Adsorption of Fibrinogen on Silica Surfaces-The Effect of Attached Nanoparticles

When a biomaterial is inserted into the body, proteins rapidly adsorb onto its surface, creating a conditioning protein film that functions as a link between the implant and adhering cells. Depending on the nano-roughness of the surface, proteins will adsorb in different amounts, with different conformations and orientations, possibly affecting the subsequent attachment of cells to the surface. Thus, modifications of the surface nanotopography of an implant may prevent biomaterial-associated infections. Fibrinogen is of particular importance since it contains adhesion epitopes that are recognized by both eukaryotic and prokaryotic cells, and can therefore influence the adhesion of bacteria. The aim of this study was to model adsorption of fibrinogen to smooth or nanostructured silica surfaces in an attempt to further understand how surface nanotopography may affect the orientation of the adsorbed fibrinogen molecule. We used a coarse-grained model, where the main body of fibrinogen (visible in the crystal structure) was modeled as rigid and the flexible α C-chains (not visible in the crystal structure) were modeled as completely disordered. We found that the elongated fibrinogen molecule preferably adsorbs in such a way that it protrudes further into solution on a nanostructured surface compared to a flat one. This implicates that the orientation on the flat surface increases its bio-availability

The Forkhead Transcription Factor Foxi1 Is a Master Regulator of Vacuolar H+-ATPase Proton Pump Subunits in the Inner Ear, Kidney and Epididymis

The vacuolar H+-ATPase dependent transport of protons across cytoplasmic membranes in FORE (forkhead related) cells of endolymphatic epithelium in the inner ear, intercalated cells of collecting ducts in the kidney and in narrow and clear cells of epididymis require expression of several subunits that assemble into a functional multimeric proton pump. We demonstrate that expression of four such subunits A1, B1, E2 and a4 all co-localize with the forkhead transcription factor Foxi1 in a subset of epithelial cells at these three locations. In cells, of such epithelia, that lack Foxi1 we fail to identify any expression of A1, B1, E2 and a4 demonstrating an important role for the transcription factor Foxi1 in regulating subunit availability. Promoter reporter experiments, electrophoretic mobility shift assays (EMSA) and site directed mutagenesis demonstrate that a Foxi1 expression vector can trans-activate an a4-promoter reporter construct in a dose dependent manner. Furthermore, we demonstrate using chromatin immunoprecipitation (ChIP) assays that Foxi1-dependent activation to a large extent depends on cis-elements at position −561/−547 in the a4 promoter. Thus, we provide evidence that Foxi1 is necessary for expression of at least four subunits in three different epithelia and most likely is a major determinant for proper assembly of a functional vacuolar H+-ATPase complex at these locations

A Positive Regulatory Loop between foxi3a and foxi3b Is Essential for Specification and Differentiation of Zebrafish Epidermal Ionocytes

BACKGROUND: Epidermal ionocytes play essential roles in the transepithelial transportation of ions, water, and acid-base balance in fish embryos before their branchial counterparts are fully functional. However, the mechanism controlling epidermal ionocyte specification and differentiation remains unknown. METHODOLOGY/PRINCIPAL FINDINGS: In zebrafish, we demonstrated that Delta-Notch-mediated lateral inhibition plays a vital role in singling out epidermal ionocyte progenitors from epidermal stem cells. The entire epidermal ionocyte domain of genetic mutants and morphants, which failed to transmit the DeltaC-Notch1a/Notch3 signal from sending cells (epidermal ionocytes) to receiving cells (epidermal stem cells), differentiates into epidermal ionocytes. The low Notch activity in epidermal ionocyte progenitors is permissive for activating winged helix/forkhead box transcription factors of foxi3a and foxi3b. Through gain- and loss-of-function assays, we show that the foxi3a-foxi3b regulatory loop functions as a master regulator to mediate a dual role of specifying epidermal ionocyte progenitors as well as of subsequently promoting differentiation of Na(+),K(+)-ATPase-rich cells and H(+)-ATPase-rich cells in a concentration-dependent manner. CONCLUSIONS/SIGNIFICANCE: This study provides a framework to show the molecular mechanism controlling epidermal ionocyte specification and differentiation in a low vertebrate for the first time. We propose that the positive regulatory loop between foxi3a and foxi3b not only drives early ionocyte differentiation but also prevents the complete blockage of ionocyte differentiation when the master regulator of foxi3 function is unilaterally compromised

Low biocide emission antifouling based on a novel route of barnacle intoxication

Marine biofouling can be defined as the colonization of man-made surfaces in seawater by microscopic and macroscopic organisms. This phenomenon can result in great loss of function and effectiveness both for cruising ships and for static constructions. Of special concern are the negative effects of hard fouler such as barnacles, which cause increased drag resistance resulting in increases in fuel consumption, and disruption of the corrosion protective layer of marine vessels and constructions. Present biocide-based antifouling strategies are based on a continuous exposure of biocides at the film/water interface and consequently release into the environment if the antifouling efficacy is to be maintained. Such biocide-based solutions can therefore not be regarded as sustainable. The aim of this thesis is to describe the possibility to design biocide antifouling coatings based on a new strategy. Instead of releasing the bioactive molecule to the bulk water the biocide will be “entrapped” in the paint matrix and only after stimuli by organism interaction with the paint surface intoxication will take place. It was shown (Paper I) that using an experimental formulation, containing ivermectin, both in static panels and on boats, long lasting protection against barnacles was obtained. Moreover, using two model surfaces (Paper II), it was possible to separate and study the different contributions to the antifouling efficacy, finding that the low leaching of ivermectin had no contribution at all while surface’s modulus of the coating was the key factor. This supports the validity of the contact active antifouling hypothesis, rather than emission based. In (Paper III) we could follow the fate of barnacle growing on ivermectin containing coatings, and both field and laboratory tests could demonstrate that the intoxication of barnacles start when the juvenile organism reach ca. 0.6-0.7mm in diameter. Electronic microscopy images on the panels after the test, demonstrate that on control paint (no biocide) the juvenile barnacles (0.6-0.7mm diameter) already leaves imprint or penetration marks on the rosin based coatings. The distribution of ivermectin in the dry film seemed to be related with enhancement of barnacles contact intoxication. This was studied by fluorescence microscopy in (Paper I) and by the use ToF-SIMS in (Paper IV). This particular analytic method gives the possibility to follow organic biocides in paint film without the need of labelling or modify the biocide molecule in any extent. The entrapped antifouling strategy opens up the possibility to achieve long term antifouling (>10 years) as there is no need to use erosive binders. Moreover, this system might also find it uses in marine constructions and other fields where maintenance is difficult

Attenuation of acute inlammatory responses by surface nanotopography

The interaction between biology and non-viable surfaces is crucial for many organisms and cells. For example, bacterial cells need to adhere to mineral surfaces in the soil, plants climb and adhere to walls and marine organisms produce adhesives to cling to underwater rocks etc. In the human body, tissue needs to firmly adhere to the mineral surface of bone, but also to foreign materials when for example a biomaterial is implanted. The knowledge of how biology interacts with surfaces is hence important and interesting in many aspects. Within seconds after implantation of a biomaterial, proteins from the immune complement and coagulation systems adsorbs to the surface with possible adverse consequences for the patient. To overcome this, chemical surface modifications are readily employed. However, recently the significance of surface nanotopography for the adsorption of proteins, and attachment of cells have been acknowledged. To facilitate research on the interactions between biology and nanostructured substrates novel experimental surfaces with defined nanotopography and surface chemistry were developed. The surfaces are fabricated by binding gold nanoparticles to a gold surface, using a non-lithographic method and standard laboratory equipment. The surface chemistry was evaluated using XPS and ToF-SIMS. On these surfaces, the effect of surface nanotopography on the activation of the immune complement and activation of blood platelets was studied using QCM-D, SEM and fluorescence microscopy. It was found that although nanostructured surfaces adsorbed greater amount of serum proteins, activation of the immune complement was attenuated by surface nanotopography. A suggested mechanism is that the curvature of the nanoparticles prevents interaction between complement proteins. It was also found that blood platelets were activated to a lower degree on nanostructured surfaces and were sensitive to changes in nanoparticle size and inter-particle distance. These nanostructures surfaces can hopefully facilitate research on protein/cell interactions on nanostructured surfaces

SLC26A4 Targeted to the Endolymphatic Sac Rescues Hearing and Balance in Slc26a4 Mutant Mice

Mutations of SLC26A4 are a common cause of human hearing loss associated with enlargement of the vestibular aqueduct. SLC26A4 encodes pendrin, an anion exchanger expressed in a variety of epithelial cells in the cochlea, the vestibular labyrinth and the endolymphatic sac. Slc26a4[superscript Δ/Δ] mice are devoid of pendrin and develop a severe enlargement of the membranous labyrinth, fail to acquire hearing and balance, and thereby provide a model for the human phenotype. Here, we generated a transgenic mouse line that expresses human SLC26A4 controlled by the promoter of ATP6V1B1. Crossing this transgene into the Slc26a4[superscript Δ/Δ] line restored protein expression of pendrin in the endolymphatic sac without inducing detectable expression in the cochlea or the vestibular sensory organs. The transgene prevented abnormal enlargement of the membranous labyrinth, restored a normal endocochlear potential, normal pH gradients between endolymph and perilymph in the cochlea, normal otoconia formation in the vestibular labyrinth and normal sensory functions of hearing and balance. Our study demonstrates that restoration of pendrin to the endolymphatic sac is sufficient to restore normal inner ear function. This finding in conjunction with our previous report that pendrin expression is required for embryonic development but not for the maintenance of hearing opens the prospect that a spatially and temporally limited therapy will restore normal hearing in human patients carrying a variety of mutations of SLC26A4

Immune complement activation is attenuated by surface nanotopography

Mats Hulander1, Anders Lundgren1, Mattias Berglin1, Mattias Ohrlander2, Jukka Lausmaa3,4, Hans Elwing1 1Department of Cell and Molecular Biology/Interface Biophysics, University of Gothenburg, Medicinaregatan 9E, Gothenburg, 2Bactiguard AB, Stockholm, 3SP Technical Research Institute, Boras, 4Biomatcell, The Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden Abstract: The immune complement (IC) is a cell-free protein cascade system, and the first part of the innate immune system to recognize foreign objects that enter the body. Elevated activation of the system from, for example, biomaterials or medical devices can result in both local and systemic adverse effects and eventually loss of function or rejection of the biomaterial. Here, the researchers have studied the effect of surface nanotopography on the activation of the IC system. By a simple nonlithographic process, gold nanoparticles with an average size of 58 nm were immobilized on a smooth gold substrate, creating surfaces where a nanostructure is introduced without changing the surface chemistry. The activation of the IC on smooth and nanostructured surfaces was viewed with fluorescence microscopy and quantified with quartz crystal microbalance with dissipation monitoring in human serum. Additionally, the ability of pre-adsorbed human immunoglobulin G (IgG) (a potent activator of the IC) to activate the IC after a change in surface hydrophobicity was studied. It was found that the activation of the IC was significantly attenuated on nanostructured surfaces with nearly a 50% reduction, even after pre-adsorption with IgG. An increase in surface hydrophobicity blunted this effect. The possible role of the curvature of the nanoparticles for the orientation of adsorbed IgG molecules, and how this can affect the subsequent activation of the IC, are discussed. The present findings are important for further understanding of how surface nanotopography affects complex protein adsorption, and for the future development of biomaterials and blood-contacting devices. Keywords: nanostructure, protein adsorption, gold nanoparticles, QCM-D, IgG, innate immunit

Inflammatory and lipemic response to red meat intake in women with and without Rheumatoid Arthritis: a single meal study within a randomized controlled trial

Abstract Background The risk of atherosclerotic cardiovascular disease (ASCVD) is increased in Rheumatoid Arthritis (RA). Previous research has suggested that lipid metabolism is altered in RA, but research under postprandial conditions is scarce. The aim of this study was to investigate whether women with RA have a different lipemic and inflammatory response to a mixed meal containing red meat compared to women without RA. Methods Twenty-two women with RA, with modest disease activity, and 22 women without RA matched for age and body mass index (BMI) at the group level consumed a hamburger meal containing ca. 700 kcal (53 E% from fat, 27 E% from carbohydrate). Venous blood was sampled in the fasted state and after 30 min, 1, 2, 3 and 5 h and analysed for lipid species using nuclear magnetic resonance spectroscopy. Postprandial inflammation was measured by interleukin- 6 (IL- 6). The postprandial lipid response was calculated as the incremental area under the curve minimal value, and serial measurements were analysed by repeated measures analysis of variance. Lipid and inflammatory responses were compared by linear regression analysis, adjusted for age, BMI, physical activity, and baseline plasma concentration. Results Plasma concentrations of IL- 6, triglycerides (TGs) and very low-density lipoprotein (VLDL) particles increased significantly after the meal compared to baseline within both groups, but no differences were observed between groups. However, the women with RA had a less pronounced response in cholesterol carried in VLDL particles (p = 0.03) and in TGs in the subfraction of VLDL particles with highest density (p = 0.03). No association was found between the response in TGs and IL- 6. Conclusion This study does not provide compelling evidence for any difference in the lipemic or inflammatory response in women with RA compared with age- and BMI-matched women without RA following ingestion of a mixed, high-fat meal containing red meat. The modest disease activity in women with RA should be considered when interpreting these findings. Subtle group differences found in the lipids carried by VLDL particles warrant further investigation. Trial registration The PIRA (Postprandial Inflammation in Rheumatoid Arthritis) trial was registered 2020–01 - 28 at Clinicaltrials.gov (NCT04247009)