Three-dimensional layer-by-layer strategies for tissue engineering and nanomedicine

Layer-by-layer (LbL) is a self-assembly-driven surface modification strategy that allows the construction of nanostructured films onto substrates of any geometry, from simple bidimensional surfaces to more complex three-dimensional porous scaffolds. The principle behind LbL lies in the existence of multiple intermolecular interactions, such as electrostatic contacts, hydrophobic interactions, and hydrogen bonding, where the cooperative effects of multipoint attractions play the most important role. It is a technique that offers ease of preparation, versatility, fine control over the materials structure and robustness under physiological conditions. Although LbL has been mostly limited to the modification of planar surfaces, its potential lies in the capability to be extrapolated to 3D structures and coat increasingly complex geometries. Currently trending is the use of spherical templates – sacrificial or non-sacrificial – for applications in Nanomedicine, such as the construction of drug carriers or for the encapsulation of cells. The nanostructured nature of multilayered coatings makes it possible to build containers which permeability to molecules may be tuned simply by varying the number of involving layers or the class of materials involved. This way, in drug delivery it would be possible to construct structures in which the permeability of a drug to the exterior could be adjusted to a specific application or therapy, such as non-systemic approaches to cancer. In cell encapsulation, multilayer films could be employed to grant immune protection to the encapsulated biological materials, such as pancreatic islet cells, and enhanced control of both transport properties and surface physicochemical characteristics. Therefore, LbL presents an ambitious step in the development of effective encapsulating barriers for both active agents and cells

Nanostructured multilayers in the production of new devices for biomedical applications

Surface engineering is of the utmost significance in the conception of devices with an improved biological performance. By addressing physical and chemical features of interfaces, it has been possible to develop patterned and stimuli-responsive devices with tunable wettability and protein/cell adhesion properties with application in biomedicine and tissue engineering. While several surface engineering approaches exist, there is an increasing emphasis to non-harmful and versatile techniques to modify polymeric substrates: the sequential adsorption of proteins and polysaccharides, known as layer-by-layer (LbL) adsorption, is one of the most promising today. It is a simple and versatile technique where the cooperative effects of multipoint attractions allowing to produce robust coatings, even in substrates with complex geometries. Because it discards the need of organic and harmful solvents, it is an attractive technique for tissue engineering applications. Multilayer systems have already been proposed for different biomedical applications, including for biomimetic composite-like coatings, surfaces with smart properties, and to manipulate the adhesion and proliferation of cells

Multilayered films produced by layer-by-layer assembly of chitosan and alginate as a potential platform for the formation of human adipose-derived stem cell aggregates

The construction of multilayered films with tunable properties could offer new routes to produce biomaterials as a platform for 3D cell cultivation. In this study, multilayered films produced with five bilayers of chitosan and alginate (CHT/ALG) were built using water-soluble modified mesyl and tosylâ CHT via layer-by-layer (LbL) self-assembly. NMR results demonstrated the presences of mesyl (2.83 ppm) and tosyl groups (2.39, 7.37 and 7.70 ppm) in the chemical structure of modified chitosans. The buildup of multilayered films was monitored by quartz-crystal-microbalance (QCM-D) and film thickness was estimated using the Voigt-based viscoelastic model. QCM-D results demonstrated that CHT/ALG films constructed using mesyl or tosyl modifications (mCHT/ALG) were significantly thinner in comparison to the CHT/ALG films constructed with unmodified chitosan (p < 0.05). Adhesion analysis demonstrated that human adipose stem cells (hASCs) did not adhere to the mCHT/ALG multilayered films and formed aggregates with sizes between ca. 100â 200 µm. In vitro studies on cell metabolic activity and live/dead staining suggested that mCHT/ALG multilayered films are nontoxic toward hACSs. Multilayered films produced via LbL assembly of ALG and off-the-shelf, water-soluble modified chitosans could be used as a scaffold for the 3D aggregates formation of hASCs in vitro.This paper was partially financed by the ELASTISLET H2020-NMP-2014-646075 project, Fundo Social Europeu (FSE) and Programa Operacional de Potencial Humano (POPH). Javad Hatami, Rui R. Costa and Mariana B. Oliveira acknowledge the Fundação para a Ciência e Tecnologia (FCT) for grants SFRH/BPD/117202/2016, SFRH/BPD/95446/2013 and SFRH/BPD/111354/2015, respectively. Sandra G. Silva acknowledges the grant from ComplexiTE project.info:eu-repo/semantics/publishedVersio

Tuberculosis Presenting as Acute Sepsis and Secondary Hemophagocytic Lymphohistiocytosis

We report the case of a 61-year-old man admitted to our emergency department with fever. At admission, he was hypotensive and tachycardic. In the initial investigation, elevation of inflammatory parameters, acute kidney injury (Kidney Disease Improving Global Outcomes (KDIGO) 3), hyperbilirubinemia, and hepatic cytocholestasis were evident. Empirical antibiotic therapy was started, after sepsis was assumed without an identifiable cause. His condition took an unfavorable clinical course, with respiratory failure, hepatosplenomegaly, pancytopenia, hyperferritinemia and hypofibrinogenemia. Microbial culture studies and a general immunological study were negative and lymphoproliferative disease was therefore excluded. Bone marrow aspirate revealed hemophagocytosis without granulomas. A diagnosis of hemophagocytic lymphohistiocytosis was assumed and pulse methylprednisolone therapy initiated. As this resulted in only a transient improvement, immunoglobulin and rituximab were initiated as a second-line therapy. The patient sadly had an unfavorable outcome despite all measures undertaken. In the postmortem study, Mycobacterium tuberculosis complex was isolated in the bone marrow aspirate, which led to the postmortem diagnosis of disseminated tuberculosis and angioinvasive pulmonary aspergillosis. The clinical presentation of disseminated tuberculosis is non-specific and hemophagocytic lymphohistiocytosis is one of its rare presentations. The mortality rate of hemophagocytic lymphohistiocytosis is high and increases with delayed diagnosis of the underlying condition and respective treatment

Nanostructured thin coatings from chitosan and an elastin-like recombinamer with acute stimuli-responsive behavior

Publicado em: Materials Science Forum, vols. 730-732; Online available since 2012/Nov/12In the present work, chitosan (CHI) and elastin-like recombinamers (ELRs) were used to conceive nanostructured thin films driven by sequential electrostatic layer-by-layer (LbL), a simple and versatile technique that discards the use of harmful reagents. Two similar ELRs were engineered to contain negatively charged aminoacids and organized and a single monoblock or a triblock. The buildup of the films was monitored in real time using a quartz-crystal microbalance with dissipation monitoring (QCM-D). Wettability transitions were observed from a moderate hydrophobic surface to an extremely wettable upon increasing the temperature to 50 ºC, accompanied by topography changes at the nanoscale as assessed by atomic force microscopy (AFM). Furthermore, the dependence on time for the surface molecular rearrangement was studied for the films with each ELR. The potential of this technology may stimulate the development of devices and biomaterials for biomedical applications in the near future, such as surfaces with tunable and patterned cell adhesion, while the use of ELRs will allow developing polypeptides with biological significance.The authors acknowledge the financial support through Fundação para a Ciência e Tecnologia (FCT, PhD grant SFRH/BD/61126/2009, ‘‘Junta de Castilla y Leon’’ (VA034A09), the MICINN (MAT 2009-14195-C03-03, ACI2009-0890, IT2009-0089, MAT2010-15310, and MAT2010- 15982), the CIBER-BBN and the ‘‘Network Center of Regenerative Medicine and Cellular Therapy of Castilla y León’’.MICINNFundação para a Ciência e a Tecnologia (FCT)Junta de Castilla y LeónCIBER-BBNNetwork Center of Regenerative Medicine and Cellular Therapy of Castilla y Leó

Total and acute uterine inversion after delivery: a case report

INTRODUCTION: Uterine inversion is a rare obstetric emergency that can lead to hypovolemic shock or even maternal death. There are many management strategies, but they are poorly described and dispersed in the medical literature. The purpose of this article is to describe a case of complete acute uterine inversion and a review of the literature. CASE PRESENTATION: The authors describe a case of complete uterine inversion after a normal delivery with fundal placenta and without cord traction, in a 33-year-old Caucasian woman. After the diagnosis was made and after several attempts of manual correction of the inversion, the patient was taken immediately to the operating room and a laparotomy was performed. With opposing pressures in the cervical ring through the abdominal cavity and on the uterus fundus through her vagina, the inversion was resolved. An incision on the cervical ring was unnecessary. Due to incomplete detachment of the placenta the bleeding was mild. She recovered without complications and the histological examination of placenta was unremarkable. In this case, the only risk factor for uterine inversion was the fundal implantation of the placenta. CONCLUSIONS: The low incidence of uterine inversion leads to sparse experience in resolving this obstetrical emergency. The best prognosis occurs in situations where the diagnosis and maneuvers for uterine reversal are made at an early stage. The authors concluded that opposing pressures in the cervical ring through the abdominal cavity and on the uterus fundus through the vagina can resolve the inversion without the need of other surgical techniques. It is essential to keep in mind this diagnosis, and be updated about the strategies required to solve this complication

Layer-by-layer assembly of chitosan and recombinant biopolymers into biomimetic coatings with multiple stimuli-responsive properties

In this work, biomimetic smart thin coatings using chitosan and a recombinant elastin-like recombinamer (ELR) containing the cell attachment sequence arginine–glycine–(aspartic acid) (RGD) are fabricated through a layer-by-layer approach. The synthetic polymer is characterized for its molecular mass and composition using mass spectroscopy and peptide sequencing. The adsorption of each polymeric layer is followed in situ at room temperature and pH 5.5 using a quartz-crystal microbalance with dissipation monitoring, showing that both polymers can be successfully combined to conceive nanostructured, multilayered coatings. The smart properties of the coatings are tested for their wettability by contact angle (CA) measurements as a function of external stimuli, namely temperature, pH, and ionic strength. Wettability transitions are observed from a moderate hydrophobic surface (CAs approximately from 62° to 71°) to an extremely wettable one (CA considered as 0°) as the temperature, pH, and ionic strength are raised above 50 °C, 11, and 1.25 m, respectively. Atomic force microscopy is performed at pH 7.4 and pH 11 to assess the coating topography. In the latter, the results reveal the formation of large and compact structures upon the aggregation of ELRs at the surface, which increase water affinity. Cell adhesion tests are conducted using a SaOs-2 cell line. Enhanced cell adhesion is observed in the coatings, as compared to a coating with a chitosan-ending film and a scrambled arginine–(aspartic acid)–glycine (RDG) biopolymer. The results suggest that such films could be used in the future as smart biomimetic coatings of biomaterials for different biomedical applications, including those in tissue engineering or in controlled delivery systems.EUJCyL - VA034A09, VA030A08Fundação para a Ciência e Tecnologia (FCT) - SFRH/BD/61126/2009, SFRH/BD/61390/2009MICINN - MAT 2007-66275-C02-01, MAT 2007-61604, MAT 2009-14195-C03-03, PSE-300100-2006-1European regional development fund (ERDF)Junta de Castilla y LeonNetwork Center of Regenerative Medicine and Cellular Therapy of Castilla and LeónCIBER-BBN (project CB06-01-0003

Nanostructured multilayer compartments : towards multifunctionality and ‘‘cell-like’’ hierarchical complexity

In living organisms, there are phenomena that require the presence of specific biomolecules with distinct function and in variable concentrations at a given time, such as the healing and regeneration of tissue and organ lesions. In this work, we propose the use of a compartmented drug delivery device for the multiple release of bioactive agents. It consists of nanostructured microcapsules confined within a millimetric container that can be easily handled, mimicking the concept of cells which possess organelles with specialized functions. Each hierarchical structure was conceived using the layer-by-layer (LbL) method to form micro and macrocapsules that could individually carry either molecules and release them with distinct kinetics or magnetic nanoparticles (MNPs) to be used in targeted therapies. Furthermore, the internal microcontainers were constructed with a temperature-responsive elastin-like recombinamer (ELR) to further add smart properties to the proposed system. Sacrificial CaCO3 microparticles empty or entrapping either rhodamine or Fe3O4 MNPs were incubated in chitosan and ELR solutions using LbL for the conception of the microcapsules. Then, the microcapsules were suspended in alginate which was ionically crosslinked in CaCl2 drop-wise. Rhodamine could be encapsulated at this point in the alginate. The bead was coated with chitosan and alginate to build the external macrocapsule compartment. All structures were coated with 3 bilayers. The CaCO3 cores were chelated and the alginate beads liquefied using EDTA. Fluorescence microscopy using FITC and rhodamine markers showed a uniform distribution of the microcapsules within the macroreservoir. The release of rhodamine from either in the micro or macrocapsule was assessed at 25 and 37 °C in PBS. While the release from the macrocapsule follows a profile similar to that of traditional drug delivery systems, it is more sustained and delayed when released from the internal compartments. Such retention is more pronounced at 37 ºC (65% of release in comparison to 90%). This is due to the temperature responsive behavior of ELRs, which undergo a phase transition and make the LbL shell less permeable. For the magnetic response, the incorporation of the MNPs was observed by transmitted light microscopy. The attraction of the devices was observed by applying an external magnetic field along a defined trajectory. The results let foresee the use of such multilayer devices as compartmented structures to encapsulate growth factors, MNPs and stem cells for their controlled differentiation and maintenance or for guided regeneration of tissues and organs.Fundo Social Europeu (FSE)Fundação para a Ciência e a Tecnologia (FCT)Programa Diferencial de Potencial Humano (POPH