Now showing items 1-20 of 194

    • Extreme climate response to marine cloud brightening in the arid Sahara-Sahel-Arabian Peninsula zone

      Zhu, Yuanzhuo; Zhang, Zhihua; Crabbe, M. James C.; Shandong University; Beijing Normal University; Oxford University; University of Bedfordshire; Shanxi University (Emerald, 2021-02-08)
      Purpose Climatic extreme events are predicted to occur more frequently and intensely and will significantly threat the living of residents in arid and semi-arid regions. Therefore, this study aims to assess climatic extremes’ response to the emerging climate change mitigation strategy using a marine cloud brightening (MCB) scheme. Design/methodology/approach Based on Hadley Centre Global Environmental Model version 2-Earth System model simulations of a MCB scheme, this study used six climatic extreme indices [i.e. the hottest days (TXx), the coolest nights (TNn), the warm spell duration (WSDI), the cold spell duration (CSDI), the consecutive dry days (CDD) and wettest consecutive five days (RX5day)] to analyze spatiotemporal evolution of climate extreme events in the arid Sahara-Sahel-Arabian Peninsula Zone with and without MCB implementation. Findings Compared with a Representative Concentration Pathways 4.5 scenario, from 2030 to 2059, implementation of MCB is predicted to decrease the mean annual TXx and TNn indices by 0.4–1.7 and 0.3–2.1°C, respectively, for most of the Sahara-Sahel-Arabian Peninsula zone. It would also shorten the mean annual WSDI index by 118–183 days and the mean annual CSDI index by only 1–3 days, especially in the southern Sahara-Sahel-Arabian Peninsula zone. In terms of extreme precipitation, MCB could also decrease the mean annual CDD index by 5–25 days in the whole Sahara and Sahel belt and increase the mean annual RX5day index by approximately 10 mm in the east part of the Sahel belt during 2030–2059. Originality/value The results provide the first insights into the impacts of MCB on extreme climate in the arid Sahara-Sahel-Arabian Peninsula zone.
    • COVID-19 in Wuhan, China: pressing realities and city management

      Li, Rita Yi Man; Yue, Xiao-Guang; Crabbe, M. James C.; Hong Kong Shue Yan University; European University of Cyprus; Oxford University; University of Bedfordshire; Shanxi University (Frontiers, 2021-02-17)
      To most economists around the World, Covid-19 has provided an objective lesson in market failure. In the absence of complete information and sometimes even fake news, nobody knew what kind of pandemic it was at the beginning. Yet, there were 32,583 patients with laboratory-confirmed Covid-19 in Wuhan between December 8, 2019, and 8 March 8, 2020. The pandemic crippled and continues to cripple many health systems and has created unprecedented pressure on the psychological and physical aspects of millions of people's lives around the world. Over 200 countries and territories suffer from an acute shortage of medical personnel and medical equipment. The responses of different countries to Covid-19 has involved a range of measures that reflect national values, politics, and variations in scientific advice provided by local experts. Political considerations have often become more important than science. The Covid-19 outbreak in Wuhan was one of the most serious cases amongst all cities in the world, yet Wuhan managed and gain control of this pandemic. Health care systems and policies are important aspects that affect the control of infectious diseases like Covid-19. In China, 98% of primary health-care is complemented by traditional Chinese medicine (TCM) with allopathic approaches. Previous research has found that the cure rate increased by 33% among mild cases after adopting TCM with allopathic approaches. The hospital stay of severe patients with TCM's and nucleic acid turning negative was shortened by over 2 days. Prior to Covid-19, the government's Healthy China 2030 plan was already addressing chronic diseases in the aging population by raising healthcare expenditure. This is in sharp contrast to other countries with aging population problems such as Italy, where the government cut the healthcare budget substantially after the economic downturn. Hospital bed allocation went down from a maximum of four for every thousand inhabitants to a maximum of 3.7, despite the fact that 23.1% of the Italian population were aged 65 years and older in 2020. Likewise, the post-2008 financial crisis in Spain forced severe cuts to healthcare costs, which caused pressure on the system when there was an increase in demand for healthcare services. These measures particularly affected the elderly and disabled who are more vulnerable to Covid-19. Healthcare costs become underfunded at the level of 6.4% of GDP. Apart from scientific evidence on the effectiveness of TCM in curing covid, financial expenditures on health care is an important distal factor that helped Wuhan overcome Covid-19 quickly. In the following sections of this paper, we review the three city management stages adopted in Wuhan, to study proximal causes of success in combating the virus: (1) strong government intervention early in the outbreak; (2) the city lockdown; and, (3) the use of digital measures, such as a health code, when the city reopened.
    • MicroRNAs for virus pathogenicity and host responses, identified in SARS-CoV-2 genomes, may play roles in viral-host co-evolution in putative zoonotic host species

      Lange, Sigrun; Arisan, Elif Damla; Grant, Guy H.; Uysal-Onganer, Pinar; University of Westminster; Gebze Technical University; University of Bedfordshire (MDPI, 2021-01-16)
      Our recent study identified seven key microRNAs (miR-8066, 5197, 3611, 3934-3p, 1307-3p, 3691-3p, 1468-5p) similar between SARS-CoV-2 and the human genome, pointing at miR-related mechanisms in viral entry and the regulatory effects on host immunity. To identify the putative roles of these miRs in zoonosis, we assessed their conservation, compared with humans, in some key wild and domestic animal carriers of zoonotic viruses, including bat, pangolin, pig, cow, rat, and chicken. Out of the seven miRs under study, miR-3611 was the most strongly conserved across all species; miR-5197 was the most conserved in pangolin, pig, cow, bat, and rat; miR-1307 was most strongly conserved in pangolin, pig, cow, bat, and human; miR-3691-3p in pangolin, cow, and human; miR-3934-3p in pig and cow, followed by pangolin and bat; miR-1468 was most conserved in pangolin, pig, and bat; while miR-8066 was most conserved in pangolin and pig. In humans, miR-3611 and miR-1307 were most conserved, while miR-8066, miR-5197, miR-3334-3p and miR-1468 were least conserved, compared with pangolin, pig, cow, and bat. Furthermore, we identified that changes in the miR-5197 nucleotides between pangolin and human can generate three new miRs, with differing tissue distribution in the brain, lung, intestines, lymph nodes, and muscle, and with different downstream regulatory effects on KEGG pathways. This may be of considerable importance as miR-5197 is localized in the spike protein transcript area of the SARS-CoV-2 genome. Our findings may indicate roles for these miRs in viral-host co-evolution in zoonotic hosts, particularly highlighting pangolin, bat, cow, and pig as putative zoonotic carriers, while highlighting the miRs' roles in KEGG pathways linked to viral pathogenicity and host responses in humans. This in silico study paves the way for investigations into the roles of miRs in zoonotic disease.
    • Targeted editing of SlMAPK6 using CRISPR/Cas9 technology to promote the development of axillary buds in tomato plants

      Li, Yunzhou; Yue, Ningbo; Basit, Abdul; Li, Yulong; Zhang, Dalong; Qin, Lei; Crabbe, M. James C.; Xu, Wen; Wang, Yong; Yan, Jianmin; et al. (Canadian Center of Science and Education, 2021-01-15)
      The mitogen-activated protein kinase (MAPK) cascade signaling system has been relatively conserved throughout the evolution of eukaryotes and is involved in the regulation of growth and development and metabolism. In this study, dwarf tomato plants were used as the research material. First, the tissue-specific expression of SlMAPK6 was measured in wild-type plants by quantitative RT-PCR. The results showed that SlMAPK6 was highly expressed in the tissues of the stems, leaves and flowers but was expressed at low levels in the tissues of the roots, sepals and fruits. Second, SlMAPK6-knockout lines CRISPR-3 and CRISPR-7 were obtained by CRISPR-Cas9 technology and Agrobacterium-mediated transformation. Compared with wild-type, the mutant lines CRISPR-3 and CRISPR-7 showed significant phenotypic characteristics, such as increased numbers of axillary buds and true leaves, thickened stems, and longer leaflets. In addition, to explore the molecular mechanism by which MAPK regulates axillary bud growth, we also showed that SlMAPK6 positively regulates the strigolactone synthesis genes SlCCD7 and SlCCD8 and the gibberellin (GA) synthesis genes GA20ox3 and GA3ox1 and negatively regulates the axillary bud development-related genes Ls, BL and BRC1b/TCP8 and the GA synthesis inhibitory gene GAI. Therefore, SlMAPK6 appears to regulate the synthesis of strigolactone and GA to induce the growth and development of tomato axillary buds.
    • Bovine milk fat globule epidermal growth factor Ⅷ activates PI3K/Akt signaling pathway and attenuates sarcopenia in rat model induced by D-galactose

      Li, He; Wang, Rongchun; Wang, Lifeng; Li, Lin; Ma, Ying; Zhou, Shaobo; Jiangsu Normal University; Harbin Institute of Technology; Northeast Agriculture University; University of Bedfordshire (Elsevier, 2020-12-17)
      To develop a more effective and safer treatment for sarcopenia, this research investigated the anti-sarcopenia mechanism of Milk Fat Globule Epidermal Growth Factor Ⅷ (MFG-E8) from the liver function and metabolism in sarcopenic model rat. After 4 weeks nutritional intervention experiment, MFG-E8 can significantly increase the gastrocnemius mass in rat. The mechanism of MFG-E8 in improving sarcopenia was related to its promotional capacity to the activities of superoxide dismutase (SOD) activity in serum, Glutamic-oxaloacetic transaminase (GOT) and glutamic-pyruvic transaminase (GPT) in liver. Meanwhile, MFG-E8 could also down-regulate obesity-related indicators, such as triglyceride (TG) and Non-esterified fatty acid (NEFA). The analysis of liver and gastrocnemius histopathology found that MFG-E8 could reduce the accumulation of fatty vesicles, improve liver function, thereby alleviating gastrocnemius tissue inflammation. In vitro experiments, myoblasts obtained from gastrocnemius tissue showed that MFG-E8 could reduce mitochondrial autophagy and inhibit cell apoptosis. In addition, MFG-E8 could up-regulate the phosphorylation level of PI3K via activating PI3K/Akt signaling pathway in gastrocnemius tissue, and promote the formation of muscle fibers, thereby increasing muscle mass. Moreover, MFG-E8 could also promote the formation of neuromuscular junctions by up-regulating the mRNA and protein expression of MusK in gastrocnemius.
    • Effect and mechanism of Ganoderma lucidum spores on alleviation diabetic cardiomyopathy in a pilot in vivo study

      Shaher, Fahmi; Wang, Shuqiu; Qiu, Hong-Bin; Hu, Yu; Zhang, Yu; Wang, Weiqun; AL-ward, Hisham; Abdulghani, Mahfoudh A. M.; Baldi, Salem; Zhou, Shaobo; et al. (Dove Press, 2020-12-07)
      Background: Ganoderma lucidum spores (GLS) exhibit disease prevention properties, but no study has been carried out on the anti-diabetic cardiomyopathy property of GLS. The aim of this study is to evaluate the hyperglycemia-mediated cardiomyopathy protection and mechanisms of GLS in diabetic rats induced by streptozotocin (STZ). Methods: Male SD rats were randomly divided into three groups. Two groups were given STZ (50 mg/kg, i.p.) treatment and when their fasting plasma glucose was above 16.7 mmol/L, one group was given placebo, as diabetic group; and another group was given GLS (300 mg/kg) treatment. The group without STZ treatment was given placebo as a control group. The experiment lasted 70 days. The histology of myocardium and biomarkers of antioxidant, myocardial injury, pro-inflammatory cytokines, pro-apoptotic proteins and phosphorylation of key proteins in PI3K/AKT pathway were assessed. Results: Biochemical analysis showed that GLS treatment significantly reduced the blood glucose (-20.3%) and triglyceride (-20.4%) levels compared to diabetic group without treatment. GLS treatment decreased the content of MDA (-25.6%) and activity of lactate dehydrogenase (-18.9%) but increased the activity of GSH-Px (65.4%). Western blot analysis showed that GLS treatment reduced the expression of both alpha-smooth muscle actin and brain natriuretic peptide. Histological analysis on the cardiac tissue micrographs showed that GLS treatment reduced the collagen fibroses and glycogen reactivity in myocardium. Both western blot and immunohistochemistry analyses showed that GLS treatment decreased the expression levels of pro-inflammatory factors (cytokines IL-1β, and TNF-α) as well as apoptosis regulatory proteins (Bax, caspase-3 and -9), but increased the Bcl-2. Moreover, GLS treatment significantly increased the phosphorylation of key proteins involved in PI3K/AKT pathway, e.g. p-AKT p-PI3K and mTOR. Conclusion: The results indicated that GLS treatment alleviates diabetic cardiomyopathy by reducing hyperglycemia, oxidative stress, inflammation, apoptosis and further attenuating the fibrosis and myocardial dysfunction induced by STZ through the stimulation of the PI3K/Akt/mTOR signaling pathway.
    • Lamellipodin promotes invasive 3D cancer cell migration via regulated interactions with Ena/VASP and SCAR/WAVE

      Carmona, G.; Perera, U.; Gillett, C.; Naba, A.; Law, Ah-Lai; Sharma, V.P.; Wang, J.; Wyckoff, J.; Balsamo, M.; Mosis, F.; et al. (Nature Publishing Group, 2016-03-21)
      Cancer invasion is a hallmark of metastasis. The mesenchymal mode of cancer cell invasion is mediated by elongated membrane protrusions driven by the assembly of branched F-actin networks. How deregulation of actin regulators promotes cancer cell invasion is still enigmatic. We report that increased expression and membrane localization of the actin regulator Lamellipodin correlate with reduced metastasis-free survival and poor prognosis in breast cancer patients. In agreement, we find that Lamellipodin depletion reduced lung metastasis in an orthotopic mouse breast cancer model. Invasive 3D cancer cell migration as well as invadopodia formation and matrix degradation was impaired upon Lamellipodin depletion. Mechanistically, we show that Lamellipodin promotes invasive 3D cancer cell migration via both actin-elongating Ena/VASP proteins and the Scar/WAVE complex, which stimulates actin branching. In contrast, Lamellipodin interaction with Scar/WAVE but not with Ena/VASP is required for random 2D cell migration. We identified a phosphorylation-dependent mechanism that regulates selective recruitment of these effectors to Lamellipodin: Abl-mediated Lamellipodin phosphorylation promotes its association with both Scar/WAVE and Ena/VASP, whereas Src-dependent phosphorylation enhances binding to Scar/WAVE but not to Ena/VASP. Through these selective, regulated interactions Lamellipodin mediates directional sensing of epidermal growth factor (EGF) gradients and invasive 3D migration of breast cancer cells. Our findings imply that increased Lamellipodin levels enhance Ena/VASP and Scar/WAVE activities at the plasma membrane to promote 3D invasion and metastasis.
    • Cleavage of mer tyrosine kinase (MerTK) from the cell surface contributes to the regulation of retinal phagocytosis

      Law, Ah-Lai; Parinot, Celia; Chatagnon, Jonathan; Gravez, Basile; Sahel, Jose-Alain; Bhattacharya, Shomi S.; Nandrot, Emeline F.; ; Sorbonne Universite; University College London; et al. (American Society for Biochemistry and Molecular Biology Inc., 2014-12-23)
      Background: The MerTK receptor is necessary for retinal phagocytosis and its daily rhythm. Results: MerTK is cleaved from the apical cell surface in vitro and in vivo, reducing the phagocytic capacity of RPE cells. Conclusion: MerTK cleavage might help control the duration of the daily phagocytic peak. Significance: Our data show that extracellular cleavage of MerTK partly regulates retinal phagocytosis.
    • Stimulation of GLP-1 secretion downstream of the ligand-gated ion channel TRPA1

      Emery, Edward C.; Diakogiannaki, Eleftheria; Gentry, Clive; Psichas, Arianna; Habib, Abdella M.; Bevan, Stuart; Fischer, Michael J.M.; Reimann, Frank; Gribble, Fiona M.; ; et al. (American Diabetes Association Inc., 2014-10-16)
      Stimulus-coupled incretin secretion from enteroendocrine cells plays a fundamental role in glucose homeostasis and could be targeted for the treatment of type 2 diabetes. Here, we investigated the expression and function of transient receptor potential (TRP) ion channels in enteroendocrine L cells producing GLP-1. By microarray and quantitative PCR analysis, we identified trpa1 as an L cell-enriched transcript in the small intestine. Calcium imaging of primary L cells and the model cell line GLUTag revealed responses triggered by the TRPA1 agonists allyl-isothiocyanate (mustard oil), carvacrol, and polyunsaturated fatty acids, which were blocked by TRPA1 antagonists. Electrophysiology in GLUTag cells showed that carvacrol induced a current with characteristics typical of TRPA1 and triggered the firing of action potentials. TRPA1 activation caused an increase in GLP-1 secretion from primary murine intestinal cultures and GLUTag cells, an effect that was abolished in cultures from trpa1-/- mice or by pharmacological TRPA1 inhibition. These findings present TRPA1 as a novel sensory mechanism in enteroendocrine L cells, coupled to the facilitation of GLP-1 release, which may be exploitable as a target for treating diabetes.
    • Fbp17 and cip4 recruit ship2 and lamellipodin to prime the plasma membrane for fast endophilin-mediated endocytosis

      Hak, Laura Chan Wah; Khan, Shaheen; Meglio, Ilaria Di; Law, Ah-Lai; Häsler, Safa Lucken-Ardjomande; Quintaneiro, Leonor M.; Ferreira, Antonio P.A.; Krause, Matthias; McMahon, Harvey T.; Boucrot, Emmanuel; et al. (Nature Publishing Group, 2018-07-30)
      Endocytosis mediates the cellular uptake of micronutrients and the turnover of plasma membrane proteins. Clathrin-mediated endocytosis is the major uptake pathway in resting cells 1 , but several clathrin-independent endocytic routes exist in parallel 2,3 . One such pathway, fast endophilin-mediated endocytosis (FEME), is not constitutive but triggered upon activation of certain receptors, including the β 1 adrenergic receptor 4 . FEME activates promptly following stimulation as endophilin is pre-enriched by the phosphatidylinositol-3,4-bisphosphate-binding protein lamellipodin 4,5 . However, in the absence of stimulation, endophilin foci abort and disassemble after a few seconds. Looking for additional proteins involved in FEME, we found that 20 out of 65 BAR domain-containing proteins tested colocalized with endophilin spots. Among them, FBP17 and CIP4 prime the membrane of resting cells for FEME by recruiting the 5′-lipid phosphatase SHIP2 and lamellipodin to mediate the local production of phosphati-dylinositol-3,4-bisphosphate and endophilin pre-enrichment. Membrane-bound GTP-loaded Cdc42 recruits FBP17 and CIP4, before being locally deactivated by RICH1 and SH3BP1 GTPase-activating proteins. This generates the transient assembly and disassembly of endophilin spots, which lasts 5–10 seconds. This mechanism periodically primes patches of the membrane for prompt responses upon FEME activation.
    • Lipid derivatives activate GPR119 and trigger GLP-1 secretion in primary murine L-cells

      Moss, Catherine E.; Glass, Leslie L.; Diakogiannaki, Eleftheria; Pais, Ramona; Lenaghan, Carol; Smith, David M.; Wedin, Marianne; Bohlooly-Y, Mohammad; Gribble, Fiona M.; Reimann, Frank; et al. (Elsevier Inc., 2015-07-02)
      Aims/hypothesis Glucagon-like peptide-1 (GLP-1) is an incretin hormone derived from proglucagon, which is released from intestinal L-cells and increases insulin secretion in a glucose dependent manner. GPR119 is a lipid derivative receptor present in L-cells, believed to play a role in the detection of dietary fat. This study aimed to characterize the responses of primary murine L-cells to GPR119 agonism and assess the importance of GPR119 for the detection of ingested lipid. Methods GLP-1 secretion was measured from murine primary cell cultures stimulated with a panel of GPR119 ligands. Plasma GLP-1 levels were measured in mice lacking GPR119 in proglucagon-expressing cells and controls after lipid gavage. Intracellular cAMP responses to GPR119 agonists were measured in single primary L-cells using transgenic mice expressing a cAMP FRET sensor driven by the proglucagon promoter. Results L-cell specific knockout of GPR119 dramatically decreased plasma GLP-1 levels after a lipid gavage. GPR119 ligands triggered GLP-1 secretion in a GPR119 dependent manner in primary epithelial cultures from the colon, but were less effective in the upper small intestine. GPR119 agonists elevated cAMP in ∼70% of colonic L-cells and 50% of small intestinal L-cells. Conclusions/interpretation GPR119 ligands strongly enhanced GLP-1 release from colonic cultures, reflecting the high proportion of colonic L-cells that exhibited cAMP responses to GPR119 agonists. Less GPR119-dependence could be demonstrated in the upper small intestine. In vivo, GPR119 in L-cells plays a key role in oral lipid-triggered GLP-1 secretion.
    • Gut hormone regulation and secretion via FFA1 and FFA4

      Gribble, Fiona M.; Diakogiannaki, Eleftheria; Reimann, Frank (Springer New York LLC, 2016-11-22)
      The digestion, absorption and utilisation of dietary triglycerides are controlled by gut hormones, released from enteroendocrine cells along the length of the gastrointestinal tract. Major players in the detection of ingested lipids are the free fatty acid receptors FFA1 and FFA4, which are highly expressed on enteroendocrine cells. These receptors are activated when free fatty acids (FFA) are absorbed across the intestinal epithelium, and provide a dynamic hormonal signal indicating that lipids are arriving in the bloodstream from the gut. This review addresses our current knowledge of how ingested triglycerides modulate gut hormone release via FFA1 and FFA4.
    • A novel orvinol analog, BU08028, as a safe opioid analgesic without abuse liability in primates

      Ding, Huiping; Czoty, Paul W.; Kiguchi, Norikazu; Cami-Kobeci, Gerta; Sukhtankar, Devki D.; Nader, Michael A.; Husbands, Stephen M.; Ko, Mei-Chuan; ; Wake Forest University; et al. (National Academy of Sciences, 2016-08-29)
      Despite the critical need, no previous research has substantiated safe opioid analgesics without abuse liability in primates. Recent advances in medicinal chemistry have led to the development of ligands with mixed mu opioid peptide (MOP)/nociceptin-orphanin FQ peptide (NOP) receptor agonist activity to achieve this objective. BU08028 is a novel orvinol analog that displays a similar binding profile to buprenorphine with improved affinity and efficacy at NOP receptors. The aim of this preclinical study was to establish the functional profile of BU08028 in monkeys using clinically used MOP receptor agonists for side-by-side comparisons in various wellhoned behavioral and physiological assays. Systemic BU08028 (0.001-0.01 mg/kg) produced potent long-lasting (i.e., >24 h) antinociceptive and antiallodynic effects, which were blocked by MOP or NOP receptor antagonists. More importantly, the reinforcing strength of BU08028 was significantly lower than that of cocaine, remifentanil, or buprenorphine in monkeys responding under a progressive-ratio schedule of drug self-administration. Unlike MOP receptor agonists, BU08028 at antinociceptive doses and ?10-to 30-fold higher doses did not cause respiratory depression or cardiovascular adverse events as measured by telemetry devices. After repeated administration, the monkeys developed acute physical dependence on morphine, as manifested by precipitated withdrawal signs, such as increased respiratory rate, heart rate, and blood pressure. In contrast, monkeys did not show physical dependence on BU08028. These in vivo findings in primates not only document the efficacy and tolerability profile of bifunctional MOP/NOP receptor agonists, but also provide a means of translating such ligands into therapies as safe and potentially abusefree opioid analgesics.
    • Conservation of inner nuclear membrane targeting sequences in mammalian Pom121 and yeast Heh2 membrane proteins

      Kralt, Annemarie; Basheer, Noorjahan Jagalur; Van Den Boom, Vincent; Lokareddy, Ravi K.; Steen, Anton; Cingolani, Gino; Fornerod, Maarten; Veenhoff, Liesbeth M.; ; University of Groningen; et al. (American Society for Cell Biology, 2015-07-15)
      Endoplasmic reticulum-synthesized membrane proteins traffic through the nuclear pore complex (NPC) en route to the inner nuclear membrane (INM). Although many membrane proteins pass the NPC by simple diffusion, two yeast proteins, ScSrc1/ScHeh1 and ScHeh2, are actively imported. In these proteins, a nuclear localization signal (NLS) and an intrinsically disordered linker encode the sorting signal for recruiting the transport factors for FG-Nup and RanGTP-dependent transport through the NPC. Here we address whether a similar import mechanism applies in metazoans. We show that the (putative) NLSs of metazoan HsSun2, MmLem2, HsLBR, and HsLap2β are not sufficient to drive nuclear accumulation of a membrane protein in yeast, but the NLS from RnPom121 is. This NLS of Pom121 adapts a similar fold as the NLS of Heh2 when transport factor bound and rescues the subcellular localization and synthetic sickness of Heh2ΔNLS mutants. Consistent with the conservation of these NLSs, the NLS and linker of Heh2 support INM localization in HEK293T cells. The conserved features of the NLSs of ScHeh1, ScHeh2, and RnPom121 and the effective sorting of Heh2-derived reporters in human cells suggest that active import is conserved but confined to a small subset of INM proteins.
    • Mastermind-Like 3 controls proliferation and differentiation in neuroblastoma

      Heynen, Guus J.J.E.; Nevedomskaya, Ekaterina; Palit, Sander; Basheer, Noorjahan Jagalur; Lieftink, Cor; Schlicker, Andreas; Zwart, Wilbert; Bernards, René; Bajpe, Prashanth Kumar; ; et al. (American Association for Cancer Research Inc., 2016-01-19)
      Neuroblastoma cell lines can differentiate upon treatment with retinoic acid (RA), a finding that provided the basis for the clinical use of RA to treat neuroblastoma. However, resistance to RA is often observed, which limits its clinical utility. Using a gain-of-function genetic screen, we identified an unexpected link between RA signaling and mastermindlike 3 (MAML3), a known transcriptional coactivator for NOTCH. Our findings indicate that MAML3 expression leads to the loss of activation of a subset of RA target genes, which hampers RA-induced differentiation and promotes resistance to RA. The regulatory DNA elements of this subset of RA target genes show overlap in binding of MAML3 and the RA receptor, suggesting a direct role for MAML3 in the regulation of these genes. In addition, MAML3 has RA-independent functions, including the activation of IGF1R and downstream AKT signaling via upregulation of IGF2, resulting in increased proliferation. These results demonstrate an important mechanistic role for MAML3 in proliferation and RA-mediated differentiation.
    • BU08073 a buprenorphine analogue with partial agonist activity at μ-receptors in vitro but long-lasting opioid antagonist activity in vivo in mice

      Khroyan, Taline V.; Wu, J.; Polgar, Willma E.; Cami-Kobeci, Gerta; Fotaki, N.; Husbands, Stephen M.; Toll, Lawrence; (John Wiley and Sons Inc., 2014-11-05)
      Background and Purpose Buprenorphine is a potent analgesic with high affinity at μ, δ and κ and moderate affinity at nociceptin opioid (NOP) receptors. Nevertheless, NOP receptor activation modulates the in vivo activity of buprenorphine. Structure activity studies were conducted to design buprenorphine analogues with high affinity at each of these receptors and to characterize them in in vitro and in vivo assays. Experimental Approach Compounds were tested for binding affinity and functional activity using [35S]GTPγS binding at each receptor and a whole‐cell fluorescent assay at μ receptors. BU08073 was evaluated for antinociceptive agonist and antagonist activity and for its effects on anxiety in mice. Key Results BU08073 bound with high affinity to all opioid receptors. It had virtually no efficacy at δ, κ and NOP receptors, whereas at μ receptors, BU08073 has similar efficacy as buprenorphine in both functional assays. Alone, BU08073 has anxiogenic activity and produces very little antinociception. However, BU08073 blocks morphine and U50,488‐mediated antinociception. This blockade was not evident at 1 h post‐treatment, but is present at 6 h and remains for up to 3–6 days. Conclusions and Implications These studies provide structural requirements for synthesis of ‘universal’ opioid ligands. BU08073 had high affinity for all the opioid receptors, with moderate efficacy at μ receptors and reduced efficacy at NOP receptors, a profile suggesting potential analgesic activity. However, in vivo, BU08073 had long‐lasting antagonist activity, indicating that its pharmacokinetics determined both the time course of its effects and what receptor‐mediated effects were observed.
    • Effects of stimulation of mu opioid and nociceptin/orphanin FQ peptide (NOP) receptors on alcohol drinking in rhesus monkeys

      Flynn, Shawn M.; Epperly, Phillip M.; Davenport, April T.; Cami-Kobeci, Gerta; Husbands, Stephen M.; Ko, Mei-Chuan; Czoty, Paul W. (Nature Publishing Group, 2019-04-10)
      Alcohol use disorder (AUD) persists as a devastating public health problem; widely effective pharmacological treatments are needed. Evidence from rodent models suggests that stimulating brain receptors for the neuropeptide nociceptin/orphanin FQ (NOP) can decrease ethanol drinking. We characterized the effects of the mu opioid peptide (MOP) receptor agonist buprenorphine and the buprenorphine analog (2S)-2-[(5R,6R,7R,14S)-N-cyclopropylmethyl-4,5-epoxy-6,14-ethano-3-hydroxy-6 methoxymorphinan-7-yl]-3,3-dimethylpentan-2-ol (BU08028), which stimulates MOP and NOP receptors, in a translational nonhuman primate model of AUD. Rhesus monkeys drank a 4% ethanol solution 6 h per day, 5 days per week via an operant behavioral panel in their home cages. To assess behavioral selectivity, monkeys responded via a photo-optic switch to earn food pellets. After characterizing the acute effects of BU08028 (0.001–0.01 mg/kg, i.m.) and buprenorphine (0.003–0.056 mg/kg, i.m.), the drugs were administered chronically using a model of pharmacotherapy assessment that incorporates clinical aspects of AUD and treatment. Acutely, both drugs decreased ethanol drinking at doses that did not affect food-maintained responding. During chronic treatment, effects of BU08028 and buprenorphine were maintained for several weeks without development of tolerance or emergence of adverse effects. BU08028 was ~0.5 and 1.0 log units more potent in acute and chronic studies, respectively. The selective NOP receptor agonist SCH 221510 also selectively decreased ethanol intakes when given acutely (0.03–1.0 mg/kg, i.m.), whereas the MOP antagonist naltrexone (1.7–5.6 mg/kg, i.m.) decreased both ethanol intake and food pellets delivered. These data demonstrate that bifunctional MOP/NOP agonists, which may have therapeutic advantages to MOP-selective drugs, can decrease alcohol drinking in nonhuman primates.
    • Synthesis, biological evaluation, and SAR studies of 14β-phenylacetyl substituted 17-cyclopropylmethyl-7, 8-dihydronoroxymorphinones derivatives: Ligands with mixed NOP and opioid receptor profile

      Kumar, Vinod; Polgar, Willma E.; Cami-Kobeci, Gerta; Thomas, Mark P.; Khroyan, Taline V.; Toll, Lawrence; Husbands, Stephen M.; ; Central University of Punjab; SRI International; et al. (Frontiers Media S.A., 2018-09-19)
      A series of 14β-acyl substituted 17-cyclopropylmethyl-7,8-dihydronoroxymorphinone compounds has been synthesized and evaluated for affinity and efficacy for mu (MOP), kappa (KOP), and delta (DOP) opioid receptors and nociceptin/orphanin FQ peptide (NOP) receptors. The majority of the new ligands displayed high binding affinities for the three opioid receptors, and moderate affinity for NOP receptors. The affinities for NOP receptors are of particular interest as most classical opioid ligands do not bind to NOP receptors. The predominant activity in the [35S]GTPγS assay was partial agonism at each receptor. The results are consistent with our prediction that an appropriate 14β side chain would access a binding site within the NOP receptor and result in substantially higher affinity than displayed by the parent compound naltrexone. Molecular modeling studies, utilizing the recently reported structure of the NOP receptor, are also consistent with this interpretation.
    • BU10038 as a safe opioid analgesic with fewer side-effects after systemic and intrathecal administration in primates

      Kiguchi, Norikazu; Ding, Huiping; Cami-Kobeci, Gerta; Sukhtankar, Devki D.; Czoty, Paul W.; DeLoid, Heather B.; Hsu, Fang-Chi; Toll, Lawrence; Husbands, Stephen M.; Ko, Mei-Chuan; et al. (Elsevier Ltd, 2019-03-01)
      Background: The marked increase in mis-use of prescription opioids has greatly affected our society. One potential solution is to develop improved analgesics which have agonist action at both mu opioid peptide (MOP) and nociceptin/orphanin FQ peptide (NOP) receptors. BU10038 is a recently identified bifunctional MOP/NOP partial agonist. The aim of this study was to determine the functional profile of systemic or spinal delivery of BU10038 in primates after acute and chronic administration. Methods: A series of behavioural and physiological assays have been established specifically to reflect the therapeutic (analgesia) and side-effects (abuse potential, respiratory depression, itch, physical dependence, and tolerance) of opioid analgesics in rhesus monkeys. Results: After systemic administration, BU10038 (0.001–0.01 mg kg−1) dose-dependently produced long-lasting antinociceptive and antihypersensitive effects. Unlike the MOP agonist oxycodone, BU10038 lacked reinforcing effects (i.e. little or no abuse liability), and BU10038 did not compromise the physiological functions of primates including respiration, cardiovascular activities, and body temperature at antinociceptive doses and a 10–30-fold higher dose (0.01–0.1 mg kg−1). After intrathecal administration, BU10038 (3 μg) exerted morphine-comparable antinociception and antihypersensitivity without itch scratching responses. Unlike morphine, BU10038 did not cause the development of physical dependence and tolerance after repeated and chronic administration. Conclusions: These in vivo findings demonstrate the translational potential of bifunctional MOP/NOP receptor agonists such as BU10038 as a safe, non-addictive analgesic with fewer side-effects in primates. This study strongly supports that bifunctional MOP/NOP agonists may provide improved analgesics and an alternative solution for the ongoing prescription opioid crisis.
    • Architecture and self-assembly of Clostridium sporogenes and Clostridium botulinum spore surfaces illustrate a general protective strategy across spore formers

      Janganan, Thamarai K.; Mullin, Nic; Dafis-Sagarmendi, Ainhoa; Brunt, Jason; Tzokov, Svetomir B.; Stringer, Sandra; Moir, Anne; Chaudhuri, Roy R.; Fagan, Robert P.; Hobbs, Jamie K.; et al. (American Society for Microbiology, 2020-07-01)
      Spores, the infectious agents of many Firmicutes, are remarkably resilient cell forms. Even distant relatives can have similar spore architectures although some display unique features; they all incorporate protective proteinaceous envelopes. We previously found that Bacillus spores can achieve these protective properties through extensive disulfide cross-linking of self-assembled arrays of cysteine-rich proteins. We predicted that this could be a mechanism employed by spore formers in general, even those from other genera. Here, we tested this by revealing in nanometer detail how the outer envelope (exosporium) in Clostridium sporogenes (surrogate for C. botulinum group I), and in other clostridial relatives, forms a hexagonally symmetric semipermeable array. A cysteine-rich protein, CsxA, when expressed in Escherichia coli, self-assembles into a highly thermally stable structure identical to that of the native exosporium. Like the exosporium, CsxA arrays require harsh “reducing” conditions for disassembly. We conclude that in vivo, CsxA self-organizes into a highly resilient, disulfide cross-linked array decorated with additional protein appendages enveloping the forespore. This pattern is remarkably similar to that in Bacillus spores, despite a lack of protein homology. In both cases, intracellular disulfide formation is favored by the high lattice symmetry. We have identified cysteine-rich proteins in many distantly related spore formers and propose that they may adopt a similar strategy for intracellular assembly of robust protective structures.