[Hsa_circ_0000670 promoted the progression of gastric cancer through the miR-515-5p/SIX1 molecular axis].

Objective: To explore whether hsa_circ_0000670 promotes the progression of gastric cancer by regulating the miR-515-5p/SIX1 molecular axis. Methods: The gastric cancer and adjacent normal tissues of 35 gastric cancer patients admitted to Rugao Hospital Affiliated to Nantong University from 2014 to 2015 were collected. The expression levels of circ_0000670, miR-515-5p and Sine oculis homeobox 1 (SIX1) in gastric cancer tissues and cells were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and western blot. The correlations between circ_0000670 and miR-515-5p, miR-515-5p and SIX1, circ_0000670 and SIX1 were analyzed by the Pearson method. Patients were divided into low circ_0000670 expression group (17 cases) and high circ_0000670 expression group (18 cases) based on the median of circ_0000670 expression level, and Kaplan-Meier was used to analyze the 5-year survival of patients. Cell proliferation was assessed via clone formation assay. Cell cycle and apoptosis were detected by flow cytometry. Wound healing and Transwell assays were used to detect cell migration and invasion ability. The targeting relationship between miR-515-5p and circ_0000670 or SIX1 was confirmed by the dual luciferase reporter assay. Nude mice were injected into HGC-27 cells transfected with sh-NC or sh-circ_0000670, and the volume and weight of the transplanted tumor were measured, also, the levels of circ_0000670, miR-515-5p and SIX1 in the transplanted tumor tissue were detected. Results: The expression levels of circ_0000670 and SIX1 in gastric cancer tissues and cell lines were significantly increased (P<0.05), while the expression levels of miR-515-5p were significantly decreased (P<0.05). The survival rate of patients in the low circ_0000670 expression group (82.4%) was significantly higher than that in the high circ_0000670 expression group (28.7%, P=0.034). Circ_0000670 was negatively correlated with miR-515-5p (r=-0.846, P<0.001), and miR-515-5p was negatively correlated with SIX1 (r=-0.615, P<0.001), but circ_0000670 was positively correlated with SIX1 (r=0.814, P<0.001). Transfection of si-circ_0000670 or miR-515-5p mimic could significantly reduce the number of clone-forming cells, migration distance, migration and invasion cells (P<0.05), and increase the ratio of G(0)/G(1) phase cells, apoptosis rate and the protein level of E-cadherin (P<0.05), decreased the proportion of S-phase cells and the protein level of Vimentin (P<0.05). The dual luciferase report assay confirmed that circ_0000670 could target miR-515-5p, and miR-515-5p could bind to SIX1. Co-transfection of si-circ_0000670 and miR-515-5p inhibitor could significantly attenuate the effects of si-circ_0000670 on cell proliferation, migration, invasion, cell cycle and apoptosis (P<0.05). Co-transfection of miR-515-5p mimic and pcDNA-SIX1 could significantly reduce the effects of miR-515-5p mimic on cell proliferation, migration, invasion, cell cycle and apoptosis (P<0.05). Compared with the sh-NC group [volume=(596.20±125.46) mm(3) and weight=(538.00±114.39) g], the volume and weight of transplanted tumors in the sh-circ_0000670 group [volume=(299.20±47.58) mm 3 and weight=(289.80±48.73 g)] were significantly reduced (P<0.05), the expression levels of circ_0000670 and SIX1 were significantly reduced (P<0.05), and the expression level of miR-515-5p was significantly increased (P<0.05). Conclusion: Knockdown of circ_0000670 could inhibit cell proliferation, migration, invasion of gastric cancer cells, induce cell cycle arrest in G(0)/G(1) phase and promote cell apoptosis by regulating the miR-515-5p/SIX1 axis.

Associations of TNF-α -308 G>A and TNF-ß 252 A>G with Physical Function and BNP-Rugao Longevity and Ageing Study.

OBJECTIVES: To explore the associations of TNF-α -308 G>A (rs1800629) and TNF-ß 252 A>G (rs909253) with physical function and plasma B-type natriuretic peptide (BNP). METHODS: Data of 1747 community-dwelling elders from the ageing arm of the Rugao Longevity and Ageing Study was used. Physical function was measured by handgrip strength, Timed Up and Go (TUG) test and 5-meter walking test (5MWT). RESULTS: AA genotype of the TNF-α -308 G>A was associated with higher mean time of TUG test and 5MWT (multivariable adjusted ß=5.75 and 5.70, respectively, p<0.05), compared with GG genotype. For the TNF-ß 252 A>G polymorphism, GG genotype was associated with higher mean time of TUG test and 5MWT (multivariable adjusted ß=1.55 and 0.83, respectively, p<0.05) and lower handgrip strength (multivariable adjusted ß=-0.69, p<0.05), compared with AA genotype. Further, GG was associated with greater odds of low handgrip strength (OR=1.47, 95% CI=1.06-2.04), low speed of TUG test (OR=1.87, 95% CI=1.20-2.01) and elevated BNP (OR=1.30, 95% CI=1.08-1.84). GG also interacted with elevated BNP to be associated with greater odds of low handgrip strength and 5MWT. CONCLUSIONS: TNF-ß 252 A>G was associated with physical function measurements, plasma BNP level, and odds of elevated BNP in an elderly population. TNF-ß 252 A>G also interacted with elevated BNP to be associated with greater odds of physical function measurements.

LncRNA LINC00673 inhibits p53 expression by interacting with EZH2 and DNMT1 in papillary thyroid carcinoma.

OBJECTIVE: This study aims to elucidate the regulatory role of long noncoding RNA (lncRNA) LINC00673 in proliferative, invasive and metastatic capacities of papillary thyroid carcinoma (PTC), and to investigate the possible underlying mechanism. PATIENTS AND METHODS: LINC00673 expression in PTC tissues and cell lines was detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). The correlation between LINC00673 expression and PTC prognosis was analyzed. By plasmid transfection, we constructed PTC cell lines with stable knockdown of LINC00673. The regulatory effect of LINC00673 on the proliferation of K1 and TPC-1 cells were determined by Cell Counting Kit-8 (CCK-8) assay and colony formation assay. The transwell assay was conducted to evaluate the effect of LINC00673 on the metastatic ability of PTC cells. The binding condition between LINC00673 with enhancer of zeste homolog 2 (EZH2) and DNA Methyltransferase 1 (DNMT1) was verified by RNA immunoprecipitation (RIP), chromatin immunoprecipitation (ChIP) and Western blot. The cellular mechanism of p53 in regulating biological behaviors of PTC cells was explored by Western blot. Finally, the gain-of-function experiment was performed to elucidate whether LINC00673 could regulate PTC development by inhibiting p53 expression. RESULTS: LINC00673 expression in PTC tissues was significantly higher than that of the adjacent normal tissues. Besides, higher expression of LINC00673 indicated worse prognosis of PTC. The knockdown of LINC00673 in K1 and TPC-1 cells markedly reduced the proliferative rate. Meanwhile, LINC00673 down-regulation remarkably inhibited the migratory and invasive capacities of K1 and TPC-1 cells. RIP and ChIP assay demonstrated that LINC00673 could bind to EZH2 and DNMT1. Besides, Western blot analysis showed that LINC00673 negatively regulated p53 expression. In addition, the knockdown of p53 in K1 and TPC-1 cells partially reversed the inhibitory effect of LINC00673 deficiency on the proliferation and metastasis of PTC cells. CONCLUSIONS: High expression of LINC00673 in PTC predicts a poor prognosis. LINC00673 remarkably promotes the proliferation and invasion of PTC cells by inhibiting p53 expression by binding to EZH2 and DNMT1.