Specific Interactions versus Counterion Condensation. 2. Theoretical Treatment within the Counterion Condensation Theory

Polyuronates such as pectate and alginate are very well-known examples of biological polyelectrolytes undergoing,upon addition of divalent cations, an interchain association that acts as the junction of an eventually formed stable hydrogel. In the present paper, a thermodynamic model based on the counterion condensation theory has been developed to account for this cation-induced chain pairing of negatively charged polyelectrolytes. The strong interactions between cross-linking ions and uronate moieties in the specific binding site have been described in terms of chemical bonding, with complete charge annihilation between the two species. The chain-pairing process is depicted as progressively increasing with the concentration of cross-linking counterions and is thermodynamically defined by the fraction of each species. On these bases, the total Gibbs energy of the system has been expressed as the sum of the contributions of the Gibbs energy of the (single) chain stretches and of the (associated) dimers, weighted by their respective fractions 1 - õ and õ. In addition, the model assumes that the condensed divalent counterions exhibit an affinity free-energy for the chain, GCaff,0, and the junction, GDaff,0, respectively. Moreover, a specific Gibbs energy of chemical bonding, Gbond,0, has been introduced as the driving force for the formation of dimers. The model provides the mathematical formalism for calculating the fraction, õ, of chain dimers formed and the amount of ions condensed and bound onto the polyelectrolyte when two different types of counterions (of equal or different valence) are present. The effect of the parameter Gbond,0 has been investigated and, in  icular, its difference from GC,Daff,0 was found to be crucial in determining the distribution of the ions into territorial condensation and chemical bonding, respectively. Finally, the effect of the variation of the molar ratio between cross-linking ions and uronic groups in the specific binding sites, ó0, was evaluated. In particular, a remarkable decrease in the amount of condensed counterions has been pointed out in the case of ó0 ) 1/3, with respect to the value of ó0 ) 1/4, characterizing the traditional “egg-box” structure, as a result of the drop of the charge density of the polyelectrolyte induced by complete charge annihilation.- õ and õ. In addition, the model assumes that the condensed divalent counterions exhibit an affinity free-energy for the chain, GCaff,0, and the junction, GDaff,0, respectively. Moreover, a specific Gibbs energy of chemical bonding, Gbond,0, has been introduced as the driving force for the formation of dimers. The model provides the mathematical formalism for calculating the fraction, õ, of chain dimers formed and the amount of ions condensed and bound onto the polyelectrolyte when two different types of counterions (of equal or different valence) are present. The effect of the parameter Gbond,0 has been investigated and, in  icular, its difference from GC,Daff,0 was found to be crucial in determining the distribution of the ions into territorial condensation and chemical bonding, respectively. Finally, the effect of the variation of the molar ratio between cross-linking ions and uronic groups in the specific binding sites, ó0, was evaluated. In particular, a remarkable decrease in the amount of condensed counterions has been pointed out in the case of ó0 ) 1/3, with respect to the value of ó0 ) 1/4, characterizing the traditional “egg-box” structure, as a result of the drop of the charge density of the polyelectrolyte induced by complete charge annihilation.Fil: Donati, Ivan. University of Trieste; ItaliaFil: Benegas, Julio Ciro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - San Luis. Instituto de Matemática Aplicada de San Luis "Prof. Ezio Marchi". Universidad Nacional de San Luis. Facultad de Ciencias Físico, Matemáticas y Naturales. Instituto de Matemática Aplicada de San Luis "Prof. Ezio Marchi"; ArgentinaFil: Cesàro, Attilio. University of Trieste; ItaliaFil: Paolettin, Sergio. University of Trieste; Itali

Insights into Mechanical Behavior and Biological Properties of Chia Seed Mucilage Hydrogels

In this contribution we report insights on the rheological properties of chia (Salvia hispanica) seed mucilage hydrogels. Specifically, we studied the influence of temperature and polymer concentration on the viscoelastic properties of resulting networks. Creep experiments performed in steady-state conditions allowed calculating Newtonian viscosities for chia hydrogels at different polymer concentrations, pointing at inter-chain interactions as the main responsible for the different behavior toward network slipping under constant stress. The combination of oscillatory frequency and stress sweep tests highlighted a moderate effect of temperature in influencing hydrogel mechanics. The latter results prompted us to investigate potential biological functions for this set of biomaterials. Lactate Dehydrogenase assay proved lack of cytotoxicity of chia suspensions toward Human Mesenchymal Stem Cells from adipose tissue here used as cell model. Differentiation experiments were finally undertaken to verify the influence of chia samples on osteo-induction triggered by chemical differentiation factors. Alkaline Phosphatase enzyme activity assay and Alizarin red staining demonstrated that chia mucilage did not alter in vitro stem cell differentiation. Collectively, this set of experiments revealed an almost inert role associated with chia suspensions, indicating a possible application of chia-based networks as scaffold models to study osteogenesis in vitro

Hyaluronan delivery by polymer demixing in polysaccharide-based hydrogels and membranes for biomedical applications

Alginate-based membranes containing hyaluronic acid (HA) were manufactured by freeze-drying calcium-reticulated hydrogels. The study of the distribution of the two macromolecules within the hydrogel enabled to highlight a polymer demixing mechanism that tends to segregate HA in the external parts of the constructs. Resistance and pliability of the membranes were tuned, while release and degradation studies enabled to quantify the diffusion of both polysaccharides in physiological solution and to measure the viable lifetime of the membranes. Biological studies in vitro proved that the liquid extracts from the HA-containing membranes stimulate wound healing and that fibroblasts are able to colonize the membranes. Overall, such novel alginate-HA membranes represent a promising solution for several medical needs, in particular for wound treatment, giving the possibility to provide an in situ administration of HA from a resorbable device

Controlled Quenching of Agarose Defines Hydrogels with Tunable Structural, Bulk Mechanical, Surface Nanomechanical, and Cell Response in 2D Cultures

: The scaffolding of agarose hydrogel networks depends critically on the rate of cooling (quenching) after heating. Efforts are made to understand the kinetics and evolution of biopolymer self-assembly upon cooling, but information is lacking on whether quenching might affect the final hydrogel structure and performance. Here, a material strategy for the fine modulation of quenching that involves temperature-curing steps of agarose is reported. Combining microscopy techniques, standard and advanced macro/nanomechanical tools, it is revealed that agarose accumulates on the surface when the curing temperature is set at 121 °C. The inhomogeneity can be mostly recovered when it is reduced to 42 °C. This has a drastic effect on the stiffness of the surface, but not on the viscoelasticity, roughness, and wettability. When hydrogels are strained at small/large deformations, the curing temperature has no effect on the viscoelastic response of the hydrogel bulk but does play a role in the onset of the non-linear region. Cells cultured on these hydrogels exhibit surface stiffness-sensing that affects cell adhesion, spreading, F-actin fiber tension, and assembly of vinculin-rich focal adhesions. Collectively, the results indicate that the temperature curing of agarose is an efficient strategy to produce networks with tunable mechanics and is suitable for mechanobiology studies

Antibacterial-Nanocomposite Bone Filler Based on Silver Nanoparticles and Polysaccharides

Injectable bone fillers represent an attractive strategy for the treatment of bone defects. These injectable materials should be biocompatible, capable of supporting cell growth and possibly able to exert antibacterial effects. In this work, nanocomposite microbeads based on alginate, chitlac, hydroxyapatite and silver nanoparticles were prepared and characterized. The dried microbeads displayed a rapid swelling in contact with simulated body fluid and maintained their integrity for more than 30\ua0days. The evaluation of silver leakage from the microbeads showed that the antibacterial metal is slowly released in saline solution, with less than 6% of silver released after 1\ua0week. Antibacterial tests proved that the microbeads displayed bactericidal effects toward S. aureus, P. aeruginosa and S. epidermidis and were also able to damage pre-formed bacterial biofilms. On the other hand, the microbeads did not exert any cytotoxic effect towards osteoblast-like cells. After characterization of the bioactive microbeads, a possible means to embed them in a fluid medium was explored in order to obtain an injectable paste. Upon suspension of the particles in alginate solution or alginate/hyaluronic acid mixtures, a homogenous and time-stable paste was obtained. Mechanical tests enabled to quantify the extrusion forces from surgical syringes, pointing out the proper injectability of the material. This novel antibacterial bone-filler appears as a promising material for the treatment of bone defects, in particular when possible infections could compromise the bone-healing process

Nucleation, reorganization and disassembly of an active network from lactose-modified chitosan mimicking biological matrices

Developing synthetic materials able to mimic micro- and macrorheological properties of natural networks opens up to novel applications and concepts in materials science. The present contribution describes an active network based on a semi-synthetic polymer, a lactitol-bearing chitosan derivative (Chitlac), and a transient inorganic cross-linker, boric acid. Due to the many and diverse anchoring points for boric acid on the flanking groups of Chitlac, the cross-links constantly break and reform in a highly dynamic fashion. The consequence is a network with unusual non-equilibrium and mechanical properties closely resembling the rheological behavior of natural three-dimensional arrangements and of cytoskeleton. Concepts like network nucleation, reorganization and disassembly are declined in terms of amount of the cross-linker, which acts as a putative motor for remodeling of the network upon application of energy. The out-of-equilibrium and non-linear behavior render the semi-synthetic system of great interest for tissue engineering and for developing in-vitro mimics of natural active matrices

Red and Blue Light Differently Influence Actinidia chinensis Performance and Its Interaction with Pseudomonas syringae pv. Actinidiae

Light composition modulates plant growth and defenses, thus influencing plant–pathogen interactions. We investigated the effects of different light-emitting diode (LED) red (R) (665 nm) and blue (B) (470 nm) light combinations on Actinidia chinensis performance by evaluating biometric parameters, chlorophyll a fluorescence, gas exchange and photosynthesis-related gene expression. Moreover, the influence of light on the infection by Pseudomonas syringae pv. actinidiae (Psa), the etiological agent of bacterial canker of kiwifruit, was investigated. Our study shows that 50%R–50%B (50R) and 25%R–75%B (25R) lead to the highest PSII efficiency and photosynthetic rate, but are the least effective in controlling the endophytic colonization of the host by Psa. Monochromatic red light severely reduced ΦPSII, ETR, Pn, TSS and photosynthesis-related genes expression, and both monochromatic lights lead to a reduction of DW and pigments content. Monochromatic blue light was the only treatment significantly reducing disease symptoms but did not reduce bacterial endophytic population. Our results suggest that monochromatic blue light reduces infection primarily by modulating Psa virulence more than host plant defenses

Possible Fruit Protein Effects on Primate Communities in Madagascar and the Neotropics

The ecological factors contributing to the evolution of tropical vertebrate communities are still poorly understood. Primate communities of the tropical Americas have fewer folivorous but more frugivorous genera than tropical regions of the Old World and especially many more frugivorous genera than Madagascar. Reasons for this phenomenon are largely unexplored. We developed the hypothesis that Neotropical fruits have higher protein concentrations than fruits from Madagascar and that the higher representation of frugivorous genera in the Neotropics is linked to high protein concentrations in fruits. Low fruit protein concentrations in Madagascar would restrict the evolution of frugivores in Malagasy communities.We reviewed the literature for nitrogen concentrations in fruits from the Neotropics and from Madagascar, and analyzed fruits from an additional six sites in the Neotropics and six sites in Madagascar. Fruits from the Neotropical sites contain significantly more nitrogen than fruits from the Madagascar sites. Nitrogen concentrations in New World fruits are above the concentrations to satisfy nitrogen requirements of primates, while they are at the lower end or below the concentrations to cover primate protein needs in Madagascar.Fruits at most sites in the Neotropics contain enough protein to satisfy the protein needs of primates. Thus, selection pressure to develop new adaptations for foods that are difficult to digest (such as leaves) may have been lower in the Neotropics than in Madagascar. The low nitrogen concentrations in fruits from Madagascar may contribute to the almost complete absence of frugivorous primate species on this island

Exploiting natural polysaccharides to enhance in vitro bio-constructs of primary neurons and progenitor cells

Current strategies in Central Nervous System (CNS) repair focus on the engineering of artificial scaffolds for guiding and promoting neuronal tissue regrowth. Ideally, one should combine such synthetic structures with stem cell therapies, encapsulating progenitor cells and instructing their differentiation and growth. We used developments in the design, synthesis, and characterization of polysaccharide-based bioactive polymeric materials for testing the ideal composite supporting neuronal network growth, synapse formation and stem cell differentiation into neurons and motor neurons. Moreover, we investigated the feasibility of combining these approaches with engineered mesenchymal stem cells able to release neurotrophic factors. We show here that composite bio-constructs made of Chitlac, a Chitosan derivative, favor hippocampal neuronal growth, synapse formation and the differentiation of progenitors into the proper neuronal lineage, that can be improved by local and continuous delivery of neurotrophins. Statement of Significance In our work, we characterized polysaccharide-based bioactive platforms as biocompatible materials for nerve tissue engineering. We show that Chitlac-thick substrates are able to promote neuronal growth, differentiation, maturation and formation of active synapses. These observations support this new material as a promising candidate for the development of complex bio-constructs promoting central nervous system regeneration. Our novel findings sustain the exploitation of polysaccharide-based scaffolds able to favour neuronal network reconstruction. Our study shows that Chitlac-thick may be an ideal candidate for the design of biomaterial scaffolds enriched with stem cell therapies as an innovative approach for central nervous system repair

Predicting lymphoma in Sjögren's syndrome and the pathogenetic role of parotid microenvironment through precise parotid swelling recording

Objective: Parotid swelling (PSW) is a major predictor of non-Hodgkin lymphoma (NHL) in primary Sjögren's syndrome (pSS). However, since detailed information on the time of onset and duration of PSW is scarce, this was investigated to verify whether it may lead to further improved prediction. NHL localisation was concomitantly studied to evaluate the role of the parotid gland microenvironment in pSS-related lymphomagenesis. Methods: A multicentre study was conducted among patients with pSS who developed B cell NHL during follow-up and matched controls that did not develop NHL. The study focused on the history of salivary gland and lachrymal gland swelling, evaluated in detail at different times and for different durations, and on the localisation of NHL at onset. Results: PSW was significantly more frequent among the cases: at the time of first referred pSS symptoms before diagnosis, at diagnosis, and from pSS diagnosis to NHL. The duration of PSW was evaluated starting from pSS diagnosis, and the NHL risk increased from PSW of 2-12 months to > 12 months. NHL was prevalently localised in the parotid glands of the cases. Conclusion: A more precise clinical recording of PSW can improve lymphoma prediction in pSS. PSW as a very early symptom is a predictor, and a longer duration of PSW is associated with a higher risk of NHL. Since lymphoma usually localises in the parotid glands, and not in the other salivary or lachrymal glands, the parotid microenvironment appears to be involved in the whole history of pSS and related lymphomagenesis