Role of PGE2 Receptors in Mouse Skin Cancer

PGE2 受体在小鼠皮肤癌中的作用

• 批准号: 8553785
• 负责人: Robert Langenbach
• 金额: $ 42.94万
• 依托单位: NATIONAL INSTITUTE OF ENVIRONMENTAL HEALTH SCIENCES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8553785
• 关键词: Affect; Agonist; Agreement; Alcohols; Alprostadil; Apoptosis; CREB1 gene; Cell Surface Receptors; Cell Survival; Chemoprevention; Complex; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Data; Development; Dinoprostone; Dose; Epidermal Growth Factor Receptor; Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor; Epidermis; Family; G-Protein-Coupled Receptors; GTP-Binding Proteins; Goals; Indomethacin; Investigation; MAPK3 gene; Malignant Neoplasms; Measures; Mediating; Modeling; Mus; PKA inhibitor; PTGS2 gene; Pathway interactions; Play; Production; Prostaglandin-Endoperoxide Synthase; Receptor Activation; Receptor Inhibition; Regulation; Reporting; Role; STAT3 gene; Signal Pathway; Signal Transduction; Skin; Skin Cancer; Skin Neoplasms; Therapeutic; Time; Topical application; Transducers; UVB induced; Ultraviolet B Radiation; Wild Type Mouse; arrestin B; butaprost; carcinogenesis; celecoxib; chemotherapy; cyclooxygenase 1; human WFDC2 protein; inhibitor/antagonist; keratinocyte; protective effect; receptor; response; survivin; transcription factor; tumor

We previously reported that the prostaglandin E2 (PGE2) G protein coupled receptor (GPCR), EP2, plays important roles in mouse skin tumor development using the initiation promotion model (Chun et al., Carcinogenesis, 30:1620, 2009). Because keratinocyte proliferation is essential for skin tumor development, EP2-mediated signaling pathways that could contribute to keratinocyte proliferation were investigated. A single topical application of the EP2 agonist, butaprost, dose-dependently increased keratinocyte replication and activated EGFR and PKA; and inhibition of either EGFR or PKA (AG1478 or H89, respectively) decreased butaprost-induced keratinocyte proliferation by 70%. Because previous studies indicated that GPCR activation of EGFR involved a b-arrestin1/Src complex, the possibility of EP2 acting via this mechanism in keratinocytes was investigated. It was observed that butaprost increased Src and EGFR activation and induced b-arrestin1/Src complex formation. b-arrestin1-deficiency reduced Src and EGFR activation, confirming b-arrestin1s role in their activation. In agreement with b-arrestin1 contributing to Src and EGFR activation, studies with Src and EGFR inhibitors (PP2 and AG1478, respectively) indicated Src to be upstream of EGFR. Butaprost also induced the activation of Akt, ERK1/2, and STAT3; and either b-arrestin1-deficiency or EGFR inhibition decreased their activation, confirming roles for b-arrestin1 and EGFR in their activation. In addition to EGFR activation, butaprost also increased cAMP levels and PKA activation, as measured by p-GSK3b and p-CREB formation. The PKA inhibitor, H89, decreased butaprost-induced GSK3b, CREB, and ERK activation, but did not affect EGFR activation. Thus, our recent results indicate that EP2 contributed to mouse keratinocyte proliferation by G protein independent, barrestin1 dependent activation of EGFR, and G protein dependent activation of PKA. In addition to EP2 contributing to skin tumor formation in the initiation/promotion model (Chun et al., Carcinogenesis, 30:1620, 2009), we previously reported that EP2 played a role in an anti-apoptotiic response in UVB-exposed mouse skin (Chun et al. Cancer Res., 67:2015, 2007). Because survivin is a regulator of cell survival, the possible regulation of survivin by COX-2 and EP2 in UVB-exposed mouse skin was investigated. In wild type mouse skin UVB time-dependently increased the levels of COX-2, survivin and phosphorylated-signal transducer and activator of transcription3 (p-STAT3), a transcription factor that regulates survivin expression; and COX-2- or EP2-deficiency significantly reduced their induction. Topical application of the COX-2 selective inhibitor, celecoxib, also reduced UVB-induced survivin levels. To further investigate the roles of PGE2 and EP2 in the regulation of survivin, indomethacin was used to inhibit UVB-induced endogenous PG production. Indomethacin reduced UVB-induced survivin levels, while PGE2 and the EP2 agonist, butaprost, partially restored survivin levels. The epidermal growth factor receptor (EGFR) is a downstream effector of EP2 and inhibition of EGFR (AG1478) significantly reduced UVB induction of survivin and activation of STAT3. UVB-induced epidermal apoptosis in COX-2-/- mice was reduced by butaprost and EGFR inhibition blocked butaprosts protective effects. Furthermore, butaprost in the absence of UVB exposure time-dependently increased p-EGFR, p-STAT3 and survivin levels in nave mouse skin, whereas the EP4 agonist, PGE1 alcohol, did not significantly increase p-STAT3 or survivin levels. These data suggest that COX-2-generated PGE2 regulates survivin expression in mouse skin via an EP2-mediated EGFR/STAT3 pathway, and inhibiting the EP2/survivin pathway may provide a therapeutic strategy for the chemoprevention/chemotherapy of UVB-induced skin cancer. Recent studies have indicated that B-arrestin1 deficiency reduced skin tumor multiplicity and tumor size in the initiation/ promotion model, whereas B-arrestin2 deficiency increased tumor multiplicity and tumor sizes. The differing effects of B-arrestin1/2 deficiency are currently being investigated.

我们之前报道过前列腺素 E2 (PGE2) G 蛋白偶联受体 (GPCR) EP2 在启动促进模型中在小鼠皮肤肿瘤发展中发挥重要作用 (Chun 等人, Carcinogenesis, 30:1620, 2009)。由于角质形成细胞增殖对于皮肤肿瘤的发展至关重要，因此研究了可能有助于角质形成细胞增殖的 EP2 介导的信号通路。单次局部应用 EP2 激动剂布他前列素，可剂量依赖性地增加角质形成细胞复制并激活 EGFR 和 PKA；抑制 EGFR 或 PKA（分别为 AG1478 或 H89）可使布他前列素诱导的角质形成细胞增殖减少 70%。因为之前的研究表明 EGFR 的 GPCR 激活涉及 b-arrestin1/Src 复合物，所以研究了 EP2 通过这种机制在角质形成细胞中发挥作用的可能性。据观察，布他前列素增加了 Src 和 EGFR 的激活并诱导 b-arrestin1/Src 复合物的形成。 b-arrestin1 缺陷减少了 Src 和 EGFR 的激活，证实了 b-arrestin1 在其激活中的作用。与 b-arrestin1 促进 Src 和 EGFR 激活一致，Src 和 EGFR 抑制剂（分别为 PP2 和 AG1478）的研究表明 Src 位于 EGFR 的上游。布他前列素还诱导 Akt、ERK1/2 和 STAT3 的激活； b-arrestin1 缺陷或 EGFR 抑制都会降低其激活，证实了 b-arrestin1 和 EGFR 在其激活中的作用。除了 EGFR 激活之外，布他前列素还增加了 cAMP 水平和 PKA 激活（通过 p-GSK3b 和 p-CREB ​​形成测量）。 PKA 抑制剂 H89 减少了布他前列素诱导的 GSK3b、CREB ​​和 ERK 激活，但不影响 EGFR 激活。因此，我们最近的结果表明，EP2 通过 G 蛋白独立的、barrestin1 依赖性的 EGFR 激活和 G 蛋白依赖性的 PKA 激活，促进小鼠角质形成细胞增殖。 除了 EP2 在起始/促进模型中促进皮肤肿瘤形成之外 (Chun et al., Carcinogenesis, 30:1620, 2009)，我们之前报道过 EP2 在 UVB 暴露小鼠皮肤的抗凋亡反应中发挥作用 (Chun et al. Cancer Res., 67:2015, 2007)。由于生存素是细胞存活的调节剂，因此研究了 COX-2 和 EP2 在 UVB 暴露的小鼠皮肤中对生存素的可能调节作用。在野生型小鼠皮肤中，UVB 时间依赖性地增加了 COX-2、生存素和磷酸化信号转导器和转录激活剂 3 (p-STAT3)（一种调节生存素表达的转录因子）的水平。 COX-2-或EP2-缺陷显着降低了它们的诱导。局部应用 COX-2 选择性抑制剂塞来考昔也可降低 UVB 诱导的生存素水平。为了进一步研究PGE2和EP2在生存素调节中的作用，使用吲哚美辛抑制UVB诱导的内源性PG产生。吲哚美辛降低了 UVB 诱导的生存素水平，而 PGE2 和 EP2 激动剂布他前列素则部分恢复了生存素水平。表皮生长因子受体 (EGFR) 是 EP2 的下游效应子，抑制 EGFR (AG1478) 可显着降低 UVB 对生存素的诱导和 STAT3 的激活。布他前列素可减少 COX-2-/- 小鼠中 UVB 诱导的表皮细胞凋亡，并且抑制 EGFR 会阻断布他前列素的保护作用。此外，在没有 UVB 暴露的情况下，布他前列素会随时间依赖性地增加小鼠皮肤中 p-EGFR、p-STAT3 和生存素的水平，而 EP4 激动剂、PGE1 酒精则不会显着增加 p-STAT3 或生存素的水平。这些数据表明COX-2产生的PGE2通过EP2介导的EGFR/STAT3途径调节小鼠皮肤中生存素的表达，抑制EP2/生存素途径可能为UVB诱导的皮肤癌的化学预防/化疗提供治疗策略。 最近的研究表明，B-arrestin1 缺乏在启动/促进模型中降低了皮肤肿瘤的多样性和肿瘤大小，而 B-arrestin2 缺乏则增加了肿瘤的多样性和肿瘤大小。目前正在研究 B-arrestin1/2 缺陷的不同影响。