Stem cells undergo cytokine-driven differentiation, but this process often takes longer than several weeks to complete. A novel mechanism for somatic stem cell differentiation via phagocytosing ‘model cells’ (apoptotic differentiated cells) was found to require only a short time frame. Pluripotent-like Muse cells, multipotent mesenchymal stem cells (MSCs), and neural stem cells (NSCs) phagocytosed apoptotic differentiated cells via different phagocytic receptor subsets than macrophages. The phagocytosed-differentiated cell-derived contents (e.g., transcription factors) were quickly released into the cytoplasm, translocated into the nucleus, and bound to promoter regions of the stem cell genomes. Within 24 ~ 36 h, the cells expressed lineage-specific markers corresponding to the phagocytosed-differentiated cells, both in vitro and in vivo. At 1 week, the gene expression profiles were similar to those of the authentic differentiated cells and expressed functional markers. Differentiation was limited to the inherent potential of each cell line: triploblastic-, adipogenic-/chondrogenic-, and neural-lineages in Muse cells, MSCs, and NSCs, respectively. Disruption of phagocytosis, either by phagocytic receptor inhibition via small interfering RNA or annexin V treatment, impeded differentiation in vitro and in vivo. Together, our findings uncovered a simple mechanism by which differentiation-directing factors are directly transferred to somatic stem cells by phagocytosing apoptotic differentiated cells to trigger their rapid differentiation into the target cell lineage.

Multilineage-differentiating stress enduring (Muse) cells, non-tumorigenic endogenous pluripotent stem cells, reside in the bone marrow (BM), peripheral blood, and connective tissue as pluripotent surface marker SSEA-3(+) cells. They express other pluripotent markers, including Nanog, Oct3/4, and Sox2 at moderate levels, differentiate into triploblastic lineages, self-renew at a single cell level, and exhibit anti-inflammatory effects. Cultured mesenchymal stromal cells (MSCs) and fibroblasts contain several percent of SSEA-3(+)-Muse cells. Circulating Muse cells, either endogenous or administered exogenously, selectively accumulate at the damaged site by sensing sphingosine-1-phosphate (S1P), a key mediator of inflammation, produced by damaged cells and replace apoptotic and damaged cells by spontaneously differentiating into multiple cells types that comprise the tissue and repair the tissue. Thus, intravenous injection is the main route for Muse cell treatment, and surgical operation is not necessary. Furthermore, gene introduction or cytokine induction are not required for generating pluripotent or differentiated states prior to treatment. Notably, allogenic and xenogenic Muse cells escape host immune rejection after intravenous injection and survive in the tissue as functioning cells over 6 and ∼2 months, respectively, without immunosuppressant treatment. Since Muse cells survive in the host tissue for extended periods of time, therefore their anti-inflammatory, anti-fibrotic, and trophic effects are long-lasting. These unique characteristics have led to the administration of Muse cells via intravenous drip in clinical trials for stroke, acute myocardial infarction, epidermolysis bullosa, spinal cord injury, neonatal hypoxic ischemic encephalopathy, amyotrophic lateral sclerosis, and COVID-19 acute respiratory distress syndrome without HLA-matching or immunosuppressive treatment.

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Multilineage-differentiating stress-enduring (Muse) cells are non-tumorigenic pluripotent-like stem cells that exhibit triploblastic differentiation and self-renewability at the single-cell level, and are collectable as pluripotent surface marker SSEA-3(+) from the bone marrow (BM), peripheral blood, and organ connective tissues. SSEA-3(+) cells from human amniotic membrane mesenchymal stem cells (hAMSCs) were compared with hBM-Muse cells. Similar to hBM-Muse cells, hAMSC-SSEA-3(+) cells expressed pluripotency genes (OCT3/4, NANOG, and SOX2), differentiated into triploblastic cells from a single cell, self-renewed, and exhibited non-tumorigenicity. Notably, however, they exhibited unique characteristics not seen in hBM-Muse cells, including higher expression of genes related to germline- and extraembryonic cell-lineages compared with those in hBM-Muse cells in single-cell RNA-sequencing; and enhanced expression of markers relevant to germline- (PRDM14, TFAP2C, and NANOS3) and extraembryonic cell- (CDX2, GCM1, and ID2) lineages when induced by cytokine subsets, suggesting a broader differentiation potential similar to naïve pluripotent stem cells. t-SNE dimensionality reduction and Gene ontology analysis visualized hAMSC-SSEA-3(+) cells comprised a large undifferentiated subpopulation between epithelial- and mesenchymal-cell states and a small mesenchymal subpopulation expressing genes relevant to the placental formation. The AM is easily accessible by noninvasive approaches. These unique cells are a potentially interesting target naïve pluripotent stem cell-like resource without tumorigenicity.

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