Effect of ceramic calcium-phosphorus ratio on chondrocyte-mediated biosynthesis and mineralization.

The osteochondral interface functions as a structural barrier between cartilage and bone, maintaining tissue integrity postinjury and during homeostasis. Regeneration of this calcified cartilage region is thus essential for integrative cartilage healing, and hydrogel-ceramic composite scaffolds have been explored for calcified cartilage formation. The objective of this study is to test the hypothesis that Ca/P ratio of the ceramic phase of the composite scaffold regulates chondrocyte biosynthesis and mineralization potential. Specifically, the response of deep zone chondrocytes to two bioactive ceramics with different calcium-phosphorus ratios (1.35 ± 0.01 and 1.41 ± 0.02) was evaluated in agarose hydrogel scaffolds over two weeks in vitro. It was observed that the ceramic with higher calcium-phosphorus ratio enhanced chondrocyte proliferation, glycosaminoglycan production, and induced an early onset of alkaline phosphorus activity, while the ceramic with lower calcium-phosphorus ratio performed similarly to the ceramic-free control. These results underscore the importance of ceramic bioactivity in directing chondrocyte response, and demonstrate that Ca/P ratio is a key parameter to be considered in osteochondral scaffold design. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2694-2702, 2017.

An insight on human skin penetration of diflunisal: lipogel versus hydrogel microemulsion.

Diflunisal is a NSAID used in acute and long term management of pain and inflammation associated with osteoarthritis, rheumatoid arthritis and symptoms of primary dysmenorrhea. However, its oral use is associated with side effects such as peptic ulceration, dyspepsia, gastrointestinal disturbances and bleeding. The aim of this work was to develop lecithin organogels (LO) transdermal delivery system for diflunisal and to study its human skin penetration ability in comparison with an optimized microemulsion-based hydrogel. Ternary phase diagrams were constructed using butyl lactate as an organic solvent and two commercial grades of lecithin. The formation of gel phase was lecithin concentration dependent with Phosholipion 85 G being capable of forming organogels at lower lecithin concentration than Lipoid S75. The gels prepared using butyl lactate were able to tolerate higher amounts of water than could be incorporated in the lipogels prepared with other organic solvents. All the investigated gels possessed acceptable physical properties and were able to deliver diflunisal through human skin. The lipogels delivered higher total drug amount through the skin than the hydrogel. The composition of lecithin seemed to have some effect on the skin permeability enhancement ability of the lipogel. Lecithin containing higher amount of phosphatidyl ethanolamine could provide better transdermal delivery. The elaborated lecithin organogels are potential carriers that create a good opportunity for transdermal delivery of diflunisal overcoming the side effects associating its oral route.

Exploration of the biomacromolecular interactions of an interpenetrating proteo-saccharide hydrogel network at the mucosal interface.

The relationship between mucin (MUC) and pectin (PEC) was explored in an attempt to understand the biomacromolecular interactions that occur at mucosal surfaces when mucus membranes are exposed to PEC-based materials. These interactions were explored through techniques, such as attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy, SEM imagery of lyophilized MUC-PEC blends, thermodynamic analysis, rheology investigations, and in silico static lattice atomistic simulations using a molecular mechanics energy relationships (MMER) approach. Three types of PEC that had different degrees of esterification and degrees of amidation were investigated at different MUC-PEC mass ratios (1:0, 1:1, 1:4, 1:9, and 0:1). The effect PEG 400 and Ca(2 +) in the MUC-PEC interactions were also studied. ATR-FTIR spectroscopy revealed broadening and strengthening of FTIR peaks at 3363 cm(-1) and between 3000-3650 cm(-1) due to stretching vibrations of the -OH, -COOH groups on MUC and PEC as well as the -N-H group on MUC. This suggested significant intra- and inter-molecular H-bonding. Morphologically, MUC-rich scaffolds were porous, thin, and multidirectional compared with the smooth, rigid, and unidirectional PEC-rich scaffolds. The Flory-Huggins interaction parameter (χ12 ) for all MUC-PEC mass ratios was negative, thus confirming MUC-PEC miscibility and interactions. UV absorbance increased with increasing relative concentration of PEC in the aqueous MUC-PEC dispersions. Furthermore, rheology investigations demonstrated synergistic enhancement in viscosity (η) and dynamic moduli upon the addition of PEG 400 and Ca(2 +) . MMER analysis revealed several key MUC-PEC interactions that corroborated well with the experimental data. Notably, higher esterification and larger mass ratios of PEC yielded greater MUC-PEC interactions.

Revegetation of a uranium mine dump by using fertilizer treated sessile oaks.

The rehabilitation of contaminated sites and the establishment of suitable trees for revegetation purposes is often problematic due to the mostly suboptimal nutrient supply and the poor humus reservoir. For these reasons hydrogels (Stockosorb) and novel humus substitutes (NOVIHUM), serving as long lasting fertilizer (LLF), were recently tested successfully. At the beginning of this multiyear study, those LLFs were administered to the root zone of young sessile oaks (Quercus petraea (Mattuschka) Liebl.), growing in test trials on a uranium mine dump in Schlema (Germany). To quantify the effect of LLFs on plant vitality, chlorophyll a fluorescence measurements and JIP test analyses were used. The results revealed up to 49% higher average photosynthetic vitality (PI(ABS)) of the LLF treated plants compared to controls. Particularly in the first test year, the efficiency of photosynthetic electron transport was strongly increased. This stimulation of photosynthetic activity was supported by direct measurements showing up to 129% increased diameter growth of the treated plants after a four year experimental period. Furthermore an increase of the maximum water holding capacity of the dump soil was attained by using LLFs. Overall, the findings reported here represent a feasible, ecologically justifiable reforestation method with a low environmental hazard potential.

Differentiation of bone marrow stromal cells into osteoblasts in a self-assembling peptide hydrogel: in vitro and in vivo studies.

A prerequisite of tissue engineering approaches with regard to autograft is a suitable scaffold that can harbor cells and signals. Conventionally, such scaffolds have been prepared as 3D scaffolds prefabricated from synthetic or natural biomaterials. RAD16 has been introduced as a new biomaterial, where synthetic peptides self-assemble to form a hydrogel. In this study, RAD16 was examined in terms of osteogenic efficacy and feasibility of ectopic mineralization. Two hundred and seventy-one RAD16 was cocultured with 1 × 10(6) bone marrow cells from the femurs of 6-week-old Wistar male rats in alpha minimum essential medium supplemented with or without dexamethasone. Second, the same volume of the RAD16 construct hosting the cells with or without hydroxyapatite (HA) particles was treated in the dexamethasone medium as well, prepared in a Teflon tube, and implanted subcutaneously. Cell proliferation was prominent in the RAD16 coculture with dexamethasone at 1 week and significantly decreased by 2 weeks, whereas the other combinations remained or inclined, and their osteogenic differentiation was accelerated up to 2 weeks, as seen in increasing alkaline phosphatase (ALP) activity and mRNAs of ALP, OPN, and OCN. The RAD16 implant prepared with HA particles allowed more osteoblast-like cells and blood cells to grow inside, which was accompanied by elevating OPN gene expression and the stronger peak of VEGF gene expression at 2 weeks. Furthermore, more OPN mRNA signal was detected around the RAD16 containing HA particles by 4 weeks. On the other hand, the RAD16 alone represented lower expression of OPN gene. During the experiment, however, no ectopic mineralization was observed in both groups. Conclusively, it was suggested that the RAD16 showed feasibility of serving as a matrix for osteogenic differentiation of cocultured bone marrow cells in vitro and in vivo. Proceeding of exploration and modification of RAD16 are continuously required for cell-based tissue engineering.

A solid-phase Bcr-Abl kinase assay in 96-well hydrogel plates.

Regulated phosphorylation by protein tyrosine kinases (PTKs), such as c-Abl, is critical to cellular homeostasis. In turn, once deregulated as in the chronic myeloid leukemia (CML) fusion protein Bcr-Abl, PTKs can promote cancer onset and progression. The dramatic success of the Bcr-Abl inhibitor imatinib as therapy for CML has inspired interest in other PTKs as targets for cancer drug discovery. Here we report a novel PTK activity and inhibition screening method using hydrogel-immobilized peptide substrates. Using acrylate crosslinkers, we tether peptides via terminal cysteines to thiol-presenting hydrogels in 96-well plates. These surfaces display low background and high reproducibility, allowing semiquantitative detection of peptide phosphorylation by recombinant c-Abl or by Bcr-Abl activity in cell extracts using traditional anti-phosphotyrosine immunodetection and chemifluorescence. The capabilities of this assay are demonstrated by performing model screens for inhibition with several commercially available PTK inhibitors and a collection of pyridopyrimidine Src/Abl dual inhibitors. This assay provides a practical method to measure the activity of a single kinase present in a whole cell lysate with high sensitivity and specificity as a valuable means for efficient small molecule screening.

Adsorption of bile acid by chitosan salts prepared with cinnamic acid and analogue compounds.

A chitosan (CS) powder treated with cinnamic acid and an analogue compound (CN) was prepared as CS-CN. Using it, bile acid adsorption by CS-CN and the release of CN were investigated in vitro. When CS-CN was soaked in a taurocholate solution, it released CN and simultaneously adsorbed the bile acid. For CS-CN prepared with cinnamic acid, the amount of CN released was 0.286 +/- 0.001 mmol/g CS-CN; the amount of taurocholate adsorbed was 0.284 +/- 0.003 mmol/g CS-CN. These two functions were recognized on alginate or pectin gel beads containing CS-CN. The amount of released CN was altered extensively by the species of CN used for gel-bead preparation. Results suggest that CS-CN is a candidate for complementary medicine to prevent lifestyle-related diseases.

The regrowth of axons within tissue defects in the CNS is promoted by implanted hydrogel matrices that contain BDNF and CNTF producing fibroblasts.

In this study we demonstrate the potential for combining biocompatible polymers with genetically engineered cells to elicit axon regrowth across tissue defects in the injured CNS. Eighteen- to 21-day-old rats received implants of poly N-(2-hydroxypropyl)-methacrylamide (HPMA) hydrogels containing RGD peptide sequences that had been infiltrated with control (untransfected) fibroblasts (n = 8), fibroblasts engineered to express brain-derived neurotrophic factor (BDNF) (n = 5), ciliary neurotrophic factor (CNTF) (n = 5), or a mixture of BDNF and CNTF expressing fibroblasts (n = 11). Fibroblasts were prelabeled with Hoechst 33342. Cell/polymer constructs were inserted into cavities made in the left optic tract, between thalamus and superior colliculus. After 4-8 weeks, retinal projections were analyzed by injecting right eyes with cholera toxin (B-subunit). Rats were perfused 24 h later and sections were immunoreacted to visualize retinal axons, other axons (RT97 antibody), host astrocytes and macrophages, donor fibroblasts, and extracellular matrix molecules. The volume fraction (VF) of each gel that was occupied by RT97(+) axons was quantified. RT-PCR confirmed expression of the transgenes prior to, and 5 weeks after, transplantation. Compared to control rats (mean VF = 0.02 +/- 0.01% SEM) there was increased ingrowth of RT97(+) axons into implants in CNTF (mean VF = 0.33 +/- 0.19%) and BDNF (mean VF = 0.62 +/-0.19%) groups. Axon growth into hydrogels in the mixed BDNF/CNTF group (mean VF = 3.58 +/- 0.92%) was significantly greater (P < 0.05) than in the BDNF or CNTF fibroblast groups. Retinal axons exhibited a complex branching pattern within gels containing BDNF or BDNF/CNTF fibroblasts; however, they regrew the greatest distances within implants containing both BDNF and CNTF expressing cells.