MiR-29b level-mediated regulation of Klotho methylation via DNMT3A targeting in chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is a chronic lung disease characterized by chronic bronchitis and emphysema. Cigarette smoke extract (CSE) is the predominant cause of COPD. This study aimed to investigate the effects of miR-29b and their underlying mechanisms in a COPD cell model. MiR-29b and DNMT3A expression in lung tissue samples (taken at least 5 cm away from the tumor lesion) of NSCLC cases with smoking (n = 30), without smoking (n = 30), and with COPD (with smoking) (n = 30) was researched by qRT-PCR. A medium containing 10 % CSE was employed to induce murine alveolar macrophage MH-S cells to establish COPD cells. 5-Aza-cdr (5-AZA-2'-deoxycytidine) was used to block DNMT3A. The relationship and interaction between miR-29b and DNMT3A were validated through the dual luciferase reporter assay. The expression levels of macrophage M1 polarization marker proteins iNOS and TNF-α, DNMT3A, and Klotho protein were monitored using western blotting. The methylation levels of the miR-29b precursor gene and Klotho promoter were detected by quantitative methylation-specific PCR (MS-qPCR). The levels of IL-1ß, IL-6, and TNF-α in cell culture medium were detected via ELISA. It was found that the expression of miR-29b was downregulated, as a result of increased DNA methylation, and that of DNMT3A was upregulated in the lung tissues of NSCLC cases with COPD (with smoking). DNMT3A expression was negatively correlated with miR-29b expression in the lung tissues of NSCLC cases with COPD (with smoking). In addition, miR-29b expression was distinctly downregulated in CSE-induced MH-S cells and inhibited CSE-induced M1 polarization and inflammation. Importantly, DNMT3A was identified as a direct target gene of miR-29b. MiR-29b is negatively regulated by DNMT3A-mediated DNA methylation. Moreover, Klotho expression was downregulated and the Klotho promoter methylation level was increased in lung tissues of NSCLC cases with COPD (with smoking). The negative feedback between miR-29b and DNMT3A modulates CSE-induced M1 polarization and inflammation in macrophages as well as Klotho promoter methylation in CSE-mediated MH-S. Collectively, these findings indicate that the miR-29b level in COPD controls Klotho methylation via DNMT3, which maybe a promising target for the treatment of COPD.

Electrode Layout Optimization and Numerical Simulation of Cast Conductive Asphalt Concrete Steel Bridge Deck Pavement.

In order to obtain the optimal electrode layout and ice melting effect of cast conductive asphalt concrete steel bridge deck pavement, firstly, pouring conductive asphalt concrete was prepared; secondly, different electrode materials and layout methods were selected to test the heating rate of the specimen from start to 120 min, and the electrode materials and layout methods were optimized. Then, the finite element analysis software ANSYS was used to build the model for heating and ice melting simulation, and the indoor test was used to further verify the ice melting effect of the cast conductive asphalt coagulation with or without the insulation layer. Finally, the thermal-structural coupling analysis of cast conductive asphalt concrete steel bridge deck pavement was carried out using ANSYS finite element software. The results showed that the stainless steel electrode material had the best heating effect, and the electrode thickness in the range of 0.1~3 mm had no effect on the heating effect. The intermediate heating rate of the upper surface of the stainless steel sheet electrode cast conductive asphalt concrete in the left and right external electrodes was 8 ∘C/h, while the intermediate heating rate of the upper surface of the stainless steel mesh electrode cast conductive asphalt concrete was 12.9 ∘C/h. The layout of the left and right buried stainless steel metal mesh was able to effectively improve the snow melting efficiency; ANSYS finite element ice melting simulation was used to obtain the variation law of ice melting efficiency and a temperature field of cast conductive asphalt concrete. The indoor ice melting test showed that when melting the same thickness ice layer at 50 V voltage, it took 240 min with an insulation layer and 720 min without an insulation layer, which was three times that of the ice with an insulation layer, which further verifies the superiority of its ice melting effect. The most unfavorable load position of pavement under load and temperature field was determined. The maximum tensile stress and compressive stress of the pavement surface were transverse, and the maximum shear stress of the pavement bottom was transverse.

Silibinin activates AMP-activated protein kinase to protect neuronal cells from oxygen and glucose deprivation-re-oxygenation.

In this study, we explored the cytoprotective potential of silibinin against oxygen-glucose deprivation (OGD)-induced neuronal cell damages, and studied underling mechanisms. In vitro model of ischemic stroke was created by keeping neuronal cells (SH-SY5Y cells and primary mouse cortical neurons) in an OGD condition followed by re-oxygenation. Pre-treatment of silibinin significantly inhibited OGD/re-oxygenation-induced necrosis and apoptosis of neuronal cells. OGD/re-oxygenation-induced reactive oxygen species (ROS) production and mitochondrial membrane potential (MMP) reduction were also inhibited by silibinin. At the molecular level, silibinin treatment in SH-SY5Y cells and primary cortical neurons led to significant AMP-activated protein kinase (AMPK) signaling activation, detected by phosphorylations of AMPKα1, its upstream kinase liver kinase B1 (LKB1) and the downstream target acetyl-CoA Carboxylase (ACC). Pharmacological inhibition or genetic depletion of AMPK alleviated the neuroprotective ability of silibinin against OGD/re-oxygenation. Further, ROS scavenging ability by silibinin was abolished with AMPK inhibition or silencing. While A-769662, the AMPK activator, mimicked silibinin actions and suppressed ROS production and neuronal cell death following OGD/re-oxygenation. Together, these results show that silibinin-mediated neuroprotection requires activation of AMPK signaling.