Moreover, the cost of treatment with skin substitutes is high but the only one function, protective, can be replaced with them [209]: all these tissue-engineered constructs cannot restore thermoregulation, sensation, UV-protection, excretion, perspiration, etc. Nowadays, in the design of skin substitute, there are three main approaches: cell-based, biomaterial-based, and delivery-based. wound and burn healing [41]. Keratinocytes are the major cell component of the epidermis and responsible for its stratified structure and form numerous tight intercellular junctions. Fibroblasts are the main cell type of the dermis and produce ECM components and secrete various growth factors (TGF-), cytokines (TNF-), and matrix metalloproteinases, which ensure the ECM formation and keratinocyte proliferation and differentiation [16]. Commercial products such as Epicel, Cryoskin, and BioSeed-S contain keratinocytes; Dermagraft, TransCyte and Hyalograft 3Dfibroblasts; and Apligraf, Theraskin, and OrCella combination. The use of these cells enables the large-scale production of standardized product batches. However, these materials are mostly non-permanent bioactive dressings, which provide cytokines, ECM, and growth factors for the successful skin reparation [41C43]. Immune rejection is commonly reported with allogeneic fibroblasts and keratinocytes, [44] but this is mostly shown for allogeneic keratinocytes that can be explained by the difference in HLA expression and cytokine production [45]. Fetal fibroblasts are of particular interest because they can significantly improve skin repair due to the high expansion ability, low immunogenicity, and intense secretion of bioactive substances such as basic fibroblast growth factor, vascular endothelial growth factor, and keratinocyte growth factor. However, ethical IL4 issues limit their application [46C49]. Epidermal stem cells (ESC) are of particular interest for skin tissue regeneration as they have favorable features such as high proliferation XL-228 rate and easy access and keep their potency and XL-228 differentiation potential for long periods [65, 82]. They are one of the skin stem cell types, either heterogeneous or autogenous origins (Table?2). ESC are mostly connected to the process of skin regeneration [17]. XL-228 They are rare, infrequently divide and generate short-lived and rapidly dividing cells, which are involved in the regeneration process [65]. Their main population, responsible for skin repair, is located in the basal layer of the epidermis; however, they can also be revealed in the base of sebaceous glands and the bulge region of hair follicles [6, 65, 82]. However, while working with ESC culture, we may face progressive aneuploidy or polyploidy and mutation accumulation after several passages. Moreover, as they can be easily derived from the patients XL-228 skin and transplanted to the same patient, ESC are not restricted by ethical issues. Grafts made up of autologous holoclones ESC have proven to be effective in treating vast skin defects: epidermolysis, skin and ocular burns, etc. [83, 84]. Table 2 Subtypes of skin stem cells references Mesenchymal stromal cells (MSC) have similar (not identical) features as ESC and can be derived from various tissues, even the skin as mentioned previously [98]. They have a high differentiation potential and a certain degree of plasticity and may generate cells of mesodermal, ectodermal, and endodermal lineages [99]. Moreover, paracrine, trophic, and immunomodulatory MSC properties enable their clinical use [100, 101]. MSC can migrate to the injured tissues, differentiate, and regulate the tissue regeneration by the production of growth factors, cytokines, and chemokines [102]. Their immunomodulatory activity is based on the release of anti-inflammatory cytokines and the inhibition of proliferation of CD4+ and CD8+ natural killer cells, T cells, and B cells. MSC are considered to be hypoimmunogenic because they do not express class I and II molecules of the major histocompatibility complex (MHC) and co-stimulatory proteins (e.g., CD40, CD80, CD86). Therefore, the transplantation of allogenic MSC has a low risk of the immune rejection [103C105]. In burn therapy, adipose-derived stromal cells refined from the stromal vascular fraction are widely applied because of their easy access and isolation procedure and inspiring improvement of the healing processes [106C108]. They are showed to preserve their therapeutic effects after freezing that ensures their multiple use [109]. It is worth mentioning that even the freshly isolated stromal vascular fraction is usually showed to be effective in.