High dose insulin promotes the proliferation of vascular smooth muscle cells via AP-1/SM-α pathway.

Proliferation of vascular smooth muscle cells (VSMCs) participates in multiple cardiovascular disorders, while the mechanism remains unclear. This study aims to investigate the effects of insulin on VSMC. Insulin was used to stimulate rat VSMCs, and the effects on cell cycle and proliferation were subsequently analyzed using flow cytometry. Furthermore, AP-1 and SM-α overexpression vectors were constructed and transfected into VSMCs. AP-1 and SM-α were inhibited by SR11302 and SM-α siRNA, respectively. The mRNA and protein expression levels were subsequently detected using the reversetranscription quantitative polymerase chain reaction and western blotting, respectively. AP-1 and SM-α gene promoter binding sites were determined using luciferase and chromatin immunoprecipitation assays. As a result, we found that high dose of insulin promoted proliferation of VSMCs and increased the percentage of cells in the S phase by downregulating AP-1. AP-1 was identified to bind to the SM-α gene promoter at locus 2-177 to upregulate SM-α gene expression. Inhibition of AP-1 led to the decrease of SM-α expression. Overexpression of SM-α directly suppressed proliferation of VSMCs, while knocking it down promoted the process. Therefore, this study revealed that insulin downregulated the expression of the SM-α gene by inhibiting AP-1, which in turn facilitated proliferation of VSMCs.

UPF1 alleviates the progression of glioma via targeting lncRNA CYTOR.

OBJECTIVE: To uncover the role of UPF1 in alleviating the progression of glioma via targeting long non-coding RNA (lncRNA) CYTOR and underlying mechanism. PATIENTS AND METHODS: A total of 30 glioma tissues surgically resected from glioma patients and 30 brain tissues were collected from brain trauma patients undergoing craniotomy during the same period. Relative levels of UPF1 and CYTOR in collected tissues were detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Correlation between levels of UPF1 and CYTOR in glioma tissues was assessed, and the regulatory effects of UPF1/CYTOR on proliferative and invasive abilities in U87 and LN229 cells were evaluated by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) and transwell assay, respectively. In addition, the interaction between UPF1 and CYTOR was explored by RIP (RNA-Binding Protein Immunoprecipitation) assay. Through Actinomycin D treatment in U87 and LN229 cells, RNA stability of CYTOR influenced by UPF1 was determined. Finally, rescue experiments were conducted to ascertain the involvement of CYTOR in UPF1-regulated progression of glioma. RESULTS: UPF1 was downregulated in glioma tissues and cells. A lower level of UPF1 was observed in glioma tissues in the more advanced stage with a larger tumor size. Besides, the overexpression of UPF1 markedly suppressed proliferation and invasion abilities of U87 and LN229 cells, and CYTOR was upregulated in glioma tissues and cells, which was negatively correlated with UPF1 level. Moreover, the overexpression of UPF1 decreased the half-life of CYTOR in glioma cells. Furthermore, the RIP assay confirmed the interaction between UPF1 and CYTOR. Rescue experiments finally confirmed that the overexpression of CYTOR partially reversed the inhibitory effects of UPF1 on proliferation and invasion abilities in glioma. CONCLUSIONS: UPF1 is down-regulated in glioma and alleviates the progression of glioma via targeting CYTOR.

Spectrophotometric determination of equilibrium constants of two mutual competitive reactions by the method of isosbestic points.

We have developed a method that makes use of dual isosbestic points in a dual ligand, single metal system that allows the determination of the equilibrium constants of complexes in which two ligands compete for the same metal ion, and one complex is colourless. The competition of methyl thymol blue and citrate was used to test the model.

Determination of the stability constant of the ternary complex formed in the copper-bipyridyl-eriochrome cyanine R system, by an isosbestic point-spectrophotometric method.

This paper reports a new method for determining the stability constants of ternary complexes, based on analysis of isosbestic points. First, it is necessary to determine the absolute molar absorptivities of the various species concerned, then their concentrations and the stability constant. In this method, the approximate treatment of classical photometric methods is not required. The conditional constant can be calculated directly from the measured data.