecules, exhibit drawbacks such as severe adverse effects or an elevated risk of tumorigenesis. To overcome these limitations, we propose an innovative approach aimed at inducing cell-type-specific immune tolerance in differentiated cells. By engineering human embryonic stem cells, we enable the exclusive production of immune inhibitory molecules, PD-L1/CTLA4Ig, within differentiated cells following lineage commitment. Leveraging this approach, we have successfully generated hepatocyte-like cells expressing PD-L1 and CTLA4Ig, which effectively induced local immunotolerance. This strategy was evaluated in a humanized mouse model designed to mimic human immune system. Our results demonstrate robust and selective induction of immunotolerance specific to hepatocytes, thereby improving graft survival without tumor formation. This precise immune tolerance strategy holds promise for advancing the development of stem cell-based therapeutics in regenerative medicine.
Project description:Aged skeletal muscle is markedly affected by fatty muscle infiltration and strategies to reduce the occurrence of adipocytes within skeletal muscle, the intramuscular adipose tissue (IMAT), are urgently needed. Fibroblast growth factor-2 (FGF-2) is a critical growth factor for muscle tissue. Here, we show that FGF-2 not only stimulates muscle growth, but also promotes intramuscular adipogenesis. Using multiple screening assays for upstream and downstream signaling of microRNA (miR)-29a we located the secreted protein and adipogenic inhibitor SPARC to an FGF-2 signaling pathway that is conserved between skeletal muscle cells from mice and humans and that is activated in skeletal muscle from aged mice. FGF-2 induces the miR-29a/SPARC axis through transcriptional activation of FRA-1 which binds and activates an evolutionary conserved AP-1 site element proximal in the miR-29a promoter. Genetic deletions in muscle cells and AAV-mediated overexpression of FGF-2 or SPARC in mouse skeletal muscle revealed that this axis regulates differentiation of fibro/adipogenic progenitors in vitro and intramuscular fat formation in vivo. Thus, our data highlight an ambivalent role of FGF-2 for adult skeletal muscle and reveal a novel pathway to combat fat accumulation in aged skeletal muscle.
Project description:The content of intramuscular fat (IMF) is closely related to meat quality traits. In this study, in order to explore the candidate genes related to IMF content, the longissimus dorsi muscle of Guangling donkey was measured for intramuscular fat content. According to its intramuscular fat content, it was divided into two groups, the low fat group (L , N=3) and high-fat group (H, n=3), using RNA-seq to identify differentially expressed genes (DGEs) on the longissimus dorsi muscle tissue of Guangling donkey with high and low intramuscular fat content to reveal the possibility Gene network and metabolic pathways that help increase intramuscular fat content. A total of 167 DEGs (|log2Fold Change|>=1 and FDR<0.05) were detected in the high (H) and low (L) groups of Guangling donkeys, of which 64 were up-regulated genes and 103 were down-regulated genes. The GO enrichment and KEGG pathway analysis showed that these differential genes were enriched in several biological processes and pathways related to adipocyte differentiation, lipid biosynthesis, and neutral lipid metabolism. These results will help to further explore the molecular mechanism of IMF deposition in donkeys and provide a theoretical basis for the molecular breeding of Guangling donkeys.