Preferential Hematopoietic Differentiation in Induced Pluripotent Stem Cells Derived From Human Umbilical Cord Arterial Endothelial Cells.

BACKGROUND: The major obstacle for applications of human induced pluripotent stem cells (hiPSCs) is efficient and controlled lineage-specific differentiation. Hence, a deeper understanding of the initial populations of hiPSCs is required to instruct proficient lineage commitment. METHODS: hiPSCs were generated from somatic cells by transduction of 4 human transcription factors (OCT4, SOX2, KLF4, and C-MYC) using Sendai virus vectors. Genome-wide DNA methylation analysis and transcriptional analysis were performed to evaluate the pluripotent capacity and somatic memory state of hiPSCs. Flow cytometric analysis and colony assays were performed to assess the hematopoietic differentiation capacity of hiPSCs. RESULTS: Here, we reveal human umbilical arterial endothelial cell-derived induced pluripotent stem cells (HuA-iPSCs) exhibit indistinguishable pluripotency in comparison with human embryonic stem cells and hiPSCs derived from other tissues of origin (umbilical vein endothelial cells, cord blood, foreskin fibroblasts, and fetal skin fibroblasts). However, HuA-iPSCs retain a transcriptional memory typical of the parental human umbilical cord arterial endothelial cells, together with a strikingly similar DNA methylation signature to umbilical cord blood-derived induced pluripotent stem cells that distinguishes them from other human pluripotent stem cells. Ultimately, HuA-iPSCs are most efficient in targeted differentiation toward hematopoietic lineage among all human pluripotent stem cells based on the functional and quantitative evaluation of both flow cytometric analysis and colony assays. Application of the Rho-kinase activator significantly reduces the effects of preferential hematopoietic differentiation in HuA-iPSCs, reflected in CD34+ cell percentage of day 7, hematopoietic/endothelial-associated gene expression, and even colony-forming unit numbers. CONCLUSIONS: Collectively, our data suggest that somatic cell memory may predispose HuA-iPSCs to differentiate more amenably into hematopoietic fate, bringing us closer to generating hematopoietic cell types in vitro from nonhematopoietic tissue for therapeutic applications.

An Active Fraction of Trillium tschonoskii Promotes the Regeneration of Intestinal Epithelial Cells After Irradiation.

Despite significant scientific advances toward the development of safe and effective radiation countermeasures, no drug has been approved for use in the clinic for prevention or treatment of radiation-induced acute gastrointestinal syndrome (AGS). Thus, there is an urgent need to develop potential drugs to accelerate the repair of injured intestinal tissue. In this study, we investigated that whether some fractions of Traditional Chinese Medicine (TCM) have the ability to regulate intestinal crypt cell proliferation and promotes crypt regeneration after radiation. By screening the different supplements from a TCM library, we found that an active fraction of the rhizomes of Trillium tschonoskii Maxim (TT), TT-2, strongly increased the colony-forming ability of irradiated rat intestinal epithelial cell line 6 (IEC-6) cells. TT-2 significantly promoted the proliferation and inhibited the apoptosis of irradiated IEC-6 cells. Furthermore, in a small intestinal organoid radiation model, TT-2 promoted irradiated intestinal organoid growth and increased Lgr5+ intestinal stem cell (ICS) numbers. More importantly, the oral administration of TT-2 remarkably enhanced intestinal crypt cell proliferation and promoted the repair of the intestinal epithelium of mice after abdominal irradiation (ABI). Mechanistically, TT-2 remarkably activated the expression of ICS-associated and proliferation-promoting genes and inhibited apoptosis-related gene expression. Our data indicate that active fraction of TT can be developed into a potential oral drug for improving the regeneration and repair of intestinal epithelia that have intestinal radiation damage.

Me6TREN targets ß-catenin signaling to stimulate intestinal stem cell regeneration after radiation.

Background: Acute gastrointestinal syndrome (AGS) is one of the most severe clinical manifestations after exposure to high doses of radiation, and is life-threatening in radiological emergency scenarios. However, an unmet challenge is lacking of an FDA-approved drug that can ameliorate the damage of radiation-exposed intestinal tissues and accelerate the regeneration of injured epithelia. In this study, we investigated whether the small molecule Me6TREN (Me6) can regulate intestinal stem cell (ISC) proliferation and promote crypt regeneration after irradiation. Methods: Lethally irradiated mice were administered with Me6 or PBS to study the survival rate, and sections of their small intestine were subjected to immunostaining to evaluate epithelial regeneration. An intestinal organoid culture system was employed to detect the role of Me6 in organoid growth and ISC proliferation. We further investigated the key signaling pathways associated with Me6 using microarray, western blotting, and RNA interference techniques. Results: We identified the small molecule Me6 as a potent intestinal radiation countermeasure. Systemic administration of Me6 significantly improved ISC and crypt cell regeneration and enhanced the survival of mice after high doses of radiation. Using an in vitro intestinal organoid culture system, we found that Me6 not only induced ISC proliferation but also increased the budding rate of intestinal organoids under unirradiated and irradiated conditions. Me6 remarkably activated the expression of ISC-associated and proliferation-promoting genes, such as Ascl2, Lgr5, Myc, and CyclinD1. Mechanistically, Me6 strongly stimulated the phosphorylation of ß-catenin at the S552 site and increased the transcriptional activity of ß-catenin, a key signaling pathway for ISC self-renewal and proliferation. This is further evidenced by the fact that knockdown of ß-catenin abolished the effect of Me6 on intestinal organoid growth in vitro and crypt regeneration in irradiated mice. Conclusion: The small molecule Me6TREN induced ISC proliferation, enhanced intestinal organoid growth in vitro, and promoted intestinal tissue regeneration after radiation injury by activating ß-catenin signaling.