Дисертації: "ANTIMICROBIAL DEFENSE"

1

Frohm, Nilsson Margareta. "The human antimicrobial peptide hCAP18 in epithelial defense /." Stockholm : [Karolinska institutets bibl.], 2001. http://diss.kib.ki.se/2001/91-7349-029-6/.

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2

Rose-Martel, Megan. "Innate Mechanisms of Antimicrobial Defense Associated with the Avian Eggshell." Thesis, Université d'Ottawa / University of Ottawa, 2015. http://hdl.handle.net/10393/32299.

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During the course of evolution, the avian egg has developed multiple physical and chemical barriers in order to resist microbial challenges. These barriers are essential for the successful reproduction of avian species as well as to maintain safe and nutritious food for human consumption of the table egg. The calcified eggshell is a biomineralized barrier with an integrated organic matrix containing antimicrobial proteins, a hallmark of sophisticated biological structures. Calcium carbonate is deposited onto the outer shell membranes to form the calcified mammillary, palisade and vertical crystal layers; the final layer to be deposited is the outer eggshell cuticle. In this thesis, mass spectrometry-based technology was used to investigate the proteome of the outer cuticle, the mammillary cones and the shell membranes in order to gain insight into biomineralization and antimicrobial functions of the avian eggshell. Proteomics analysis of the eggshell cuticle revealed multiple antimicrobial proteins, supporting the hypothesis that the outermost cuticle layer is the first barrier against invading pathogens. The two most abundant cuticle proteins identified are similar to Kunitz-like protease inhibitor (ovocalyxin-25) and ovocalyxin-32. Multiple antimicrobial proteins were also revealed to be associated with the shell membrane fibres. Among the most abundant proteins were lysozyme C, avian β-defensin-11, ovotransferrin, ovocalyxin-36 and gallin. The biomineralized shell is also an important physical barrier against invading pathogens. Proteomics analysis of the mammillary cones, the initiation sites for shell calcification, revealed several candidate proteins involved in calcitic biomineralization. Promising candidates include nucleobindin-2 and SPARC, two calcium binding proteins previously shown to modulate mineralization. In-depth analysis of the comprehensive proteomes generated by this study revealed the presence of histones in the shell membranes, shell and cuticle compartments. Histones are cationic antimicrobial peptides, which are key molecules of the innate immune defense system of many species. This thesis reports the minimal inhibitory concentrations and minimal bactericidal concentrations of histones extracted from avian erythrocytes against Gram-positive, Gram-negative and antibiotic-resistant bacteria. Results suggest that the underlying antimicrobial mechanism is based on the interaction between histones and lipopolysaccharides / lipoteichoic acids, which are negatively charged components of bacterial cell membranes. Histones also inhibit the growth of Gram-positive biofilms; the minimal biofilm eradication concentrations were determined for S. aureus and methicillin-resistant S. aureus (MRSA). Sensitive proteomics analyses have provided great insight into the protein constituents of the eggshell matrix, with two primary roles in the innate immune defense of the egg: regulation of calcitic biomineralization and antimicrobial protection. Further research on these proteins and their functions can provide a new focus for selective breeding programs looking to enhance the egg’s natural defenses, or provide inspiration for alternatives to conventional antibiotics, such as the histones.

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3

Linde, Charlotte M. A. "Defense peptides against Mycobacteria /." Stockholm, 2005. http://diss.kib.ki.se/2005/91-7140-480-5/.

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4

Wang, Xinyi. "Synthesis and Characterization of Antimicrobial Polyesters by Mimicking Host Defense Peptides." University of Akron / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=akron1491508009859916.

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5

AMBIKA, KM. "ROLE OF LACTOSMART AS A NOVEL THERAPEUTIC AGENT IN ANTIMICROBIAL DEFENSE." Thesis, DELHI TECHNOLOGICAL UNIVERSITY, 2021. http://dspace.dtu.ac.in:8080/jspui/handle/repository/18433.

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The emergence of multi – drug resistance (MDR ) in microorganisms against antibiotics has become a global problem [1,2,3]. Various conventional drugs with promised efficacy and specificity are unable to withstand the threat of antibiotic drug resistance [4,5,6]. The rising crisis of MDR bacteria has led to the channelization of relevant research in the direction of antimicrobial molecules from natural sources as potential novel antibiotics. The spectrum of innate immune proteins and their potent fragments herald a promising approach to fight the problem of drug resistance. Among the natural antimicrobial proteins, Lactoferrin (LF) has been identified as a potent host defense system based on its wide spectrum bactericidal and bacteriostatic activities [7,8,9,10,11,12,13] . In the past , several studies have demonstrated the antibacterial and antifungal effects of LF and its derivative peptides, for instance, lactoferricin B [14,15,16,17,18,19] and lactoferrampin [20,21]. Structurally, LF consists of two iron bound lobes, N -lobe (1-333) and C -lobe (345-692) [22,23,24,25]. Amongst the two lobes , the highly cationic properties of N- lobe are responsible for membrane disruption by interacting with anionic components present on bacterial surface [26,27]. It has been established that the lipid A component of the LPS is a known drug target for antimicrobial therapeutics [ 28,29]. One of the mechanisms by which Lf acts as an antimicrobial agent is through binding to pathogen associated molecular patterns (PAMP) such as Lipopolysaccharide (LPS), thereby disrupting the bacterial membrane integrity and activating the chemical signaling pathway[30- 32]. This leads to the secretion of pro- inflammatory responses which down regulates the release of cytokine production [33,34]. In the past, it had been reported that LF binds to LPS with its hexameric sequence present in the 18 - loop region of the lactoferricin [35-37] . In the present study , we have performed the partial digestion of LF with trypsin which generates a potent antimicrobial molecule of the size of about 21kDa (85-281). We have proposed its name as Lactosmart due to its higher potency against pathogens when compared to native LF as a whole protein . The lactosmart has been tested for antibacterial and antifungal properties along with its inhibitory potential of biofilm formation by Pseudomonas aeruginosa through established assays [41]. Our primary focus was on the comparison of LPS binding properties of lactosmart with native LF using surface plasmon resonance technique . The docking and molecular dynamics simulations (MD) studies with LPS have also been performed to further substantiate our claims. Through our studies , we have demonstrated that LF sequesters LPS through two binding sites which are situated on the N- lobe.

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6

Negrón, Oscar A. "Fibrin(ogen)-pathogen Interactions Support Antimicrobial Host Defense following Staphylococcus Aureus Peritonitis Infection." University of Cincinnati / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=ucin150488059846864.

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7

Krynak, Katherine L. "ENVIRONMENTAL INFLUENCES ONAMPHIBIAN INNATE IMMUNE DEFENSE TRAITS." Case Western Reserve University School of Graduate Studies / OhioLINK, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=case1435590530.

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8

Cunden, Lisa Stephanie. "A molecular investigation of the antimicrobial functions of the human S100 host-defense proteins." Thesis, Massachusetts Institute of Technology, 2019. https://hdl.handle.net/1721.1/121779.

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Thesis: Ph. D., Massachusetts Institute of Technology, Department of Chemistry, 2019
Cataloged from PDF version of thesis. Vita.
Includes bibliographical references.
The human host is continually exposed to potentially harmful organisms and the innate immune response is the first line of defense against microbial invasion. One strategy employed by the human innate immune system includes the release of antimicrobial host-defense proteins (HDPs). The goal of this thesis is to understand the antimicrobial functions of four host-defense proteins of the S100 family of proteins: calprotectin (CP), S100A12, S100A7, and S100A15. In the first half of this thesis, we elucidate the Zn(lI)-binding and antimicrobial properties of S100A12 and S100A7 through the use of solution and microbiology studies. We evaluate the affinity of S100A12 for Zn(ll), the scope of its antimicrobial activity, and put forward a model whereby S100A12 uses Ca(ll) ions to tune its Zn(Il)-chelating properties and antimicrobial activity. Our work with S1 00A7 demonstrates that the protein may exist in more than one redox state under physiological conditions, and that unlike CP and S100A12, the antimicrobial properties of S100A7 are not directly modulated by Ca(ll) ions. We report a model whereby the local redox environment of S100A7 tunes its Zn(ll)-sequestration capacity through intramolecular disulfide-bond redox chemistry, and that Ca(II) ions exert an indirect modulatory effect on the Zn(Il)-binding properties of this protein. In the second half of this thesis, we examine the bactericidal properties of the four S100 proteins. Our results agree with prior work on the bactericidal properties of S100A7. Furthermore, we show that CP and S100A15, but not S100A12, possess bactericidal activity at pH 5, and that CP is a broad-spectrum Gram-negative bactericidal factor that functions through a mechanism of membrane permeabilization. Taken together, our studies provide new insights into the multifunctionality of the antimicrobial S100 HDPs.
by Lisa Stephanie Cunden.
Ph. D.
Ph.D. Massachusetts Institute of Technology, Department of Chemistry

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9

Vadapalli, Vatsala. "Role of N-Acylethanolamines in Plant Defense Responses: Modulation by Pathogens and Commercial Antimicrobial Stressors." Thesis, University of North Texas, 2010. https://digital.library.unt.edu/ark:/67531/metadc30521/.

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N-acyl ethanolamines (NAEs) are a class of lipids recently recognized as signaling molecules which are controlled, in part, by their degradation by fatty acid amide hydrolase (FAAH). On the basis of previous studies indicating increased NAE levels in a tobacco cell suspension-xylanase elicitor exposure system and the availability of FAAH mutants, overexpressor and knockout (OE and KO) genotypes in Arabidopsis thaliana, further roles of NAEs in A. thaliana plant defense was investigated. The commonly occurring urban antimicrobial contaminant triclosan (TCS) has been shown to suppress lipid signaling associated with plant defense responses. Thus, a second objective of this study was to determine if TCS exposure specifically interferes with NAE levels. No changes in steady state NAE profiles in A. thaliana-Pseudomonas syringae pv. syringae and A. thaliana-flagellin (bacterial peptide, flg22) challenge systems were seen despite evidence that defense responses were activated in these systems. There was a significant drop in enoyl-ACP reductase (ENR) enzyme activity, which catalyzes the last step in the fatty acid biosynthesis pathway in plants, on exposure of the seedlings to TCS at 10 ppm for 24 h and decreased reactive oxygen species (ROS) production due to flg22 in long term exposure of 0.1 ppm and short term exposure of 5 ppm. However, these responses were not accompanied by significant changes in steady state NAE profiles.

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10

Burkart, David. "UNDERSTANDING CHYTRIDIOMYCOSIS RESISTANCE BY INVESTIGATING THE CUTANEOUS DEFENSE MECHANISMS OF MARSUPIAL FROGS." OpenSIUC, 2015. https://opensiuc.lib.siu.edu/theses/1835.

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Anurans are declining worldwide because of the spread of Batachochytrium dendrobatidis (Bd), the fungus that causes chytridiomycosis. However, some frogs are resistant to this disease, and understanding why may be critical to saving those that are susceptible. In Peru, Gastrotheca excubitor is resistant to chytridiomycosis while Gastrotheca nebulanastes is susceptible. Two anuran skin defenses, symbiotic bacteria and antimicrobial peptides (AMPs), have demonstrated the ability to inhibit Bd in vitro when isolated from certain frogs. We tested if these defenses can explain the difference in susceptibility between the two Gastrotheca species. The cutaneous bacteria and AMPs of both species were collected, tested for their abilities to inhibit the growth of Bd, and analyzed for their compositions. Results indicate that 34%of the strains of skin bacteria from G. excubitor were able to inhibit the growth of Bd whereas only 10% isolated from G. nebulanastes were effective. Gastrotheca excubitor also has stronger anti-Bd skin bacteria. Neither frog species has peptide mixtures capable of completely inhibiting Bd, and overall species did not differ in the anti-Bd abilities of their peptides. These results suggest that the chytridiomycosis resistance experienced by G. excubitor may be attributed to its skin bacteria.

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11

Fjell, Christopher David. "The development of bioinformatic and chemoinformatic approaches for structure-activity modelling and discovery of antimicrobial peptides." Thesis, University of British Columbia, 2009. http://hdl.handle.net/2429/4613.

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The emergence of pathogens resistant to available drug therapies is a pressing global health problem. Antimicrobial peptides (AMPs) may potentially form new therapeutics to counter these pathogens. AMPs are key components in the mammalian innate immune system and are responsible for both direct killing and immunomodulatory effects in host defense against pathogenic organisms. This thesis describes computational methods for the identification of novel natural and synthetic AMPs. A bioinformatic resource was constructed for classification and discovery of gene- coded AMPs, consisting of a database of clustered known AMPs and a set of hidden Markov models (HMMs). One set of 146 clusters was based on the mature peptide sequence, and one set of 40 clusters was based on propeptide sequence. The bovine genome was analyzed using the AMPer resources, and 27 of the 34 known bovine AMPs were identified with high confidence and up to 69 AMPs were predicted to be novel peptides. One novel cathelicidin AMP was experimentally verified as up-regulated in response to infection in bovine intestinal tissue. A chemoinformatic analysis was performed to model the antibacterial activity of short synthetic peptides. Using high-throughput screening data for the activities of over 1400 peptides of diverse sequence, quantitative structure-activity relation (QSAR) models were created using artificial neural networks and physical characteristics of the peptide that included three-dimensional atomic structure. The models were used to predict the activity of a set of approximately 100,000 peptide sequence variants. After ranking the predicted activity, the models were shown to be very accurate. When 200 peptides were synthesized and screened using four levels of expected activity, 94% of the top 50 peptides expected to have the highest level of activity were found to be highly active. Several promising candidates were synthesized with high quality and tested against several multi- antibiotic-resistant pathogens including clinical strains of Pseudomonas aeruginosa, Staphylococcus aureus, Enterococcus faecalis and Escherichia coli. These peptides were found to be highly active against these pathogens as determined by minimal inhibitory concentration; this serves as independent confirmation of the effectiveness of high-throughput screening and in silico analysis for identifying peptide antibiotic drug leads.

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12

Pulido, Gómez David. "Expanding the scope of the Ribonuclease A superfamily in the host immune defense system: Structural determinants of human RNases involved in antimicrobial host defense." Doctoral thesis, Universitat Autònoma de Barcelona, 2013. http://hdl.handle.net/10803/131329.

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El meu projecte de doctorat s’enmarca en l’estudi de l’estructura i la funció de les ribonucleases antimicrobianes humanes. Actualment, es coneixen vuit ribonucleases humanes funcionals pertanyents a la superfamília de ribonucleases secretades per vertebrats. Cal remarcar, que alhora de la seva activitat per a hidrolitzar àcids ribonucleics, també presenten altres propietats biològiques com l’activitat antimicrobiana, que suggereixen una possible funció ancestral relacionada amb el sistema immunitari. La proteïna catiònica d’eosinòfils (ECP) és un dels components majoritaris dels grànuls secundaris dels eosinòfils. Degut al seu gran nombre d’arginines l’ECP és una proteïna molt catiònica i presenta una elevada activitat antimicrobiana vers una gran diversitat de patògens, com bacteris, helmints i protozous. En el nostre laboratori hem estudiat extensament el mecanisme antimicrobià de diverses ribonucleases. Concretament, el treball desenvolupat en aquesta tesi ha permès: i. Identificar el farmacòfor de l’ECP mitjançant eines de minimització racional de l’estructura. ii. Analitzar els canvis que es produeixen a la superfície bacteriana com a conseqüència de l’acció de l’ECP i dels seus pèptids derivats. Principalment, en detallar com aquests interaccionen amb els lipopolisacàrids de la paret bacteriana. iii. Descobrir que l’agregació amiloid de l’ECP a la superfície de les bactèries és fonamental per a la seva acció antimicrobiana i, en concret, per la seva activitat d’aglutinació. iv. Identificar factors estructurals i de seqüencia en el domini N-terminal necessaris per l’acció antimicrobiana de les ribonucleases conservats durant l’evolució. v. Caracteritzar els elements seqüencials de les ribonucleases que determinen les propietats antimicrobianes i d’aglutinació mitjançant el disseny de proteïnes híbrides entre les ribonucleases 2 i 3 (EDN i ECP respectivament). vi. Descobrir que tant l’ECP com la ribonucleasa 7 presenten activitat vers micobacteris a través d’un mecanisme similar a l’exibit enfront de bacteris gram negatius. En conclusió, els resultats obtinguts en aquesta tesi ens permeten conèixer millor com les ribonucleases contribueixen a l’eliminació de diferents patògens del nostre organisme i ens obren noves vies pel disseny de nous agents terapèutics per combatre microorganismes patògens.
My PhD project focuses on the structure-function analysis of human antimicrobial RNases, Currently eight functional members have been ascribed to the vertebrate secreted RNase superfamily. Together with their catalytic activity towards RNA substrates, other biological properties have been reported such as the antimicrobial activity of diverse members of the family, therefore suggesting that RNases could have an ancestral host-defence function. The eosinophil cationic protein (ECP) is one of the major components of the secondary granules of human eosinophils. ECP is a highly cationic arginine-rich protein, which displays a high antimicrobial activity against pathogens, such as bacteria, helminths and protozoa. In our laboratory the antimicrobial mechanism of action of several RNases has been extensively studied. In particular, this work enables us to determine: i. Identify the protein pharmacophore by rational structure minimization. ii. Explore the effects that take place at the bacterial cell wall upon protein-interaction. Concretely, highlight the crucial role of the lipopolysaccharide molecule in the mechanism of action of ECP, and their N-terminal peptides. iii. Determine that the amyloid formation at the bacterial surface is crucial for the antimicrobial activity of the protein, and in particular, due to the agglutinating activity. iv. Prove that the sequence and structural determinants of RNases are evolutionary conserved at the N-terminal domain. v. Characterize the sequence and structural determinants required for antimicrobial and agglutinating activities Through RNase rational mutagenesis of the homologous RNases 2 and 3 (EDN and ECP respectively) vi. Discover that either ECP or RNase 7 present antimycobacterial activity trough a similar mechanism as proved against Gram-negative bacteria. In conclusion, the results presented in this thesis work enable us to better understanding of how RNases contribute on the host pathogen clearance and open a new window in order to the design and develop new therapeutic agents as alternative antibiotics.

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13

Bergsson, Gudmundur. "Antimicrobial polypeptides and lipids as a part of innate defense mechanism of fish and human fetus /." Stockholm, 2005. http://diss.kib.ki.se/2005/91-7140-546-1/.

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14

Padhee, Shruti. "Insights into [aacute]-AA peptides and ã-AA peptides as broad spectrum antimicrobial peptidomimetics and as anti-biofilm agents." Scholar Commons, 2014. https://scholarcommons.usf.edu/etd/5091.

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The emergent resistance of bacteria against the conventional antibiotics has motivated the search for novel antimicrobial agents. Nature abounds with a number of antimicrobial peptides that are a part of our innate immune system and protect us against a variety of pathogenic bacteria. While they are broad-spectrum in their activity and show less drug-resistance induction, their intrinsic metabolic stability limits their potential therapeutic applications. Herein we describe the development of novel broad-spectrum bioactive antimicrobial peptidomimetics AA-peptides. AApeptides were designed based on chiral PNA backbone. Substitution of nucleobases yields AApeptides that are resistant to proteolysis and capable of mimicking peptides. Two types of AApeptides are discussed in this dissertation "[aacute]-AApeptides" and " ã-AApeptides" The therapeutic potential of these AApeptides were accessed by conducting antibacterial assays against a series of both gram-positive, gram-negative bacteria and fungi. These oligomers were characterized using MALDI-TOF and Circular Dichroism spectroscopy (CD). Their invitro toxicity was evaluated against human erythrocytes .We attempted to study their mechanism of action via membrane depolarization assay. We have successfully identified them as antimicrobial agents, pro-inflammatory immune response suppressing agents and as anti-biofilm agents.

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15

Lane, Amy L. "Marine natural products as antimicrobial chemical defenses and sources of potential drugs." Diss., Atlanta, Ga. : Georgia Institute of Technology, 2008. http://hdl.handle.net/1853/26556.

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Thesis (Ph. D.)--Chemistry and Biochemistry, Georgia Institute of Technology, 2009.
Committee Chair: Kubanek, Julia; Committee Member: Fernandez, Facundo M.; Committee Member: Harvey, Stephen C.; Committee Member: Hay, Mark E.; Committee Member: Hud, Nicholas V. Part of the SMARTech Electronic Thesis and Dissertation Collection.

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16

Nimmagadda, Alekhya. "Design, Synthesis, Applications of Polymers and Dendrimers." Scholar Commons, 2017. https://scholarcommons.usf.edu/etd/7430.

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WHO has reported that antibiotic resistance is the third major cause of human death all over the globe. Recent study, has focused on the development of new antibacterial resistance drugs. Herein, we tried to synthesis a series of polymers that can mimic the HDPs. HDPs can target the bacterial cell membrane and they have less chances to develop bacterial resistance. We synthesized the amphiphilic polycarbonates that are highly selective to Gram-positive bacteria, including multidrug resistant pathogens. The membrane disruption activity of these polymers was proved by fluorescence and TEM studies and the drug resistance study showed that the polymers don’t develop bacterial resistance. In order to further design the molecules that can target a broad spectrum of bacteria, we have designed a series of lipidated dendrimers that can target the Gram-positive and Gram-negative bacteria. These dendrimers mimic the HDPs and target the bacterial cell membrane. Dendrimers are reported to inhibit the formation of bacterial biofilm which makes them promising for their future development of antibiotic agents. Apart from the synthesis of polymers and dendrimers as antibacterial agents, we have designed a series of small molecular antibacterial agents that are based on the acylated reduced amide scaffold and small dimeric cyclic guanidine derivatives. These molecules display good potency against a panel of multidrug-resistant Gram-positive and Gram-negative bacterial strains. Meanwhile, they also effectively inhibit the biofilm formation. Mechanistic studies suggest that these compounds kill bacteria by compromising bacterial membranes, a mechanism analogous to that of host-defense peptides (HDPs). Lastly, we also demonstrate that these molecules have excellent in vivo activity against MRSA in a rat model. This class of compounds could lead to an appealing class of antibiotic agents combating drug-resistant bacterial strains.

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17

Uzarski, Joshua Robert. "Reflection Absorption Infrared Spectroscopic Studies of Surface Chemistry Relevant to Chemical and Biological Warfare Agent Defense." Diss., Virginia Tech, 2009. http://hdl.handle.net/10919/26107.

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Reflection absorption infrared spectroscopy was used as the primary analysis technique to study the interfacial chemistry of surfaces relevant to chemical and biological warfare agent defense. Many strategies utilized by the military to detect and decompose chemical and biological warfare agents involve their interaction with surfaces. However, much of the chemistry that occurs at the interface between the agents and surfaces of interest remains unknown. The surface chemistry plays an important role in efficacy of both detection and decontamination technology, and by obtaining a deeper understanding of that chemistry, researchers might be able to develop more sensitive detection devices and more effective decontamination strategies. Our efforts have focused on three different areas of surface chemistry relevant to chemical and biological warfare agent defense: 1) The development of a surface synthesis strategy to create and control the structure of antibacterial self-assembled monolayers (SAMs). Our work demonstrated a successful strategy for creating SAMs that contain long-chain quaternary ammonium groups, which were synthesized and subsequently characterized using RAIRS and X-ray photoelectron spectroscopy (XPS). 2) The determination of the surface conformation, orientation, and relative surface density of immobilized antimicrobial peptides. Our results revealed that the peptides consisted of tilted (50-60°), α-helices on the surface, regardless of solution conditions. 3) The design and construction of a new ultrahigh vacuum surface science instrument that allows for the study of gas-surface reactions with up to three gases simultaneously. 4) The study of the adsorption of chemical warfare agent simulants to silica nanoparticulate films. Our work demonstrated that the adsorbate structure was dependent on the number of hydrogen-bonding groups, and the adsorption consists of a pressure-dependent two part mechanism. The results presented here will help increase the understanding of the surface chemistry of three interfaces relevant to chemical and biological defense. Future researchers may apply the new information to develop more effective detection and decontamination strategies for chemical and biological warfare agents.
Ph. D.

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18

Fragoso, Miryam Araceli. "Regulation of the Lactoperoxidase System in the Airway." Scholarly Repository, 2007. http://scholarlyrepository.miami.edu/oa_dissertations/4.

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The lactoperoxidase (LPO) antimicrobial system has been shown to play an important role in maintaining sterile conditions in several tissues including the mammary gland, the salivary gland, and the airway. The LPO system in the airway consists of the enzyme LPO and its substrates hydrogen peroxide and an anion. LPO catalyzes the oxidation of a halide or pseudohalide ion for example SCN-or I- by hydrogen peroxide producing a product, OSCN- or OI- which have antibacterial, antifungal, and antiviral properties. In order to have a functional antimicrobial system all the components need to be present at appropriate concentrations. The LPO system has been suggested to be deficient in cystic fibrosis. There are three possible regulatory mechanism of this antimicrobial system and these involve the secretion and availability of the three components of the LPO system in the luminal fluid. The studies presented in this dissertation examine two of the possible regulatory mechanisms of the LPO system in the airway; the availability and transport of SCN- to the luminal surface, and the expression of LPO. The knowledge obtained from these studies could be utilized to develop treatments to control infection in diseases characterized by chronic infections such as cystic fibrosis.

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19

Gide, Mussie. "Design, Synthesis, Application of Biodegradable Polymers." Scholar Commons, 2018. https://scholarcommons.usf.edu/etd/7625.

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Bacterial infections have posed a serious threat to the public health due to the significant rise of the infections caused by antibiotic-resistant bacteria. There has been considerable interest in the development of antimicrobial agents which mimic the natural HDPs, and among them biodegradable polymers are newly discovered drug candidates with ease of synthesis and low manufacture cost compared to synthetic host defense peptides. Herein, we present the synthesis of biocompatible and biodegradable polymers including polycarbonate polymers, unimolecular micelle hyperbranched polymers and dendrimers that mimic the antibacterial mechanism of HDPs by compromising bacterial cell membranes. The developed amphiphilic polycarbonates are highly selective to Gram-positive bacteria, including multidrug-resistant pathogens and the unimolecular micelle hyperbranched polymers showed promising broad-spectrum activity. However, lipidated amphiphilic dendrimers with low molecular weight display potent and selective antimicrobial activity against both Gram-positive and Gram-negative bacteria, including multidrug-resistant strains. In addition to antibacterial activity against planktonic bacteria, these dendrimers were also shown to inhibit bacterial biofilms effectively. These class of polymers may lead to a useful generation of antibiotic agents with practical applications.

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20

Lew, Cynthia S. "Loss of the Lipopolysaccharide Core Biosynthesis rfaD Gene Increases Antimicrobial Chemokine Binding and Bacterial Susceptibility to CCL28 and Polymyxin: A Model for Understanding the Interface of Antimicrobial Chemokines and Bacterial Host Defense Avoidance Mechanisms." BYU ScholarsArchive, 2012. https://scholarsarchive.byu.edu/etd/3756.

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In order to better understand the mechanism of antimicrobial chemokine activity, including binding to and killing of bacteria, random transposon mutagenesis was performed in Yersinia pseudotuberculosis. Resulting mutants were screened for increased binding to chemokine and high binding clones were selected for further study. One mutant, designated mutant 27, was found to have a single insertion mutation in the rfaD gene. The rfaD gene product is involved in heptose biosynthesis, one of the sugars of the inner core oligosaccharide of Gram- negative lipopolysaccharide (LPS). Mutant 27 was found to bind both CCL25 and CCL28, two antimicrobial chemokines, more efficiently than the wild type bacteria. This clone was also found to be more susceptible to CCL28- mediated killing and polymyxin activity. Complementation with a plasmid bearing the full rfaDFC operon restored the wild type phenotype in both regards. These data suggest that normal LPS expression by Y. pseudotuberculosis serves to protect the bacteria from the antimicrobial function of chemokines and other antimicrobial proteins of the mammalian innate immune system.

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21

Winkle, Sean M., Andrea L. Throop та Melissa M. Herbst-Kralovetz. "IL-36γ Augments Host Defense and Immune Responses in Human Female Reproductive Tract Epithelial Cells". FRONTIERS MEDIA SA, 2016. http://hdl.handle.net/10150/617371.

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IL-36 gamma is a proinflamatory cytokine which belongs to the IL-1 family of cytokines. It is expressed in the skin and by epithelial cells (ECs) lining lung and gut tissue. We used human 3-D organotypic cells, that recapitulate either in vivo human vaginal or cervical tissue, to explore the possible role of IL-36 gamma in host defense against pathogens in the human female reproductive tract (FRT). EC were exposed to compounds derived from virus or bacterial sources and induction and regulation of IL-36 gamma and its receptor was determined. Polyinosinic-polycytidylic acid (poly I:C), flagellin, and synthetic lipoprotein (FSL-1) significantly induced expression of IL-36 gamma in a dose-dependent manner, and appeared to be TLR-dependent. Recombinant IL-36 gamma treatment resulted in self amplification of IL-36 gamma and its receptor (IL-36R) via increased gene expression, and promoted other inflammatory signaling pathways. This is the first report to demonstrate that the IL-36 receptor and IL-36 gamma are present in the human FRT EC and that they are differentially induced by microbial products at this site. We conclude that IL-36 gamma is a driver for epithelial and immune activation following microbial insult and, as such, may play a critical role in host defense in the FRT.

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22

Tavares, Tallita Cruz Lopes. "ProteÃÂnas de tintas de moluscos marinhos composiÃÂÃÂo, funÃÂÃÂo e mecanismo de aÃÂÃÂo." Universidade Federal do CearÃ, 2010. http://www.teses.ufc.br/tde_busca/arquivo.php?codArquivo=6997.

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Nos moluscos sem concha ou naqueles em que a mesma estÃÂ presente na forma vestigial, como cefalÃÂpedes e gastrÃÂpodes opistobrÃÂnquios, a falta dessa proteÃÂÃÂo externa conduziu ÃÂ evoluÃÂÃÂo de mecanismos de defesa dominados pela secreÃÂÃÂo ou incorporaÃÂÃÂo de substÃÂncias quÃÂmicas bioativas. ProteÃÂnas sintetizadas de novo sÃÂo bons exemplos dessas molÃÂculas, tendo sido encontradas nas tintas de vÃÂrias espÃÂcies de lesmas do mar, particularmente do gÃÂnero Aplysia. Muitas jÃÂ revelaram pertencer a famÃÂlias de proteÃÂnas com atividade oxidÃÂstica de L-aminoÃÂcidos (LAAOs), grupo do qual fazem parte tambÃÂm proteÃÂnas bioativas encontradas nos venenos de serpentes. A caracterizaÃÂÃÂo, portanto, das atividades biolÃÂgicas e do perfil bioquÃÂmico dessas proteÃÂnas ÃÂ muito ÃÂtil para que possamos entender como se dÃÂo os mecanismos defensivos desses animais e quÃÂo difundidos esses sÃÂo, alÃÂm de proporcionar subsÃÂdios para sua aplicaÃÂÃÂo biomedicinal. Esse estudo visou a elucidaÃÂÃÂo dos mecanismos de aÃÂÃÂo da dactilomelina-P, proteÃÂna antibacteriana purificada da tinta de Aplysia dactylomela, e a comparaÃÂÃÂo da composiÃÂÃÂo protÃÂica e das atividades antimicrobianas das tintas liberadas por dois gastrÃÂpodes, A. dactylomela e Bursatella leachii, e por um cefalÃÂpode, Octopus sp.. O estudo revelou que a dactilomelina-P ÃÂ uma L-aminoÃÂcido oxidase capaz de oxidar L-lisina e L-arginina, com maior afinidade por L- arginina, com Km de 0,22 +- 0,16 mM/L para a L-lys e de 0,015 +- 0,01 Mm/L para a L-arg. Demonstrou tambÃÂm que sua atividade antibacteriana ÃÂ medida por esta propriedade, tendo o perÃÂxido de hidrogÃÂnio, gerado na oxidaÃÂÃÂo enzimÃÂticca, grande aprtipaÃÂÃÂo na inibiÃÂÃÂo bacteriana. A proteÃÂna se comporta de maneira bacteriostÃÂtica quando em meio nÃÂo suplementado com L-lisina e L-angilina, sendo bactericida na presenÃÂa destes. AlÃÂm disso, a atividade antibacteriana mostrou ser dependente da concentraÃÂÃÂo de aminoÃÂcidos disponibilizados para oxidaÃÂÃÂo pela proteÃÂna. A atividade Staphylococcus aureus deve ocorrer por meio de mecanismos que nÃÂo induzam danoso na morfologia celular, uma vez que nÃÂo foi observada qualquer alteraÃÂÃÂo morfolÃÂgica por microscopia de forÃÂa atÃÂmica. JÃÂ a anÃÂlise das trÃÂs tintas pro PAGE-SDS revelou a presenÃÂa, em todas elas, de bandas protÃÂicas de cerca de 60 kDa, alÃÂm de bandas de peso moleculares mais baixos. Por Western blot pode-se observar a existÃÂncia de identidade imunolÃÂgica entre proteinas das trÃÂs tintas, visto que anticorpos policlonais produzidos contra a dactilomelina-P reconheceram proteÃÂna(s) de cerca de 60 kDa na tinta de B. leachii e de cerca de 30 kDa na tinta de Octopus sp. Dentre as trÃÂs tintas, somente a tinta de A. dactylomela apresentou atividades antibacterianas e antifÃÂngica
In shelled molluscs or those in which it is present as vestigial as cephalopods and gastropods opistobrÃÂnquios, the lack of external protection led to the evolution of defense mechanisms dominated by the secretion or incorporation of bioactive chemicals. Proteins synthesized again are good examples of these molecules, were found in paints of various species of sea slugs, particularly the genus Aplysia. Many have already revealed belong to protein families with oxidÃÂstica activity of L-amino acids (LAAOs), a group which comprises also bioactive proteins found in snake venoms. The characterization therefore biological activities and biochemical profile of these proteins is very useful for us to understand how to give the defense mechanisms of animals and how widespread they are, in addition to providing subsidies to its application biomedicinal. This study aimed to elucidate the mechanisms of action of P-dactilomelina, antibacterial protein purified from Aplysia ink dactylomela, and comparison of protein composition and antimicrobial activities of paint released by two gastropods, A. dactylomela and Bursatella leachii, and a cephalopod, Octopus sp .. The study revealed that dactilomelina-P is an L-amino acid oxidase capable of oxidizing L-lysine and L-arginine, with a higher affinity for L-arginine, with a Km of 0.22 + - 0.16 mM / L for L -lys and 0.015 + - 0.01 mM / L L-arg. It also demonstrated that its antibacterial activity is measured by this property, with the hydrogen peroxide generated in the oxidation enzimÃÂticca, great aprtipaÃÂÃÂo in bacterial inhibition. The protein behaves in a non-bacteriostatic when supplemented with L-lysine and L-angilina, being in the presence of bactericidal. In addition, the antibacterial activity was shown to be dependent on the concentration of amino acids available for oxidation by the protein. Staphylococcus aureus activity must occur through mechanisms that do not induce damage to cell morphology, since we did not observe any morphological change by atomic force microscopy. The analysis of the three pro paint SDS-PAGE revealed the presence in them all, protein bands of about 60 kDa, and bands of lower molecular weight. For Western blot can observe the existence of immunologic identity between proteins of the three dyes, since polyclonal antibodies produced against P-dactilomelina recognized protein (s) of about 60 kDa in ink B. leachii and approximately 30 kDa in ink Octopus sp. Among the three dyes, only the ink of A. dactylomela showed antibacterial and antifungal activities

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23

Ullal, Anirudh Jaiwant Mr. "DRAFT - Extra-erythrocytic expression of antimicrobial peptides derived from the β-subunit of hemoglobin is associated with a potent anti-parasitic defense in fish". NCSU, 2006. http://www.lib.ncsu.edu/theses/available/etd-11062006-173910/.

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Innate immunity plays a crucial role in the defense against prokaryotic and eukaryotic pathogens. Major components of this defense are antimicrobial peptides (AMPs). Some AMPs are derived from larger proteins with other recognized functions (e.g., lactoferrin, histones). In this study, we demonstrate the expression of peptides homologous to the β-chain of hemoglobin (Hb-β), one of the two major subunits of this respiratory protein. These Hb-β peptides (HbβP-1, -2 and -3), isolated from gill of the economically important channel catfish (Ictalurus punctatus), had antibacterial activity and were upregulated in gill and skin epithelium in response to parasitic (Ichthyophthirius multifiliis, ich) infection. One peptide (HbβP-1), while having relatively weak antibacterial activity, had antiparasitic activity comparable to that of other potently antiparasitic AMPs. Also, this cidal activity was specifically directed against the trophozoite (trophont) stage of ich at a low concentration (6.2 ug/ml, 1.7 mM) but had no apparent effect on the disseminative (theront) stage or the reproductive (tomont) stage at the highest concentration tested (400 mg/ml, 108 mM). In addition, HbβP-1 was not lytic to channel catfish erythrocytes at the highest concentration tested (400 mg/ml, 108 mM). Immunohistochemistry and in situ hybridization of skin and gill from fish experimentally challenged with ich indicated that the HbβP-1 sequence was both synthesized and expressed in epithelial tissues of skin and gill, which are the target tissues for ich. ?Bug blots? of gill extracts from fish recovering from a bacterial infection suggested that upregulation of these Hb-β related peptides might also occur with other infections. These findings, along with the recent discovery by others that Hb-β is expressed in mammalian macrophages and alveolar epithelium, suggest that hemoglobin-derived AMPs might play a significant role in the non-specific immune response of vertebrates.

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24

Koslowski, Maureen Jeanette [Verfasser], and Jan [Akademischer Betreuer] Wehkamp. "Disease Underlying Genetics in Ileal Crohn’s Disease : Disturbed Antimicrobial Defense and Impaired Wnt Signaling at the Epithelial Barrier / Maureen Jeanette Koslowski ; Betreuer: Jan Wehkamp." Tübingen : Universitätsbibliothek Tübingen, 2012. http://d-nb.info/1162699868/34.

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25

Koslowski, Maureen [Verfasser], and Jan [Akademischer Betreuer] Wehkamp. "Disease Underlying Genetics in Ileal Crohn’s Disease : Disturbed Antimicrobial Defense and Impaired Wnt Signaling at the Epithelial Barrier / Maureen Jeanette Koslowski ; Betreuer: Jan Wehkamp." Tübingen : Universitätsbibliothek Tübingen, 2012. http://d-nb.info/1162699868/34.

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26

Ulvila, J. (Johanna). "Functional genomic analysis of the Drosophila immune response:identification of genes essential for phagocytosis, viral defense and NF-κB signaling". Doctoral thesis, University of Oulu, 2008. http://urn.fi/urn:isbn:9789514289941.

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Abstract Innate immunity provides the first line of defense against invading, pathogenic microorganisms in all multicellular organisms. The fruit fly Drosophila melanogaster has turned out to be an excellent model organism to elucidate mechanisms of innate immune responses because of the highly conserved intracellular signaling cascades mediating these ancient immune functions in flies and mammals. In the present study, RNA interference (RNAi) -based functional genomics were utilized to identify novel components of Drosophila’s immune reactions. Mediators of bacterial phagocytosis, nuclear factor kappa B (NF-κB) signaling and the antiviral RNAi pathway were screened in hemocyte-like S2 cells. Follow-up studies were executed in mammalian cells as well as in Drosophila larvae and adult flies to gain broader significance for the results. Seven novel components essential for efficient phagocytosis of bacteria were identified. Eater was defined as Drosophila’s most important phagocytic receptor showing novel epidermal growth factor (EGF)-repeat -based microbial recognition properties. Additionally, Abelson interacting protein (Abi), capping protein alpha (cpa), 14-3-3ζ, tousled-like kinase (tlk), CG2765 and CG15609 were determined as intracellular effectors of phagocytosis, the three former ones executing their evolutionarily conserved functions through remodeling of the actin cytoskeleton. Eater, together with Scavenger receptor class C, type I (Sr-CI), was demonstrated to be responsible for double-stranded RNA (dsRNA) uptake into S2 cells and, when ectopically expressed, into mammalian cells via clathrin-mediated endocytosis. Proteasome component Pros45 and RNA helicase Belle were established as mediators of the intracellular RNAi pathway, whereas essential roles in antimicrobial signaling via the immune deficiency (Imd) pathway were addressed for Inhibitor of apoptosis 2 (Iap2) and Tak1-associated binding protein (TAB). Iap2 and TAB were shown to affect nuclear translocation of NF-κB -like transcription factor Relish. The present study identifies several novel mediators of the Drosophila immune response and provides insight into mechanisms of fly host defense. As insects serve as vectors of human diseases (e.g. malaria), knowledge about Drosophila immune mechanisms may help to better understand the transmission and pathogenesis of these diseases and develop treatments to fight these infections. Additionally, knowledge gained from model organisms serves as valuable background information, often conducting human research into new tracks
Tiivistelmä Synnynnäinen immuniteetti on elintärkeä puolustusjärjestelmä taudinaiheuttajia vastaan. Kodeissakin yleinen banaanikärpänen, Drosophila melanogaster, on osoittautunut erinomaiseksi synnynnäisen immuniteetin tutkimusmalliksi, erityisesti teknisesti yksinkertaisen ja eettisesti ongelmattoman geneettisen muunneltavuutensa ansiosta. On myös havaittu, että solunsisäiset, immunologisia signaaleja välittävät mekanismit ovat evoluutiossa hyvin säilyneitä. Hyvin usein samankaltaiset geenituotteet toimivat signaalinsiirtäjinä sekä kärpäsen että ihmisen soluissa. Tämän työn tarkoituksena oli RNA-häirintää (RNAi) sekä muita nykyaikaisia solu- ja molekyylibiologisia tutkimusmenetelmiä hyödyntäen tunnistaa uusia kärpäsen synnynnäiselle immuunipuolustukselle välttämättömiä geenituotteita. Bakteerien fagosytoosille, viruspuolustukselle ja tumatekijä nuclear factor kappa B:n (NF-κB) välittämälle signaloinnille välttämättömiä signalointimolekyylejä pyrittiin identifioimaan laajan mittakaavan RNA-häirintään perustuvilla seuloilla kärpäsen soluissa. Saatujen tulosten merkitystä nisäkkäiden immuunipuolustukselle tutkittiin myös hiiren soluissa. Seitsemän geenituotteen osoitettiin olevan bakteerien fagosytoosille tärkeitä kärpäsen soluissa. Aiemmin tuntematon geenituote, joka nimettiin Eateriksi, osoitettiin kärpäsen tärkeimmäksi bakteereja fagosytoivaksi reseptoriksi. Eaterin solun ulkoisen osan osoitettiin tunnistavan taudinaiheuttajia uudella epidermaalisen kasvutekijän (epidermal growth factor, EGF) kaltaisella toistosekvenssillä. Myös useiden solun tukirankaan, sytoskeletoniin, liittyvien proteiinien (Abi, cpa, 14-3-3ζ) sekä aiemmin vähemmän tunnettujen geenituotteiden (CG2765, CG15609, tlk) osoitettiin osallistuvan bakteerien fagosytoosiin. Näistä kolmen ensinmainitun immunologinen tehtävä havaittiin evoluutiossa säilyneeksi, kärpäsestä hiireen. Eaterin, yhdessä kärpäsen toisen scavenger reseptorin (Sr-CI) kanssa, havaittiin myös toimivan kaksijuosteisen RNA:n (dsRNA) reseptoreina kärpäsen soluissa, mahdollistaen helpon ja tehokkaan RNA-häirinnän. RNA-häirinnän, ja siten mahdollisesti myös viruspuolustuksen, välittäjiksi identifioitiin proteasomin alayksikkö Pros45 ja RNA-helikaasi Belle. Lisäksi Inhibitor of apoptosis 2 (Iap2) ja Tak1-associated binding protein (TAB) todettiin kärpäsen immune deficiency (Imd) signalointireitin komponenteiksi, jotka osallistuvat antimikrobisten peptidien tuotantoon välittämällä NF-κB:n kaltaisen kärpäsen transkriptiotekijän (Relish) siirtymisen tumaan aktivoimaan immuunipuolustusta välittävien geenien ilmentymistä. Tämän tutkimuksen tulokset valottavat banaanikärpäsen immuunipuolustuksen mekanismeja. Koska hyönteiset toimivat monien ihmisten infektiotautien välittäjinä, kärpäsen immuniteetin tuntemus luo mahdollisuuksia kehittää hoitoja näitä tauteja vastaan. Lisäksi malliorganismeista saatu tieto luo uusia teorioita ja näkökulmia, johtaen usein myös lääketieteellistä tutkimusta uusille raiteille

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Lima, Stella Maris de Freitas. "Peptídeos de defesa do hospedeiro como adjuvantes na terapia endodôntica." Universidade Católica de Brasília, 2018. https://bdtd.ucb.br:8443/jspui/handle/tede/2474.

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Failure of endodontic therapy is mainly based on the persistence of the microorganism inside the root canal system, perpetuating periradicular pathologies. The need for endodontic reintervention reduces the success rate and turns the development of new therapies into a necessity. Host defense peptides (HDPs) have a therapeutic potential because of its characteristics as broad spectrum antimicrobial activity, immunomodulatory capacity and repair induction. The present study aimed to demonstrate HDPs HHC-10, LL-37 and synoeca-MP activities compared to the commonly used chemical agents Ca(OH)2, NaOCl and chlorhexidine (CHX) in in vitro and in vivo environments. In vitro analysis involved (1) antimicrobial assays for MIC/MBC/MFC determination, antibiofilm and synergistic activities (against Candida albicans, Enterococcus faecalis and Staphylococcus aureus), (2) hemolytic assay and determination of therapeutic index, (3) immunomodulation assays (from RAW 264.7 monocytes) and (4) osteoclastogenesis assays (from mouse bone marrow cells). In vivo assays were based on periradicular lesions induced and treated in rats with subsequent analysis by radiographic, tomographic and histological examinations. Results demonstrated effective antimicrobial activity of HHC-10 (except antibiofilm activity) and synoeca-MP besides NaOCl and CHX. Higher therapeutic index was observed for peptides (HHC-10 and synoeca-MP) with low or no hemolytic activity. Synergism analysis demonstrated better interaction between synoeca-MP and CHX. Immune assays from monocyte cultures demonstrated a pro-inflammatory activity of LL-37 and synoeca-MP and a pro- and anti-inflammatory effect of HHC-10 and Ca(OH)2. However, none of the agents tested reduced osteoclastogenesis in vitro. In in vivo endodontic treatment conditions, HHC-10, synoeca-MP and Ca(OH)2 significantly reduced periapical lesions, although only peptides were able to reduce inflammation in histological analysis. Therefore, HHC-10 and synoeca-MP demonstrated in vitro and in vivo potential for application in endodontic therapy. However, further researches are needed regarding the mechanism of action of HDPs and the possible therapeutic formulations for future analysis in animal models.
O insucesso da terapia endodôntica está baseado na persistência de microrganismos principalmente no interior do sistema de canais radiculares, perpetuando as patologias perirradiculares associadas à tal infecção. A necessidade de reintervenção endodôntica reduz o percentual de sucesso e, portanto, o surgimento de novas terapias é necessário. Dessa forma, peptídeos de defesa do hospedeiro (PDHs) possuem potencial terapêutico associado às suas características de atividade antimicrobiana de amplo espectro, capacidade imunomodulatória e indução de reparo. O presente estudo objetivou demonstrar a atividade dos PDHs HHC-10, LL-37 e synoeca-MP comparado aos agentes químicos comumente utilizados Ca(OH)2, NaOCl e clorexidina (CHX) em ambientes in vitro e in vivo. Para tanto, foram realizadas análises in vitro envolvendo: (1) ensaios antimicrobianos com determinação de MIC/MBC/MFC, atividade antibiofilme e atividade sinérgica (contra Candida albicans, Enterococcus faecalis e Staphylococcus aureus), (2) ensaio hemolítico e determinação do índice terapêutico, (3) ensaios de imunomodulação (a partir de monócitos RAW 264.7) e (4) ensaios osteoclastogênese (a partir de células da medula óssea de camundongos). Os ensaios in vivo foram conduzidos a partir das lesões perirradiculares induzidas e tratadas em ratos com subsequente análise por exames radiográficos, tomográficos e histológicos. Assim, os resultados demonstraram efetiva atividade antimicrobiana dos peptídeos HHC-10 (exceto atividade antibiofilme) e synoeca-MP além do NaOCl e CHX. Observou-se maiores índices terapêuticos para os peptídeos (HHC-10 e synoeca-MP) com baixa ou nenhuma atividade hemolítica. As análises de interação demonstraram combinação sinérgica necessitando de menor concentração a partir da interação entre synoeca-MP e CHX. A partir da cultura de monócitos, observou-se uma atividade pró-inflamatória a partir dos peptídeos LL-37 e synoeca-MP e um efeito pró- e anti-inflamatório do peptídeo HHC-10 e do Ca(OH)2. No entanto, nenhum dos agentes testados reduziram a osteoclastogênese in vitro. Já em condições de tratamento endodôntico in vivo, os PDHs HHC-10, synoeca-MP e o Ca(OH)2 reduziram significativamente lesões periapicais, apesar de somente os peptídeos serem capazes de reduzir inflamação em análises histológicas. Portanto, os peptídeos HHC-10 e synoeca-MP demonstraram potencial in vitro e in vivo para aplicação na terapia endodôntica. No entanto, faz-se necessário maiores investigações sobre o mecanismo de ação dos mesmos e possíveis formulações terapêuticas para futuros testes em modelos animais.

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28

Osei-Boadi, Kate. ""An aliphatic essential amino acid influences the expression of host defense peptides in colonic epithelial cells: in vitro findings and potential clinical implications in Crohn's disease"." Diss., Kansas State University, 2014. http://hdl.handle.net/2097/17629.

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Doctor of Philosophy
Department of Human Nutrition
Tonatiuh Melgarejo
Background and Objective: Crohn’s disease (CD) patients express low levels of host defense peptides (HDPs) especially β-defensins, which may compromise intestinal barrier function. Antibiotic treatment for bacterial infections in CD is limited and rarely curative, making it necessary to find alternative therapeutic approaches. We therefore investigated to what extent an essential amino acid; L-isoleucine (L-ILE) might induce the expression of human β-defensins (HBDs) in colonic epithelial cells as an alternative approach to help patients with CD. Antimicrobial activity of HBD2 was also assessed against four bacterial isolates which can cause secondary infections in CD. Methods: HTB-37 Caco-2 cells were stimulated with L-ILE at a concentration of 0 - 500µg/ml for 6 hours. Total RNA was extracted using RNeasy Micro Kit (QIAGEN). Reverse transcription was carried out with Superscript ®III First-Strand Synthesis System. The cDNA was amplified using specific primers for HBD1-3. Antimicrobial activity of HBD2 was determined using the broth dilution assay. Results: HBD1 was constitutively expressed under all conditions. HBD2 was expressed in HTB-37 cells after stimulation with L-ILE. Below 25µg/ml L- ILE stimulation, no expression of HBD2 was observed. HBD2 exhibited antimicrobial activity against bacterial isolates tested, with a MIC of 32, 64 and 128 µg/ml for both strains of E. coli, S. aureus and P. aeruginosa respectively. Conclusions: Our results indicate that L-ILE stimulation of HTB-37 Caco-2 cells can induce HBD2 expression. Data collected from our in vitro studies might have major implications for modifying the intestinal microbiota towards a healthier state in CD patients. Promoting the expression of HBD2 by colonic cells may lead to a lower rate of infection in these patients. Future in vivo studies are warranted to determine the potential clinical use of intra colonic administration of L-ILE in CD patients. The observed antimicrobial activity of HBD2 against bacterial isolates provides evidence that it is a crucial component of mucosal epithelial defense against infections which can complicate disease symptoms in CD.

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Silva, Paulo Luiz da. "Bula da glândula metapleural de formigas cortadeiras: inferências quanto à defesa antimicrobiana." Universidade Federal de Viçosa, 2008. http://locus.ufv.br/handle/123456789/3884.

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Leafcutter ants live in a world of complex interactions, the most notable being their cultivation of a mutualistic fungus on vegetable subtrates. These ants, and their mutualistic fungus gardens, are at risk from attack by parasitic microorganisms. To defend themselves and their mutualist, they have antimicrobial defences, such as substances secreted from their metapleural glands. As this defence is subject to selective pressures according to the risk of parasite attack, variations in the amount of defence may reflect variations in the risk. Meanwhile, there may be variation in the value of individual workers to the colony, dependent in part upon the area in which they work, and investment in their defences may also reflect this. In this study, the volume of the externally visible part (bulla) of the reservoir of the metapleural gland of workers was compared betwen species, colonies and tasks, to determine variations in defence strategies against pathogens. Additionally, colony sizes of a bacterial mutualist on the workers thoraxes, were estimated, looking for a possible relationship with metapleural gland bulla size. A total of eleven laboratory colonies were used, of four species of leafcutter ants: Atta laevigata (F.Smith, 1858), Atta sexdens rubropilosa Forel, 1908, Acromyrmex subterraneus brunneus (Forel, 1911) and Acromyrmex disciger (Mayr, 1887). Workers were collected from three areas of the colonies: foraging areas, symbiotic fungus gardens and the refuse deposits. Parameters measured were: the volume of the bulla of the metapleural gland, the weight of ants and an estimate of the olony sizes of the symbiotic bacteria. Analyses of covariance were conducted using individual mass as a covariate, to determine the contributions of a range of parameters in explaining variation in the size of this organ. The two species of Atta were found to have larger bulla when compared to the two species of Acromyrmex., probably reflecting not a greater degree of exposure to parasites but a strategy of greater investment in defences. Gardeners and foragers of the species A. laevigata and Ac. subterraneus brunneus had larger bulla than did waste workers, revealing a lower quality (in terms of parasite defence) of the waste workers. In this case, we expect that these individuals, being of lower quality, are allocated to more dangerous tasks, to die more rapidly. Acromyrmex subterraneus brunneus was the only species with visible colonies of the bacterial mutualist on the thorax. There was a positive relation between the size of this colony and the volume of the bulla of the metapleural gland. These variations in degrees of defence imply that the ants have different strategies to defend their colonies against pathogens and parasites.
Formigas cortadeiras apresentam interações complexas. São cultivadoras de fungo o qual é alimentado por substratos vegetais. Tanto as formigas como o fungo do jardim correm o risco de serem parasitados por microrganismos. Para defenderem tanto o fungo quanto elas mesmas, as formigas utilizam defesas antimicrobianas, como substâncias secretadas através da glândula metapleural. Sendo que esta defesa está sujeita a pressão de seleção de acordo com o risco de ataque por parasitas, variações no investimento nela podem ser consequência deste risco. Ao mesmo tempo, pode haver indivíduos com diferentes valores para a colônia, dependendo em parte na área onde são encontradas, e podem ter maior ou menor investimento em defesas contra parasitas. Diante disso o trabalho propôs verificar a variação do volume do reservatório da glândula metapleural (bula) entre espécies, colônias e tarefas de formigas cortadeiras para verificar possíveis estratégias de defesas das operárias contra patógenos. Avaliou-se também a presença de uma bactéria mutualista, que é utilizada também como defesa contra parasitismo, e a possível relação entre esta e o volume do reservatório da glândula metapleural. Foram utilizadas um total de 11 colônias de quatro espécies de formigas cortadeiras, Atta laevigata (F.Smith, 1858), Atta sexdens rubropilosa Forel, 1908, Acromyrmex subterraneus brunneus (Forel, 1911) e Acromyrmex disciger (Mayr, 1887). Formigas foram coletadas de três áreas das colônias: as áreas de forrageamento, os jardins do fungo simbionte e o lixo. As informações tomadas foram: volume da bula da glândula metapleural, massa das formigas e uma estimativa do tamanho das colônias de bactéria mutualista. Análises de covariância foram utilizadas, com massa dos indivíduos como covariável, para determinar as contribuições de diversos fatores para explicar o tamanho desse órgão. Pelas análises obtidas, as duas espécies de Atta possuem bulas maiores em relação às duas espécies de Acromyrmex., provavelmente refletindo não uma maior exposição a parasitas mas uma estratégia de maior investimento em defesas. As jardineiras e as forrageadoras das espécies A. laevigata e Ac. subterraneus brunneus tiveram bulas maiores em relação às lixeiras, isto mostrando uma menor qualidade (em termos de defesas contra parasitas) das lixeiras. Neste caso, suponhamos que estes indivíduos, sendo de menor qualidade, são alocados a tarefas mais perigosas e morrerão rapidamente. Acromyrmex subterraneus brunneus foi a única que apresentou colônias de bactéria mutualista, dispostas no tórax. Houve uma relação positiva do tamanho desta colônia com o volume da bula da glândula metapleural. Essas variações nesses sistemas de defesas implicam que as formigas podem apresentar diferentes estratégias para defender o ninho contra patógenos e parasitas.

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30

Rose, Felicity. "The elucidation of antimicrobial activity in the human gastrointestinal tract." Thesis, University of Nottingham, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.285756.

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31

Reithmayer, Katrin. "Studies on the antimicrobial defence of human hair follicle epithelium /." Berlin : Mbv, 2009. http://d-nb.info/995877599/04.

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32

GAZZANEO, Luiz Rodrigo Saldanha. "Expressão heteróloga da defensina dehys de euphorbia hyssopifolia 32 em E. coli." Universidade Federal de Pernambuco, 2016. https://repositorio.ufpe.br/handle/123456789/19523.

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Defensinas são peptídeos antimicrobianos (AMPs) que apresentam atividade contra diversos microrganismos patogênicos, em especial fungos. Embora não totalmente elucidados, há diversos mecanismos de ação propostos para as defensinas, que incluem permeabilização seletiva ou ruptura da membrana plasmática de microorganismos, ação direta em alvos intracelulares, ativação de cascatas de sinalização e aumento da produção de espécies reativas de oxigênio. Desde a sua descoberta e, tendo em vista sua ampla atividade biológica, o uso de defensinas no melhoramento de plantas cultivadas, bem como na produção de novos medicamentos tem sido proposto. Estudos de atividade biológica e possível aplicação biotecnológica das defensinas demandam uma grande quantidade dessas proteínas. Entretanto, o processo de extração da mesma é laborioso, dispendioso e, de acordo com a população ou disponibilidades da espécie vegetal escolhida, não sustentável ecologicamente. Portanto, a utilização de sistemas heterólogos de expressão é uma importante ferramenta para obtenção de defensinas recombinantes em escala industrial. Nesse estudo, um gene de defensina “DeHys”, isolado da Euphorbia hyssopifolia, foi inserido no plasmídeo pET102/D-TOPO e células da linhagem BL21(DE3) de Escherichia coli foram transformada com essa construção. Foi produzida a defensina recombinante Dehys com tamanho aproximado de 24 kDa. Sua identidade foi confirmada por western blot e pela análise do padrão de digestão com proteases.
Defensins are antimicrobial peptides (AMPs) , which present activity against a variety of pathogenic microorganism, especially funghi. Although not completely elucidated, there are a variety of proposed mechanisms of action for defensins, which includes selective microorganisms plasmatic membrane permeabilization or rupture, straight action agains intracellular targets, activation of signaling cascades and the burst of reactive oxygen species. Since it’s discovery and due to it’s wide biological activities, it’s use in crop enhancing, as well as its use in the development of new drugs have been proposed. Defensin’s biological activity and biotecnological application studies demand a reasonable amount of purified protein. However, the extraction processes is laborious, expensive, time consuming and depending on the population or the chosen plant species supply, not ecologically sustainable. So, the use of heterologous expression sytems is an importante tool to obtain purified proteins in industrial scale. In this study, a defensing gene (Dehys) isolated from Euphorbia hyssopifolia was inserted in a p pET102/D-TOPO vector and trasnformed into BL21(DE3) Escherichia coli strains. A recombinat Dehys defensin of approximately 24 kDa was obtained. It’s identity was double-checked using by Western blot and protease digestion pattern analyses.

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33

Vargues, Thomas. "Antimicrobial peptides : structure, function and resistance." Thesis, University of Edinburgh, 2009. http://hdl.handle.net/1842/4076.

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Higher eukaryotes produce a vast range of antimicrobial peptides (AMPs) that play important roles in their defence against microbial infection. Beta defensins are small (3-5 kDa), cationic peptides that display broad, potent antimicrobial activity against a range of microbes and also act as chemoattractants of important immunomodulatory cells. To generate highly pure peptides for structural and functional studies, we developed a method to prepare recombinant human beta defensin-2 (HBD2). The HBD2 gene was synthesised by recursive PCR with codons optimised for expression in Escherichia coli. HBD2 was expressed as an insoluble fusion to a His-tagged ketosteroid isomerase. After cleavage from the fusion with cyanogen bromide, 1H NMR spectroscopy and mass spectrometry confirmed that the oxidised HBD2 was folded and possessed the correct b-defensin disulfide bond topology. The recombinant HBD2 was active against E. coli, P. aeruginosa, S. aureus and C. albicans and was also a chemoattractant against HEK293 cells expressing the chemokine receptor CCR6. 15N-labelled HBD2 was also prepared and was highly suitable for future structural studies. Since defensins are thought to interact with bacterial membranes we also tested the recombinant HBD2 in biophysical studies (surface plasmon resonance, SPR, Biacore). We observed different binding to artificial model membranes containing either E. coli Kdo2-lipid A or phospholipids. Bacterial resistance to AMPs has been linked to the covalent modification of the outer membrane lipid A by 4-amino-4-deoxy-L-arabinose (L-Ara4N). This neutralises the charge of the LPS, thereby decreasing the electrostatic attraction of cationic peptides to the bacterial membrane. The pathogen Burkholderia cenocepacia displays extremely high resistance to AMPs and other antibiotics and the Ara4N pathway appears to be essential. To explore this further we expressed recombinant forms of two enzymes (ArnB and ArnG) from the B. cenocepacia Ara4N pathway. Purified ArnB is a pyridoxal 5’-phosphate (PLP)-dependent transaminase and we tested its ability to bind amino acid substrates. We investigated the binding of inhibitors L- and D-cycloserine to ArnB and tested their antibiotic activity against Burkholderia strains. We also studied the B. cenocepacia ArnG – a proposed membrane protein undecaprenyl-L-Ara4N flippase – and showed that the protein behaved as a dimer by non-denaturing gel analysis. The B. cenocepacia ArnG failed to complement E. coli knock-out strains encoding the equivalent flippase proteins ArnE and ArnF, suggesting that ArnG is a Burkholderia-specific protein.

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Arranz, Trullén Javier. "Unveiling the multifaceted antimicrobial mechanism of action of human host defence RNases." Doctoral thesis, Universitat Autònoma de Barcelona, 2016. http://hdl.handle.net/10803/400666.

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La presente tesis doctoral se encuentra integrada dentro del estudio a gran escala de la estructura-función de las ribonucleasas antimicrobianas humanas. Estas proteínas catiónicas y de bajo peso molecular son secretadas por la mayoría de los organismos vertebrados agrupándose dentro la superfamilia de la ribonucleasa A, una de las enzimas mejor caracterizadas del siglo XX. De interés remarcable podríamos considerar su amplio abanico de propiedades biológicas, teniendo en cuenta su diverso historial de propiedades biológicas no catalíticas, convirtiéndolas en un buen modelo de proteínas multifunción. Junto a su principal característica como enzima catalizador de ácidos ribonucleicos, es importante destacar también otro tipo de propiedades biológicas no menos esenciales, como su actividad antimicrobiana, que comparten miembros distantes de la familia sugiriendo una función ancestral en el sistema inmune. Además, se ha visto que la expresión de algunas RNasas humanas puede ser inducida en procesos infecciosos. En particular, las RNasas estudiadas en este trabajo, las RNasas humanas 3, 6 y 7, se expresan principalmente en eosinófilos, monocitos y células epiteliales, respectivamente. Estas proteínas muestran una alta cationicidad debido a su alta proporción de residuos básicos y una notable actividad antimicrobiana frente a una amplia gama de patógenos humanos. Nuestro grupo de investigación posee una larga trayectoria en el estudio del mecanismo de acción de las ribonucleasas humanas y el trabajo teórico-experimental que se presenta en esta tesis ha contribuido a consolidar el actual proyecto de investigación. Los principales avances llevados a cabo por la presente tesis doctoral se enumeran a continuación: - La caracterización del mecanismo antimicrobiano de la ribonucleasa 6, evaluando sus propiedades microbicidas frente a patógenos y modelos de membrana. Concretamente se ha revelado su actividad aglutinadora además de demostrarse que su actividad antimicrobiana está localizada básicamente en su extremo Nterminal. - La resolución de la primera estructura tridimensional de la ribonucleasa 6, obtenida a 1.72 Å, que ha permitido asentar las bases estructurales para futuros estudios funcionales. Análisis complementarios sobre su caracterización cinética y predicción de complejos con diferentes ligandos han revelado sitios de unión y de catálisis que posteriormente han sido confirmados mediante mutagénesis dirigida. - El estudio de la efectiva actividad antipatogena a nivel intracelular que presentan las ribonucleasas 3,6 y 7 asi como sus péptidos derivados N-terminales frente a micobacterias en un modelo de macrófagos infectados. - La expansión del conocimiento sobre las bases antipatogenas de diferentes péptidos y proteínas antimicrobianas que participan en la erradicación de las infecciones por micobacterias, asi como las terapias derivadas. - La caracterización del mecanismo antimicrobiano de los 8 peptidos N-terminales derivados de las ribonucleasas frente a Candida albicans, como modelo de patógeno eucariota Como conclusión, los resultados presentados en esta tesis contribuyen a profundizar en la comprensión de las bases moleculares del papel que desempeñan algunas ribonucleasas en el sistema inmune y expandir el proyecto al diseño de agentes terapéuticos basados en péptidos antimicrobianos con el objetivo de erradicar enfermedades infecciosas causadas por patógenos resistentes.
The present doctoral thesis is integrated into the large-scale study of the structure and function of human antimicrobial ribonucleases. These cationic and low molecular weight proteins are grouped into the ribonuclease A superfamily, considered one of the best characterized enzymes of the twentieth century. The RNase A superfamily is specific for vertebrates and has attracted remarkable interest due to the diversity of displayed biological properties; and represents an excellent example of a multifunctional protein´s family. Together with the main enzymatic activity we must highlight other biological properties such as the angiogenic, immunomodulatory and antimicrobial activities. The reported antimicrobial activity of distantly related family members in early vertebrates suggests that the family arouse with an ancestral function in host defence. Besides, the expression of several human RNases has been reported to be induced by infection. In particular, the RNases studied in this work, the human RNases 3, 6 and 7, are mainly expressed in eosinophils, monocytes and epithelial cells respectively. These proteins show a high cationicity due to the high proportion of basic residues and a remarkable antimicrobial activity against a wide range of human pathogens. Our research group has a consolidated experience in the study of the mechanism of action of human ribonucleases and the experimental work presented in this thesis is contributing to this overall research project. The main results achieved by the present PhD study are outlined below: - The characterization of the antimicrobial mechanism of RNase 6, both in bacteria cell cultures and using membrane models. Results highlight that the antimicrobial and cell agglutinating activities are mainly located at the N-terminus. - The resolution of the first three-dimensional structure of ribonuclease 6, obtained at 1.72 Å, which has set the structural basis for future functional studies. The kinetic characterization of RNase 6 mutant variants and the prediction of protein- substrate complexes have identified the enzyme nucleotide binding sites. - The study of the intracellular activity of ribonucleases 3, 6 and 7 and their derived Nterminal peptides against intracellular resident mycobacteria using a macrophage infected model. - The analysis of the anti-pathogenic mechanism of action of human antimicrobial proteins and peptides in mycobacterial infections and their applied therapies. - The comparative characterization of the antimicrobial mechanism of action of human RNases and their N-terminal derived peptides towards Candida albicans, as an eukaryote pathogen model. The results presented in this thesis will contribute to the understanding of the role of human RNases in the immune system and provide the structure- function basis to expand the initial project into the design of novel peptide mimetic therapeutics agents towards the eradication of resistant infectious diseases.

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35

Reithmayer, Katrin [Verfasser]. "Studies on the antimicrobial defence of human hair follicle epithelium / Katrin Reithmayer." Berlin : Freie Universität Berlin, 2009. http://d-nb.info/102366383X/34.

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36

Beaumont, Paula Elizabeth. "Cathelicidin and its role in defence against bacterial infections of epithelial cells." Thesis, University of Edinburgh, 2015. http://hdl.handle.net/1842/15847.

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Cathelicidins are antimicrobial peptides (AMPs) that were first discovered to have microbicidal properties but more recently to be multifunctional immunomodulators and thus important in influencing host defence against infectious disease. Whilst roles in various organs have been demonstrated, their expression patterns in health and disease in other organs are less clear and their key immunomodulatory functions remain undefined, particularly with regard to the balance of immunomodulatory properties and microbicidal activity in their ability to promote defence against infection. I therefore set out to describe LL-37 expression (human cathelicidin) in the female reproductive tract (across the menstrual cycle) and in the lung (during specific lung diseases), to define the effects on the function of airway epithelial cells during bacterial infection and to evaluate the key in vivo roles of endogenous cathelicidin (using a knockout mouse model) as well as the effect of therapeutic administration of LL-37 in a pulmonary Pseudomonas aeruginosa infection model. I demonstrated that cathelicidin protein and transcription shows a cyclical pattern of expression in female reproductive tissues which is maintained at high levels in decidua. LL- 37 protein was also detected in hTERT endometrial epithelial cells but despite the suggestion that cathelicidin may be regulated by steroid hormones there was no direct effect of progesterone on transcription. LL-37 is barely detected in healthy airways however is well known to increase during infection or inflammation. I observed that sputum from patients with bronchiectasis showed a correlation between the level of LL-37, TNF, MPO and chronic colonisation of Pseudomonas aeruginosa. Patients with lung cancer expressed much less LL- 37 than the bronchiectasis patients but there was a trend towards increased production postsurgery compared to pre-surgery. LL-37 was previously shown by our lab to selectively promote BAX and caspase-dependant death of infected epithelial cells. I went on to show that this appears to be a partially caspase- 1 dependent mechanism and that human bronchial epithelial (HBE) cells and A549 cell lines both express several of the components required to form inflammasomes, a caspase-1 dependant form of inflammatory cell death. Finally, I showed using murine models that cathelicidin enhances bacterial clearance during pulmonary infection in vivo, a response which is defective in mice lacking endogenous cathelicidin and that administration of exogenous, synthetic LL-37 at the time of infection can promote an early protective neutrophil influx in the absence of endogenous cathelicidin production.

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Oliveira, Jaqueline Franciele Caetano de. "Lectinas vegetais: de moléculas de defesa de plantas às suas diversas aplicações biotecnológicas." Universidade Estadual do Oeste do Paraná, 2018. http://tede.unioeste.br/handle/tede/3931.

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Lectins are proteins belonging to a diverse superfamily, which has as its main characteristic, the capacity to specifically and selectively recognize carbohydrates. They are present in a great variety of microorganisms, such as viruses, bacteria, fungi, plants, and animals. Due to its ability to recognize a large number of glycans, they behave as mediators in many biological processes, as in cell migration, immunologic defense and in bacterial, protozoan and viral infections. Among all, plant lectins are the most studied, showing structural heterogeneities, mainly on its carbohydrates binding sites, what grants several biological activities to these proteins, among them, the antimicrobial, insecticide, mitogenic, anti-tumoral and anti-inflammatory activities. On the last decades, a large number of studies have demonstrated the application of lectins as a potential biotechnological tool on diverse fields like on agriculture, being used at the genetic improvement of grains; on biochemistry, where it’s used at pre-purification of glycoproteins, and on the histochemistry, applied at the distinction between normal and tumoral cells. Nowadays, one of the researched applications is the potential of lectins as a carcinogenic agent, so as its uses on the discovery of biomarkers. The present study shows a brief history of the plant lectins, their functional characteristics, and structural, their biotechnological applications, such as in the biomarkers discovery, covering too, some of the techniques employed to this function, like the Lectin Affinity Chromatography (LAC), Lectin Microarrays and the Antibody-Lectin Sandwich Array.
As lectinas são proteínas pertencentes a uma diversificada superfamília, que possui como principal característica a capacidade de reconhecer carboidratos de modo específico e seletivo. Estão presentes nos mais variados organismos, tais como, vírus, bactérias, fungos, vegetais e animais. Devido à sua habilidade de reconhecer uma grande quantidade de glicanos, são mediadoras em diversos processos biológicos, como migração celular, defesa imunológica, infecção por bactérias, vírus e protozoários. As lectinas de plantas são as mais estudadas e revelam-se com heterogeneidades estruturais principalmente na constituição de seus sítios de ligação ao carboidrato, o que lhes confere múltiplas atividades biológicas, dentre elas, a ação antimicrobiana, a ação inseticida, a ação mitogênica, a ação antitumoral e a anti-inflamatória. Nas últimas décadas, um grande número de publicações tem demonstrado a aplicação das lectinas como ferramenta biotecnológica potencial em diferentes áreas, como na agricultura, para melhoria genética de cultura de grãos; na bioquímica, utilizadas na pré-purificação de glicoproteínas e na histoquímica, para distinguir células tumorais das normais. Umas das aplicações amplamente pesquisadas na atualidade é o potencial das lectinas como agentes carcinogênicos, como também a sua utilização na descoberta de biomarcadores. Neste trabalho, apresentamos uma visão geral sobre as lectinas, incluindo um breve histórico das lectinas vegetais, suas características funcionais e estruturais básicas, suas atividades com potenciais aplicações biotecnológicas dentre elas a inseticida, a antimicrobiana e a antitumoral, bem como o emprego dessas proteínas na descoberta de biomarcadores tumorais. Algumas técnicas utilizadas para essa finalidade como a cromatografia de afinidade com lectina imobilizada (LAC - Lectin Affinity Chromatography) e os microarranjos de lectinas e lectinas/anticorpo no formato sanduíche (ALSA - Antibody-Lectin Sandwich Array) são apresentadas.

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38

Fiocco, Daniela. "α-DEFENSINS EXPRESSION IN PERIPHERAL BLOOD MONONUCLEAR CELLS FROM PATIENTS WITH HEPATITIS C VIRUS INFECTIONS". Doctoral thesis, La Sapienza, 2005. http://hdl.handle.net/11573/916796.

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39

Hilal, George E. "Antimicrobial activity of human leukocyte defensin HNP-4 against gram-negative bacteria." View the abstract Download the full-text PDF version, 2008. http://etd.utmem.edu/ABSTRACTS/2008-017-Hilal-index.html.

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Thesis (M.S. )--University of Tennessee Health Science Center, 2008.
Title from title page screen (viewed on July 31, 2008). Research advisor: Edwin L. Thomas, Ph.D. Document formatted into pages (xi, 50 p. : ill.). Vita. Abstract. Includes bibliographical references (p. 44-50).

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40

Chileveru, Haritha Reddy. "Visualizing the attack of bacteria by the antimicrobial peptide human defensin 5." Thesis, Massachusetts Institute of Technology, 2016. http://hdl.handle.net/1721.1/105021.

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Thesis: Ph. D. in Biological Chemistry, Massachusetts Institute of Technology, Department of Chemistry, 2016.
Cataloged from PDF version of thesis.
Includes bibliographical references.
Bacterial infections are a major cause of concern in healthcare because of a rise in antibiotic-resistant bacteria and an increase in hospital-acquired infections. In order to combat bacterial infections, we need fundamental understanding of the host-pathogen interaction. As a part of the innate immune response, various organisms, including humans, produce antimicrobial peptides. Human defensin 5 (HD5) is a 32-aa cysteine-rich peptide produced primarily in the small intestine that exhibits broad-spectrum activity against various bacteria, fungi and viruses. In this thesis, in order to study and understand the mechanism of action of HD5, we probed the antibacterial action of HD5 and the bacterial response. We synthesized a family of HD5 analogues with functional modifications including fluorophores. With this toolkit of HD5 derivatives, we examined the effect of HD5 on various bacteria. We demonstrated that HD5ox, the oxidized form of HD5, causes certain distinct morphological changes in Gram-negative bacteria, enters the bacterial cytoplasm and localizes near the cell poles and cell division sites. From these studies we propose that HD5ox overcomes the outer membrane permeability barrier and permeabilizes the inner membrane of Gram-negative bacteria, and it may interact with the cellular targets and interfere with the processes such as cell division. We extended these morphological studies of HD5 0x-treated bacteria and identified certain phenotypic responses in Gram-positive bacteria that further suggest that HD5ox interferes with cellular processes such as cell division.
by Haritha Reddy Chileveru.
Ph. D. in Biological Chemistry

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41

Salazar, Montoya Vivian Angélica. "Exploring the mechanism of action of human antimicrobial ribonucleases." Doctoral thesis, Universitat Autònoma de Barcelona, 2015. http://hdl.handle.net/10803/310611.

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Las ribonucleases humanas son un grupo heterogéneo de proteínas pertenecientes a la superfamilia de la Ribonucleasa A. Estas proteínas se caracterizan por su capacidad de hidrolizar ácidos ribonucleicos y por la presencia de actividad antimicrobiana frente diversos organismos patógenos como bacterias, hongos, parásitos y virus. El primer objetivo del presente estudio doctoral se centra en la caracterización de la actividad antimicrobiana y en modelos de membrana de las formas nativas de la ribonucleasa 3 purificadas a partir de eosinófilos. Las distintas formas nativas presentan modificaciones postraduccionales dadas por diversos grados de glicosilación que se correlacionan con la activación de los eosinófilos durante los procesos inflamatorios. El estudio establece la capacidad antimicrobiana de las formas nativas y su actividad en modelos de membrana. Los resultados indican que las modificaciones postrasduccionales modulan la actividad biológica de la RNasa 3, sugiriendo una contribución in vivo en su función fisiológica. Como segundo objetivo de esta tesis, se evaluó por primera vez en nuestro grupo de investigación la actividad antimicótica de las ribonucleasas 3 y 7 frente al hongo Candida albicans, el cual fue elegido como modelo patógeno eucariota. Se determinó y caracterizó la presencia de actividad frente a Candida por parte de ambas ribonucleasas humanas. Por último, el tercer objetivo de esta tesis se centra en la purificación y caracterización de la ribonucleasa 8, la más reciente ribonucleasa humana descrita, identificada inicialmente en placenta. La RNasa 8 presenta un patrón inusual de enlaces disulfuro respecto a sus proteínas homólogas. Este cambio estructural modifica la estabilidad de la proteína y expone regiones que facilitan el proceso de agregación proteica. Fue necesaria la previa optimización de un protocolo alternativo de purificación. Se analizaron sus propiedades antimicrobianas, sugiriendo su posible participación en la respuesta inmunitaria innata. Los resultados del presente estudio corroboran las propiedades antimicrobianas de diversas ribonucleasas humanas miembros de la familia de la RNasa A, sugiriendo una función ancestral en el sistema de defensa innato. El estudio contribuye a la comprensión de su mecanismo de acción y plantea su potencial uso como herramientas terapéuticas.
Human ribonucleases are a heterogeneous group of proteins belonging to the superfamily of RNase A. These proteins are characterized by their ability to hydrolyse ribonucleic acids and the presence of antimicrobial activity against various pathogens including bacteria, fungi, parasites and viruses. The first objective of this doctoral study is focused on the antimicrobial characterization of native Ribonuclease 3 forms purified from eosinophils. Native forms present posttranslational modifications giving different glycosylation grades that modulate their activity during inflammatory processes. This study aims to establish the antimicrobial properties of native forms purified from eosinophils and their activity in a membrane model system. Results indicate that post-translational modifications contribute to the the protein biological activities, suggesting a related physiological role. As a second objective, we evaluated for the first time the antifungal activity of the antimicrobial RNase 3 and RNase 7 against Candida albicans, an eukaryotic pathogen selected as a simple model to test the antimicrobial mechanism of action. Both human ribonucleases displayed a high antifungal activity. Results highlighted a dual mechanism of action, where cell lysis takes place at high protein concentration, while depolarization, cell internalization and cellular RNA degradation is achieved at sublethal doses. Finally, the last objective is focused on the characterization of ribonuclease 8, also called the placental RNase, the most recent human ribonuclease described. RNase 8 has gained and lost one cysteine residue in non-conserved positions in a mechanism called "disulphide shuffling". The protein tendency to aggregate required the design of an alternative purification protocol. We analysed its antimicrobial abilities, suggesting a possible role in innate defence. The results of this study confirmed the high antimicrobial activity of several human ribonucleases from the RNase A superfamily suggesting an ancestral role in the host immune defence response. The study contributed to the understanding of their mechanism of action and set the basis for applied drug design.

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42

Funderburg, Nicholas Thomas. "Human Beta Defensin 3: Linking Innate and Adaptive Immune Responses." Case Western Reserve University School of Graduate Studies / OhioLINK, 2008. http://rave.ohiolink.edu/etdc/view?acc_num=case1189084245.

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43

Meisch, Jeffrey P. "Human β-defensin 3 peptide is increased and redistributed in Crohn’s ileitis". Case Western Reserve University School of Graduate Studies / OhioLINK, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=case1270735950.

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Lioi, Anthony Bruno. "Monocyte Activation and Membrane Disruption Mediated by Human ß-Defensin-3." Case Western Reserve University School of Graduate Studies / OhioLINK, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=case1386255566.

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Mengesha, Bemnet Gashawbeza. "The Role of Antimicrobial Peptide Murine Beta Defensin-3 in Protection against Oropharyngeal Candidiasis." University of Toledo / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=toledo1510521465214606.

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46

Fahlgren, Anna. "Defence capabilities of human intestinal epithelial cells." Doctoral thesis, Umeå : Univ, 2003. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-151.

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47

Junior, José Carlos Bozelli. "Interação do peptídeo de defesa do hospedeiro tritrpticina (TRP3) e seus análogos com membranas modelo: efeitos na estrutura e dinâmica da membrana." Universidade de São Paulo, 2015. http://www.teses.usp.br/teses/disponiveis/46/46131/tde-07122015-113945/.

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Tritrpticina (TRP3) é um peptídeo antimicrobiano com 13 resíduos de amino ácidos com três Ws sequenciais. Com o objetivo de contribuir para a compreensão de seu mecanismo de ação, realizaram-se estudos funcionais e conformacionais da TRP3 e de dois análogos onde um (WLW) ou dois (LWL) W foram substituídos por L. Os peptídeos foram igualmente ativos contra bactérias Gram positivas e negativas. Sua atividade hemolítica requereu concentrações maiores, diminuindo na ordem TRP3>WLW>LWL. Os peptídeos permeabilizaram membranas modelo de E. coli ou contendo fosfolipídios carregados negativamente. Espectros de CD sugeriram que os peptídeos adquirem diferentes conformações ao se ligarem a bicamadas e micelas. Estudos de fluorescência mostraram que a ligação a membranas decresce na ordem: TRP3>WLW>LWL e que os peptídeos se localizam próximos à interface membrana-água. Espectros de RPE de marcadores de spin lipídicos indicaram que a ligação dos peptídeos altera a organização dos lipídios, aumentando o empacotamento molecular
Tritrpticin (TRP3) is a 13-residue antimicrobial peptide that contains three sequential Ws. With the aim of contributing to the understanding of its mechanism of action, functional and conformational studies were performed with TRP3 and two of its analogues where one (WLW) or two (LWL) of the W were replaced by L. The peptides were equally active against both Gram positive and Gram negative bacteria. Higher concentrations were required for hemolytic activity which varied in the order: TRP3>WLW>LWL. The peptides permeabilized membranes model membranes mimicking E. coli\'s lipid composition or containing different negatively charged phospholipids. CD spectra suggested the peptides acquired different conformations upon binding to bilayers or micelles. Fluorescence studies showed that membrane binding decreases in the order: TRP3>WLW>LWL and that the peptides are located close to the water-membrane interface. EPR spectra of lipid spin labels indicated that peptide binding alter lipid organization, increasing molecular packing

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48

Bailleul, Geoffrey. "Les défensines aviaires : nouveau moyen de lutte contre les entérobactéries pathogènes." Thesis, Tours, 2016. http://www.theses.fr/2016TOUR3813.

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L’utilisation intensive des antibiotiques a conduit à l’émergence inquiétante de souches bactériennes multi-résistantes, notamment chez les salmonelles et les colibacilles responsables de maladies infectieuses préoccupantes en santé publique et vétérinaire. Les peptides de défense de l’hôte (HDP) constituent les effecteurs antimicrobiens essentiels de l’immunité innée des animaux. La principale famille d’HDPs chez les oiseaux sont les défensines aviaires, dont deux membres (AvBD2 et AvBD7) isolés de la moelle osseuse du poulet présentent un large spectre antibactérien et constituent ainsi des molécules candidates alternatives aux antibiotiques. Pour démontrer l’intérêt thérapeutique de l’AvBD2 et de l’AvBD7, nous avons (1) prouvé leur efficacité antibactérienne in vitro contre des souches cliniques multirésistantes, (2) montré leur stabilité structurale et fonctionnelle face aux protéases digestives majeures, (3) mis en évidence un nouvel effet antimicrobien dans les macrophages infectés, et finalement (4) prouvé le concept thérapeutique en réduisant la mortalité et la charge bactérienne après injection d’AvBD7 dans un modèle de salmonellose systémique létale chez la souris
Large-scale use of antibiotics has leaded to the emergence of multi drug resistant bacterial strains, particularly in the genus Salmonella and Escherichia coli responsible for infectious diseases outbreaks. This constitutes a major veterinary and public health concern. Host defense peptides (HDPs) represent major effector molecules of the animal’s innate immune system. In birds, the main family of HDPs is composed of avian defensins whose two members (AvBD2 and AvBD7) isolated from chicken bone marrow exhibit large spectrum antibacterial activity. Thus, they both constitute potential candidate molecules as alternative to antibiotics. To demonstrate the therapeutic interest of AvBD2 and AvBD7, we have (1) proved their antibacterial efficiency in vitro towards clinical and multi drug resistant bacterial strains, (2) shown their functional and structural stability when facing major digestive proteases, (3) identified a novel antibacterial effect in infected macrophages, and finally (4) proved the therapeutic concept by reducing mortality and bacterial load after AvBD7 injection in a murine model of lethal salmonellosis

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Fite, Alemu. "The antimicrobial interaction of acidified nitrite with other physiologically active compounds (PAC), the potential host defence mechanism of dietary nitrate." Thesis, University of Aberdeen, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.369736.

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Transfer of nitrite at salivary concentrations enhance the antimicrobial activity of stomach acid against food borne pathogens and may be involved in the host defence of humans and animals. Studies from experimental animals, microbial tests, human ex vivo and in vivo investigations support the hypothesis that dietary nitrate is beneficial for health. The positive findings that acidified nitrite is antimicrobial at the acidic conditions of the stomach is viewed in conjunction with the potential production of nitrogen, intermediates such as NO, a molecule of significant importance in science. This led to propose that acidified nitrite can kill or inhibit enteric bacterial species that are associated with gastrointestinal diseases and could modify the antimicrobial effects of acidified nitrite by other physiologically active compounds. By a systematic exposure of selected enteric bacterial species to acidified nitrite and other PAC alone or in combinations, it was found that acidified nitrite, especially at low pH values was inhibitory to bacterial growth or killer of bacterial cells. Glutathione and vitamin C inhibited the antimicrobial effects of nitrite whilst thiocyanate and iodide enhanced it. Thiocyanate produced an effective antimicrobial activity that was comparable to nitrite but there was no synergistic antimicrobial interaction between acidified nitrite and thiocyanate. The addition of thiocyanate enhanced the antimicrobial effects of cow's milk in a pH-dependent activity. The overall results reported here suggested to conclude that dietary factors found in fruits and vegetables are not only beneficial in the supply of vitamins, antioxidants and enzymes but also antimicrobial agents that are worth investigation for therapeutic purposes.

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Ribeiro, Ana Elisa Rodrigues Alves. "Componentes salivares como fatores de defesa frente a fatores locais." Universidade de São Paulo, 2015. http://www.teses.usp.br/teses/disponiveis/58/58137/tde-16062015-111521/.

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A saliva é uma mistura de água, eletrólitos, proteínas e enzimas. A secreção diária normal é de 800-1500ml em adultos. A chamada saliva total, o fluído que realmente está presente na cavidade bucal, é produzida por diferentes glândulas salivares e contém ainda fluído crevicular e elementos transudados do plasma, e derivados da rede capilar da mucosa bucal. É importante entender o papel da saliva na proteção dos tecidos bucais, principalmente porque a co-infecção pode ser um fator importante na ativação e supressão do sistema imune, com papel importante no desenvolvimento e severidade das doenças bucais. Além disso, a cavidade bucal tem um vasto número de microrganismos e antígenos presentes, o que faz com que seja considerada em permanente estado de inflamação, em muitos casos, subclínica. Nosso estudo se propõe a observar a variação da composição salivar frente a presença de alterações locais - gengivites e periodontites. O estudo compara as citocinas salivares TNF-α, IL-1β e IL-6, e os fatores de defesa, beta defensinas 1 e 2, catelicidina e mucina 2, em três diferentes grupos de pacientes: Grupo 1 (controle) - 40 Pacientes, total ou parcialmente dentados, sem inflamação/infecção bucal; Grupo 2 - 40 Pacientes total ou parcialmente dentados, com sinais clínicos de gengivite; e Grupo 3 - 40 Pacientes total ou parcialmente dentados, com sinais clínicos de periodontite. A presença das citocinas e fatores salivares foram avaliadas por testes ELISA. Foram significativas as alterações encontradas entre os grupos para os diferentes fatores: TNF-α, e IL-6, beta defensinas 1 e 2, catelicidina e mucina 2. Apenas IL-1β não teve resultados significantes. Assim, indica-se que os componentes salivares possuem importante papel salivar frente à alterações locais.
Saliva is a mixture of water, electrolytes, proteins and enzymes. The daily secretion ranges between 800-1500mL in adults. The called whole saliva is composed by the production of different salivary glands, gingival crevicular fluid, and contain elements transudate from plasma derived from the capillary bed beneath the oral mucosa. It is important to consider the evident and important role of saliva in defense and protection of oral tissues. The effects of co-infecting pathogens have been postulated as an important factor in the activation and/or suppression of immune system, important in many situations, including the severity and rate of disease progression. The oral cavity is continually confronted with a vast number of pathogens and antigens, so, in some way, may be considered an inflammatory environment, although the level of inflammation may be sub-clinical. This study proposed to observe how the presence of local inflammation - gingivitis or periodontitis, may influence the presence of salivary cytokines or defense factors in saliva. The study compared saliva molecular components in three different groups of patients: Group 1 (as control group) - 40 Patients, total or partially dentate, without oral infectious; Group 2 - 40 Patients total or partially dentate, with clinical signs of gingivitis; Group 3 - 40 patients, total or partially dentate, with clinical signs of periodontitis. It checked the presence of TNF-α, IL-1β and IL-6 cytokines, and defense factors, 1 and 2 beta defensins, cathelicidin and mucin 2. ELISA kits determined the levels of these proteins. Found alterations were significant between groups to TNF-α, IL-6, 1 and 2 beta defensins, cathelicidin and mucin 2. Only IL-1β had not significant results. Therefore, it indicated that salivary components have important hole related to local situations.

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