ABSTRACT
SOX2 is a transcription factor essential for self-renewal and pluripotency of embryonic stem cells. Recently, SOX2 was found overexpressed in the majority of the lung squamous cell carcinoma (SQC), in which it acts as a lineage-survival oncogene. However, downstream targets/pathways of SOX2 in lung SQC cells remain to be identified. Here, we show that BMP4 is a downstream target of SOX2 in lung SQC. We found that SOX2-silencing-mediated inhibition of cell growth was accompanied by upregulation of BMP4 mRNA and its protein expression. Meta-analysis with 293 samples and qRT-PCR validation with 73 clinical samples revealed an inversely correlated relationship between levels of SOX2 and BMP4 mRNA, and significantly lower mRNA levels in tumor than in adjacent normal tissues. This was corroborated by immunohistochemistry analysis of 35 lung SQC samples showing lower BMP4 protein expression in tumor tissues. Cell-based experiments including siRNA transfection, growth assay and flow cytometry assay, further combined with a xenograft tumor model in mice, revealed that reactivation of BMP4 signaling could partially account for growth inhibition and cell cycle arrest in lung SQC cells upon silencing SOX2. Finally, chromatin immunoprecipitation analysis and luciferase reporter assay revealed that SOX2 could negatively regulate BMP4 promoter activity, possibly through binding to the promoter located in the first intron region of BMP4. Collectively, our findings suggest that BMP4 could act as a tumor suppressor and its downregulation by elevated SOX2 resulting in enhanced growth of lung SQC cells.
Subject(s)
Bone Morphogenetic Protein 4/metabolism , Carcinoma, Non-Small-Cell Lung/metabolism , Carcinoma, Squamous Cell/metabolism , Lung Neoplasms/metabolism , SOXB1 Transcription Factors/metabolism , Animals , Carcinoma, Non-Small-Cell Lung/genetics , Carcinoma, Squamous Cell/genetics , Databases, Genetic , Down-Regulation , Gene Expression Profiling , Gene Expression Regulation, Neoplastic , Gene Silencing , Humans , Lung/metabolism , Lung Neoplasms/genetics , Mice , Neoplasm Transplantation , Oligonucleotide Array Sequence Analysis , Promoter Regions, Genetic , Signal TransductionABSTRACT
PURPOSE: SOX9 is an important transcription factor required for development and has been implicated in several types of cancer. However, SOX9 has never been linked to lung cancer to date. Here, we show that SOX9 expression is upregulated in lung adenocarcinoma and show how it is associated with cancer cell growth. EXPERIMENTAL DESIGN: Data mining with five microarray data sets containing 490 clinical samples, quantitative reverse transcription-PCR validation assay in 57 independent samples, and immunohistochemistry assay with tissue microarrays containing 170 lung tissue cores were used to profile SOX9 mRNA and protein expression. Short interference RNA suppression of SOX9 in cell lines was used to scrutinize functional role(s) of SOX9 and associated molecular mechanisms. RESULTS: SOX9 mRNA and protein were consistently overexpressed in the majority of lung adenocarcinoma. Knockdown of SOX9 in lung adenocarcinoma cell lines resulted in marked decrease of adhesive and anchorage-independent growth in concordance with the upregulation of p21 (CDKN1A) and downregulation of CDK4. In agreement with higher SOX9 expression level in lung adenocarcinoma, the p21 mRNA level was significantly lower in tumors than that in normal tissues, whereas the opposite was true for CDK4, supporting the notion that SOX9 negatively and positively regulated p21 and CDK4, respectively. Finally, whereas SOX9-knockdown cells showed significantly attenuated tumorigenicity in mice, SOX9 transfectants consistently showed markedly stronger tumorigenicity. CONCLUSIONS: Our data suggest that SOX9 is a new hallmark of lung adenocarcinoma, in which SOX9 might contribute to gain of tumor growth potential, possibly acting through affecting the expression of cell cycle regulators p21 and CDK4.
Subject(s)
Adenocarcinoma/genetics , Cell Proliferation , Lung Neoplasms/genetics , SOX9 Transcription Factor/genetics , Adenocarcinoma/metabolism , Adenocarcinoma/pathology , Animals , Blotting, Western , Cell Line, Tumor , Cyclin-Dependent Kinase 4/genetics , Cyclin-Dependent Kinase 4/metabolism , Cyclin-Dependent Kinase Inhibitor p21/genetics , Cyclin-Dependent Kinase Inhibitor p21/metabolism , Down-Regulation , Gene Expression Profiling , Gene Expression Regulation, Neoplastic , Humans , Immunohistochemistry , Liver Neoplasms, Experimental/genetics , Liver Neoplasms, Experimental/metabolism , Liver Neoplasms, Experimental/pathology , Lung Neoplasms/metabolism , Lung Neoplasms/pathology , Mice , Mice, SCID , Oligonucleotide Array Sequence Analysis , RNA Interference , Reverse Transcriptase Polymerase Chain Reaction , SOX9 Transcription Factor/metabolism , Transplantation, Heterologous , Tumor Burden , Up-RegulationABSTRACT
OBJECTIVE: Patent arteriovenous fistula (AVF) is related to better prognosis and quality of life for patients on long-term dialysis. When AVF dysfunction is suspected, MDCT is a good noninvasive tool for evaluating the entire AVF structure and determining reversible conditions for treatment. The aim of this article is to introduce the scanning and interpretation techniques and to illustrate the conditions related to early and late fistula failures. CONCLUSION: MDCT is a fast, noninvasive, and accurate technique for diagnosing AVF complications. Radiologists familiar with these techniques can help to improve the prognosis and quality of life for hemodialysis patients.