The role of ERBB2 and ERBB3 receptors in skin development, homeostasis, and tumor susceptibility

表皮生长因子受体（EGFR/ERBB 1）及其配体是皮肤和毛囊稳态的既定调节剂，它们的失调迅速导致病理学改变。相比之下-虽然他们也在皮肤中表达，并已牵连在皮肤肿瘤-相关的酪氨酸激酶受体ERBB 2和ERBB 3在这个组织中发挥的功能仍然在很大程度上unexplored.We建议产生小鼠品系缺乏ERBB 2或ERBB 3专门在皮肤中。特别是，我们提出了交叉小鼠携带floxed等位基因的Erbb 2或Erbb 3基因与小鼠表达Cre重组酶的控制下的牛角蛋白5启动子（K5 cre）。得到的K5 cre-Erbb 2fl/fl（= K5-Erbb 2del）和K5 cre-Erbb 3fl/fl（= K5-Erbb 3del）小鼠将具有限于毛囊和毛囊间表皮的角质形成细胞的相应受体的早期缺失（从胚胎日e15.5左右开始），从而允许我们评估ERBB 2对皮肤发育和稳态的作用。我们将评估皮肤组织学（H&E，增殖，凋亡），表皮分化定位几个分化标志物，和毛囊形态发生和周期induction.We的动力学也提出了建立小鼠皮肤特异性损失ERBB 2或ERBB 3通过采用K14 creERT 2线和他莫昔芬治疗，并使他们受到两个阶段的化学致癌协议。在该方案中，引发剂DMBA的单次应用之后是促炎剂TPA的重复应用，导致良性乳头状瘤退化或进展为鳞状细胞癌。每周记录肿瘤的数量和大小，并且在实验结束时（DMBA应用后约20周），将肿瘤固定用于组织学分析或冷冻用于表达分析。因此，两阶段化学致癌作用提供了明确阐明ERBB 2和ERBB 3在皮肤癌发展和进展的早期和晚期事件中的影响的机会。最后，一些研究表明，ERBB 2被紫外线照射激活，并增加紫外线诱导的皮肤肿瘤发生。不幸的是，所采用的模型不允许关于ERBB 2在该过程中的作用的明确结论。因此，我们建议将ERBB 2缺陷小鼠和对照同窝仔暴露于UVA或UVB照射。24小时后，我们将评估参数，如表皮厚度，表皮细胞增殖和凋亡，晒伤celles.Finally，允许机制的研究，我们建议建立永久的角质形成细胞系缺乏ERBB 2或ERBB 3以及控制线。利用这些细胞系，我们将评估EGFR/ERBB二聚体的形成和下游信号传导途径、细胞增殖、粘附和非锚定依赖性生长，以及ERBB 2和ERBB 3与E-钙粘蛋白和LRIG 1之间的相互作用。

The epidermal growth factor receptor (EGFR/ERBB1) and its ligands are established regulators of skin and hair follicle homeostasis, and their deregulation rapidly results in pathological alterations. In contrast - although they are also expressed in the skin and have been implicated in skin tumorigenesis - the functions exerted by the related tyrosine kinase receptors ERBB2 and ERBB3 in this tissue remain largely unexplored.We propose to generate mouse lines lacking ERBB2 or ERBB3 specifically in the skin. In particular, we propose to cross mice carrying floxed alleles of the Erbb2 or Erbb3 genes with mice expressing cre recombinase under the control of the bovine keratin 5 promoter (K5cre). Resulting K5cre-Erbb2fl/fl (= K5-Erbb2del) and K5cre-Erbb3fl/fl (= K5-Erbb3del) mice will have an early deletion (starting around embryonic day e15.5) of the corresponding receptors restricted to keratinocytes of the hair follicle and interfollicular epidermis, allowing us to evaluate the role of ERBB2 for skin development and homeostasis. We will evaluate the skin histology (H&E, proliferation, apoptosis), the epidermal differentiation by localizing several differentiation markers, and the kinetics of hair follicle morphogenesis and cycle induction.We also propose to create mice with skin-specific loss of ERBB2 or ERBB3 by employing the K14creERT2 line and tamoxifen treatment and subject them to a two-stage chemical carcinogenesis protocol. In this protocol, a single application of the initiator DMBA is followed by repeated applications of the pro-inflammatory agent TPA, resulting in benign papillomas that regress or progress to squamous cell carcinoma. The number and the size of the tumors will be recorded weekly and, at the end of the experiment (~ 20 weeks after DMBA application), tumors will be fixed for histological analysis or frozen for expression analysis. Thus, the two-stage chemical carcinogenesis provides an opportunity to definitively clarify the influence of ERBB2 and ERBB3 in early and late events in skin cancer development and progression. Finally, several studies showed that ERBB2 is activated by UV irradiation and increases UV-induced skin tumorigenesis. Unfortunately, the employed models did not allow definitive conclusions about the role of ERBB2 in this process. Therefore, we propose to expose ERBB2-deficient mice and control littermates to UVA or UVB irradiation. Twenty-four hours later, we will evaluate parameters such as the epidermal thickness, the epidermal cell proliferation and apoptosis, and the number of sunburn cells.Finally, to allow mechanistic studies, we propose the establishment of permanent keratinocyte cell lines lacking ERBB2 or ERBB3 as well as control lines. With these cell lines, we will evaluate the formation of EGFR/ERBB dimers and the downstream signaling pathways, cell proliferation, adhesion, and anchorage-independent growth, as well as the interaction between ERBB2 and ERBB3 with E-cadherin and LRIG1.