Identification of Co-Existing Mutations and Gene Expression Trends Associated With K13-Mediated Artemisinin Resistance in Plasmodium falciparum

Plasmodium falciparum infects millions and kills thousands of people annually the world over. With the emergence of artemisinin and/or multidrug resistant strains of the pathogen, it has become even more challenging to control and eliminate the disease. Multiomics studies of the parasite have started to provide a glimpse into the confounding genetics and mechanisms of artemisinin resistance and identified mutations in Kelch13 (K13) as a molecular marker of resistance. Over the years, thousands of genomes and transcriptomes of artemisinin-resistant/sensitive isolates have been documented, supplementing the search for new genes/pathways to target artemisinin-resistant isolates. This meta-analysis seeks to recap the genetic landscape and the transcriptional deregulation that demarcate artemisinin resistance in the field. To explore the genetic territory of artemisinin resistance, we use genomic single-nucleotide polymorphism (SNP) datasets from 2,517 isolates from 15 countries from the MalariaGEN Network (The Pf3K project, pilot data release 4, 2015) to dissect the prevalence, geographical distribution, and co-existing patterns of genetic markers associated with/enabling artemisinin resistance. We have identified several mutations which co-exist with the established markers of artemisinin resistance. Interestingly, K13-resistant parasites harbor α-ß hydrolase and putative HECT domain–containing protein genes with the maximum number of SNPs. We have also explored the multiple, publicly available transcriptomic datasets to identify genes from key biological pathways whose consistent deregulation may be contributing to the biology of resistant parasites. Surprisingly, glycolytic and pentose phosphate pathways were consistently downregulated in artemisinin-resistant parasites. Thus, this meta-analysis highlights the genetic and transcriptomic features of resistant parasites to propel further exploratory studies in the community to tackle artemisinin resistance.</p

B Cells Regulate Neutrophilia during Mycobacterium tuberculosis Infection and BCG Vaccination by Modulating the Interleukin-17 Response

We have previously demonstrated that B cells can shape the immune response to Mycobacterium tuberculosis, including the level of neutrophil infiltration and granulomatous inflammation at the site of infection. The present study examined the mechanisms by which B cells regulate the host neutrophilic response upon exposure to mycobacteria and how neutrophilia may influence vaccine efficacy. To address these questions, a murine aerosol infection tuberculosis (TB) model and an intradermal (ID) ear BCG immunization mouse model, involving both the μMT strain and B cell-depleted C57BL/6 mice, were used. IL (interleukin)-17 neutralization and neutrophil depletion experiments using these systems provide evidence that B cells can regulate neutrophilia by modulating the IL-17 response during M. tuberculosis infection and BCG immunization. Exuberant neutrophilia at the site of immunization in B cell-deficient mice adversely affects dendritic cell (DC) migration to the draining lymph nodes and attenuates the development of the vaccine-induced Th1 response. The results suggest that B cells are required for the development of optimal protective anti-TB immunity upon BCG vaccination by regulating the IL-17/neutrophilic response. Administration of sera derived from M. tuberculosis-infected C57BL/6 wild-type mice reverses the lung neutrophilia phenotype in tuberculous μMT mice. Together, these observations provide insight into the mechanisms by which B cells and humoral immunity modulate vaccine-induced Th1 response and regulate neutrophila during M. tuberculosis infection and BCG immunization. © 2013 Kozakiewicz et al

MicroRNA-196a & microRNA-101 expression in Barrett's oesophagus in patients with medically and surgically treated gastro-oesophageal reflux

<p>Abstract</p> <p>Background</p> <p>Proton pump inhibitor (PPI) medication and surgical fundoplication are used for the control of gastro-oesophageal reflux in patients with Barrett's oesophagus, but differ in their effectiveness for both acid and bile reflux. This might impact on the inflammatory processes that are associated with progression of Barrett's oesophagus to cancer, and this may be evident in the gene expression profile and microRNA expression pattern in Barrett's oesophagus mucosa. We hypothesised that two miRNAs with inflammatory and oncogenic roles, miR-101 and miR-196a, are differentially expressed in Barrett's oesophagus epithelium in patients with reflux treated medically vs. surgically.</p> <p>Findings</p> <p>Mucosal tissue was obtained at endoscopy from patients with Barrett's oesophagus whose reflux was controlled by proton pump inhibitor (PPI) therapy (n = 20) or by fundoplication (n = 19). RNA was extracted and the expression of miR-101 and miR-196a was measured using real-time reverse transcription - polymerase chain reaction. There were no significant differences in miR-101 and miR-196a expression in Barrett's oesophagus epithelium in patients treated by PPI vs. fundoplication (p = 0.768 and 0.211 respectively). Secondary analysis showed a correlation between miR-196a expression and Barrett's oesophagus segment length (p = 0.014).</p> <p>Conclusion</p> <p>The method of reflux treatment did not influence the expression of miR-101 and miR-196a in Barrett's oesophagus. This data does not provide support to the hypothesis that surgical treatment of reflux better prevents cancer development in Barrett's oesophagus. The association between miR-196a expression and Barrett's oesophagus length is consistent with a tumour promoting role for miR-196a in Barrett's oesophagus.</p

Coronary artery calcium screening: current status and recommendations from the European Society of Cardiac Radiology and North American Society for Cardiovascular Imaging

Current guidelines and literature on screening for coronary artery calcium for cardiac risk assessment are reviewed for both general and special populations. It is shown that for both general and special populations a zero score excludes most clinically relevant coronary artery disease. The importance of standardization of coronary artery calcium measurements by multi-detector CT is discussed

Effect of amino acid substitutions at the subunit interface on the stabiity and aggregation properties of a dimeric protein: role of Arg 178 and Arg 218 at the dimer interface of thymidylate synthase

The significance of two interface arginine residues on the structural integrity of an obligatory dimeric enzyme thymidylate synthase (TS) from Lactobacillus casei was investigated by thermal and chemical denaturation. While the R178F mutant showed apparent stability to thermal denaturation by its decreased tendency to aggregate, the Tm of the R218K mutant was lowered by 5 degrees C. Equilibrium denaturation studies in guanidinium chloride (GdmCl) and urea indicate that in both the mutants, replacement of Arg residues results in more labile quaternary and tertiary interactions. Circular dichroism studies in aqueous buffer suggest that the protein interior in R218K may be less well-packed as compared to the wild type protein. The results emphasize that quaternary interactions may influence the stability of the tertiary fold of TS. The amino acid replacements also lead to notable alteration in the ability of the unfolding intermediate of TS to aggregate. The aggregated state of partially unfolded intermediate in the R178F mutant is stable over a narrower range of denaturant concentrations. In contrast, there is an exaggerated tendency on the part of R218K to aggregate in intermediate concentrations of the denaturant. The 3 A crystal structure of the R178F mutant reveals no major structural change as a consequence of amino acid substitution. The results may be rationalized in terms of mutational effects on both the folded and unfolded state of the protein. Site specific amino acid substitutions are useful in identifying specific regions of TS involved in association of non-native protein structures

Crystals of a thymidylate synthase mutant offer insights into crystal packing and plasticity of protein-protein contacts

Some crystal forms of thymidylate synthase from L. casei exhibit unit sell transformation upon irradiation by X-rays, These forms, all of which occur in the space group P6(1)22, show an elongation in the c cell dimension and in some cases stabilize to a constant cell dimension upon prolonged exposure. We present here an analysis of the possible causes of this transformation based on the crystal structures for two forms of an R178F mutant of this enzyme. We compare these structures to other structures with intermediate cell parameters reported in the literature. There are no large changes in the dimeric structure of TS in these crystal forms. Although there is a large change in the unit cell volume, the molecular contacts in the crystal structures are nearly invariant. The transformation appears to result from concerted small changes in molecular structure and intermolecular contacts. These observations corroborate the general impression that protein structures can accommodate minor changes in sequence or packing wherein the intra and intermolecular interactions are not seriously altered

Original Inventions based on Chemical scaffolds and electro-physical activity-derived biosimilars interacting with specialties in biology yielding platforms for analysis in virology and antiviral compounds

Background: Original inventions in developing countries, in terms of number of patents filed, granted and that are taken to useful applications as well as in terms of publications of high impact remain relatively lower [1,2] compared to that of developed nations. The reasons could be attributed to lack of importance given to basic research in funding, the number of institutes involved, limited technical support or expertise available etc [1]. Though such initiatives may take a long time to yield fruits, one of the parallel steps we considered worth was to take the original inventions from Japan, born out of basic research in one field, taken through an application-oriented inter disciplinary interactive research in healthcare, thereby paving way for novel solutions. Thus was conceived, the Inventions- Inter-Disciplinary Interactions and Solutions (IIDIAS), an academic session as a part of the one-day International stem cell meet organized every year in the month of October by Nichi-In Centre for Regenerative Medicine (NCRM), an academic Institute based in Chennai, India. In the IIDIAS session, based on original invention(s) presented as a prelude in brief, original interdisciplinary interactive research work based on the original invention by NCRM and/or its collaborators are presented by the faculty of the relevant institute. That will be followed by an interactive session in which the potential solutions based on the above accomplishments would be discussed. The IIDIAS session 2014 was based on the following two inventions: A bio-film-based biosimilar invented by an electro-physicist A unique polymer invented by a chemical engineer Inventions and Interdisciplinary Interactions: Invention –I: The bio-film-based biosimilar invented by an electro-physicist: An electro physicist with the Kyoto University, Japan, Dr Nobuyuki Yamaji observed during his experiments that plants and mammalian tissues secrete a layer of fluid after getting hit by lightning or electric current [3]. The significance of the secretion was unknown to him. Later when he met his friend, Dr. Sunao Kubota, who was a physician and professor of General Surgery in St. Marianna University, he shared his work and Dr. Kubota began analyzing the nature of the secretions. Dr. Kubota hypothesized that the secretion's role may be to provide a temporary barrier to the entry of pathogens through the area of skin dehiscence caused by the electrical damage, until healing process helps to cover the dehiscence. Then for the next decade Yamaji and Kubota worked to isolate the ingredients of the secretion and they together developed a citric acid-based biosimilar capable of killing a wide range of pathogens which are known to cause common as well as severe infections including nosocomial infections such as MRSA, VRE etc [4] without having to use alcohol-based disinfectants. This biosimilar named as Clinister, had the ability to kill even H5N1 influenza virus and is used in Japan as a spray disinfectant in public places [3]. Inter-Disciplinary-Interaction based on the Invention-I: Evaluation of citric acid-based granules in controlling Chickungunya Virus‏: Clinister has not only been proven as a disinfectant but is also a powerful anti-bacterial and anti-fungal agent that strongly restricts the recurrence of the microbes for a long term. In the present study, Clinister was evaluated for its virostatic efficacy against Chikungunya virus (CHIKV) in vitro for the first time. It showed expressive effect on virus grown in BHK cell cultures by inhibiting the development of virus induced cytopathic effect at an effective concentration of 1.5mg/ml in comparison to 70% ethanol, a common disinfectant. The effects on the virus were further checked by Polymerase Chain Reaction (PCR) at different regular post infective intervals from 0 to 72 hrs. The observations suggested that, other than the anti-microbial activity, Clinister also have specific effect on viruses that may cater the need to prevent the spreading of contagious viruses during epidemics which may occur in the future. However it is essential to study the specific targets of Clinister in viruses at a molecular level, and the same is under progress. Invention – II : A unique polymer invented by a chemical engineer A chemical engineer Prof. Yuichi Mori jointly with polymer scientist Dr Hiroshi Yoshioka developed a thermoreversible polymer (TGP) hydrogel composed of thermoresponsive polymer block [poly (Nisopropylacrylamide- co-n-butyl methacrylate) (poly NIPAAm-co-BMA)] and the hydrophilic polymer block [polyethylene glycol (PEG)] (commercial name :Mebiol gel) [5]. This Mebiol gel provides a suitable in vitro environment enabling culture expansion of cells in the lab without the use of biological components such as amniotic membrane or feeder layers. TGP has been earlier employed for the three-dimensional culture of many cell types like corneal limbal stem cells [6], chondrocytes [7], embryonic stem cells [8], hepatocytes [9], induced pluripotent stem (iPS) cells [8] and bone marrow mononuclear cells [10]. TGP has been used for transportation of corneal endothelial precursor cells over long distances without cool preservation [11], for micro-encapsulation of islet cells [12] and as a wound dressing [13]. TGP has been intralesionally applied along with stem cells in an animal model of spinal cord in injury [10]. TGP is a unique polymer capable of maintaining stem cells in an undifferentiated manner for a longer period of time [14]. It does not affect the gene expression profile [15] and the karyotype of the cells is maintained even after long term culture [16]. The safety and efficacy of TGP as a valuable scaffold material has been established in the various in vitro and translational studies [5]. Inter-Disciplinary-Interaction based on the Invention-II: Mebiol Gel as a 3D scaffold for hepatocytes and viral replication Mebiol Gel was used as a 3D scaffold for growing hepatocyte cell line. Mebiol Gel based hepatocytes had better differentiation and the cells were susceptible to hepatitis C virus replication. The Mebiol Gel based 3D culture system can be used as a better cell culture system for viral studies. Potential Solutions: The works presented in the IIDIAS session have portrayed that original invention in any field when subjected to multi-disciplinary interaction after carefully evaluating the potentials can lead to novel solutions. Though the experts in various disciplines in the reported work have got to interact by chance or due to geographical proximity to each other, when a networking platform to throw such ideas to a forum with mutually rewarding opportunities given for discussion and planning of interactive researches is made possible, novel solutions may not be impossible, if the target platform is well understood within the limitations of each stake holder. Creation of a niche or a suitable environment for such multi disciplinary interaction may prevail and work to its best in a physical state of interaction but advances in information technology has broken the need for such physical interaction among peers to bring out novel solutions. Interactions across specialties in the IIDIAS session help in identifying roles for the invention in other fields which may not be directly related to healthcare. The probable solutions and advantages by combining the electro-physical activity-derived biosimilar and the chemically synthesized polymer described in the study include: A novel platform for in vitro culture of HCV having been described, similar platforms for not so easily cultivable viruses can also be developed. The 3D viral culture technology can be exploited to study the replication process and other intricacies of the virus in in vitro culture. When in vitro expansion of viruses in an appropriate manner becomes feasible, anti-viral agents, either virostatic or virucidal, herbal or synthetic biosimilars can be analysed to come out with effective anti viral agents. A biosimilar-based compound having been proven against Chickungunya virus, the same could be tried against life threatening viruses in appropriate and safe environments. Clinister being a food additive-based material, is considered to be safe to human beings [4] and hence its inclusion as an additive to cell culture medium could be studied further in order to check whether the use of common antibiotics and anti-fungals which may be detrimental to cell growth can be avoided yielding safer anti-bacterial and anti-viral agents for cell culture. Conclusion: Multidisciplinary interaction within and across various domains of science has several potentials in bringing out novel solutions and this has been proven by this study of interaction among physicists, chemists and biologists. The authors recommend that establishment of barrier-free interaction platforms within and among institutes and various specialties at varied stages of research activities as portrayed here like the IIDIAS session to bring about novel solutions such as the ones described in this article. References Sumathipala A, Siribaddana S, Patel V. Under-representation of developing countries in the research literature: ethical issues arising from a survey of five leading medical journals. BMC Med Ethics. 2004;5: E5. The World Bank (2008). Global Economic Prospects 2008: Technology Diffusion in the Developing World. Washington, DC: The World Bank. Retrieved February 18, 2008 from http://siteresources.worldbank.org/INTGEP2008/Resources/complete-report.pdf Abraham S. The "Electric Biology" Duo. Trade Secrets- Nature Biotechnology Blog 2011. Full text can be accessed at http://blogs.nature.com/tradesecrets/2011/12/06/the-electric-biology-duo. Kubota S, Matsuzawa K, Wada M, Yamaji N. Bactericidal Effect of a Disinfectant (Bio Io Nurse) Prepared by Adding Citric Acid and Low Concentration Alcohol to Strongly Acidic Electrolyzed Water. New Food Industry 2008; 50:9. Dedeepiya VD, William JB, Parthiban JKBC, Yoshioka H, Mori Y, Kuroda S, Iwasaki M, Preethy S, Abraham SJK. Scaffolds for Cell Transplantation in Neurology: The Suitability of a Thermoreversible Gelation Polymer—Our Perspectives. Journal of Spinal Surgery 2014;1(1). Sitalakshmi G, Sudha B, Madhavan HN, Vinay S, Krishnakumar S, Mori Y, Yoshioka H, Abraham S. Ex Vivo Cultivation of Corneal Limbal Epithelial Cells in a Thermoreversible Polymer (Mebiol Gel) and Their Transplantation in Rabbits: An Animal Model. Tissue Eng Part A. 2009;15(2):407-15. Yasuda A, Kojima K, Tinsley KW, Yoshioka H, Mori Y, Vacanti CA. In vitro culture of chondrocytes in a novel thermoreversible gelation polymer scaffold containing growth factors. Tissue Eng. 2006;12(5):1237-45. Kataoka K, Huh N. Application of a Thermo-Reversible Gelation Polymer, Mebiol Gel, for Stem Cell Culture and Regenerative Medicine. J Stem Cells Regen Med. 2010 ;6(1); 10-14. Parveen N, Khan AA, Baskar S, Habeeb MA, Babu R, Abraham S, Yoshioka H, Mori Y, Mohammed HC. Intraperitoneal Transplantation of Hepatocytes Embedded in Thermoreversible Gelation Polymer (Mebiol Gel) in Acute Liver Failure Rat Model. Hepatitis Monthly 2008; 8(4): 275-280. William BJ, Prabakaran R, Ayyappan S, Puskhinraj H, Rao D, Manjunath S, Thamaraikannan P, Dedeepiya VD, Kuroda S, Yoshioka H, Mori Y, Preethy, Abraham S. Functional Recovery of Spinal Cord Injury Following Application of Intralesional Bone Marrow Mononuclear Cells Embedded in Polymer Scaffold - Two Year Follow-up in a Canine. J Stem Cell Res Ther.2011; 1:110. Rao SK, Sudhakar J, Parikumar P, Natarajan S, Insaan A, Yoshioka H, Mori Y, Tsukahara S, Baskar S, Manjunath SR, Senthilkumar R, Thamaraikannan P, Srinivasan T, Preethy S, Abraham SJ. Successful transportation of human corneal endothelial tissues without cool preservation in varying Indian tropical climatic conditions and in vitro cell expansion using a novel polymer. Indian J Ophthalmol. 2014;62(2):130-5. Shimizu S, Yamazaki M, Kubota S, Ozasa T, Moriya H, Kobayashi K, Mikami M, Mori Y, Yamaguchi S. In vitro studies on a new method for islet microencapsulation using a thermoreversible gelation polymer, N-isopropylacrylamide-based copolymer. Artif Organs. 1996;20(11):1232-7. Yoshioka H, Mori Y, Kubota S. Wound dressing of newly developed thermogelling thermoreversible hydrogel. Jpn J Artif Organs. 1998; 27: 503-506. Arumugam S, Manjunath S , Senthilkumar R, Srinivasan V, Rajendiran S, Yoshioka H, Mori Y, Abraham S. In vitro expansion and characterization of human chondrocytes using a novel Thermoreversible Gelation Polymer (TGP). Journal of orthopaedics. 2011; 8(3); 1-10. Hishikawa K, Miura S, Marumo T, Yoshioka H, Mori Y, Takato T, Fujita T. Gene expression profile of human mesenchymal stem cells during osteogenesis in three-dimensional thermoreversible gelation polymer. Biochem Biophys Res Commun. 2004;317(4):1103-7. Lei Y, Schaffer DV. A fully defined and scalable 3D culture system for human pluripotent stem cell expansion and differentiation. Proc Natl Acad Sci U S A. 2013;110(52):E5039-48

Synthesis, structural and thermoluminescence properties of YAlO3:Dy3+ nanophosphors

Thermoluminescence properties of YAlO3:Dy3+ nanophosphor prepared by a low temperature solution combustion (SC) method using oxalyl dihydrazide as a fuel were studied and the results were compared to bulk phosphor prepared by solid state (SS) synthesis. Powder X-ray diffraction patterns confirm the orthorhombic phase of SC and SS methods. Rietveld refinement was used to estimate the cell parameters of undoped and Dy3+ doped YAlO3. Scanning electron micrographs reveal dumbbell shape particles. Electron paramagnetic resonance spectra of YAlO3:Dy3+ nanophosphors were studied at 293 K, 77 K and 10 K. Thermoluminescence responses of SC and SS prepared phosphor were studied using gamma irradiation in the dose range 0.1-6 kGy at a warming rate of 1 degrees C s (1) at room temperature (RT). The optimized concentrations of Dy3+ ions in YAlO3 was found to be 3 mol%. The trapping parameters (i. e. activation energy, frequency factor, order of kinetic) of all the individual peaks of the glow curves have been analysed by using Chen's method. The low fading and linear response in the wide range (0.1-1 kGy) suggests the possibility of usage of SC prepared phosphor in dosimeter applications. (C) 2013 Elsevier B. V. All rights reserved