Riassunto analitico
The common neurotrophin receptor CD271, also called p75NTR, is the transmembrane low-affinity receptor for neurotrophins, belonging to the TNF superfamily. By interacting with various ligands and partners to form multimeric complexes, it is able to mediate different cellular responses, including both apoptosis and cell survival. In the skin, CD271 is involved in the differentiation process and seems like to be implicated in cutaneous pathogenesis, including psoriasis and squamous cell carcinoma. So far, different animal models have been developed to obtain null mutations of CD271, however they defy to explain CD271 precise role in keratinocytes.We developed an epidermal-specific inducible mouse model, hereafter named CD271iKO, where the expression of Cre recombinase is under the control of the keratin (KRT) 14 promoter. Three weeks CD271iKO mice were topically treated with Tamoxifen (TAM) or the vehicle for 5 consecutive days in order to promote CD271 KO in keratinocytes. Mouse skin was analysed at different time points (5, 15, and 30 days after treatment) to determine the effect of CD271 deletion. Skin abnormalities, including thickness and hair loss, were observed. Histological analysis of the CD271iKO mice demonstrates phenotypic changes in the skin, including epidermal hyperplasia and alteration of keratinocyte cell morphology. TAM-treated CD271iKO dermis showed an increase in the number of hair follicles (HF) as compared to control. Deletion of CD271 showed increased proliferation and activation of keratinocytes, as indicated by increased expression of markers such as KRT5, Ki67, and KRT6. Moreover, altered differentiation and increased inflammation were observed through changes in KRT1, Filaggrin, and CD45 expression. Given such phenotype, we analyze the expression of proteins belonging to the mitogenic pathways in CD271iKO skin, finding a significant upregulation in all the different time-points. In particular, CD271 deletion provokes a strong downregulation of DLX3, which is crucial for keratinocyte differentiation and cell death regulation, as indicated by the analysis of markers such as KRT10 and Filaggrin. The potential axis between CD271 and DLX3 is also indicated by the alteration of HF development, where DLX3 plays a critical role. In fact, CD271 deletion also leads to the downregulation of WNT5A, which is necessary for HF differentiation and demonstrated being an upstream regulator of DLX3. To confirm these data, we performed additional analyses on primary mouse keratinocytes (PMK) extracted from the skin of p75NTRFlox mice and subsequently treated with a vector expressing the Cre recombinase, using GFP-only expressing vector as control. We found that CD271 deletion promotes an increase in proliferation as compared to the control and CD271-KO PMK also showed misregulation of the previously mentioned pathways and markers. The CD271iKO mouse model developed and characterized in this study will be a useful tool for investigating the role of the NTs network in the skin, both in healthy and pathological conditions. It will clarify the involvement of CD271 in hyperproliferative skin disease, including susceptibility to skin cancer development.
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