Development of a novel beta-glucan supplemented hydrogel spray formulation and wound healing efficacy in a db/db diabetic mouse model.

To relieve the severe economic and social burdens and patient suffering caused by the increasing incidence of chronic wounds, more effective treatments are urgently needed. In this study, we focused on developing a novel sprayable wound dressing with the active ingredient ß-1,3/1,6-glucan (ßG). Since ßG is already available as the active ingredient in a commercial wound healing product provided as a hydrogel in a tube (ßG-Gel), the sprayable format should bring clinical benefit by being easily sprayed onto wounds; whilst retaining ßG-Gel's physical stability, biological safety and wound healing efficacy. Potentially sprayable ßG hydrogels were therefore formulated, based on an experimental design setup. One spray formulation, named ßG-Spray, was selected for further investigation, as it showed favorable rheological and spraying properties. The ßG-Spray was furthermore found to be stable at room temperature for more than a year, retaining its rheological properties and sprayability. The cytotoxicity of ßG-Spray in keratinocytes in vitro, was shown to be promising even at the highest tested concentration of 100 µg/ml. The ßG-Spray also displayed favorable fluid affinity characteristics, with a capacity to both donate and absorb close to 10% fluid relative to its own weight. Finally, the ßG-Spray was proven comparably effective to the commercial product, ßG-Gel, and superior to both the water and the carrier controls (NoßG-Spray), in terms of its ability to promote wound healing in healing-impaired animals. Contraction was found to be the main wound closure mechanism responsible for the improvement seen in the ßG-treatment groups (ßG-Spray and ßG-Gel). In conclusion, the novel sprayable ßG formulation, confirmed its potential to expand the clinical use of ßG as wound dressing.

Beta-glucan-loaded nanofiber dressing improves wound healing in diabetic mice.

The increased prevalence of chronic wounds requires novel treatment options. The aim of this study was to develop a beta-glucan (ßG)-loaded nanofiber wound dressing. Nanofibers were prepared using the needle-free Nanospider™ technology, an electrospinning method which enables the production of nanofibers at an industrial scale. The ßG was selected as active ingredient based on its confirmed wound healing potential in both animals and humans. Hydroxypropyl methylcellulose (HPMC) and polyethylene oxide (PEO) were included as copolymers. Rheological profiles of spinning solutions containing HPMC, PEO, ßG, ethanol and water, were optimized. The nanofiber formation was confirmed by Field Emission Scanning Electron Microscopy (FE-SEM), and both nanofibers with (ßG-nanofibers) or without ßG (NoßG-nanofibers) were evaluated by their swelling index and FT-IR spectroscopy. The formulations, active ingredient and excipients were tested for their possible in vitro toxicity in keratinocytes. Finally, the wound healing potential of the nanofibers was tested in externally induced excisional wounds in male diabetic db/db mice. Three different doses of ßG-nanofibers and the ßG-free, NoßG-nanofibers, were evaluated for their in vivo wound healing efficacy. All nanofiber-treatments provided improved wound healing as compared to the negative control (water). All ßG-nanofiber treated groups exhibited significantly improved wound healing as compared to the NoßG-nanofiber treated group, indicating the potential of ßG-nanofibers as wound dressing.

Sprayable Carbopol hydrogel with soluble beta-1,3/1,6-glucan as an active ingredient for wound healing - Development and in-vivo evaluation.

Chronic wounds represent a significant health problem worldwide. There is a need for advanced- and cost-efficient wound healing products able to increase patient comfort and reduce the healing time. The aim of this study was to develop a sprayable hydrogel dressing with beta-glucan (ßG) as the active ingredient, targeting future application in the treatment of both chronic and burn wounds. The ßG was chosen as an active ingredient because of its promising wound healing capabilities, whereas Carbopol 971P NF (Carbopol) was chosen as the thickening agent in the formulation due to several attractive characteristics such as its low viscosity, low toxicity, high transparency and good ion tolerance. Four different hydrogel formulations were prepared with varying Carbopol concentrations. The higher Carbopol concentration, 0.5% (w/w), was used to prepare three formulations comprising the HighCP:NoßG, HighCP:LowßG and the HighCP:MediumßG formulation, respectively. Lower Carbopol concentration, 0.25% (w/w), was used to prepare the LowCP:HighßG formulation. The content of ßG varied from 0.25% in the HighCP:LowßG, 0.5% in the HighCP:MediumßG and 1.0% (w/w) in the LowCP:HighßG formulation, respectively. The first part of the study focused on the rheological characterization of the hydrogels and the fluid affinity testing. All formulations were confirmed to be stable gels; the ßG was shown to augment the gel strength by increasing the yield strength of the gel in a dose dependent manner. The stability of the formulations containing either Carbopol alone or in a combination with ßG did not deteriorate over 26weeks, and the fluid donation and absorption study indicated a fluid donation profile, which favors healing of dry wounds. The in vivo efficacy of the formulations, evaluated in the modified diabetic male mice (db/db mice), showed that Carbopol alone was unable to induce improved healing and caused adverse reactions in some wounds. The inclusion of ßG increased the epithelialization and wound contraction in the db/db mice when given at high ßG:Carbopol ratio. The positive effect of ßG was, however, not sufficient to counteract the adverse effect of Carbopol, thus a more suitable thickening agent should be investigated for further development of a sprayable wound care product.

PLGA/PLA micro- and nanoparticle formulations serve as antigen depots and induce elevated humoral responses after immunization of Atlantic salmon (Salmo salar L.).

Novel vaccine delivery systems are highly needed to improve the salmon aquaculture industry. Although particles of biocompatible polymers such as poly(lactic-co-glycolic) acid (PLGA) have long been considered promising candidates for delivery of immunogenic compounds, few studies have addressed their use as vaccine carriers in Atlantic salmon (Salmo salar L.). Investigating their ability to retain/depot antigen and induce time and dosage dependent adaptive humoral responses to immunization, we here present a basic study of the adjuvantic properties PLGA and PLA particles may have in salmon vaccines. A model antigen (human gamma globulin, HGG) was co-encapsulated with ß-glucan in nanoparticles (<1000nm) and microparticles (∼8µm) of different chemical compositions. Atlantic salmon were immunized with (a) PLGA or PLA particle entrapped antigen (12 different treatment groups), (b) antigen and ß-glucan in PBS, (c) an oil-based formulation or (d) nanoparticles (NPs) or microparticles (MPs) combined with the oil-adjuvanted formulation. ELISA analysis showed that NPs and MPs were capable of inducing elevated antibody responses at day 60 and 75 post immunization, but the antibody levels were reduced at day 90 and 120. In contrast, oil-based formulations, either alone or in combination with NPs or MPs resulted in strong antibody responses at all sampling time points. Comparable dosage dependent increase in antibody responses was observed when administering antigen with ß-glucan either in PBS, entrapped in NPs or MPs, or in an oil-adjuvanted formulation. However, as the antigen doses were increased, MPs and the oil-based formulation gave the strongest responses. Antigen presence in the blood, organ package/injection site, kidney, carcass and the whole body was quantified by radiotracing of I(125)-labelled HGG at day 7 and 36 post immunization. At both sampling time points, the highest radioactivity levels were measured from the whole-body and organ package/injection site in groups injected with MPs and oil-based formulations, indicating that these formulations resulted in superior antigen retention. Interestingly, NPs were found to accumulate in the kidney, a result that corroborated with in vitro uptake of NPs in a DC/Mφ-like cell line from Atlantic salmon.