Biomimetic poly(amidoamine) hydrogels as synthetic materials for cell culture

<p>Abstract</p> <p>Background</p> <p>Poly(amidoamine)s (PAAs) are synthetic polymers endowed with many biologically interesting properties, being highly biocompatible, non toxic and biodegradable. Hydrogels based on PAAs can be easily modified during the synthesis by the introduction of functional co-monomers. Aim of this work is the development and testing of novel amphoteric nanosized poly(amidoamine) hydrogel film incorporating 4-aminobutylguanidine (agmatine) moieties to create RGD-mimicking repeating units for promoting cell adhesion.</p> <p>Results</p> <p>A systematic comparative study of the response of an epithelial cell line was performed on hydrogels with agmatine and on non-functionalized amphoteric poly(amidoamine) hydrogels and tissue culture plastic substrates. The cell adhesion on the agmatine containing substrates was comparable to that on plastic substrates and significantly enhanced with respect to the non-functionalized controls. Interestingly, spreading and proliferation on the functionalized supports are slower than on plastic exhibiting the possibility of an easier control of the cell growth kinetics. In order to favor the handling of the samples, a procedure for the production of bi-layered constructs was also developed by means the deposition via spin coating of a thin layer of hydrogel on a pre-treated cover slip.</p> <p>Conclusion</p> <p>The obtained results reveal that PAAs hydrogels can be profitably functionalized and, in general, undergo physical and chemical modifications to meet specific requirements. In particular the incorporation of agmatine warrants good potential in the field of cell culturing and the development of supported functionalized hydrogels on cover glass are very promising substrates for applications in cell screening devices.</p

Conversion of nanoscale topographical information of cluster-assembled zirconia surfaces into mechanotransductive events promotes neuronal differentiation

Additional file 4: Table S1. Proteomic data for upregulated proteins. Proteins upregulated (compared to flat-Zr) or present only in cells grown on ns-Zr15. Adhesome proteins and proteins with roles in mechanobiological processes are marked in dark and light grey, respectively

An Easy Path for Correlative Electron and Super-Resolution Light Microscopy

A number of new Correlative Light and Electron Microscopy approaches have been developed over the past years, offering the opportunity to combine the specificity and bio-compatibility of light microscopy with the high resolution achieved in electron microscopy. More recently, these approaches have taken one step further and also super-resolution light microscopy was combined with transmission or scanning electron microscopy. This combination usually requires moving the specimen between different imaging systems, an expensive set-up and relatively complicated imaging workflows. Here we present a way to overcome these difficulties by exploiting a commercially available wide-field fluorescence microscope integrated in the specimen chamber of a Scanning Electron Microscope (SEM) to perform correlative LM/EM studies. Super-resolution light microscopy was achieved by using a recently developed algorithm - the Super-Resolution Radial Fluctuations (SRRF) - to improve the resolution of diffraction limited fluorescent images. With this combination of hardware/software it is possible to obtain correlative super-resolution light and scanning electron microscopy images in an easy and fast way. The imaging workflow is described and demonstrated on fluorescently labelled amyloid fibrils, fibrillar protein aggregates linked to the onset of multiple neurodegenerative diseases, revealing information about their polymorphism.ISSN:2045-232

Interfaces Determine the Fate of Seeded α‐Synuclein Aggregation

ISSN:2196-735

The influence of Mediterraneandiet in acne pathogenesis and the correlation with insulin-like growth factor-1 serum levels: implications and results

Acne is a chronic inflammatory disease of the pilosebaceous unit and its etiology is complex and multifactorial. The role of the diet in its pathogenesis is still debated. The purpose of this study was to assess the association between MD and IGF-1 in acne patients and, as secondary objective, the role of systemic treatment on IGF-1 serum levels, in accordance to the patients’ diet. This study included 35 patients aged 14-30 years affected by acne and treated in line with the EDF guidelines. Patients were divided into 2 groups based on a questionnaire score assessing the adherence to the Mediterranean diet: the Mediterranean Group (score ≥ 6) and the Western Group (score &lt; 5). IGF-1 serum levels were measured in all patients before and after treatment and then compared to healthy population. IGF-1 levels were higher in patients than in controls and in the Western group than in the Mediterranean group. We speculate that the Mediterranean diet can have a protective role in the pathogenesis of acne by acting on the systemic route of IGF-1

K+ channel cAMP activated in guinea pig gallbladder epithelial cells

In guinea pig gallbladder epithelial cells, an increase in intracellular cAMP levels elicits the rise of anion channel activity. We investigated by patchclamp techniques whether K1 channels were also activated. In a cell-attached configuration and in the presence of theophylline and forskoline or 8-Br-cAMP in the cellular incubation bath, an increase of the open probability (Po) values for Ca21-activated K1 channels with a single-channel conductance of about 160 pS, for inward current, was observed. The increase in Po of these channels was also seen in an inside-out configuration and in the presence of PKA, ATP, and cAMP, but not with cAMP alone; phosphorylation did not influence single-channel conductance. In the inside-out configuration, the opioid loperamide (1025 M) was able to reduce Po when it was present either in the microelectrode filling solution or on the cytoplasmic side. Detection in the epithelial cells by RT-PCR of the mRNA corresponding to the a subunit of largeconductance Ca21-activated K1 channels (BKCa) indicates that this gallbladder channel could belong to the BK family. Immunohistochemistry experiments con- firm that these cells express the BK asubunit, which is located on the apical membrane. Other K1 channels with lower conductance (40 pS) were not activated either by 8-Br-cAMP (cell-attached) or by PKA 1 ATP 1 cAMP (inside-out). These channels were insensitive to TEA1 and loperamide. The data demonstrate that under conditions that induce secretion, phosphorylation activates anion channels as well as Ca21- dependent, loperamide-sensitive K1 channels present on the apical membrane

Functional assessment of allelic variants in the SLC26A4 gene involved in Pendred syndrome and nonsyndromic EVA

Pendred syndrome is an autosomal recessive disorder characterized by sensorineural hearing loss, with malformations of the inner ear, ranging from enlarged vestibular aqueduct (EVA) to Mondini malformation, and deficient iodide organification in the thyroid gland. Nonsyndromic EVA (ns-EVA) is a separate type of sensorineural hearing loss showing normal thyroid function. Both Pendred syndrome and ns-EVA seem to be linked to the malfunction of pendrin (SLC26A4), a membrane transporter able to exchange anions between the cytosol and extracellular fluid. In the past, the pathogenicity of SLC26A4 missense mutations were assumed if the mutations fulfilled two criteria: low incidence of the mutation in the control population and substitution of evolutionary conserved amino acids. Here we show that these criteria are insufficient to make meaningful predictions about the effect of these SLC26A4 variants on the pendrin-induced ion transport. Furthermore, we functionally characterized 10 missense mutations within the SLC26A4 ORF, and consistently found that on the protein level, an addition or omission of a proline or a charged amino acid in the SLC26A4 sequence is detrimental to its function. These types of changes may be adequate for predicting SLC26A4 functionality in the absence of direct functional tests

Volume-regulated Cl- channels in human pleural mesothelioma cells

Anion channels in human mesothelial and mesothelioma cell lines were characterized by patch-clamp and biomolecular approaches. We found an outwardly rectifying anionic current which was inactivated at positive voltages and inhibited by extracellular adenosine 5P-triphosphate (ATP). Mesothelial and mesothelioma cells behaved differently concerning current inactivation properties. Inactivation is more pronounced and has a steeper onset in mesothelial cells. Different reversal potentials, in asymmetrical Cl- solutions, that could be attributed to a different selectivity of the channel, have been observed in the two cell lines. Mesothelioma cell single-channel analysis indicates that the number of the same active anion channel (3-4 pS)increased under hypoosmotic conditions. Immunocytochemistry experiments showed the presence of ICln protein in the cytosol and in the plasma membrane. Western blot analysis revealed an increase of ICln in the membrane under hypotonic conditions, an event possibly related to the activation of Cl- channels