Multifuncional biopolymer-bases materials for modulatig the activites of chromic wound enzymes

This thesis focuses on the development of active multifunctional dressing materials and nanoparticle formulations with suitable exploitation characteristics for chronic wounds treatment. Chronic wounds a growing clinical challenge in the aging and/or reduced mobility population include pressure, venous, arterial and diabetic neuropathic ulcers. Due to the nonhealing character of these ulcers their management requires an intensive medical intervention at huge healthcare costs. The prolonged inflammation and elevated concentrations of oxidative and proteolytic enzymes in all chronic wounds, imposes the need for novel functional dressing materials to actively modulate the wound environment at molecular level and stimulate the healing process. Based on an extensive analysis of the current state-of-the-art in chronic wound healing, the proper dressings should combine both antimicrobial and enzyme inhibitory functions coupled to optimal hydrophilicity. Such integrated approach would allow for the suppression of the persistent inflammation and stimulation of the synthesis of the dermal tissue components. Biopolymers with intrinsic antimicrobial and wound repair properties appear as appropriate matrix materials to be further upgraded with bioactive molecules (therapeutics) that couple high reactivity with the ability to address specific targets in the biochemical environment of chronic wounds. Therapeutic devices can be designed in different forms depending on the particular clinical application, i.e. wound type and its characteristics. During the thesis realisation biopolymer-based platforms were generated in various designs and functionalised with active agents for controlled inhibition of major chronic wound enzymes. The capacity of all developed materials to inhibit proteolytic (e.g. collagenase) and oxidative (e.g. myeloperoxidase) enzymes involved in chronic inflammation was evaluated in vitro. In the first approach sponge-like biopolymer matrices were produced via freeze-drying technique and controlled chemical cross-linking. These matrices were further impregnated with natural polyphenolic compounds. Modulation of the deleterious wound enzyme activities was achieved upon release of active agent from the platform. The exploitation characteristics of the sponges, i.e. mechanical properties, biostability, biocompatibility, extent and duration of wound enzymes inhibition, were tuned by: the biopolymer composition, concentration of the cross-linking agent, and the proper selection of the bioactive phenolic compounds. The second approach aimed at the permanent functionalisation of the biopolymeric platforms with thiol-bearing compounds. In this case the active agent is expected to act from the platform, without being released into the wound. The obtained thiolated biopolymers were further processed into functional nanomaterials of different design: *Nanoscale films/coatings were built using a layer-by-layer approach for alternate deposition of oppositely charged polyelectrolytes. Naturally occurring glycosaminoglycans were used as counterions to cationic thiolated conjugates. *Nanoparticle formulations were obtained from thiolated conjugates in a one-step sonochemical process. In both cases the biopolymer thiolation degree was identified as a key factor for the successful fabrication of the multilayered coatings and nanoparticles, as well as to achieve control of the thickness/size of the functional nanomaterials. In addition, tuneable inhibition/adsorption of the deleterious enzymes coupled to fibroblast attachment/proliferation was observed by ruling the biopolymer modification degree.Postprint (published version

Multifuncional biopolymer-bases materials for modulatig the activites of chromic wound enzymes

This thesis focuses on the development of active multifunctional dressing materials and nanoparticle formulations with suitable exploitation characteristics for chronic wounds treatment. Chronic wounds a growing clinical challenge in the aging and/or reduced mobility population include pressure, venous, arterial and diabetic neuropathic ulcers. Due to the nonhealing character of these ulcers their management requires an intensive medical intervention at huge healthcare costs. The prolonged inflammation and elevated concentrations of oxidative and proteolytic enzymes in all chronic wounds, imposes the need for novel functional dressing materials to actively modulate the wound environment at molecular level and stimulate the healing process. Based on an extensive analysis of the current state-of-the-art in chronic wound healing, the proper dressings should combine both antimicrobial and enzyme inhibitory functions coupled to optimal hydrophilicity. Such integrated approach would allow for the suppression of the persistent inflammation and stimulation of the synthesis of the dermal tissue components. Biopolymers with intrinsic antimicrobial and wound repair properties appear as appropriate matrix materials to be further upgraded with bioactive molecules (therapeutics) that couple high reactivity with the ability to address specific targets in the biochemical environment of chronic wounds. Therapeutic devices can be designed in different forms depending on the particular clinical application, i.e. wound type and its characteristics. During the thesis realisation biopolymer-based platforms were generated in various designs and functionalised with active agents for controlled inhibition of major chronic wound enzymes. The capacity of all developed materials to inhibit proteolytic (e.g. collagenase) and oxidative (e.g. myeloperoxidase) enzymes involved in chronic inflammation was evaluated in vitro. In the first approach sponge-like biopolymer matrices were produced via freeze-drying technique and controlled chemical cross-linking. These matrices were further impregnated with natural polyphenolic compounds. Modulation of the deleterious wound enzyme activities was achieved upon release of active agent from the platform. The exploitation characteristics of the sponges, i.e. mechanical properties, biostability, biocompatibility, extent and duration of wound enzymes inhibition, were tuned by: the biopolymer composition, concentration of the cross-linking agent, and the proper selection of the bioactive phenolic compounds. The second approach aimed at the permanent functionalisation of the biopolymeric platforms with thiol-bearing compounds. In this case the active agent is expected to act from the platform, without being released into the wound. The obtained thiolated biopolymers were further processed into functional nanomaterials of different design: *Nanoscale films/coatings were built using a layer-by-layer approach for alternate deposition of oppositely charged polyelectrolytes. Naturally occurring glycosaminoglycans were used as counterions to cationic thiolated conjugates. *Nanoparticle formulations were obtained from thiolated conjugates in a one-step sonochemical process. In both cases the biopolymer thiolation degree was identified as a key factor for the successful fabrication of the multilayered coatings and nanoparticles, as well as to achieve control of the thickness/size of the functional nanomaterials. In addition, tuneable inhibition/adsorption of the deleterious enzymes coupled to fibroblast attachment/proliferation was observed by ruling the biopolymer modification degree

Assessment of attitudes among medicine students at the University of Mainz, Germany towards communication skills training regarding the treatment of chronic musculoskeletal pain

Chronische Schmerzen stellen eine große Herausforderung auf mehreren Ebenen für das Gesundheitssystem dar, welche in Industrieländern zunimmt und zu den Top 10 der globalen Krankheitslasten gehört (Hoy et al., 2014). Chronische muskuloskelettale Schmerzen wirken sich negativ auf die Funktion des Bewegungsapparates aus und beeinträchtigen so das allgemeine Funktionsniveau und die Fähigkeit zur Teilhabe von Patienten, indem sie die Mobilität und Lebensqualität dieser einschränken (Jordan et al., 2010). Chronische muskuloskelettale Schmerzen stellen eine multidimensionale Herausforderung für das Gesundheitssystem dar (Hadi et al., 2017). Die Belastung auf verschiedensten Ebenen, die chronische Schmerzen in der Gesellschaft darstellen, macht es zu einer kritischen Angelegenheit, die untersucht und behandelt werden muss. Eine unzureichende Kommunikation zwischen Primärversorgern und Patienten stellt eines der Haupthindernisse für eine wirksame Behandlung chronischer Schmerzen dar (Butow und Sharpe, 2013). Die Wirksamkeit der Behandlung von chronischen muskuloskelettalen Schmerzen durch Primärversorger wird positiv durch den Kommunikationsstil zwischen dem Arzt und dem Patienten beeinflusst (Frantsve und Kerns, 2007). Insbesondere unterstützt die Forschung die Implementierung eines Shared-Decision-Making-Ansatzes (SDM) (Parsons et al., 2012). SDM ist ein kollaborativer Prozess, welcher die gemeinsame Verantwortung für die medizinische Versorgung des Patienten fördert und gleichzeitig die Präferenzen des Patienten in den Entscheidungsprozess integriert (Frantsve und Kerns, 2007). Daher ist die Lehre und der Erwerb von Kommunikationsfähigkeiten in der medizinischen Ausbildung notwendig, damit die Studierenden ausreichend qualifizierte Ärzte werden können (Alotaibi und Alsaeedi, 2016). Das Verständnis der Einstellungen von Medizinstudierenden zum Erlernen von Kommunikationsfähigkeiten ist für die Fakultätsangehörigen, Bildungsprogrammgestaltenden und Pädagogen von zentraler Bedeutung (Alotaibi und Alsaeedi, 2016). Die Einstellung von Medizinstudierenden zum Training kommunikativer Fähigkeiten hängt mit einer Vielzahl von Faktoren zusammen, die mit ausbildungs- und demografischen Variablen verbunden sind, einschließlich des Semesters, in welchem sich die Studierenden befinden, ihres Geschlechts und ob die Eltern der Studierenden Ärzte sind oder nicht (Rees und Sheard, 2002). Insgesamt ist ein erhöhter Bedarf an der Integration und Optimierung bestehender Lehrinhalte welche Kommunikationsfähigkeiten adressieren in den medizinischen Fakultäten zu beobachten (Frantsve und Kerns, 2007). Kommunikationstraining ist in den von deutschen Hochschulen verwendeten Lernzielkatalogen unzureichend vertreten und wird daher nicht ausreichend in die Prüfungen für Medizinstudierende mit einbezogen (Jünger, 2017). Eine umfassende Neustrukturierung der Lernzielkatalogs ist daher ein zentraler Schritt zur Verbesserung der kommunikativen Ausbildung von angehenden Ärzten (Jünger, 2017). Gemäß dem in Deutschland verabschiedeten Masterplan Medizinstudium 2020 soll das Medizinstudium stärker auf die Vermittlung kommunikativer Kompetenzen ausgerichtet werden (wissenschaftsrat.de, 2018). Die vorliegende Dissertation untersuchte, ob zwischen Medizinstudierenden der Universität Mainz des ersten und zweiten Semesters (d.h. Studienanfängern) im Vergleich zu Studierenden des neunten und zehnten Semesters (d.h. Studierenden im fünften Studienjahr) ein Unterschied in der Einstellung zur Kommunikationsausbildung besteht und ob das Geschlecht der Studierenden und die Ärzteschaft als Eltern ihre Einstellung zum Kommunikationstraining beeinflussen. Insgesamt N = 270 (67,8% weiblich; M Alter = 23,62, SD = 3,84) Medizinstudierende nahmen an der Universität Mainz an der Umfrage dieser Dissertation teil. Der Communication Skills Attitude Scale (CSAS) wurde verwendet. Die Einstellung zur Ausbildung kommunikativer Fähigkeiten Studierender unterscheidet sich nicht signifikant zwischen Studierenden in weniger fortgeschrittenen Semestern und höheren Semestern. Darüber hinaus wurden keine signifikanten geschlechtsspezifischen Unterschiede in Bezug auf die Einstellung zum Training kommunikativer Fähigkeiten gefunden. Weiterhin unterscheidet sich die Einstellung zum Kommunikationstraining nicht signifikant zwischen Studierenden, deren Eltern Ärzte sind, und solchen, deren Eltern keine Ärzte sind. Es besteht ein klarer Bedarf, die Einstellungen zur Kommunikationsfähigkeit von Studierenden an der Universität Mainz sowie Faktoren, die diese Einstellungen beeinflussen, weiter zu untersuchen, um behandlungsfördernde Kommunikationsstile zukünftiger Ärzten zu fördern. Die vorliegende Dissertation zeigt Möglichkeiten zur systematischen Verbesserung des medizinischen Curriculums an der Universität Mainz auf, um die kommunikative Ausbildung von Studierenden und zukünftigen Ärzten zu verbessern.ca. 55 Seite

Hydrogel dressings for advanced wound management

The published manuscript is available at EurekaSelect via http://www.eurekaselect.com/openurl/content.php?genre=article&doi=10.2174/0929867324666170920161246Composed in a large extent of water and due to their non-adhesiveness, hydrogels found their way to the wound dressing market as materials that provide a moisture environment for healing while being comfortable to the patient. Hydrogels’ exploitation is constantly increasing after evidences of their even broader therapeutic potential due to resemblance to dermal tissue and ability to induce partial skin regeneration. The innovation in advanced wound care is further directed to the development of so-called active dressings, where hydrogels are combined with components that enhance the primary purpose of providing a beneficial environment for wound healing. The aim of this mini-review is to concisely describe the relevance of hydrogel dressings as platforms for delivery of active molecules for improved management of difficult-to-treat wounds. The emphasis is on the most recent advances in development of stimuli-responsive hydrogels, which allow for control over wound healing efficiency in response to different external modalities. Novel strategies for monitoring of the wound status and healing progress based on incorporation of sensor molecules into the hydrogel platforms are also discussed.Peer ReviewedPostprint (author's final draft

Immobilization of antimicrobial core-shell nanospheres onto silicone for prevention of Escherichia coli biofilm formation

Escherichia coli (E. coli) strains are among the most frequently isolated microorganisms in urinary tract infections able to colonize the surface of urinary catheters and form biofilms. These biofilms are highly resistant to antibiotics and host immune system, resulting in increased morbidity and mortality rates. Strategies to prevent biofilm development, especially via restricting the initial stages of bacteria attachment are therefore urgently needed. Herein, a common urinary catheter material – polydimethylsiloxane (PDMS) – was covalently functionalized with antibacterial aminocellulose nanospheres (ACNSs) using the epoxy/amine grafting chemistry. The PDMS surface was pre-activated with (3-glycidyloxypropyl)-triethoxysilane to introduce epoxy functionalities prior to immobilization of the intact ACNSs via its amino groups. The AC biopolymer was first sonochemically processed into NSs improving by up to 80% its potential to prevent the E. coli biofilm formation on a polystyrene surface. The silicone surface decorated with these NSs demonstrated efficient inhibition of E. coli biofilms, reducing the total biomass when compared with pristine silicone material. Therefore, the functionalization of silicone-based materials with ACNSs shows promise as potential platform for prevention of biofilm-associated infections caused by E. coli.Peer ReviewedPostprint (author's final draft

Sonochemical synthesis and stabilization of concentrated antimicrobial silver-chitosan nanoparticle dispersions

This work reports on a green synthetic route to produce concentrated aqueous dispersions of silver nanoparticles (AgNP) employing high-intensity ultrasound (US) and chitosan (CS) as a non-toxic reducing agent for Ag1 salts and AgNP stabilizer. The sonication simultaneously boosted the synthesis and improved the stability of the AgNP, capping them with CS. Hybrid AgNP-CS antimicrobial dispersions, stable for at least 6 months, were synthesized in a simple single step process. The use of US allowed for applying relatively mild processing temperatures (608C) and reaction time between 30 min and 3 h to obtain concentrated disper- sions of AgNP that otherwise could not be obtained even after 72 h under mechanical stirring at the same reaction conditions. Upon sonication spherical AgNP-CS with a size between 60 and 100 nm were generated, in contrast to the average diameter of 200 nm of the particles obtained by stirring. The antibacterial efficiency of the AgNP-CS hybrids was evaluated against the medically relevant pathogens Staphylococcus aureus and Escherichia coli. The US-synthesized AgNP-CS showed more than 3-fold higher antibacterial activity compared to the particles obtained under stirring, due to their higher concentration and smaller size.Postprint (author's final draft

Nanotransformation of vancomycin overcomes the intrinsic resistance of Gram-negative bacteria

The increased emergence of antibiotic-resistant bacteria is a growing public health concern, and although new drugs are constantly being sought, the pace of development is slow compared with the evolution and spread of multidrug- resistant species. In this study, we developed a novel broad-spectrum antimicrobial agent by simply transforming vancomycin into nanoform using sonochemistry. Vancomycin is a glycopeptide antibiotic largely used for the treatment of infections caused by Gram-positive bacteria but inefficient against Gram-negative species. The nanospherization extended its effect toward Gram-negative Escherichia coli and Pseudomonas aeruginosa, making these bacteria up to 10 and 100 times more sensitive to the antibiotic, respectively. The spheres were able to disrupt the outer membranes of these bacteria, overcoming their intrinsic resistance toward glycopeptides. The penetration of nanospheres into a Langmuir monolayer of bacterial membrane phospholipids confirmed the interaction of the nanoantibiotic with the membrane of E. coli cells, affecting their physical integrity, as further visualized by scanning electron microscopy. Such mechanism of antibacterial action is unlikely to induce mutations in the evolutionary conserved bacterial membrane, therefore reducing the possibility of acquiring resistance. Our results indicated that the nanotransformation of vancomycin could overcome the inherent resistance of Gram-negative bacteria toward this antibiotic and disrupt mature biofilms at antibacterial-effective concentrations.Peer ReviewedPostprint (author's final draft

Functional biopolymer-based matrices for modulation of chronic wound enzyme activities

Collagen, collagen/hyaluronic acid (HA) and collagen/HA/chitosan (CS) sponges loaded with epigallocatechin gallate (EGCG), catechin (CAT) and gallic acid (GA) were developed and evaluated as active chronic wound dressings. Their physico-mechanical properties, biostability, biocompatibility and ability to inhibit in vitro myeloperoxidase (MPO) and collagenase—major enzymes related with the persistent inflammation in chronic wounds—were investigated as a function of the biopolymer composition and the polyphenolic compound used. The results demonstrated that the molecular weight of HA influences significantly the bulk properties of the obtained materials: higher elastic modulus, swelling ability and biostability against collagenase were measured when HA with higher molecular weights (830 and 2000 kDa) were added to the collagen matrices. The addition of CS and the polyphenols increased further the biostability of the sponges. Preliminary in vitro tests with fibroblasts revealed that the cells were able to adhere to all sponges. Cell viability was not affected significantly by the addition of the polyphenols; however, the presence of CS or high molecular weight HA in the sponge composition was associated with lower cellular viability. Finally, all specimens containing polyphenols efficiently inhibited the MPO activity. The highest inhibition capacity was observed for EGCG (IC50 = 15 ± 1 lM) and it was coupled to the highest extent of binding to the biopolymers (>80%) and optimal release profile from the sponges that allowed for prolonged (up to 3–5 days) effects.Spanish Ministerio de Ciencia e Innovació

GAGs-thiolated chitosan assemblies for chronic wounds treatment: control of enzyme activity and cell attachment

Multilayered polyelectrolyte coatings comprising thiolated chitosan (TC) and glycosaminoglycans (GAGs), chondroitin sulphate and hyaluronic acid, were built using a layer-by-layer approach. The surface activity of these coatings for binding and inhibition of enzymes related to chronic inflammation, such as collagenase and myeloperoxidase, was assessed. The build-up of five bi-layers of TC/GAGs onto gold surfaces was monitored in situ by QCM-D. All experimental groups showed exponential growth of the coatings controlled by the degree of chitosan thiolation and the molecular weight of the GAGs. The degree of chitosan modification was also the key parameter influencing the enzyme activity: increasing the thiols content led to more efficient myeloperoxidase inhibition and was inversely proportional to the adsorption of collagenase. Enhanced fibroblast attachment and proliferation were observed when the multilayered polyelectrolyte constructs terminated with GAGs. The possibility to control either the activity of major wound enzymes by the thiolation degree of the coating or the cell adhesion and proliferation by proper selection of the ultimate layer makes these materials potentially useful in chronic wounds treatment and dermal tissue regeneration.EU projects Lidwine (contract no. FP6-026741)Ministerio de Ciencia e Innovaci on de España - (MICINN) - bolsa BES-2008-0037

Thiolated chitosan/glycosaminoglycans multilayered films : QCM-D study on the films formation and their biological properties

Layer-by-layer technique is widely used to produce polyelectrolyte multilayered films for material surface functionalization. The technique simplicity coupled with the biological potential of biopolymers, e.g. polysaccharides, make such assemblies a suitable choice for many biomedical applications. In this study the formation of the films comprising of thiolated chitosan and glycosaminoglycans (GAGs) by alternate deposition was assessed in situ using QCM-D. Thiolated chitosan was used under the hypothesis of disulfide formation between its molecules to increase stability and/or stiffness of the films. The effects of the chitosan modification degree and GAGs molecular weight on the film thickness were investigated. All experimental groups showed exponential film growth, while the thickness increased with the chitosan thiolation degree and molecular weight of GAGs. Cellular behavior on the assemblies was found to be tunable by the appropriate selection of the terminate layer. Antimicrobial activity and protein adsorption on the new constructs are also commented