Kir2.1 channel regulates macrophage polarization via the Ca2+/CaMK II/ERK/NF-κB signaling pathway.

Macrophage polarization plays a key role in the inflammatory response. Various ion channels expressed in macrophages have been documented, but very little is known about their roles in macrophage polarization. We found that knockdown or blockade of the Kir2.1 (also known as KCNJ2) channel significantly inhibited M1 macrophage polarization, but promoted M2 macrophage polarization. Lipopolysaccharide (LPS)-induced M1 polarization was also remarkably suppressed in high extracellular K+ solutions (70 mM K+), and this inhibition was partially abolished by adding Ca2+ to the culture medium. Ca2+ imaging showed that Ca2+ influx was dependent on the hyperpolarized membrane potential generated by the Kir2.1 channel. The upregulation of phospho (p)-CaMK II, p-ERK, and p-NF-κB proteins in macrophages from the RAW264.7 cell line that were stimulated with LPS was significantly reversed by blocking the Kir2.1 channel or culturing the cells with 70 mM K+ medium. Furthermore, in vivo studies showed that mice treated with a Kir2.1 channel blocker were protected from LPS-induced peritonitis. In summary, our data reveal the essential role of the Kir2.1 channel in regulating macrophage polarization via the Ca2+/CaMK II/ERK/NF-κB signaling pathway.

Que Zui tea ameliorates hepatic lipid accumulation and oxidative stress in high fat diet induced nonalcoholic fatty liver disease.

In this study, the protective effects of hot water (QW) and aqueous-ethanol extracts (QA) from Que Zui tea on non-alcoholic fatty liver disease (NAFLD) were investigated. Quantitative and qualitative analysis revealed that QW and QA were rich in polyphenols, especially 6'-O-caffeoylarbutin. Both QW and QA significantly reduced body weight and liver index, increased serum levels of high density lipoprotein cholesterol (HDL-C), and decreased the levels of total cholesterol (TC), triglyceride (TG), nonesterified free fatty acids (NEFA) and low density lipoprotein cholesterol (LDL-C) in NAFLD rats induced high fat diet. Furthermore, the contents of TC, TG, NEFA, alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in the liver tissues were inhibited after QW and QA administration. Histopathological examination showed that QW and QA significantly reduced liver lipid accumulation of NAFLD rats. In addition, QW and QA could enhance increase the activity of antioxidant (glutathione, superoxide dismutase and catalase) in the liver by regulation Nrf2 signaling pathway, thereby alleviating liver damage caused by lipid peroxidation. QW and QA activated AMPK/PPAR-α signaling pathway by increasing the expression of adiponectin and its receptor AdipoR2, thereby reducing fat production and enhancing fatty acid ß oxidation. These data suggested that QW and QA had the potential to in the prevention and treatment of NAFLD.

Macrophage Paired Immunoglobulin-Like Receptor B Deficiency Promotes Peripheral Atherosclerosis in Apolipoprotein E-Deficient Mice.

Background: Peripheral atherosclerotic disease (PAD) is the narrowing or blockage of arteries that supply blood to the lower limbs. Given its complex nature, bioinformatics can help identify crucial genes involved in the progression of peripheral atherosclerosis. Materials and Methods: Raw human gene expression data for 462 PAD arterial plaque and 23 normal arterial samples were obtained from the GEO database. The data was analyzed using an integrated, multi-layer approach involving differentially-expressed gene analysis, KEGG pathway analysis, GO term enrichment analysis, weighted gene correlation network analysis, and protein-protein interaction analysis. The monocyte/macrophage-expressed leukocyte immunoglobulin-like receptor B2 (LILRB2) was strongly associated with the human PAD phenotype. To explore the role of the murine LILRB2 homologue PirB in vivo, we created a myeloid-specific PirB-knockout Apoe -/- murine model of PAD (PirB MΦKO) to analyze femoral atherosclerotic burden, plaque features of vulnerability, and monocyte recruitment to femoral atherosclerotic lesions. The phenotypes of PirB MΦKO macrophages under various stimuli were also investigated in vitro. Results: PirB MΦKO mice displayed increased femoral atherogenesis, a more vulnerable plaque phenotype, and enhanced monocyte recruitment into lesions. PirB MΦKO macrophages showed enhanced pro-inflammatory responses and a shift toward M1 over M2 polarization under interferon-γ and oxidized LDL exposure. PirB MΦKO macrophages also displayed enhanced efferocytosis and reduced lipid efflux under lipid exposure. Conclusion: Macrophage PirB reduces peripheral atherosclerotic burden, stabilizes peripheral plaque composition, and suppresses macrophage accumulation in peripheral lesions. Macrophage PirB inhibits pro-inflammatory activation, inhibits efferocytosis, and promotes lipid efflux, characteristics critical to suppressing peripheral atherogenesis.

Statin downregulation of miR-652-3p protects endothelium from dyslipidemia by promoting ISL1 expression.

BACKGROUND: Aberrant endothelial function is a major contributing factor in cardiovascular disease. Dyslipidemia leads to decreased nitric oxide (NO) bioavailability, an early sign of endothelial failure. Low insulin gene enhancer protein (ISL1) levels decrease healthy NO bioavailability. We hypothesized that the microRNA miR-652-3p negatively regulates endothelial ISL1 expression and that dyslipidemia-induced miR-652-3p upregulation induces aberrant endothelial functioning via ISL1 downregulation. METHODS: Various in vitro experiments were conducted in human umbilical vein endothelial cells (HUVECs). Luciferase assays were performed in HEK293 cells. We constructed a high-fat diet (HFD) Apoe-/- murine model of dyslipidemia and a rat model of low-density lipoprotein (LDL)-induced dyslipidemia to conduct in vivo and ex vivo experiments. RESULTS: Luciferase assays confirmed miR-652-3p's targeting of the ISL1 3'-untranslated region (3'-UTR). Simvastatin blocked oxidized LDL (ox-LDL)-induced increases in miR-652-3p and ox-LDL-induced decreases in ISL1 protein expression, endothelial NO synthase (eNOS) activation, and NO production. Simvastatin's effects were abrogated by miR-652-3p overexpression and phenocopied by miR-652-3p inhibition. The dyslipidemic mouse model exhibited increased miR-652-3p and decreased ISL1 protein levels in the endothelium, effects opposed by simvastatin or miR-652-3p inhibition. The impact of simvastatin in vivo was abolished by overexpressing miR-652-3p or knocking-down ISL1. The rat model of dyslipidemia exhibited a similar pattern of miR-652-3p upregulation, attenuated ISL1 protein levels, decreased eNOS activation, and decreased NO production, effects mitigated by simvastatin. CONCLUSIONS: Dyslipidemia upregulates endothelial miR-652-3p, which decreases ISL1 protein levels, eNOS activation, and NO production. Simvastatin therapy lowers endothelial miR-652-3p expression to protect endothelial function under dyslipidemic conditions.

Potential ameliorative effects of epigallocatechin­3­gallate against testosterone-induced benign prostatic hyperplasia and fibrosis in rats.

Green tea is among the most popular beverages in the world and is an important source of phytoestrogens. Epigallocatechin­3­gallate (EGCG) is the major polyphenol in green tea. The aim of this study was to investigate the anti-benign prostatic hyperplasia (BPH) activity and underling mechanisms of EGCG in testosterone-induced BPH rats and in BPH-1 cells. Prostatic levels of oxidative stress and inflammation makers, as well as angiogenesis related growth factors were measured. Additionally, the prostatic levels of sex hormonal mediators (androgen receptor (AR), estrogen receptor (ER)-α and ER-ß), hypoxia-inducible factor (HIF)-1α, transforming growth factor-ß1 (TGF-ß1), type I TGF-ß receptor (TGF-ßRI), Smad3, phosphorylation-Smad3 (p-Smad3), epithelial-mesenchymal transition (EMT) markers (E-cadherin, collagen-I, fibronectin and α-SMA) and microRNA (miR)-133a/b were analyzed by immunohistochemistry assay, western blot and/or quantitative RT-PCR. It was observed that EGCG attenuated the prostatic oxidative stress and inflammatory microenvironment, ameliorated prostatic hyperplasia and collagen deposition, reduced the levels of angiogenesis related growth factors, inhibited the over-expression of AR, ER-α, HIF-1α, TGF-ß1, TGF-ßRI and p-Smad3, enhanced the expression of ER-ß, increased the levels of miR-133a/b, as well as relieved prostatic EMT in rats. Both HIF-1α inhibitor and EGCG decreased the expression of HIF-1α and TGF-ß1, as well as attenuated EMT in BPH-1 cells. It indicated that EGCG could attenuate testosterone-induced BPH and fibrosis.

Upregulation of microRNA-351 exerts protective effects during sepsis by ameliorating skeletal muscle wasting through the Tead-4-mediated blockade of the Hippo signaling pathway.

Sepsis-induced skeletal muscle wasting may lead to various severe clinical consequences. Understanding molecular mechanisms of the regulation of the loss of skeletal muscle mass in septic patients remains a significant clinical challenge. The current study was conducted to establish septic mice models to explore the relationship between microRNA (miR)-351 and the transcription element apical (TEA) domain transcription factor (Tead)-4 gene and to investigate its effects on the skeletal muscle through mediating the Hippo signaling pathway in mice with acute sepsis. A total of 60 mice were collected to establish mouse models of acute sepsis. The positive expression rate of Tead-4 and the apoptotic index (AI) were measured. A dual-luciferase reporter gene assay was conducted to verify the targeting relationship between miR-351 and Tead-4. Furthermore, the muscle fiber diameter (MFD) and area (MFA) and the content of 3-methylhistidine (3-MH) and tyrosine (Tyr) were assessed. The expression levels of miR-351, p38-MAPK, Yes-associated protein, Tead-4, B-cell lymphoma X protein (Bax), and Caspase-3 were determined with quantitative RT-PCR and Western blot analysis. Finally, cell viability, apoptosis, and levels of inflammatory factors, including IL-1ß, IL-6, IGF-1, TNF-α, and monocyte chemoattractant protein-1 were detected by 3-(4,5-dimethylthiazol-2- yl)-2,5-diphenyltetrazolium bromide assay, flow cytometry, and ELISA. Initially, Tead-4 protein expression was higher in skeletal muscle tissues of mice with acute sepsis. Tead-4 was identified to negatively regulate miR-351. Upregulation of miR-351 increased MFA and MFD, muscle weight water content, Bcl-2 expression levels, and cell viability. Up-regulation of miR-351 reduced AI; 3-MH and Tyr content; positive expression of Tead-4 protein; the expression levels of p38-MAPK, Yap, Tead-4, Bax, and Caspase-3; apoptosis; and inflammatory responses. The current study demonstrated that up-regulation of miR-351 inhibits the degradation of skeletal muscle protein and the atrophy of skeletal muscle in mice with acute sepsis by targeting Tead-4 through suppression of the Hippo signaling pathway. Thus, miR-351 overexpression may be a future therapeutic strategy for acute sepsis.-Zhang, L.-N., Tian, H., Zhou, X.-L., Tian, S.-C., Zhang, X.-H., Wu, T.-J. Upregulation of microRNA-351 exerts protective effects during sepsis by ameliorating skeletal muscle wasting through the Tead-4-mediated blockade of the Hippo signaling pathway.

Epigallocatechin-3-Gallate Ameliorates Angiotensin II-Induced Oxidative Stress and Apoptosis in Human Umbilical Vein Endothelial Cells through the Activation of Nrf2/Caspase-3 Signaling.

INTRODUCTION: This study aimed to investigate whether epigallocatechin-3-gallate (EGCG) shows antioxidant activity against angiotensin II (Ang II)-induced human umbilical vein endothelial cell (HUVEC) apoptosis. MATERIALS AND METHODS: The viability of HUVECs was revealed by MTT and LDH assay. The cell apoptosis was detected by FITC-PI assay. A fluorescent probe assay was used to measure the reactive oxygen species (ROS) generation in HUVECs. Mitochondrial permeability transition pore (MPTP) opening, mitochondrial membrane potential, and caspase-3, -4, -8, -9 activities were also measured. RESULTS: We found that Ang II treatment increased the generation of ROS, enhanced MPTP opening and cytochrome c release, activated caspase-3/9, and consequently induced HUVEC apoptosis. EGCG treatment-suppressed Ang II induces the oxidative stress of HUVECs and mitochondria-related cell apoptosis. We also showed that the antioxidant activity pathway, including cytochrome c release, MPTP opening, and caspase-3/9 activation, is a key endogenous defensive system in HUVECs, provoking Ang II exposure. Our study revealed that increased expression of Nrf2 by EGCG could partially repress Ang II-induced injury effects. CONCLUSIONS: All of our findings indicated that EGCG treatment provides a protective effect for Ang II-induced HUVEC apoptosis by decreasing oxidative stress and ameliorating mitochondrial injury.

Separation and identification of oligomeric ethyl silicates by liquid chromatography with electrospray ionization mass spectrometry.

Reversed-phase liquid chromatography coupled with electrospray ionization mass spectrometry was used to study the molecular structures of components and molar mass distributions in ethyl silicate-40, a versatile liquid precursor for silicon-based materials. Identity testing by standard spectroscopic techniques showed that a commercial sample of ethyl silicate-40 was composed of linear/branched ethoxysiloxane oligomers with the silicon atoms ranging from 2 to 12 together with minor monocyclic species. Analysis of the sample by liquid chromatography coupled with evaporative light scattering detection resulted in an elution profile consisting of a series of peak clusters. Peak identification showed that the linear/branched homologous series of oligomers were eluted in the order of increasing number of silicon atoms in the molecules and the time duration (width) of the resulting peak clusters increased in the same fashion corresponding to increasing number of geometric isomers. In addition, small amounts of monocyclic oligomers present in the sample were found to be less retained than each linear/branched counterpart. Finally, the molar mass distribution parameters for ethyl silicate-40 determined by the developed method were in good agreement with the literature values. Overall, this work demonstrates that reversed-phase liquid chromatography coupled with electrospray ionization mass spectrometry is an indispensable tool for the comprehensive characterization of complex mixtures of this type.

Optical Properties of GaSb Nanofibers.

Amorphous GaSb nanofibers were obtained by ion beam irradiation of bulk GaSb single-crystal wafers, resulting in fibers with diameters of ~20 nm. The Raman spectra and photoluminescence (PL) of the ion irradiation-induced nanofibers before and after annealing were studied. Results show that the Raman intensity of the GaSb LO phonon mode decreased after ion beam irradiation as a result of the formation of the amorphous nanofibers. A new mode is observed at ~155 cm(-1) both from the unannealed and annealed GaSb nanofiber samples related to the A1g mode of Sb-Sb bond vibration. Room temperature PL measurements of the annealed nanofibers present a wide feature band at ~1.4-1.6 eV. The room temperature PL properties of the irradiated samples presents a large blue shift compared to bulk GaSb. Annealed nanofibers and annealed nanofibers with Au nanodots present two different PL peaks (400 and 540 nm), both of which may originate from Ga or O vacancies in GaO. The enhanced PL and new band characteristics in nanostructured GaSb suggest that the nanostructured fibers may have unique applications in optoelectronic devices.

Experimental investigation of superfocusing of plasmonic lens with chirped circular nanoslits.

A plasmonic lens with metallic chirped circular nanoslits corrugated on Au film supported on quartz substrate for the purpose of superfocusing was put forth and fabricated by means of focused ion beam direct milling technique. Topography of the lens was imaged using an atomic force microscope. After that a near-field scanning optical microscope was employed for optical characterization of focusing performance of the lens. Our experimental results verify the focusing performance and further demonstrate that they are in agreement with the theoretical calculation results. Focusing performance is significantly improved in comparison to that of the non-chirped lens. The lenses are possible to be used for the applications of bioimaging, detection, and inspection in submicron scale resolution.

Practical method to calculate post-LASIK corneal power: the Actual K(a+p) method.

AIM: To evaluate the accuracy of a practical method (the Actual K(a+p) method) of corneal power measurement for post-LASIK eyes undergoing cataract surgery. METHODS: Ten eyes of 7 patients (4 male, 3 female, average age 50.10±4.01 years, with -11.01±3.55D mean refraction before LASIK), underwent post-LASIK phaco+IOL cataract surgery. We used the posterior corneal curvature as measured by the Pentacam in a method we named Actual K(a+p) to calculate the post-LASIK corneal power for IOL calculation. The refractive outcomes after cataract surgery were evaluated. The Actual K(a+p) was compared with the back- calculated corneal power (BCK), which was thought to be the benchmark of true corneal power. The corneal power estimated by other published methods, including Maloney, Shammas, Koch-Maloney, Savini, and McCulley, together with the true net power and equivalent K reading (EKR) as found by the Pentacam were also compared with the BCK. RESULTS: All eyes achieved satisfied refractive status after cataract surgery. The difference between the postoperative refraction and the target refraction was 0.04±0.40D, range from -0.63D and +0.85D. Among all the methods we studied, although the Bonferroni multiple comparison tests did not detect significant differences between any two of them, the Actual K(a+p) yielded the highest agreement with the BCK, with 80% of the eyes falling within ±0.5D and 100% within ±1.0D from the BCK values. CONCLUSION: The Actual K(a+p) method can provide encour- aging results in post-LASIK eyes undergoing cataract surgery.

[The accuracy of posterior curvature method in corneal power calculation after LASIK surgery].

OBJECTIVE: To estimate the accuracy of posterior curvature method in corneal power calculation after LASIK surgery. METHODS: Corneal power calculation in 11 eyes that underwent Intraocular Lens (IOL) implantation after LASIK surgery (10 cases of Phaco + IOL, 1 case of IOL displacement), all of which used posterior curvature method, was analyzed retrospectively. The differences between post-operative stable refraction and target refraction were calculated, the actual corneal powers were deduced, and the expected refractive errors using other corneal power evaluation methods (auto-keratometry, corneal topography, spherical equivalent method, anterior curvature method, Equivalent K Reading method provided by Pentacam) were analyzed. In addition, refraction of 23 eyes underwent LASIK surgery were done on their 6 months follow-up. The theoretical corneal powers were deduced by subtracting the change of refraction before and after LASIK surgery from the pre-operative corneal powers. The differences between calculated corneal powers using posterior curvature method and the theoretical corneal powers were analyzed, and were compared with other corneal power evaluation methods. RESULTS: The mean uncorrected post-operative visual acuity of IOL implantation eyes using posterior curvature method was 0.8 +/- 0.2, with mean absolute refractive error from target of (0.36 +/- 0.36) D (-0.63 to +0.85 D). The ratio of eyes with absolute error within 0.25 D, 0.50 D, and 1.00 D was 55%, 73%, and 91% respectively. This result was significantly lower than that of the auto-keratometry (2.50 +/- 1.08) D, corneal topography (1.90 +/- 0.88) D, and those obtained from spherical equivalent method (2.09 +/- 1.62) D (P < 0.01) or anterior curvature method (1.45 +/- 1.10) D (P < 0.05). It also showed less bias (-1.13 to 0.85 D) when compared to the Equivalent K Reading (-1.10 to 1.80 D), but the difference was not significant (P > 0.05). For the 23 post LASIK eyes, the absolute difference between the corrected corneal power using posterior curvature method and theoretical power was (0.67 +/- 0.45) D, also showed least bias compared with other methods. CONCLUSION: It is a practical and accurate way to calculate the corneal power after LASIK surgery using posterior curvature method.