A Stable Niche Supports Long-Term Maintenance of Human Epidermal Stem Cells in Organotypic Cultures

Abstract

Stem cells in human interfollicular epidermis are still difficult to identify, mainly because of a lack of definitive markers and the inability to label human beings for label-retaining cells (LRCs). Here, we report that LRCs could be identified and localized in organotypic cultures (OTCs) made with human cells. Labeling cultures for 2 weeks with iododeoxyuridine (IdU) and then chasing for 6 -10 weeks left <1% of basal cells retaining IdU label. Whole mounts demonstrated that LRCs were individually dispersed in the epidermal basal layer. Some LRCs, but not all, colocalized with cells expressing melanoma chondroitin sulfate proteoglycan, a putative stem cell marker. Although we found LRCs in both collagen-and scaffold-based OTCs, only the scaffold-OTCs supported long-term survival and regeneration. LRCs ' short survival in collagen-OTCs was not due to loss of appropriate growth factors from fibroblasts. Instead, it was due to expression of metalloproteinases, especially matrix metalloproteinase (MMP)-2 and MMP-14, which caused collagen fragmentation, matrix degradation, and dislocation of specific basement membrane components bound to epidermal integrins. Blocking MMP activation not only abrogated MMP-dependent matrix degradation but also increased longevity of the epidermis and the LRCs in these cultures. Such findings indicate that the stem cell niche, the microenvironment surrounding and influencing the stem cell, is essential for stem cell survival and function, including long-term tissue regeneration.

Modeling the Prostate Stem Cell Niche: An Evaluation of Stem Cell Survival and Expansion In Vitro

“…In the prostate, monolayer culture provides the most useful model so far for the expansion of adult PSCs. Nonetheless, alternative strategies are still required to study the stem cell niche, such as those used to model skin [44,45]. Organotypic cultures of stroma and keratinocytes in 2D have been developed that are able to maintain label retaining cells for over 10 weeks, reminiscent of vivo-like architecture.…”

Keratinocyte stem cells: Friends and foes

“…Actually, one could comment that such a random distribution throughout the entire basal layer better fulfills the tasks of IF stem cells that would thus be ready to reconstitute epidermis wherever it is needed. While stem cells should reside in protected locations, factors, adhesion molecules, and cells participating in the niche are evenly expressed throughout the epidermis and not just at the tip or in the upper part of the rete ridges (Muffler et al, 2008).…”

Nuevos modelos experimentales para el estudio de la homeostasis y la enfermedad cutánea

“…Las principales desventajas de los cultivos organotípicos son: su corta duración y la ausencia de gran parte de los componentes celulares mesenquimáticos presentes en la piel natural. Aunque la utilización del ácido hialurónico como un componente adicional de la matriz ha permitido prolongar la vida útil de los equivalentes cutáneos con matrices de fibrina 42 , los modelos in vitro de piel bioingenierizada no son los adecuados para estudios o ensayos terapéuticos de larga duración, que requieren un recambio celular continuo a expensas de ciclos de proliferación-diferenciación a partir de células madre o de progenitores epidérmicas tempranos. En estos casos, es imprescindible recurrir a los modelos in vivo de ratón con piel humanizada.…”