The cellular and molecular microenvironment of epithelial stem and progenitor cells is poorly characterized despite well-documented roles in homeostatic tissue renewal, wound healing, and cancer progression. Here, we demonstrate that, in organotypic cocultures, dermal pericytes substantially enhanced the intrinsically low tissue-regenerative capacity of human epidermal cells that have committed to differentiate and that this enhancement was independent of angiogenesis. We used microarray analysis to identify genes expressed by human dermal pericytes that could potentially promote epidermal regeneration. Using this approach, we identified as a candidate the gene LAMA5, which encodes laminin α5, a subunit of the ECM component laminin-511/521 (LM-511/521). LAMA5 was of particular interest as we had previously shown that it promotes skin regeneration both in vitro and in vivo. Analysis using immunogold localization revealed that pericytes synthesized and secreted LAMA5 in human skin. Consistent with this observation, coculture with pericytes enhanced LM-511/521 deposition in the dermal-epidermal junction of organotypic cultures. We further showed that skin pericytes could also act as mesenchymal stem cells, exhibiting the capacity to differentiate into bone, fat, and cartilage lineages in vitro. This study suggests that pericytes represent a potent stem cell population in the skin that is capable of modifying the ECM microenvironment and promoting epidermal tissue renewal from non-stem cells, a previously unsuspected role for pericytes.

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Although homeostatic renewal of human skin epidermis is achieved by the combined activity of quiescent stem cells (SCs) and their actively cycling progeny, whether these two populations are equipotent in their capacity to regenerate tissue has not been determined in biological assays that mimic lifelong renewal. Using fluorescence activated cell separation strategy validated previously by us, human epidermis was fractionated into three distinct subsets: that is, α 6briCD71dim, α 6briCD71bri, and α 6dim with characteristics of keratinocyte stem, transient amplifying, and early differentiating cells, respectively. The global gene expression profile of these fractions was determined by microarray, confirming that the α 6briCD71dim subset was quiescent, the α 6briCD71bri was actively cycling, and the α 6dim subset expressed markers of differentiation. More importantly, functional evaluation of these populations in an in vivo model for tissue reconstitution at limiting cell dilutions revealed that the quiescent α 6briCD71dim fraction was the most potent proliferative and tissue regenerative population of the epidermis, capable of long‐term (LT) epidermal renewal from as little as 100 cells for up to 10 weeks. In contrast, the cycling α 6briCD71bri fraction was the first to initiate tissue reconstitution, although this was not sustained in the LT, while differentiating α 6dim cells possessed the lowest demonstrable tissue regenerative capacity. Our data suggest that in human skin, the epidermal proliferative compartment is not composed of equipotent cells, but rather is organized in a functionally hierarchical manner with the most potent quiescent SCs at its apex (i.e., α 6briCD71dim) followed by cycling progenitors (i.e., α 6briCD71bri) and finally early differentiating keratinocytes (i.e., α 6dim). STEM CELLS 2011;29:1256–1268

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Purpose: The aim of this study was to investigate the role of pericytes in regulating malignant ovarian cancer progression.Experimental Design: The pericyte mRNA signature was used to interrogate ovarian cancer patient datasets to determine its prognostic value for recurrence and mortality. Xenograft models of ovarian cancer were used to determine if co-injection with pericytes affected tumor growth rate and metastasis, whereas coculture models were utilized to investigate the direct effect of pericytes on ovarian cancer cells. Pericyte markers were used to stain patient tissue samples to ascertain their use in prognosis.Results: Interrogation of two serous ovarian cancer patient datasets [the Australian Ovarian Cancer Study, n ¼ 215; and the NCI TCGA (The Cancer Genome Atlas), n ¼ 408] showed that a high pericyte score is highly predictive for poor patient prognosis. Co-injection of ovarian cancer (OVCAR-5 & -8) cells withpericytes in a xenograft model resulted in accelerated ovarian tumor growth, and aggressive metastases, without altering tumor vasculature. Pericyte co-culture in vitro promoted ovarian cancer cell proliferation and invasion. High aSMA protein levels in patient tissue microarrays were correlated with more aggressive disease and earlier recurrence.Conclusions: High pericyte score provides the best means to date of identifying patients with ovarian cancer at high risk of rapid relapse and mortality (mean progression-free survival time < 9 months). The stroma contains rare yet extremely potent locally resident mesenchymal stem cells-a subset of "cancer-associated fibroblasts" that promote aggressive tumor growth and metastatic dissemination, underlying the prognostic capacity of a high pericyte score to strongly predict earlier relapse and mortality.

Keratins are cytoskeletal intermediate filament proteins that are increasingly being recognised for their diverse cellular functions. Here we report the consequences of germ line inactivation of Keratin 76 (Krt76) in mice. Homozygous disruption of this epidermally expressed gene causes neonatal skin flaking, hyperpigmentation, inflammation, impaired wound healing, and death prior to 12 weeks of age. We show that this phenotype is associated with functionally defective tight junctions that are characterised by mislocalization of the integral protein CLDN1. We further demonstrate that KRT76 interacts with CLDN1 and propose that this interaction is necessary to correctly position CLDN1 in tight junctions. The mislocalization of CLDN1 has been associated in various dermopathies, including the inflammatory disease, psoriasis. These observations establish a previously unknown connection between the intermediate filament cytoskeleton network and tight junctions and showcase Krt76 null mice as a possible model to study aberrant tight junction driven skin diseases.

BackgroundThe establishment and maintenance of polarity is vital for embryonic development and loss of polarity is a frequent characteristic of epithelial cancers, however the underlying molecular mechanisms remain unclear. Here, we identify a novel role for the polarity protein Scrib as a mediator of epidermal permeability barrier acquisition, skeletal morphogenesis, and as a potent tumor suppressor in cutaneous carcinogenesis.MethodsTo explore the role of Scrib during epidermal development, we compared the permeability of toluidine blue dye in wild-type, Scrib heterozygous and Scrib KO embryonic epidermis at E16.5, E17.5 and E18.5. Mouse embryos were stained with alcian blue and alizarin red for skeletal analysis. To establish whether Scrib plays a tumor suppressive role during skin tumorigenesis and/or progression, we evaluated an autochthonous mouse model of skin carcinogenesis in the context of Scrib loss. We utilised Cre-LoxP technology to conditionally deplete Scrib in adult epidermis, since Scrib KO embryos are neonatal lethal.ResultsWe establish that Scrib perturbs keratinocyte maturation during embryonic development, causing impaired epidermal barrier formation, and that Scrib is required for skeletal morphogenesis in mice. Analysis of conditional transgenic mice deficient for Scrib specifically within the epidermis revealed no skin pathologies, indicating that Scrib is dispensable for normal adult epidermal homeostasis. Nevertheless, bi-allelic loss of Scrib significantly enhanced tumor multiplicity and progression in an autochthonous model of epidermal carcinogenesis in vivo, demonstrating Scrib is an epidermal tumor suppressor. Mechanistically, we show that apoptosis is the critical effector of Scrib tumor suppressor activity during skin carcinogenesis and provide new insight into the function of polarity proteins during DNA damage repair.ConclusionsFor the first time, we provide genetic evidence of a unique link between skin carcinogenesis and loss of the epithelial polarity regulator Scrib, emphasizing that Scrib exerts a wide-spread tumor suppressive function in epithelia.Electronic supplementary materialThe online version of this article (doi:10.1186/s12943-015-0440-z) contains supplementary material, which is available to authorized users.

The effects of different doses of oestrogens in constitutionally tall girls were evaluated in two centres for paediatric endocrinology. In one centre, 38 girls were treated with a high oestrogen dose of 0.3 to 0.5 mg ethinyloestradiol (EE) daily. In the other, 44 girls received a comparably low dose of 0.1 mg EE per day. Height prediction (HP), chronological age (CA), and height at the onset of treatment were comparable in both groups. Although the duration of treatment was significantly longer in those receiving the low dose, the cumulative oestrogen dose was still significantly lower. The dose of EE had no effect on final height reduction (high dose group: 4.9 +/- 2.6 cm, low dose group: 5.1 +/- 2.4 cm). Final height was more reduced in both groups when treatment was started at an early bone age (BA) (less than or equal to 13 years). No serious side effects were observed in either group, however weight gain was more pronounced in girls receiving the higher dose. We conclude that treatment of constitutionally tall girls with low doses of oestrogens is equally effective in reducing the final height as the usually administered high doses. The lowest effective dose has to be determined in a randomized, prospective clinical trial.