Defining the role of miR-30 in human skin

定义 miR-30 在人类皮肤中的作用

• 批准号: 8813976
• 负责人: Hasan Mukhtar
• 金额: $ 19.87万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-08 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8813976
• 关键词: 3' Untranslated Regions; 3-Dimensional; Adverse effects; Apoptosis; Basal cell carcinoma; Binding; Bioinformatics; Biological; Biological Assay; Biological Process; Carcinogens; Cell Culture Techniques; Cell Cycle Regulation; Colon Carcinoma; Cutaneous; DNA Damage; Data; Data Analyses; Defense Mechanisms; Dermal; Diagnosis; Diagnostic; Disease; Dose; Down-Regulation; Epidermal Growth Factor; Evaluation; Fibroblasts; Formalin; Functional RNA; Gene Expression Regulation; Gene Mutation; Gene Targeting; Genes; Hepatocyte Growth Factor; Human; Immune response; In Situ Hybridization; In Vitro; Light; Luciferases; Malignant Neoplasms; Malignant neoplasm of lung; Malignant neoplasm of urinary bladder; Mediating; Messenger RNA; MicroRNAs; Modeling; Oncogenic; Paraffin Embedding; Pathogenesis; Pathway interactions; Patients; Pattern; Prevention; Prognostic Marker; Proteins; RNA; Reaction; Reporter; Role; Series; Signal Transduction; Signal Transduction Pathway; Simulate; Site; Skin; Skin Cancer; Skin Tissue; Specimen; System; Techniques; Testing; Thyroid carcinoma; Time; Tissue Sample; Tissues; Transcriptional Regulation; UV induced; UVB induced; Ultraviolet B Radiation; Ultraviolet Rays; Validation; Western Blotting; carcinogenesis; cell growth; epithelial to mesenchymal transition; gene function; in vivo; irradiation; keratinocyte; loss of function; mRNA Expression; malignant breast neoplasm; monolayer; novel strategies; public health relevance; research study; response; skin disorder; skin morphogenesis; ultraviolet; ultraviolet irradiation

DESCRIPTION (provided by applicant): Solar ultraviolet (UV) radiation is a major environmental skin carcinogen that induces DNA damage and modulates a variety of genes that regulate cell growth, proliferation and apoptosis. The transcriptional regulation of genes is part of the cellular reaction that operates as a defense mechanism against the adverse effects of UV radiation. MicroRNAs (miRNA) are a group of small non-coding RNAs which regulate gene functions by targeting sequences in their 3' untranslated regions. Disruption of miRNA expression has been observed in various malignancies including skin cancers. Very little is known about the role of the miRNAs in the regulation of gene expression in response to UV irradiation in human skin. We generated a miRNA profile of UVB-irradiated normal human epidermal keratinocytes (NHEKs) selecting the physiologically relevant UVB dose of 40mJ/cm2. In our preliminary microarray experiments, we identified a subset of 44 miRNAs that were significantly (p< 0.05) differentially expressed in NHEKs, 4 h post UVB exposure. This data was further validated by qPCR which revealed a total of 22 miRNAs that were modulated by UVB. Additional statistical testing showed miR-30, miR-24 and miR-222 to be significantly modulated in both experimental systems. Studies in 3-D epidermal constructs exposed to UVB irradiation confirmed monolayer cell culture findings and demonstrated significant downregulation of miR-30. Our data indicate a potential role of miR-30 in the UVB exposed human skin. The hypothesis we test here is that UVB-mediated decrease in miR-30 expression results in loss of inhibitory control of proliferative pathways implicated in the pathogenesis of human skin cancer. After establishing the involvement of miR-30 in UVB induced skin response, we will shortlist the target genes of miR-30, employing mRNA microarray platform and computational target prediction sites. Functional studies will be conducted to verify miR-30 targets with potential role in UV-induced response using miRNA gain- and loss-of-function experiments. These results will be validated in a multilayered, highly differentiated, 3-D human epidermal skin model closely simulating human skin. For in vivo relevance of our in vitro data, we will examine miR-30 and its target genes in human skin specimens and study a possible correlation between miR-30 expression levels and the occurrence of skin cancer. Our study will provide in-depth understanding of the functions of miR-30 in the human skin and delineate its role in UV- induced skin cancer.

描述（由申请人提供）：太阳能紫外线（UV）辐射是一种主要的环境皮肤致癌物，可诱导DNA损伤并调节各种调节细胞生长，增殖和凋亡的基因。基因的转录调节是细胞反应的一部分，该反应是针对紫外线辐射的不利影响的防御机制。 microRNA（miRNA）是一组小的非编码RNA，通过靶向其3'未翻译区域中的序列来调节基因功能。在包括皮肤癌在内的各种恶性肿瘤中已经观察到miRNA表达的破坏。关于miRNA在人类皮肤中紫外线辐射的调节中的作用知之甚少。我们生成了一个UVB辐射的正常人表皮角质形成细胞（NHEKS）的miRNA谱，选择了40MJ/cm2的生理相关的UVB剂量。在我们的初步微阵列实验中，我们确定了44个miRNA的子集，这些miRNA在NHEKS中显着表达（p <0.05），在UVB暴露后4小时。 QPCR进一步验证了该数据，该数据揭示了由UVB调节的22个miRNA。其他统计测试表明，在两个实验系统中，miR-30，miR-24和miR-222均受到显着调节。暴露于UVB辐照的3-D表皮构建体中的研究证实了单层细胞培养结果，并显示了miR-30的显着下调。我们的数据表明miR-30在UVB暴露的人皮肤中的潜在作用。我们在这里检验的假设是，UVB介导的miR-30表达降低导致抑制与人皮肤癌发病机理有关的增生途径的抑制作用。在建立miR-30参与UVB引起的皮肤反应之后，我们将使用mRNA微阵列平台和计算目标预测位点入围miR-30的靶基因。将进行功能研究以验证具有潜在作用的miR-30目标 在使用miRNA增益和功能丧失实验的紫外线诱导响应中。这些结果将在多层，高度分化的3-D人表皮皮肤模型中得到验证，以密切模拟人类皮肤。对于我们的体外数据的体内相关性，我们将检查miR-30及其在人体皮肤标本中的靶基因，并研究miR-30表达水平与皮肤癌的发生之间的可能相关性。我们的研究将对miR-30在人皮肤中的功能进行深入了解，并描述其在紫外线诱导的皮肤癌中的作用。