Equine Epidermis: A Source of Epithelial-Like Stem/Progenitor Cells with In Vitro and In Vivo Regenerative Capacities

Abstract

Besides the presence of somatic stem cells in hair follicles and dermis, the epidermis also contains a subpopulation of stem cells, reflecting its high regenerative capacity. However, only limited information concerning epidermis-derived epithelial-like stem/progenitor cells (EpSCs) is available to date. Nonetheless, this stem cell type could prove itself useful in skin reconstitution after injury. After harvesting from equine epidermis, the purified cells were characterized as EpSCs by means of positive expression for CD29, CD44, CD49f, CD90, Casein Kinase 2b, p63, and Ki67, low expression for cytokeratin (CK)14 and negative expression for CD105, CK18, Wide CK, and Pan CK. Furthermore, their self-renewal capacity was assessed in adhesion as well as in suspension. Moreover, the isolated cells were differentiated toward keratinocytes and adipocytes. To assess the regenerative capacities of EpSCs, six full-thickness skin wounds were made: three were treated with EpSCs and platelet-rich-plasma (EpSC/PRP-treated), while the remaining three were administered carrier fluid alone (PRP-treated). The dermis of EpSC/PRP-treated wounds was significantly thinner and exhibited more restricted granulation tissue than did the PRP-treated wounds. The EpSC/PRP-treated wounds also exhibited increases in EpSCs, vascularization, elastin content, and follicle-like structures. In addition, combining EpSCs with a PRP treatment enhanced tissue repair after clinical application.

Sphere-Forming Capacity as an Enrichment Strategy for Epithelial-Like Stem Cells from Equine Skin

“…The isolation and characterization of skin-derived EpSCs was performed as previously described [26,9] with some minor modifications. Briefly, horses were sedated with an intravenous injection of detomidine hydrochloride (0.01mg/kg, Medesedan®, VIRBAC, Belgium) and the analgesic butorphanol (0.01mg/kg, Dolorex®, Intervet, Belgium).…”

“…Differentiation of the isolated cells towards keratinocytes was induced using a distinct differentiation protocol for EpSCs developed by Broeckx et al [9]. Briefly, adherent cells from P 4 and P 10 were trypsinized using 0.25% trypsin-EDTA and EpSCs were seeded into 24-well plates at a density of 2.5x10 3 per cm 2 .…”

“…Furthermore, their surprisingly diverse differentiation potential has also contributed to a significant recent interest in adult skin stem cells [8]. Permanently residing EpSCs in mammalian skin are able to differentiate into multiple lineages and have been reported in the epidermis [9], hair follicle [10] as well as dermis [6]. …”

“…Many of the methods developed for the isolation and analysis of specific cell types cannot be used for medical experimentation on living materials because of the damaging nature of these techniques, such as isotope radiation and cell fixation [21]. For isolation of EpSCs, in vitro methods describe the use of (i) colony forming culture to localize multipotent stem cells in the hair follicle bulge region [22], (ii) Hoechst exclusion technique to isolate a side population in the hair follicles [23], (iii) cell sorting using CD71 and α6-integrin expression markers for harvesting keratinocyte stem cells [24] and (v) sphere formation [9]. …”

Mesenchymal Stem Cell Therapy Alleviates Interstitial Cystitis by Activating Wnt Signaling Pathway

“…Epithelial integrity is dynamically regulated via the homeostatic proliferation and differentiation of organ-specific stem and progenitor cells, which generally reside in the basal layer of the epithelium [6,7]. The activity of these cells is dependent on the turnover of these tissues.…”