Tissue-engineered dermal substitutes constructed by human embryonic stem cell-derived fibroblasts facilitate the repair of skin wounds.

Tissue engineering is an efficient method for constructing functional skin equivalents to treat large-area skin wounds. However, the source of seed cells is limited. Human embryonic stem cell (hESC) derivatives play an important role in tissue engineering. However, whether hESC-derived fibroblasts can be used as seed cells of tissue-engineered dermal substitutes has not been reported. In this study, we bypassed traditional induction methods, but rather directly induced chHES-8 cells to differentiate into mesenchymal stem cells (hESC-MSCs) without going through the embryoid body (EB) stage. hESC-MSCs were enriched and treated with connective tissue growth factor (CTGF) to induce their differentiation into fibroblasts (hESC-Fbs). Finally, we seeded hESC-Fbs into collagen gels to construct tissue-engineered dermal substitutes and evaluated their efficacy in treating skin wound in vivo. The hESCs line chHES-8 expressed ALP, NANOG, OCT3/4, and SSEA-3 and formed teratomas containing three germ layers. FACS analysis results showed that hESC-MSCs expressed mesenchymal cell surface markers, including CD29, CD44, and CD105, but not CD34 or CD45, and these cells could be induced to differentiate into adipogenic, osteogenic, and chondrogenic lineages. qRT-PCR and ELISA assays showed that hESC-Fbs expressed high levels of growth factors (including FGF and TGF-β1), extracellular matrix components (such as COL-III, MMP-1, and FN), and the interstitial cell marker vimentin (VIM), similar to human skin fibroblasts (HSF). The mouse skin wounds were successfully repaired by day 20 post-transplantation of tissue-engineered dermal substitutes derived from hESC-Fbs. hESCs could be directly induced to differentiate into fibroblasts, which could be applied to the construction of tissue-engineered dermal substitutes and skin defect repair. Moreover, this EB-free induction strategy could offer significant advantages for clinical translation, including higher efficiency and better purity.