Riassunto analitico
The common neurotrophin receptor CD271, also called p75NTR, is the low-affinity receptor for neurotrophins. Depending on the specific ligand and co-receptor interacting with it, CD271 mediates various cellular responses in human keratinocytes. In detail, CD271 plays a key role during epidermal differentiation. The expression of CD271 is implicated in the transition from keratinocyte stem cells to transient-amplifying cells, which represent the epidermal progenitors.
CD271 is also implicated in skin pathogenesis. In fact, it was found to be absent in psoriasis, a skin pathology characterized by hyperproliferation and reduced apoptosis of keratinocytes, while in cutaneous squamous cell carcinoma (cSCC) its function has not yet been elucidated. Recent studies have shown that the exogenous expression of CD271 in cSCC spheroids derived from cSCC patients increases differentiation and reduces proliferation, as well as the level of markers associated with an aggressive phenotype, independently of stadiation.
Given the role of CD271 in the skin, the aim of our research is to characterize a novel mouse model where the expression of CD271 is absent in keratinocytes. To do so, a novel mouse model, named CD271cKO (hereafter referred to as cKO) in which the deletion is skin-specific, was developed by mating mice carrying the "floxed" CD271 gene with animals expressing Cre recombinase under the control of the KRT14 promoter. Since our previous research identified skin abnormalities starting from postnatal day 1, particular attention was given to this stage.
By the histological analysis of the skin isolated from cKO or WT mice, substantial changes in the skin phenotype were detected. In particular, an increase in epidermal thickness and a higher number of hair follicles were found.
In order to evaluate the potential molecular mechanisms associated with the development of the cKO skin phenotype, a transcriptome analysis of cKO neonatal skin was performed by RNAseq. Data analysis revealed 444 differentially expressed genes in cKO vs WT. To further understand the functions of these genes, a KEGG enrichment pathway analysis was performed through the DAVID GO database. PI3K-AKT, MAPK, and RAS signaling pathways were found being to top significant terms, thus explaining the observed phenotype.
In addition, given that cKO model developed spontaneous lesions at 4 months, with a higher number of inflammatory cells infiltrating the skin, a mouse inflammation array was performed on P1 skin, revealing that cKO produces a higher amount of specific cytokines, such as MIP1a, MIG, and IL6, which has been found upregulated in different tumor microenvironments.
Therefore, to analyze the role of CD271 in skin diseases characterized by hyperproliferation and inflammation, topical treatments with TPA (tetradecanoylphorbol acetate), a tumor promoter, was performed. TPA treatment allows to recapitulate the morphological changes occurring in keratinocytes after an inflammatory stimulus.cKO skin exhibits increased susceptibility to hyperproliferation and inflammation, as indicated by the modulation of KRT6, Ki67, KRT1 and CD45 expression. Similarly, analyses conducted on primary mouse keratinocytes isolated from cKO and WT skin at P1, show increased "activation" and proliferative capacity, as demonstrated by the previously indicated markers.
Globally, these results confirm that the CD271cKO model represents a useful tool for the analysis of skin pathophysiology. The use of the TPA protocol simulates more effectively the development of features of skin diseases characterized by hyperproliferation and inflammation, and demonstrates a major susceptibility of the CD271cKO skin to developing skin diseases, becoming a means for studying the efficacy of therapies.
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