Microglia inflammatory response contributes to chronic constriction injury-induced neuropathic pain via miR-339/PFKFB3 axis

Purpose: To investigate the effect of miR-339 on neuropathic pain. Methods: A rat neuropathic pain model was established through chronic constriction injury (CCI). Expression of miR-339 in spinal cord was determined 14 days later. Microglial inflammatory response was evaluated using immunofluorescence analysis of ionized calcium binding adaptor molecule 1 (Iba1), while IL-6 and TNF-α were assessed by enzyme-linked immunosorbent assay (ELISA). Pain- associated behavioral effects and microglia-related inflammation were investigated after intrathecal administration of miR-339 agomir into rats post-CCI. The target gene of miR-339 involved in neuropathic pain was evaluated by a luciferase reporter assay. Microglia cells were isolated from rats, then treated with lipopolysaccharide (LPS). The LPS-induced inflammatory response in microglia cells was determined using quantitative reverse transcription PCR analysis of IL-6 and TNF-α. Results: CCI decreased mechanical allodynia and thermal hyperalgesia thresholds, but increased Iba1, IL-6, and TNF-α in rats. MiR-339 was reduced in rat spinal cord after CCI induction while intrathecal injection of miR-339 agomir alleviated CCI-induced changes in mechanical and thermal hyperalgesia in rats, and reversed expression of Iba1, IL-6, and TNF-α. Furthermore, 6-phosphofructo-2- kinase/fructose-2,6-bisphosphatase-3 (PFKFB3) was identified as a miR-339 target gene, and over- expression of miR-339 decreased the expression of PFKFB3, IL-6, and TNF-α in LPS-induced microglia cells. Conclusion: The miR-339/PFKFB3 axis ameliorates CCI-induced neuropathic pain by suppression of microglia inflammatory response, suggesting a novel strategy for neuropathic pain management

Characterization of the Biosynthesis, Processing and Kinetic Mechanism of Action of the Enzyme Deficient in Mucopolysaccharidosis IIIC

Heparin acetyl-CoA:alpha-glucosaminide N-acetyltransferase (N-acetyltransferase, EC 2.3.1.78) is an integral lysosomal membrane protein containing 11 transmembrane domains, encoded by the HGSNAT gene. Deficiencies of N-acetyltransferase lead to mucopolysaccharidosis IIIC. We demonstrate that contrary to a previous report, the N-acetyltransferase signal peptide is co-translationally cleaved and that this event is required for its intracellular transport to the lysosome. While we confirm that the N-acetyltransferase precursor polypeptide is processed in the lysosome into a small amino-terminal alpha- and a larger ß- chain, we further characterize this event by identifying the mature amino-terminus of each chain. We also demonstrate this processing step(s) is not, as previously reported, needed to produce a functional transferase, i.e., the precursor is active. We next optimize the biochemical assay procedure so that it remains linear as N-acetyltransferase is purified or protein-extracts containing N-acetyltransferase are diluted, by the inclusion of negatively charged lipids. We then use this assay to demonstrate that the purified single N-acetyltransferase protein is both necessary and sufficient to express transferase activity, and that N-acetyltransferase functions as a monomer. Finally, the kinetic mechanism of action of purified N-acetyltransferase was evaluated and found to be a random sequential mechanism involving the formation of a ternary complex with its two substrates; i.e., N-acetyltransferase does not operate through a ping-pong mechanism as previously reported. We confirm this conclusion by demonstrating experimentally that no acetylated enzyme intermediate is formed during the reaction

Pemetrexed plus Platinum as the First-Line Treatment Option for Advanced Non-Small Cell Lung Cancer: A Meta-Analysis of Randomized Controlled Trials

To compare the efficacy and toxicities of pemetrexed plus platinum with other platinum regimens in patients with previously untreated advanced non-small cell lung cancer (NSCLC). Methods: A meta-analysis was performed using trials identified through PubMed, EMBASE, and Cochrane databases. Two investigators independently assessed the quality of the trials and extracted data. The outcomes included overall survival (OS), progression-free survival (PFS), response rate (RR), and different types of toxicity. Hazard ratios (HRs), odds ratios (ORs) and their 95% confidence intervals (CIs) were pooled using RevMan software. Results: Four trials involving 2,518 patients with previously untreated advanced NSCLC met the inclusion criteria. Pemetrexed plus platinum chemotherapy (PPC) improved survival compared with other platinum-based regimens (PBR) in patients with advanced NSCLC (HR = 0.91, 95% CI: 0.83–1.00, p = 0.04), especially in those with non-squamous histology (HR = 0.87, 95% CI: 0.77–0.98, p = 0.02). No statistically significant improvement in either PFS or RR was found in PPC group as compared with PBR group (HR = 1.03, 95% CI: 0.94–1.13, p = 0.57; OR = 1.15, 95% CI: 0.95–1.39, p = 0.15, respectively). Compared with PBR, PPC led to less grade 3–4 neutropenia and leukopenia but more grade 3–4 nausea. However, hematological toxicity analysis revealed significant heterogeneities. Conclusion: Our results suggest that PPC in the first-line setting leads to a significant survival advantage with acceptable toxicities for advanced NSCLC patients, especially those with non-squamous histology, as compared with other PRB. PPC could be considered as the first-line treatment option for advanced NSCLC patients, especially those with non-squamous histology

A C19MC-LIN28A-MYCN Oncogenic Circuit Driven by Hijacked Super-enhancers Is a Distinct Therapeutic Vulnerability in ETMRs: A Lethal Brain Tumor

© 2019 Elsevier Inc. Embryonal tumors with multilayered rosettes (ETMRs) are highly lethal infant brain cancers with characteristic amplification of Chr19q13.41 miRNA cluster (C19MC) and enrichment of pluripotency factor LIN28A. Here we investigated C19MC oncogenic mechanisms and discovered a C19MC-LIN28A-MYCN circuit fueled by multiple complex regulatory loops including an MYCN core transcriptional network and super-enhancers resulting from long-range MYCN DNA interactions and C19MC gene fusions. Our data show that this powerful oncogenic circuit, which entraps an early neural lineage network, is potently abrogated by bromodomain inhibitor JQ1, leading to ETMR cell death. Sin-Chan et al. uncover a C19MC-LIN28A-MYCN super-enhancer-dependent oncogenic circuit in embryonal tumors with multilayered rosettes (ETMRs). The circuit entraps an early neural lineage network to sustain embryonic epigenetic programming and is vulnerable to bromodomain inhibition, which promotes ETMR cell death

Investigation of extracellular esterase activities from thermophilic fungi, and purification and characterization of novel small enzymes

The existence of naturally occurring, very small enzymes (<10 kDa, microenzymes) has been established with the findings that some enzyme activities are associated with proteins smaller than 10 kDa. This work investigated the extracellular esterase activities observed in some thermophilic fungi, and purified and characterised three novel thermostable microenzymes with esterase activities. Among the nine thermophilic fungi studied, two esterase-positive organisms, Emericella nidulans and Talaromyces emersonii, exhibited the enzyme activities in less than 10 kDa fraction when grown in either malt extract medium or a synthetic medium. Two small enzymes (E40 and E32) from E. nidulans and one (T40) from T emersonii were identified, and purified to homogeneity by ultrafiltration, gel filtration and reverse-phase HPLC. Nondenaturing gel filtration showed that MWs of E40 and T40 were 1.6 kDa, and E32 was 4.1 kDa, while electrospray and MALDI mass spectrometry indicated monomeric MWs of 510.3,609.3 and 1424.7 Da for E40, T40 and E32 respectively. This was consistent with a trimer structure in solution. Sequence analysis revealed that all the three esterases had the (Gly-Pro-Hyp)n repeating unit, the characteristic of collagen. The esterase activities were associated with small diffusible factor(s). X-ray microanalysis indicated the esterases contained Zn and Al. The esterases exhibited extremely high thermostability and unusual pH stability. They were more active against short chain-length fatty acids than long ones and hydrolysed glycerol esters with 1,3 specificity. An attempt at chemical synthesis of the enzymes showed that the synthetic peptide itself was inactive. Circular dichroism spectra showed that the native esterase possessed more triple-helical conformation than the synthetic peptide.The existence of naturally occurring, very small enzymes (<10 kDa, microenzymes) has been established with the findings that some enzyme activities are associated with proteins smaller than 10 kDa. This work investigated the extracellular esterase activities observed in some thermophilic fungi, and purified and characterised three novel thermostable microenzymes with esterase activities. Among the nine thermophilic fungi studied, two esterase-positive organisms, Emericella nidulans and Talaromyces emersonii, exhibited the enzyme activities in less than 10 kDa fraction when grown in either malt extract medium or a synthetic medium. Two small enzymes (E40 and E32) from E. nidulans and one (T40) from T emersonii were identified, and purified to homogeneity by ultrafiltration, gel filtration and reverse-phase HPLC. Nondenaturing gel filtration showed that MWs of E40 and T40 were 1.6 kDa, and E32 was 4.1 kDa, while electrospray and MALDI mass spectrometry indicated monomeric MWs of 510.3,609.3 and 1424.7 Da for E40, T40 and E32 respectively. This was consistent with a trimer structure in solution. Sequence analysis revealed that all the three esterases had the (Gly-Pro-Hyp)n repeating unit, the characteristic of collagen. The esterase activities were associated with small diffusible factor(s). X-ray microanalysis indicated the esterases contained Zn and Al. The esterases exhibited extremely high thermostability and unusual pH stability. They were more active against short chain-length fatty acids than long ones and hydrolysed glycerol esters with 1,3 specificity. An attempt at chemical synthesis of the enzymes showed that the synthetic peptide itself was inactive. Circular dichroism spectra showed that the native esterase possessed more triple-helical conformation than the synthetic peptide

An Improved Adaptive-Torque-Gain MPPT Control for Direct-Driven PMSG Wind Turbines Considering Wind Farm Turbulences

Maximum power point tracking (MPPT) plays an important role in increasing the efficiency of a wind energy conversion system (WECS). In this paper, three conventional MPPT methods are reviewed: power signal feedback (PSF) control, decreased torque gain (DTG) control, and adaptive torque gain (ATG) control, and their potential challenges are investigated. It is found out that the conventional MPPT method ignores the effect of wind turbine inertia and wind speed fluctuations, which lowers WECS efficiency. Accordingly, an improved adaptive torque gain (IATG) method is proposed, which customizes adaptive torque gains and enhances MPPT performances. Specifically, the IATG control considers wind farm turbulences and works out the relationship between the optimal torque gains and the wind speed characteristics, which has not been reported in the literature. The IATG control is promising, especially under the ongoing trend of building wind farms with large-scale wind turbines and at low and medium wind speed sites

Puf mediates translation repression of transmission-blocking vaccine candidates in malaria parasites.

Translational control of gene expression plays an essential role in development. In malaria parasites, translational regulation is critical during the development of specialized transition stages between the vertebrate host and mosquito vector. Here we show that a Pumilio/FBF (Puf) family RNA-binding protein, PfPuf2, is required for the translation repression of a number of transcripts in gametocytes including two genes encoding the transmission-blocking vaccine candidates Pfs25 and Pfs28. Whereas studies to date support a paradigm of Puf-mediated translation regulation through 3' untranslated regions (UTRs) of target mRNAs, this study, for the first time, identifies a functional Puf-binding element (PBE) in the 5'UTR of pfs25. We provide both in vitro and in vivo evidence to demonstrate that PfPuf2 binds to the PBEs in pfs25 and pfs28 to mediate translation repression. This finding provides a renewed view of Pufs as versatile translation regulators and sheds light on their functions in the development of lower branches of eukaryotes

Zebrafish prmt7 negatively regulates antiviral responses by suppressing the retinoic acid-inducible gene-I-like receptor signaling

Arginine methylation is a post-translational modification in histone and nonhistone proteins that can affect numerous cellular activities. Protein arginine methyltransferase 7 (Prmt7), a type III arginine methyltransferase, catalyzes the formation of stable monomethylarginines of histones. The role of PRMT7 in virus-induced innate immunity signaling, however, remains largely unknown. We demonstrate that zebrafish prmt7 could be inhibited by spring viremia of carp virus (SVCV) and grass carp reovirus (GCRV) infection. The overexpression of prmt7 suppresses cellular antiviral responses that are partially dependent on the arginine methyltransferase activity of prmt7. Consistently, prmt7-null zebrafish were more resistant to SVCV or GCRV infection, exhibiting enhanced expression of key antiviral genes and fewer necrotic cells in the liver and kidney upon viral infection. Furthermore, we established a zebrafish model to investigate grass carp hemorrhagic disease. Our findings suggest that by suppressing the RIG-I-like receptors signaling, zebrafish prmt7 negatively regulates antiviral responses, indicating the vital role of prmt7 and its arginine methyltransferase activity in innate immunity