Assessing glucose and oxygen diffusion in hydrogels for the rational design of 3D stem cell scaffolds in regenerative medicine.

Hydrogels are attractive biomaterials for replicating cellular microenvironments, but attention needs to be given to hydrogels diffusion properties. A large body of literature shows the promise of hydrogels as 3D culture models, cell expansion systems, cell delivery vehicles, and tissue constructs. Surprisingly, literature seems to have overlooked the important effects of nutrient diffusion on the viability of hydrogel-encapsulated cells. In this paper, we present the methods and results of an investigation into glucose and oxygen diffusion into a silated-hydroxypropylmethylcellulose (Si-HPMC) hydrogel. Using both an implantable glucose sensor and implantable oxygen sensor, we continuously monitored core glucose concentration and oxygen concentration at the centre of hydrogels. We demonstrated that we could tune molecular transport in Si-HPMC hydrogel by changing the polymer concentration. Specifically, the oxygen diffusion coefficient was found to significantly decrease from 3.4 × 10-10 to 2.4 × 10-10  m2  s-1 as the polymer concentration increased from 1% to 4% (w/v). Moreover, it was revealed during in vitro culture of cellularized hydrogels that oxygen depletion occurred before glucose depletion, suggesting oxygen diffusion is the major limiting factor for cell survival. Insight was also gained into the mechanism of action by which oxygen and glucose diffuse. Indeed, a direct correlation was found between the average polymer crosslinking node size and glucose parameters, and this correlation was not observed for oxygen. Overall, these experiments provide useful insights for the analysis of nutrient transport and gas exchange in hydrogels and for the development of future cellular microenvironments based on Si-HPMC or similar polysaccharide hydrogels.

Enriching a cellulose hydrogel with a biologically active marine exopolysaccharide for cell-based cartilage engineering.

The development of biologically and mechanically competent hydrogels is a prerequisite in cartilage engineering. We recently demonstrated that a marine exopolysaccharide, GY785, stimulates the in vitro chondrogenesis of adipose stromal cells. In the present study, we thus hypothesized that enriching our silated hydroxypropyl methylcellulose hydrogel (Si-HPMC) with GY785 might offer new prospects in the development of scaffolds for cartilage regeneration. The interaction properties of GY785 with growth factors was tested by surface plasmon resonance (SPR). The biocompatibility of Si-HPMC/GY785 towards rabbit articular chondrocytes (RACs) and its ability to maintain and recover a chondrocytic phenotype were then evaluated in vitro by MTS assay, cell counting and qRT-PCR. Finally, we evaluated the potential of Si-HPMC/GY785 associated with RACs to form cartilaginous tissue in vivo by transplantation into the subcutis of nude mice for 3 weeks. Our SPR data indicated that GY785 was able to physically interact with BMP-2 and TGFß. Our analyses also showed that three-dimensionally (3D)-cultured RACs into Si-HPMC/GY785 strongly expressed type II collagen (COL2) and aggrecan transcripts when compared to Si-HPMC alone. In addition, RACs also produced large amounts of extracellular matrix (ECM) containing glycosaminoglycans (GAG) and COL2. When dedifferentiated RACs were replaced in 3D in Si-HPMC/GY785, the expressions of COL2 and aggrecan transcripts were recovered and that of type I collagen decreased. Immunohistological analyses of Si-HPMC/GY785 constructs transplanted into nude mice revealed the production of a cartilage-like extracellular matrix (ECM) containing high amounts of GAG and COL2. These results indicate that GY785-enriched Si-HPMC appears to be a promising hydrogel for cartilage tissue engineering. Copyright © 2015 John Wiley & Sons, Ltd.

Low polydispersity (N-ethyl pyrrolidine methacrylamide-co-1-vinylimidazole) linear oligomers for gene therapy applications.

Nonviral methods for gene delivery are becoming ever more prevalent along with the need to design new vectors that are highly effective, stable in biological fluids, inexpensive, and facile to produce. Here, we synthesize our previously reported monomer N-ethyl pyrrolidine methacrylamide (EPA) and evaluate its effectiveness in gene vector applications when copolymerized with 1-vinylimidazole (VI). A range of these novel linear cationic copolymers were synthesized via free radical polymerization with low molecular weights (oligomers) and low polydispersities showing two pK(a) values as the two co-monomers are cationic. DNA-polymer polyplexes had average sizes between 100 and 250nm and zeta-potentials between 10 and 25mV, and a strong dependence of composition on the size on the zeta-potential was observed. The cytotoxicity of the homopolymers, oligomers, and polyplexes toward human fibroblasts and 3T3 mouse fibroblasts was evaluated using the MTT and AlamarBlue™ assays, proving that formulations could be made with toxicity as low as low molecular weight linear poly (dimethylaminoethyl methacrylate) (PDMAEMA). The transfection capability of the polyplexes measured using the G-luciferase marker gene far superseded PDMAEMA when evaluated in biological conditions. Furthermore, blood compatibility studies showed that these new oligomers exhibit no significant hemolysis or platelet activation above PBS controls. These new EPA based oligomers with low toxicity and ease of scalability show high transfection abilities in serum conditions, and blood compatibility showing its potential for systemic gene delivery applications.

Archaeosomes based on synthetic tetraether-like lipids as novel versatile gene delivery systems.

Novel cationic liposomes, termed "archaeosomes", based on mixtures of neutral/cationic bilayer-forming lipids and archaeobacterial synthetic tetraether-type bipolar lipids show efficient in vitro gene transfection properties and represent a new approach for modulating the lipidic membrane fluidity of the complexes they form with DNA.

Archaeosomes based on novel synthetic tetraether-type lipids for the development of oral delivery systems.

The in vitro stability of archaeosomes made from novel synthetic membrane-spanning tetraether lipids was evaluated in conditions mimicking those of oral route application in terms of bile salts, serum and low pH..