Novel Transcripts of EMT Driving the Malignant Transformation of Oral Submucous Fibrosis [Whole transcriptome]

OSF is a chronic, progressive, fibrotic condition of the oral mucosa that carries an elevated risk of undergoing malignant transformation. We aimed to identify and validate novel genes associated with the regulation of epithelial to mesenchymal transition (EMT) in Oral submucous fibrosis (OSF). Genes regulating EMT were identified through differential gene expression analysis, with a LogFC threshold of -1 and +1, and padj value < 0.05, utilizing data retrieved from GEO datasets, TCGA-HNSC dataset and whole transcriptome data generated from tissue samples of OSF, OSFSCC (OSF associated with OSCC) and OSCC (Oral squamous cell carcinoma). The curated EMT genes were correlated with functional states of cancer, subjected to clustering to identify the candidate genes. The EMT genes, namely MMP9, SPARC and ITGA5 were identified to be the novel candidate genes. Comprehensive pathway analysis and immunohistochemical analysis confirmed their role in regulating EMT in OSF, OSCC, and OSFSCC. Integration of bioinformatics and proteomics led to the discovery of novel candidate genes regulating EMT. The significant role of candidate genes MMP9, SPARC, and ITGA5 observed in fibrosis and malignancy indicates a novel mechanism of transition from fibrosis associated type 2 EMT to type 3 EMT, driving OSF to malignancy. Whole transcriptome sequencing was performed for normal(NOM) (n=2), oral submucous fibrosis (OSF)(n=2) and oral squamous cell carcinoma(OSCC) (n=2) with the parameters of 2x150bp, 60M reads/sample on HiSeq X Ten/Novaseq.

口腔黏膜下纤维化（Oral submucous fibrosis, OSF）是一种慢性、进展性的口腔黏膜纤维化疾病，且存在较高的恶性转化风险。本研究旨在识别并验证与OSF中上皮-间质转化（Epithelial to Mesenchymal Transition, EMT）调控相关的新型基因。研究人员通过差异基因表达分析，以LogFC阈值为-1和+1、padj值<0.05为筛选标准，结合从GEO数据集、TCGA-HNSC数据集以及OSF、OSFSCC（OSF associated with OSCC，即OSF相关性口腔鳞状细胞癌）、口腔鳞状细胞癌（Oral squamous cell carcinoma, OSCC）的组织样本中获取的全转录组数据，识别出调控EMT的基因；随后将筛选得到的EMT相关基因与癌症功能状态进行关联分析，并通过聚类算法识别候选基因，最终确定MMP9、SPARC及ITGA5为新型候选调控基因。全面的通路分析与免疫组化分析证实了这三个基因在OSF、OSCC及OSFSCC的EMT调控中发挥关键作用。通过生物信息学与蛋白质组学的整合分析，本研究发现了一批调控EMT的新型候选基因。候选基因MMP9、SPARC与ITGA5在纤维化病变与恶性转化过程中的显著作用，提示了一条全新的机制：由纤维化相关的2型EMT向3型EMT转变，进而推动OSF向恶性表型进展。本研究对正常口腔黏膜（normal oral mucosa, NOM，n=2）、OSF（n=2）及OSCC（n=2）的组织样本开展全转录组测序，测序参数为2×150bp、单样本测序reads数为60M，测序平台为HiSeq X Ten/Novaseq。