ABSTRACT
Interferon regulatory factor-7 (IRF7) is an essential regulator of both innate and adaptive immunity. It is also expressed in the otic vesicle of zebrafish embryos. However, any role for irf7 in hair cell development was uncharacterized. Does it work as a potential deaf gene to regulate hair cell development? We used whole-mount in situ hybridization (WISH) assay and morpholino-mediated gene knockdown method to investigate the role of irf7 in the development of otic vesicle hair cells during zebrafish embryogenesis. We performed RNA sequencing to gain a detailed insight into the molecules/genes which are altered upon downregulation of irf7. Compared to the wild-type siblings, knockdown of irf7 resulted in severe developmental retardation in zebrafish embryos as well as loss of neuromasts and damage to hair cells at an early stage (within 3 days post fertilization). Coinjection of zebrafish irf7 mRNA could partially rescued the defects of the morphants. atp1b2b mRNA injection can also partially rescue the phenotype induced by irf7 gene deficiency. Loss of hair cells in irf7-morphants does not result from cell apoptosis. Gene expression profiles show that, compared to wild-type, knockdown of irf7 can lead to 2053 and 2678 genes being upregulated and downregulated, respectively. Among them, 18 genes were annotated to hair cell (HC) development or posterior lateral line (PLL) development. All results suggest that irf7 plays an essential role in hair cell development in zebrafish, indicating that irf7 may be a member of deafness gene family.
Subject(s)
Hair Cells, Auditory/cytology , Interferon Regulatory Factor-7 , Zebrafish Proteins , Zebrafish , Animals , Embryonic Development , Gene Expression Regulation, Developmental , Gene Knockdown Techniques , Interferon Regulatory Factor-7/genetics , Interferon Regulatory Factor-7/metabolism , Zebrafish/embryology , Zebrafish Proteins/genetics , Zebrafish Proteins/metabolismABSTRACT
OBJECTIVES/HYPOTHESIS: Transformer-2 protein homolog beta (Tra2ß) generally plays an important role in various human cancers, but its role and the underlying mechanisms in laryngeal squamous cell carcinoma (LSCC) remained unknown. So this study aimed to assess the clinical significance and regulatory mechanisms of Tra2ß in LSCC. STUDY DESIGN: Laboratory analysis. METHODS: Expression of Tra2ß was compared in human LSCC tissue samples and paired adjacent normal tissue samples. The in vitro effects of Tra2ß expression in Hep-2 cells on their proliferation, invasion, and migration were assessed by CCK-8 assays, Matrigel invasion, and transwell migration assays. In addition, the effects of downregulation of Tra2ß on the activation of PI3K/AKT signaling pathway were measured using Western blot analysis. The effect of Tra2ß on the growth of tumors was detected in the Hep-2-injected xenograft models in vivo. RESULTS: Reverse-transcription quantitative polymerase chain reaction analysis and immunochemistry analysis indicated that the increased expression of Tra2ß in LSCC was significantly associated with poor differentiation, lymph node metastasis, and advanced clinical stage. In vitro knockdown of Tra2ß caused a significant decrease in the proliferation, invasion, and migration of Hep-2 cells. Tra2ß silencing decreased the expression of Bcl-2 but increased Bax and Caspase-3 both in mRNA and protein levels. Furthermore, knockdown of Tra2ß eliminated the suppressive effects of activation of PI3K/AKT signaling. In vivo knockdown of Tra2ß significantly inhibited the tumor growth of Hep-2-injected xenograft mice. CONCLUSIONS: The results of the present study demonstrated that knockdown of Tra2ß inhibits the proliferation and invasion of LSCC cells, at least partly via inhibiting PI3K/AKT signaling. LEVEL OF EVIDENCE: NA Laryngoscope, 129:E318-E328, 2019.
Subject(s)
Carcinoma, Squamous Cell/genetics , Gene Silencing/physiology , Laryngeal Neoplasms/genetics , Nerve Tissue Proteins/metabolism , Serine-Arginine Splicing Factors/metabolism , Signal Transduction/genetics , Cell Line, Tumor , Cell Proliferation/genetics , Humans , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolismABSTRACT
Vascular endothelial growth factor 2 (VEGFR2) was initially identified as a receptor of VEGF on endothelial cells with a role in regulating angiogenesis during organism development and tumorigenesis. Previously, in cancer tissue, VEGFR2 has been reported to be expressed in endothelial cells. In our research, we found that VEGFR2 was expressed not only in endothelial cells but also cancer cells in head and neck squamous cell carcinomas (HNSCCs). Knockdown of VEGFR2 in Hep2 cells could arrest the cell cycle in G0/G1, leading to a decrease in proliferation. We also present evidence that MAPK/ERK signal pathways and expression of CDK1 downstream of VEGFR2 might regulate proliferation and cell cycle arrest. Furthermore, we discovered that down-regulate VEGRF2 in Hep2 cells could significantly affect the invasion ability. Taken together, our data suggest that VEGFR2 might regulate proliferation and invasion in HNSCC cancer cells in vivo.
