135 research outputs found

    Design and synthesis of peptides that neutralize bacterial endotoxins as therapeutic agents for the treatment of sepsis

    Get PDF
    [spa] La sepsis es un síndrome muy complejo de definir, diagnosticar y tratar. La patología de la sepsis se deriva de una respuesta perjudicial del organismo a una infección. La exposición a endotoxinas bacterianas induce una respuesta sistémica inflamatoria que tiene como finalidad luchar contra la infección. Esta respuesta se puede describir con una serie de síntomas clínicos como la fiebre o un aumento en la frecuencia cardiaca y respiratoria, entre otros. Este proceso, necesario para la defensa del organismo, conduce a la destrucción de los patógenos. Pero si todo y activar estos mecanismos, la infección persiste así como la exposición del organismo a productos bacterianos nocivos, los síntomas clínicos descritos pueden agravarse derivando en una septicemia. Las septicemias en un gran número de casos progresan a estadios más graves, como las septicemias severas o el shock séptico, que acostumbran a comportar un fallo generalizado de órganos vitales y la muerte del paciente. Las septicemias son la primera causa de mortalidad en las unidades de cuidados intensivos, con una mortalidad que varia entre el 30 y el 70%. En los Estados Unidos, el número de pacientes con septicemia supera ya los 750.000 casos anuales, siendo la décima causa de mortalidad. Estas cifras son parecidas en Europa. El lipopolisacárido (LPS) es la molécula que origina la septicemia y todas sus variantes, y es responsable de un gran nombre de enfermedades infecciosas. Esta molécula es el componente mayoritario de la pared celular de les bacterias Gram-negativas. La estructura química del LPS se basa en dos dominios unidos covalentemente y claramente diferenciados. Una parte hidrofílica formada por unidades repetitivas de oligosacáridos, y una parte lipídica, conocida como el lípido A que confiere toxicidad a la molécula. La presente Tesis Doctoral explora el diseño, la síntesis y la evaluación biológica de péptidos con capacidad para neutralizar el LPS, a partir de proteínas de unión al LPS. También se examina el diseño de inhibidores de tipo peptídico, con actividad biológica y propiedades farmacocinéticas mejoradas. Finalmente se estudia una familia de peptidomiméticos, peptoides, para neutralizar el LPS. Estos compuestos resultaron ser incapaces para neutralizar esta endotoxina, pero demostraron ser citotóxicos frente diversas líneas tumorales

    Cilengitide: The First Anti-Angiogenic Small Molecule Drug Candidate. Design, Synthesis and Clinical Evaluation

    Get PDF
    Cilengitide, a cyclic RGD pentapeptide, is currently in clinical phase III for treatment of glioblastomas and in phase II for several other tumors. This drug is the first anti-angiogenic small molecule targeting the integrins αvβ3, αvβ5 and α5β1. It was developed by us in the early 90s by a novel procedure, the spatial screening. This strategy resulted in c(RGDfV), the first superactive αvβ3 inhibitor (100 to 1000 times increased activity over the linear reference peptides), which in addition exhibited high selectivity against the platelet receptor αIIbβ3. This cyclic peptide was later modified by N-methylation of one peptide bond to yield an even greater antagonistic activity in c(RGDf(NMe)V). This peptide was then dubbed Cilengitide and is currently developed as drug by the company Merck-Serono (Germany)

    Antimicrobial peptides: Powerful biorecognition elements to detect bacteria in biosensing technologies

    Get PDF
    Bacterial infections represent a serious threat in modern medicine. In particular, biofilm treatment in clinical settings is challenging, as biofilms are very resistant to conventional antibiotic therapy and may spread infecting other tissues. To address this problem, biosensing technologies are emerging as a powerful solution to detect and identify bacterial pathogens at the very early stages of the infection, thus allowing rapid and effective treatments before biofilms are formed. Biosensors typically consist of two main parts, a biorecognition moiety that interacts with the target (i.e., bacteria) and a platform that transduces such interaction into a measurable signal. This review will focus on the development of impedimetric biosensors using antimicrobial peptides (AMPs) as biorecognition elements. AMPs belong to the innate immune system of living organisms and are very effective in interacting with bacterial membranes. They offer unique advantages compared to other classical bioreceptor molecules such as enzymes or antibodies. Moreover, impedance-based sensors allow the development of label-free, rapid, sensitive, specific and cost-effective sensing platforms. In summary, AMPs and impedimetric transducers combine excellent properties to produce robust biosensors for the early detection of bacterial infectionsPeer ReviewedPostprint (published version

    A molecular toolkit for the functionalization of titanium-based biomaterials that selectively control integrin-mediated cell adhesion

    No full text
    We present a click chemistry- based molecular toolkit for the bio- functionalization of materials to selec- tively control integrin-mediated cell ad- hesion. To this end, a 5 b 1-selective RGD peptidomimetics were covalently immobilized on Ti-based materials, and the capacity to promote the selective binding of a 5 b 1 was evaluated using a solid-phase integrin binding assay. This functionalization strategy yielded surfa- ces with a nine-fold increased affinity for a 5 b 1, in comparison to control samples, and total selectivity against the binding of the closely related integ- rin a v b 3. Moreover, our methodology allowed the screening of several phos- phonic acid containing anchoring units to find the best spacer–anchor moiety required for establishing an efficient binding to titanium and to promote se- lective integrin binding. The integrin subtype specificity of these biofunc- tionalized surfaces was further exam- ined in vitro by inducing selective ad- hesion of genetically modified fibro- blasts, which express exclusively the a 5 b 1 integrin. The versatility of our molecular toolkit was proven by shift- ing the cellular specificity of the mate- rials from a 5 b 1- to a v b 3-expressing fi- broblasts by using an a v b 3-selective peptidomimetic as coating molecule. The results shown here represent the first functionalization of Ti-based mate- rials with a 5 b 1- or a v b 3-selective pep- tidomimetics that allow an unprece- dented control to discriminate between a 5 b 1- and a v b 3-mediated adhesions. The role of these two integrins in dif- ferent biological events is still a matter of debate and is frequently discussed in literature. Thus, such bioactive titanium surfaces will be of great relevance for the study of integrin-mediated cell ad- hesion and the development of new bi- omaterials targeting specific cell typesPeer ReviewedPostprint (published version

    Impedimetric antimicrobial peptide-based sensor for the early detection of periodontopathogenic bacteria

    Get PDF
    Peri-implantitis, an inflammation caused by biofilm formation, constitutes a major cause of implant failure in dentistry. Thus, the detection of bacteria at the early steps of biofilm growth represents a powerful strategy to prevent implant-related infections. In this regard, antimicrobial peptides (AMPs) can be used as effective biological recognition elements to selectively detect the presence of bacteria. Thus, the aim of the present study was to combine the use of miniaturized and integrated impedimetric transducers and AMPs to obtain biosensors with high sensitivity to monitor bacterial colonization. Streptococcus sanguinis, which is one of the most prevalent strains in the onset of periodontal diseases, was used as a model of oral bacteria. To this end, a potent AMP derived from human lactoferrin was synthesized and covalently immobilized on interdigitated electrode arrays (IDEA). X-ray photoelectron spectroscopy (XPS) and electrochemical impedance spectroscopy (EIS) were employed to optimize and characterize the method of immobilization. Noteworthy, the interaction of Streptococcus sanguinis with AMP-coated sensors provoked significant changes in the impedance spectra, which were univocally associated with the presence of bacteria, proving the feasibility of our method. In this regard, the developed biosensor permits to detect the presence of bacteria at concentrations starting from 101 colony forming units (CFU) mL-1 in KCl and from 102 CFU mL-1 in artificial saliva. Moreover, the system was devoid of cytotoxicity for human fibroblasts. These results indicate that the proposed approach can be effective in the detection of initial stages of biofilm formation, and may be useful in the early prevention and treatment of peri-implantitisPeer ReviewedPostprint (author's final draft

    Biofunctionalization strategies on tantalum-based materials for osseointegrative applications

    Get PDF
    The use of tantalum as biomaterial for orthopedic applications is gaining considerable attention in the clinical practice because it presents an excellent chemical stability, body fluid resistance, biocompatibility, and it is more osteoconductive than titanium or cobalt-chromium alloys. Nonetheless, metallic biomaterials are commonly bioinert and may not provide fast and long-lasting interactions with surrounding tissues. The use of short cell adhesive peptides derived from the extracellular matrix has shown to improve cell adhesion and accelerate the implant’s biointegration in vivo. However, this strategy has been rarely applied to tantalum materials. In this work, we have studied two immobilization strategies (physical adsorption and covalent binding via silanization) to functionalize tantalum surfaces with a cell adhesive RGD peptide. Surfaces were used untreated or activated with either HNO3 or UV/ozone treatments. The process of biofunctionalization was characterized by means of physicochemical and biological methods. Physisorption of the RGD peptide on control and HNO3-treated tantalum surfaces significantly enhanced the attachment and spreading of osteoblast-like cells; however, no effect on cell adhesion was observed in ozone-treated samples. This effect was attributed to the inefficient binding of the peptide on these highly hydrophilic surfaces, as evidenced by contact angle measurements and X-ray photoelectron spectroscopy. In contrast, activation of tantalum with UV/ozone proved to be the most efficient method to support silanization and subsequent peptide attachment, displaying the highest values of cell adhesion. This study demonstrates that both physical adsorption and silanization are feasible methods to immobilize peptides onto tantalum-based materials, providing them with superior bioactivity.Peer Reviewe

    Blocking methods to prevent non-specific adhesion of mesenchymal stem cells to titanium and evaluate the efficiency of surface functionalization: albumin vs poly(ethylene glycol) coating

    Get PDF
    Premio SIBB 2014Surface modification of biocompatible materials with biologically-active molecules is a well-known strategy to enhance the osteointegration of implantable devices. In order to evaluate the efficiency of these treatments, an in vitro study of cell behavior on the modified surface is usually carried out. A key point to evaluate the efficacy of this strategy is to avoid non specific protein adsorption by creating a non-fouling background. This blocking step ensures that the observed response of cells can be exclusively ascribed to the modification treatment applied. Several techniques are available to create this neutral background. Thus, the aim of this study is to compare two different blocking methods, namely adsorption of albumin from bovine serum (BSA) and grafting of small polyethylene glycol (PEG) chains to titanium, which is the gold standard metal for orthopedics and dentistry. To this end, titanium surfaces were coated with a recently synthesized cell adhesive peptide-based molecule and subjected to a blocking procedure with either BSA or PEG.  Non-functionalized titanium samples were also blocked and used as controls. The biological response of human mesenchymal stem cells was evaluated by measuring the number of attached cells and studying the degree of cell spreading on the substrate. Both aspects of cell behavior are not affected significantly by the blocking method: cells adhere and spread significantly more on the functionalized samples, regardless of the blocking method used. This confirms that the surface feature that defines cell response is the presence/absence of the biomolecule, and not the anti-fouling layer. These results, together with the reduction of variability of results observed in presence of a blocking layer, demonstrate the efficacy and necessity of blocking the surface. PEG grafting is demonstrated as effective as BSA coating in reducing non-specific interactions and not hindering the effect of the biomolecule. However, taken into account the numerous advantages of a synthetic and customizable polymer chain over a complex natural protein, PEG blocking stands out as a very good alternative to albumin adsorption.Peer ReviewedAward-winnin

    Laser-assisted surface modification of zirconia-based materials to guide osteoblast responses for dental applications

    Get PDF
    The project is aimed at investigating the effect of laser surface modification of dental-grade zirconia (3Y-TZP) to guide cellular behaviour. A nanosecond laser was employed to fabricate groove patterns on the surface of 3Y-TZP, yielding three different topographies according to the groove periodicity (30, 50 and 100µm). The resulting topography, microstructure, hardness and hydrothermal degradation resistance of the modified zirconia was assessed by means of advanced characterization techniques. A cellular study was conducted to evaluate the behaviour of human mesenchymal stem cells on the patterned samples in terms of adhesion and cell morphology. The results showed grooves of approximately 1.7µm heigh with pronounced pile-up due to the melting and re-solidification of the material. The phase analysis revealed that no phase transformation is induced during the laser modification; however, a non-homogeneous monoclinic phase distribution was observed on the patterned surfaces after hydrothermal degradation. Finally, all the patterned surfaces allowed cell attachment, increased cell area and favoured cell elongation and alignment in the direction of the laser patterns.Postprint (published version

    Polyethylene glycol pulsed electrodeposition for the development of antifouling coatings on titanium

    Get PDF
    Titanium dental implants are widely used for the replacement of damaged teeth. However, bacterial infections at the interface between soft tissues and the implant can impair the functionality of the device and lead to failure. In this work, the preparation of an antifouling coating of polyethylene glycol (PEG) on titanium by pulsed electrodeposition was investigated in order to reduce Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) adhesion while maintaining human fibroblast adhesion. Different pulsed conditions were prepared and characterized by contact angle, Focused Ion Beam (FIB), Fourier Transformed Infrared Spectroscopy in the Attenuated Total Reflectance mode (ATR-FTIR), and X-ray photoelectron spectroscopy (XPS). XPS tested fibronectin adsorption. S. aureus, E. coli and human fibroblast adhesion was tested in vitro in both mono and co-culture settings. Physicochemical characterization proved useful for confirming the presence of PEG and evaluating the efficiency of the coating methods. Fibronectin adsorption decreased for all of the conditions, but an adsorption of 20% when compared to titanium was maintained, which supported fibroblast adhesion on the surfaces. In contrast, S. aureus and E. coli attachment on coated surfaces decreased up to 90% vs. control titanium. Co-culture studies with the two bacterial strains and human fibroblasts showed the efficacy of the coatings to allow for eukaryotic cell adhesion, even in the presence of pre-adhered bacteria.Peer ReviewedPostprint (published version

    Reducció de l’adhesió i formació de biofilm bacterià sobre titani mitjançant immobilització d’un pèptid antibacterià

    Get PDF
    L’ús d’implants de titani per a traumatologia i odontologia és una pràctica molt estesa actualment. El seu èxit, però, es veu afectat pels casos de infeccions que pateixen. En el cas concret dels implants dentals, el 14% dels implants pateixen infeccions bacterianes i entre el 5 i el 8% presenten infecció bacteriana que desenvolupa una peri-implantitis i una conseqüent pèrdua de l’implant. Per aquest motiu, en els darrers anys s’ha estudiat el desenvolupament de superfícies antibacterianes. L'objectiu d'aquest estudi va ser desenvolupar un tractament de la superfície de titani per formar un recobriment antibacterià sobre titani basat en un pèptid antibacterià derivat de lactoferrina i comparar l'eficàcia dels mètodes emprats per immobilitzar el pèptid a la superfície del titani, i avaluar el seu efecte en l'activitat antibacteriana in vitro. S’han estudiat les característiques fisicoquímiques de la superfície que en resulta, la seva biocompatibilitat i capacitat per evitar l’adhesió de proteïnes i bacteris
    • …