Inhibition of microRNA-543 alleviates sepsis-induced acute kidney injury via targeting Bcl-2.

OBJECTIVE: Sepsis has a high morbidity and mortality and is prone to cause acute kidney injury (AKI). Here, we aimed to demonstrate the function and molecular mechanism of microRNA-543 (miR-543) in septic AKI. MATERIALS AND METHODS: MiR-543 inhibitor or NC was transfected into LPS-treated HK-2 cells to observe lipopolysaccharide (LPS)-induced inflammation and apoptosis. The detection of inflammation and apoptosis of HK-2 cells relies on Western blot, quantitative Reverse-Transcription Polymerase Chain Reaction (qRT-PCR), enzyme-linked immunosorbent assay (ELISA), Cell Counting Kit-8 (CCK-8) assay, flow cytometry, and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) staining. RESULTS: MiR-543 expression was increased in LPS-treated HK-2 cells. By transfecting miR-543 inhibitor into HK-2 cells, miR-543 expression was dramatically reduced. The downregulation of miR-543 remarkably inhibited the inflammation and apoptosis, which was manifested by the reduction of inflammatory cytokines (TNF-α, IL-6, IL-1ß), the reversal of apoptosis-related proteins expression (Bcl-1, Bax), the increase of cell viability and the decrease of the proportion of apoptotic cells. The result of Luciferase activity assay demonstrated that miR-543 directly targets Bcl-2. CONCLUSIONS: MiR-543 expression was increased in LPS-treated HK-2 cells, and silencing miR-543 could inhibit LPS-induced inflammation and apoptosis in HK-2 cells via targeting Bcl-2.

Inhibition of microRNA-495 inhibits hypoxia-induced apoptosis in H9c2 cells via targeting NFIB.

OBJECTIVE: Acute myocardial infarction (AMI) is a serious cardiovascular disease that threatens human life. MicroRNA is considered to be an important participant in the pathophysiology of AMI. This article focused on the role of microRNA-495 (miR-495) in regulating apoptosis after myocardial infarction (MI) and its underlying mechanisms. MATERIALS AND METHODS: H9c2 cells were cultured in an incubator containing 1% O2 to establish a cell model of MI. Quantitative reverse-transcription polymerase chain reaction (RT-PCR) was utilized to detect miR-495 expression in H9c2 cells. The effects of miR-495 and NFIB on hypoxia-treated H9c2 cells were observed by Western blot, lactate dehydrogenase (LDH) detection, MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay, flow cytometry, and terminal dexynucleotidyl transferase(TdT)-mediated dUTP nick end labeling (TUNEL) staining. Luciferase reporter gene experiment was used to prove the regulatory relationship between miR-495 and NFIB. RESULTS: Hypoxia induced injury to H9c2 cells, which was manifested by decreased cell viability, increased LDH release, increased pro-apoptotic proteins (Bax, Cleaved Caspase-3) expression, decreased anti-apoptotic protein (Bcl-2) expression, and increased in the rate of apoptosis and TUNEL positive cells. MiR-495 expression was remarkably increased in H9c2 cells treated with hypoxia. Inhibiting miR-495 expression markedly alleviated the hypoxia-induced injury in H9c2 cells, while silencing NFIB aggravated the hypoxia-induced damage. In addition, NFIB was confirmed to be the target of miR-495. CONCLUSIONS: MiR-495 expression was increased in hypoxia-treated H9c2 cells. Silencing miR-495 could significantly inhibit hypoxia-induced apoptosis of H9c2 cells by targeting NFIB.

Identification and characterization of differentially expressed lncRNA in 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced cleft palate.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a ubiquitous environmental pollutant and also a strong teratogen for cleft palate (CP). But up to now, the underlying molecular mechanisms of TCDD-induced CP are largely unknown. More recently, accumulating evidences are revealing important roles of long noncoding RNAs (lncRNAs) in all kinds of diseases including CP. However, the role and molecular mechanism of lncRNAs in TCDD-induced CP are still largely unexplored. Thus, identification of differentially expressed lncRNA (DEL) might help figuring out the mechanism of CP induced by TCDD. In this study, a CP offspring model of C57BL/6 female mice was generated by TCDD (64 µg/kg body weight) induce on embryo day 10 (E10). The incidence rate of CP was 100% in the TCDD group (105) after cervical dislocation on E16. Then, the high-throughput RNA sequencing (RNA-seq) was established to search a comprehensive profile of the lncRNAs. In addition, a coexpression network of lncRNA and messenger RNA (mRNA) was performed to discern potential mechanism. The result showed that 26,246 novel lncRNAs and 9635 known lncRNAs were screened out, and 413 lncRNA transcripts and 65 mRNA transcripts were identified as being significantly different between the CP group and control group. Notably, we found that there are seven lncRNAs that can target Smad1 and Smad5, which are key molecules of bone morphogenetic protein (BMP) signaling pathway, which suggested that they may be concerned with BMP signaling in TCDD-induced CP. In addition, some lncRNAs targeted the important molecules of Hippo and Wnt signaling pathways. These results suggested that characteristic lncRNA alterations may play a critical role in TCDD-induced CP, which provided a theoretical basis for further research.

[An analysis of the characteristics of perioperative cardiac troponin elevation in orthopaedic surgical patients].

Objective: Perioperative myocardial infarction remains a severe complication in non-cardiac surgery and is one of the major causes of death. Cardiac troponin (cTn) I elevation is associated with short-term and long-term mortality. The aim of the study was to assess the proportion rate of cTnI elevation and its clinical characteristics among patients admitted for orthopaedic surgery with or without cardiovascular events. Methods: This is a retrospective study including 27 744 patients aged 50 years or older who admitted for orthopaedic surgery from 2009-2015 in Beijing Jishuitan Hospital. Results: Two hundred and sixty-five patients [age (71.7±9.9) years] had cTnI level> 0.04 µg/L with 66% (175 patients) of them being female. Among them, 59 patients were isolated troponin rise (ITR) (n=59), 13 were preoperative acute myocardial infarction (AMI), and 193 were postoperative AMI. The proportion of postoperative AMI was 0.69%. Those patients were more likely to have a history of coronary artery disease or hypertension. Non-ST-segment elevation myocardial infarction (NSTEMI) was more common (93.3%) than ST-segment elevation myocardial infarction in these patients. Most of them did not experience ischemic symptoms. Totally 76.7% of the AMI occurred within 3 days of surgery; and the in-hospital mortality rate was 10.4%. Conclusions: Perioperative elevation of troponin is common in patients undergoing orthopaedic surgery. Most postoperative AMI were NSTEMI and with absent or atypical ischemia symptoms. Monitoring troponin levels and electrocardiograph in at-risk patients is needed to find most of the AMI.

Stimulation of dendritic cell maturation and induction of apoptosis in lymphoma cells by a stable lectin from buckwheat seeds.

The present study aims to purify and characterize lectin from tartary buckwheat seeds and study its properties as well as biological activities to determine its possible biomedical applications in promoting maturation and proliferation of peripheral blood DCs derived from healthy donors and to study the effect of inducing apoptosis in human leukemia U937 cells. A novel tartary buckwheat lectin (TBL) protein, purified from tartary buckwheat seeds, showed a single band with a molecular mass of 65 kDa in SDS-PAGE. The purified TBL hemagglutinated both human and animal erythrocytes and showed preference for blood type O and the rabbit blood type. TBL is active at up to 60°C, and it is acid- and alkali-stable. TBL (25 µg/mL) combined with 5 x 10(-5) M rhIL-4 promotes maturation and proliferation of peripheral blood dendritic cells (DCs), which is stronger than that promoted by rhTNF-α (20 ng/mL). Exposure of DCs to 50 µg/mL TBL for 48 h resulted in extensive upregulation of maturation markers CD83 and CD40. These TBL-DCs were capable of producing several pro-inflammatory cytokines such as interleukin-10 (IL-10) and interleukin-12 (IL-12). The results of the treatment of human leukemia U937 cells with TBL in doses of 12.5, 25, 50, and 100 µg/mL showed that tartary buckwheat-derived lectin induces apoptosis in a dose-dependent manner. Our results encourage the use of tartary buckwheat and tartary buckwheat-derived lectins as immunopotentiating foods, targeted to strengthen immune responses and display a potential dietary supplement for cancer prevention.

Rutin, quercetin, and free amino acid analysis in buckwheat (Fagopyrum) seeds from different locations.

In this study, five common buckwheats and nine tartary buckwheats grown at different locations were analyzed for the contents of rutin, quercetin, and amino acids by high-performance liquid chromatography and spectrophotometry. The rutin content was higher than quercetin in buckwheat seeds. Rutin content was in the range from 0.05 (0.05 g per 100 g dry seeds) to 1.35% of buckwheat seeds. Quercetin content varied from 0.01 to 0.17% and in some common buckwheats it was even difficult to detect. Comparatively, tartary buckwheat seeds contained more rutin and quercetin than common buckwheat seeds. Meanwhile, the bran has higher rutin content than the farina in tartary buckwheat seeds, with a respective content of 0.45 to 1.19% and 0.14 to 0.67%. It was found that amino acid contents were around 1.79 to 12.65% (farina) and 5.74 to 7.89% (bran) in common buckwheats, and 1.73 to 5.63% (farina) and 2.64 to 16.78% (bran) in tartary buckwheat seeds. The highest total rutin content was found to be 1.35% in tartary buckwheat seeds from Sichuan, China. The highest total amounts of amino acid were detected to be 20.13% in tartary buckwheat seeds from Changzhi, Shanxi Province (China). Our results suggested that food products made of whole-buckwheat flour are healthier than those made of fine white flour.

Homology cloning, sequence characterization, and expression analysis of cDNA encoding electron transfer flavoprotein beta polypeptide in mud crab (Scylla paramamosain).

Electron transfer flavoproteins (ETFs) are αß-heterodimers found in eukaryotic mitochondria and bacteria. Herein we report a full-length complementary DNA of a mud crab (Scylla paramamosain) ETF ß subunit (Scpa-ETFB) isolated with a homology cloning strategy. The complete complementary DNA of the Scpa-ETFB contains a 17-nt 5'-untranslated region, a 765-nt open reading frame encoding 254 amino acids, and a 248-nt 3'-untranslated region. The high identity of Scpa-ETFB with ETFB in other organisms indicated that Scpa-ETFB is a new member of the ETFB family. Although the conserved motif associated with flavin adenine dinucleotide binding is absent in Scpa-ETFB, the signature sequences of the ETF superfamily were identified. Using reverse transcriptase polymerase chain reaction, we detected the messenger RNA transcript of Scpa-ETFB in high levels in the tissues of the hepatopancreas, ovary, heart, and muscle. Phylogenetic analysis showed that Scpa-ETFB is most closely related to the ETFB genes of Caligus rogercresseyi and Lepeophtheirus salmonis. These results provided basic information for elucidating the molecular mechanism of energy production in the mud crab.

Characteristic spectrum of very low-energy photoelectron from above-threshold ionization in the tunneling regime.

We report an experimental and theoretical study of very low-energy photoelectrons in tunneling ionization process from noble gas atoms interacting with ultrashort intense infrared laser pulses. A universal peak structure with electron energy well below 1 eV in the photoelectron spectrum, corresponding to the double-hump structure in the longitudinal momentum distribution, is identified experimentally for all atomic species. Our quantum and semiclassical analysis reveal the role of long-range Coulomb potential in the production of this very low-energy peak structure.

Rapid determination of 19 quinolone residues in spiked fish and pig muscle by high-performance liquid chromatography (HPLC) tandem mass spectrometry.

A multiresidue method has been developed for the confirmation and quantification of 19 quinolones (enrofloxacin, ciprofloxacin, norfloxacin, ofloxacin, flumequine, oxolinic acid, difloxacin, sarafloxacin, sparfloxacin, danofloxacin, fleroxacin, marbofloxacin, enoxacin, orbifloxacin, pefloxacin, nalidixic acid, pipemidic acid, lomefloxacin and cinoacin) in pig and fish by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The samples were extracted with acetonitrile, analytes separated by LC on a C18 column using 0.1% formic acid-methanol with a linear gradient elution programme, and detected by MS/MS. The linear range was 0.3-50 microg kg(-1) with correlation coefficients (r) more than 0.9956. The limits of detection were 0.1 microg kg(-1). Mean recoveries for each analyte in pig muscle and fish ranged from 75.3% to 96.3% and from 79.7% to 94.2% with relative standard deviations below 10%. The method is fast, safe, sensitive and precise, and can be used simultaneously to analyse residual quinolones.

Development of an enzyme-linked immunosorbent assay for the alpha-cyano pyrethroids multiresidue in Tai lake water.

A general and broad class selective competitive enzyme-linked immunosorbent assay (ELISA) for the detection of the alpha-cyano pyrethroids was developed. One class specific hapten, the 5-(3-benzylphenyl)-5-cyanopentanoic acid (CPBA-BE), was conjugated with BSA as an immunogen. Polyclonal antibodies, PY-antibodies, were generated. The assay with the most selectivity for the family alpha-cyano pyrethroids such as cyphenothrin was further optimized and tested for tolerance to cosolvent, pH, and ionic strength changes. The IC(50) values of the optimized immunoassay were 4.58 microg L(-1) for cyphenothrin, 5.62 microg L(-1) for fenpropathrin, 7.08 microg L(-1) for deltamethrin, 10.72 microg L(-1) for cypermethrin, 19.95 microg L(-1) for flucythrinate, and 28.18 microg L(-1) for esfenvalerate. The cross-reactivities of cyphenothrin, fenpropathrin, deltamethrin, cypermethrin, flucythrinate, and esfenvalerate were 100%, 81.49%, 64.68%, 42.72%, 22.96%, and 16.25%, respectively. The low detection limit (LDL) of this assay was 0.107 microg L(-1) for cyphenothrin. This method was used for the detection of the alpha-cyano pyrethroids in Tai lake water.