MYEOV with High Frequencies of Mutations in Head and Neck Cancers Facilitates Cancer Cell Malignant Behaviors.

Cancer driver genes (CDGs) and the driver mutations disrupt the homeostasis of numerous critical cell activities, thereby playing a critical role in tumor initiation and progression. In this study, integrative bioinformatics analyses were performed based on a series of online databases, aiming to identify driver genes with high frequencies of mutations in head and neck cancers. Higher myeloma overexpressed (MYEOV) genetic variation frequency and expression level were connected to a poorer prognosis in head and neck cancer patients. MYEOV was dramatically upregulated within head and neck tumor samples and cells. Consistently, MYEOV overexpression remarkably enhanced the aggressiveness of head and neck cancer cells by promoting colony formation, cell invasion, and cell migration. Conversely, MYEOV knockdown attenuated cancer cell aggressiveness and inhibited tumor growth and metastasis in the oral orthotopic tumor model. In conclusion, MYEOV is overexpressed in head and neck cancer, with greater mutation frequencies correlating to a poorer prognosis in head and neck cancer patients. MYEOV serves as an oncogene in head and neck cancer through the promotion of tumor cell colony formation, invasion, and migration, as well as promoting tumor growth and metastasis in the oral orthotopic tumor model.

miR-340-5p affects oral squamous cell carcinoma (OSCC) cells proliferation and invasion by targeting endoplasmic reticulum stress proteins.

Lip and oral cancer is the 12th most common malignancy and oral squamous cell carcinoma (OSCC) represents about 90% of all oral malignant tumors, with an annual mortality rate exceeding 50%. Recent studies have concluded that endoplasmic reticulum stress may have a close link to tumor genesis, progression, and prognosis. As an epigenetic regulatory factor, miRNA exerts a substantial effect on tumor development. This study found that transcription factor 6 (ATF6) and Protein kinase R-like endoplasmic reticulum kinase (PERK) were abnormally increased within OSCC tissue samples and oral cancer cell lines. The biological functions of ATF6 and PERK within CAL-27 and SCC-9 oral cancer cell lines were investigated. In vitro experiments revealed that silencing ATF6 and PERK suppressed the ability of cells to proliferate and to invade and mitigated cell endoplasmic reticulum (ER) stress. As predicted by bioinformatics analyses and experiments, miR-340-5p could simultaneously bind to ATF6 and PERK 3' untranslated region (UTR) and inhibit ATF6 and PERK expression. miR-340-3p overexpression inhibited while down-regulation of miR-340-5p boosted the invading and proliferating ability of oral cancer cells, and miR-340-3p also affects ER stress. When co-transfected in oral cancer cells, dynamic effects of miR-340-5p and its targets PERK and ATF6 on cell phenotypes in vitro and in vivo were investigated. PERK or ATF6 overexpression dramatically attenuated phenotypes of miR-340-5p up-regulation. Altogether, miR-340-5p targets the endoplasmic reticulum stress proteins PERK and ATF6 to affect OSCC cell proliferation and invasion.

[Effects of LIMK2 RNA interference on the mechanosensitivity of c-fos gene in osteoblast].

OBJECTIVE: To study the effects of cytoskeleton reorganization inhibition with LIMK2 RNAi upon the mechanosensitivity of c-fos gene in osteoblast. METHODS: Mouse primary osteoblast was treated with LIMK2 specific siRNA (RNAi Group), negative control siRNA (NC Group), and then were loaded or unloaded by fluid shear stress. Real-time PCR and immunofluorescence were used to detect the c-fos expression levels and statistics analysis was performed. RESULTS: When the cytoskeleton reorganization was inhibited with RNAi only, the c-fos mRNA (0.0108 +/- 0.0074 and 0.0042 +/- 0.0018, t = -1.86, P > 0.05) and protein (121 +/- 7 and 119 +/- 6, t = -1.272, P > 0.05) expression levels of each unloaded group had no significant difference; Fluid shear stress could up-regulate the c-fos mRNA (0.2203 +/- 0.1532 vs 0.0042 +/- 0.0018, t = -707.35, P < 0.05)and protein (178 +/- 12 vs 119 +/- 6, t = -30.761, P < 0.05) expression; After the cytoskeleton reorganization was inhibited with RNAi, the c-fos mRNA (0.5280 +/- 0.0879 vs 0.2203 +/- 0.1532, t = -1007.00, P < 0.05) and protein (224 +/- 46 vs 178 +/- 12, t = -6.853, P < 0.05) expression induced by fluid shear stress had significant difference. Cytoskeleton reorganization inhibition with RNAi had synergistic effect upon the expression of c-fos mRNA (F = 84.388, P < 0.05) and protein (F = 42.409, P < 0.05) induced by fluid shear stress. CONCLUSION: Using RNAi against LIMK2 to inhibit the cytoskeleton reorganization can promote the expression of c-fos gene and thus enhance the mechanosensitivity of c-fos gene in osteoblast.

Resident Microglia Activate before Peripheral Monocyte Infiltration and p75NTR Blockade Reduces Microglial Activation and Early Brain Injury after Subarachnoid Hemorrhage.

Early brain injury (EBI) after aneurysmal subarachnoid hemorrhage (SAH) contributes to high morbidity and mortality. Although it is well recognized that acute neuroinflammation reaction is one of the most important triggers of EBI, pharmacotherapy proved to be clinically effective against the initiating of neuroinflammation after SAH is lacking. The resident microglia and infiltrated peripheral monocyte are two main types of immune cells in central nervous system (CNS) and control the inflammation process in brain after SAH. But the time course and relative contributions of these two immune cell activations after SAH are unknown. The p75 neurotrophin receptor (p75NTR), member of TNF receptor superfamily, expresses on infiltrated peripheral monocytes and suppresses their proinflammatory action after brain insults. But the p75NTR expression on resident microglia in vivo is rarely explored and their function keeps elusive. Therefore, we designed this study to investigate the time course of resident microglia activation and peripheral monocyte infiltration, as well as the microglial expression of p75NTR by using CX3C-chemokine receptor 1 (Cx3cr1) and chemokine receptor 2 (Ccr2) double transgenic mice (Cx3cr1GFP/+Ccr2RFP/+) after SAH. The results showed activated microglia was observed in cortex as early as 24 h and further increased at 48 and 72 h post SAH, while the infiltrated monocyte was not found until 72h. In addition, activated microglia expressed p75NTR acutely and p75NTR specific antagonist TAT-Pep5 significantly reduced microglia activation, neuroinflammation and EBI from 24 to 72 h. Together, these data suggest that the early neuroinflammation reaction might be initiated and intensified mainly by resident microglia rather than infiltrated monocyte at least in the first 48 h after SAH and p75NTR blockading by TAT-Pep5P might alleviate EBI through mediating microglial activation.

[Effect of cytoskeleton reorganization inhibition on the activation of extracellular signal-regulated kinase in osteoblasts by fluid shear stress].

OBJECTIVE: To investigate the effect of cytoskeleton reorganization inhibition with RNA interference on the activation of extracellular signal-regulated kinase (ERK1/2) in primary osteoblasts induced by fluid shear stress (FSS). METHODS: BALB/c mouse primary cultured osteoblasts were isolated by enzyme digestion technique. Osteoblasts were treated with LIM domain kinase 2 (LIM-2) specific siRNA or negative control siRNA, and then were loaded or unloaded by FSS of 1.2 Pa for 0, 5, 15, 30 and 60 min, respectively. The Western blotting was performed to detect the protein expression levels of P-ERK1/2 and ERK1/2, respectively. Two-way ANOVA and one-way ANOVA were used in data analysis. RESULTS: FSS loading for different time (0, 5, 15, 30, 60 min) treated with negative RNA inteference had significant effect on the levels of P-ERK/ERK ratio (0.047 ± 0.031, 0.253 ± 0.137, 0.390 ± 0.155, 0.613 ± 0.123, 0.680 ± 0.108, respectively, P < 0.01). Statistical analysis showed that there was significant interaction between FSS and cytoskeleton reorganization inhibition treated with RNA inteference on the levels of P-ERK/ERK ratio (P < 0.01). The levels of P-ERK/ERK ratio increased when osteoblasts were loaded for 5 - 15 min (0.623 ± 0.129 and 0.623 ± 0.064, respectively, P < 0.05) and returned to baseline at 30 min (0.333 ± 0.086), and then reached the peak at 60 min (0.667 ± 0.064, P < 0.01). CONCLUSIONS: FSS could activate ERK1/2 rapidly in primary cultured osteoblasts. Cytoskeleton reorganization inhibition treated with RNA interference speeded-up the activation of ERK1/2 by FSS, which could increase the sensitivity of ERK1/2 to FSS.

[Effects of cofilin phosphorylation on the actin cytoskeleton reorganization induced by shear stress].

OBJECTIVE: To explore the effects of cofilin on the actin cytoskeleton reorganization in osteoblasts induced by fluid shear stress. METHODS: Fluid shear stress (1.2 Pa) was applied to osteoblasts for 0 (control group), 15, 30, 45, 60, 120 min in vitro. Cells were stained with fluorescein isothiocyanate (FITC)-phalloidin for fiber-actin, and confocal laser scanning microscope(CLSM) was used to observe the fluorescence of fiber-actin. Western blotting was used to detect the expression of the cofilin and the phospho-cofilin. RESULTS: Actin filaments became organized into stress fibers that were thicker and more abundant than those in non-flowed cells. The fluorescence intensity (38.00 ± 6.88) of fiber-actin after 120 min (42.93 ± 6.41) loading it was 2.8 times as much as that in control group (15.41 ± 3.60, P < 0.05). Additionally, the level of phospho-cofilin protein was dramatically elevated after loading. Fluid shear stress induced an initial decrease of cofilin at 60 min. However, at 120 min cofilin (0.254 ± 0.026) increased to 1.5 times as much as that at 60 min (0.162 ± 0.004). CONCLUSIONS: The results indicate that cofilin phosphorylation mediates fiber-actin reorganization in the osteoblasts induced by fluid shear stress.