Characterization of a nanoparticulate exopolysaccharide from Leuconostoc holzapfelii KM01 and its potential application in drug encapsulation.

Fermentation of Lactic Acid Bacteria (LAB) is considered to be a sustainable approach for polysaccharide production. Herein, exopolysaccharide (EPS)-producing LAB strain KM01 was isolated from Thai fermented dessert, Khao Mak, which was then identified as Leuconostoc holzapfelii. High-performance anion-exchange chromatography, nuclear magnetic resonance spectroscopy and Fourier-transform infrared spectroscopy suggested that the KM01 EPS comprises α-1,6-linked glucosides. The molecular weight of KM01 EPS was around 500 kDa, but it can form large aggregates formation (MW > 2000 kDa) in an aqueous solution, judged by transmission electron microscopy and dynamic light scattering to be around 150 nm in size. Furthermore, this KM01 EPS form highly viscous hydrogels at concentrations above 5% (w/v). The formation of hydrogels and nanoparticle of KM01 EPS was found to be reversible. Finally, the suitability of KM01 EPS for biomedical applications was demonstrated by its lack of cytotoxicity and its ability to form complexes with quercetin. Unlike the common α-1,6-linked dextran, KM01 EPS can enhance the solubility of quercetin significantly.

Evaluation of natural gum-based cryogels for soft tissue engineering.

In this study, three natural biomaterials, Locust bean gum (LBG), Xanthan gum (XG), and Mastic gum (MG), were combined to form cryogel scaffolds. Thermal and chemical characterizations revealed the successful blend formation from LBG-XG (LX) and LBG-XG-MG (LXM) polymers. All blends resulted in macro-porous scaffolds with interconnected pore structures under the size of 400 µm. The swollen cryogels had similar mechanical properties compared with other polysaccharide-based cryogels. The mean tensile and compressive modulus values of the wet cryogels were in the range of 3.5-11.6 kPa and 82-398 kPa, respectively. The sustained release of the small molecule Kartogenin from varying concentrations and ratios of cryogels was in between 32 and 66% through 21 days of incubation. Physical, mechanical, and chemical properties make LX and LXM polysaccharide-based cryogels promising candidates for cartilage and other soft tissue engineering, and drug delivery applications.

Phosphorylated xanthan gum-Ag(I) complex as antibacterial viscosity enhancer for eye drops formulation.

Topical instillation of eye drops represents the treatment of choice for many ocular diseases. Ophthalmic formulations must meet general requirements, i.e. pH, osmolality, transparency and viscosity to ensure adequate retention without inducing irritation and the development of eye infections. We developed a phosphorylated xanthan gum-Ag(I) complex (XGP-Ag) showing pH (pH = 7.1 ± 0.3) and osmolality values (311 ± 2 mOsm/kg) close to that of human tears (pH = 6.5-7.6 and 304 ± 23 mOsm/kg) thanks to the presence of phosphate moieties along the chain. The presence of phosphate groups covalently bound to the XG chains avoids their dispersion in fluid, thus reducing the risk of corneal calcification. 0.02% w/v XGP-Ag solution showed high transparency (higher than 95% along the entire visible range), adequate refractive index (1.334 ± 0.001) and viscosity in the range: γ 1 s-1-10,000 s- 1 (26.4 ± 0.8-2.1 ± 0.4 mPa·s). Its cytotoxicity and capability to hinder bacterial proliferation was also verified.

Production and characterization of exopolysaccharide from the sponge-associated Bacillus subtilis MKU SERB2 and its in-vitro biological properties.

In this study, the sponge-associated a potential endosymbiotic bacterium, Bacillus subtilis MKU SERB2 was identified and optimized the production of exopolysaccharide (EPS) by using response surface methodology (RSM). The central composite rotatable design (CCRD) exhibited the highest yield of EPS (617.81 µg/mL) obtained from the optimized medium containing 11.5 g/L of sucrose, 3.5 g/L of yeast extract, 3.0 g/L of peptone, and 2.5 g/L of calcium chloride. Fourier transform infrared (FTIR) spectrum of purified EPS indicated that the presence of carboxyl, hydroxyl, and amide as functional groups, and their structural composition was confirmed by 1H and 13C nuclear magnetic resonance (NMR) analysis. Moreover, the fibrous, porous and semi-crystalline nature of EPS was confirmed by SEM and X-ray powder diffraction (XRD) analysis and the EDX inferred demonstrated the presence of C, Na, O, N, S, and Cl respectively. Further, the isolated EPS exhibited potent antioxidant activity and moderate anticoagulant efficacy whereas there was no hemolytic and lymphocytes toxicity. Overall, our result suggests that the functional and biological properties of the EPS imply the potential applications in food and pharmaceutical industries in the future.

A fully human anti-BMP6 antibody reduces the need for erythropoietin in rodent models of the anemia of chronic disease.

Recombinant erythropoietin (EPO) and iron substitution are a standard of care for treatment of anemias associated with chronic inflammation, including anemia of chronic kidney disease. A black box warning for EPO therapy and concerns about negative side effects related to high-dose iron supplementation as well as the significant proportion of patients becoming EPO resistant over time explains the medical need to define novel strategies to ameliorate anemia of chronic disease (ACD). As hepcidin is central to the iron-restrictive phenotype in ACD, therapeutic approaches targeting hepcidin were recently developed. We herein report the therapeutic effects of a fully human anti-BMP6 antibody (KY1070) either as monotherapy or in combination with Darbepoetin alfa on iron metabolism and anemia resolution in 2 different, well-established, and clinically relevant rodent models of ACD. In addition to counteracting hepcidin-driven iron limitation for erythropoiesis, we found that the combination of KY1070 and recombinant human EPO improved the erythroid response compared with either monotherapy in a qualitative and quantitative manner. Consequently, the combination of KY1070 and Darbepoetin alfa resulted in an EPO-sparing effect. Moreover, we found that suppression of hepcidin via KY1070 modulates ferroportin expression on erythroid precursor cells, thereby lowering potentially toxic-free intracellular iron levels and by accelerating erythroid output as reflected by increased maturation of erythrocyte progenitors. In summary, we conclude that treatment of ACD, as a highly complex disease, becomes more effective by a multifactorial therapeutic approach upon mobilization of endogenous iron deposits and stimulation of erythropoiesis.

A 3D porous microsphere with multistage structure and component based on bacterial cellulose and collagen for bone tissue engineering.

Collagen (COL) and bacterial cellulose (BC) were chemically recombined by Malaprade and Schiff-base reactions. A three-dimensional (3D) porous microsphere of COL/BC/Bone morphogenetic protein 2 (BMP-2) with multistage structure and components were prepared by the template method combined with reverse-phase suspension regeneration. The microspheres were full of pores and had a rough surface. The particle size ranged from 8 to 12 microns, the specific surface area (SBET) was 123.4 m2/g, the pore volume (VPore) was 0.59 cm3/g, and the average pore diameter (DBJH) was 198.5 nm. The adsorption isotherm of the microspheres on the N2 molecule belongs to that of mesoporous materials. The microspheres showed good biocompatibility, and the 3D porous microspheres with multiple structures and components effectively promoted the adhesion, proliferation, and osteogenic differentiation of mice MC3T3-E1 cells. The study can provide a theoretical basis for the application of COL/BC porous microspheres in the field of bone tissue engineering.

Process control and scale-up of modified bacterial cellulose production for tailor-made anti-inflammatory drug delivery systems.

Bacterial cellulose (BC) has proven its high potential as active wound dressing and drug delivery system in many scientific studies, but the transferability of the methods to efficient manufacturing still needs to be demonstrated. This study presents a technically feasible, straightforward and efficient approach to modify BC according to specific medical requirements, to scale-up the cultivation and to load the active pharmaceutical ingredient of interest. By means of in situ-modification of the network structure using water-soluble poly(ethylene glycol) 400 and 4000 on pilot-scale, up to 41.5 ±â€¯3.0 % higher transparency of the dressing, 40.6 ±â€¯3.8 % increased loading capacity and 9% increased total release of the anti-inflammatory model drug diclofenac sodium could be obtained. Spray loading was investigated as material efficient alternative to absorption loading allowing a significant reduction in loading time.

Strength-tunable printing of xanthan gum hydrogel via enzymatic polymerization and amide bioconjugation.

Amide bioconjugation and interfacial enzyme polymerization are designed to provide a general strategy for regulating the mechanical strength (storage modulus from 3 kPa to 100 kPa) of printable hydrogel inks.

Ruminococcus gnavus, a member of the human gut microbiome associated with Crohn's disease, produces an inflammatory polysaccharide.

A substantial and increasing number of human diseases are associated with changes in the gut microbiota, and discovering the molecules and mechanisms underlying these associations represents a major research goal. Multiple studies associate Ruminococcus gnavus, a prevalent gut microbe, with Crohn's disease, a major type of inflammatory bowel disease. We have found that R. gnavus synthesizes and secretes a complex glucorhamnan polysaccharide with a rhamnose backbone and glucose sidechains. Chemical and spectroscopic studies indicated that the glucorhamnan was largely a repeating unit of five sugars with a linear backbone formed from three rhamnose units and a short sidechain composed of two glucose units. The rhamnose backbone is made from 1,2- and 1,3-linked rhamnose units, and the sidechain has a terminal glucose linked to a 1,6-glucose. This glucorhamnan potently induces inflammatory cytokine (TNFα) secretion by dendritic cells, and TNFα secretion is dependent on toll-like receptor 4 (TLR4). We also identify a putative biosynthetic gene cluster for this molecule, which has the four biosynthetic genes needed to convert glucose to rhamnose and the five glycosyl transferases needed to build the repeating pentasaccharide unit of the inflammatory glucorhamnan.

Legionella feeleii: pneumonia or Pontiac fever? Bacterial virulence traits and host immune response.

Gram-negative bacterium Legionella is able to proliferate intracellularly in mammalian host cells and amoeba, which became known in 1976 since they caused a large outbreak of pneumonia. It had been reported that different strains of Legionella pneumophila, Legionella micdadei, Legionella longbeachae, and Legionella feeleii caused human respiratory diseases, which were known as Pontiac fever or Legionnaires' disease. However, the differences of the virulence traits among the strains of the single species and the pathogenesis of the two diseases that were due to the bacterial virulence factors had not been well elucidated. L. feeleii is an important pathogenic organism in Legionellae, which attracted attention due to cause an outbreak of Pontiac fever in 1981 in Canada. In published researches, it has been found that L. feeleii serogroup 2 (ATCC 35849, LfLD) possess mono-polar flagellum, and L. feeleii serogroup 1 (ATCC 35072, WRLf) could secrete some exopolysaccharide (EPS) materials to the surrounding. Although the virulence of the L. feeleii strain was evidenced that could be promoted, the EPS might be dispensable for the bacteria that caused Pontiac fever. Based on the current knowledge, we focused on bacterial infection in human and murine host cells, intracellular growth, cytopathogenicity, stimulatory capacity of cytokines secretion, and pathogenic effects of the EPS of L. feeleii in this review.

Production of hydroxyapatite-bacterial nanocellulose scaffold with assist of cellulose nanocrystals.

Bacterial nanocellulose/hydroxyapatite/cellulose nanocrystal (BHC) composites were synthesized via in-situ synthesis using cellulose nanocrystals (CNCs) to improve colloidal stability and the dispersion of hydroxyapatite (HA) during the bacterial nanocellulose (BNC) cultivation period. Transmission electron microscopy images and energy dispersive spectroscopy (EDS) results confirmed the dispersion of HA on the CNC particles with a Ca/P ratio of 1.66 corresponding to that of the stoichiometric HA. The SEM images and EDS results showed that the integration of the HA and BNC network without CNC assistance (BHA (0.25 and 0.5 wt.%) composites) was less than that for BHC at both concentrations. Fourier-transform infrared analysis, XRD and thermal degradation revealed the effect of HA on the BHC composites with a decreased CrI% and improved thermal property. Cytotoxicity proved the potential for using BHC composites for bone tissue engineering scaffold with cell viability up to 83.4 ± 3.6% compared to the negative control (99.2 ± 0.08%).

Hydrazone-Linkage-Based Self-Healing and Injectable Xanthan-Poly(ethylene glycol) Hydrogels for Controlled Drug Release and 3D Cell Culture.

Polymeric hydrogels have been extensively explored for controlled drug-delivery applications, but there is an increasing demand for smart drug delivery combined with tunable physicochemical attributes and tissue engineering potential. In this work, novel xanthan-poly(ethylene glycol) (PEG) hydrogels were developed by cross-linking polysaccharide, oxidized xanthan, and 8-arm PEG hydrazine through dynamic, pH-responsive, and biodegradable hydrazone linkages. Aqueous solutions (pH 6.5) of oxidized xanthan and PEG hydrazine were mixed together at 37 °C to obtain hydrogels within minutes, and the formation of hydrazone linkages was ascertained using Fourier transform infrared spectroscopy. Fabrication of xanthan-PEG hydrogels using hydrazone linkages has not been reported previously. The 3% hydrogels exhibited the storage modulus of 194 Pa, which increased to 770 Pa for 5% hydrogels. When subjected to alternating cycles of varying strains of 1 and 800% (5 cycles), hydrogels demonstrated instant recovery each time the extreme strain was relieved, thus suggesting excellent self-healing capabilities. Doxorubicin (DOX), chemotherapeutic agent, was loaded onto hydrogels, and release studies were carried out at pH 5.5 (tumoral) and 7.4 (physiological). The cumulative release from 3, 4, and 5% hydrogels at pH 5.5 was 81.06, 61.98, and 41.67%, whereas the release at pH 7.4 was 47.43, 37.01, and 35.34% at 30 days. MTT assay showed that oxidized xanthan and PEG hydrazine are not toxic to mammalian cells (NIH-3T3), as the cell viabilities were found to be 84.66 and 102% for concentrations up to 1 mg/mL. The live/dead assay with encapsulated NIH-3T3 cells showed no significant dead cell population, suggesting excellent compatibility of hydrogels in 2D and 3D culture. DOX-loaded hydrogels exhibited cytotoxicity against A549 cells when exposed to media released from hydrogels. Overall, hydrogels developed in this work may have potential applications in drug delivery and 3D cell culture for cell delivery.

Radiation fabrication of Xanthan-based wound dressing hydrogels embedded ZnO nanoparticles: In vitro evaluation.

Depending on the biocompatibility of Xanthan, polyvinyl alcohol and the antibacterial efficiency of zinc oxide nanoparticles (ZnO), a series of (Xanthan-polyvinyl alcohol)/ZnO nanocomposite hydrogels were prepared as wound dressing using eco-friendliness 60Co γ-ray irradiation facility. ZnO nanoparticles were characterized using X-ray diffraction, UV-vis spectroscopy, transmission electron microscopy and energy dispersive X-ray analysis. The size of ZnO nanoparticles was ranged between 15 and 25 nm. The presence of ZnO nanoparticles reconstructed the internal structure of the hydrogel network which aid in a homogenous porous structure as indicated by scanning electron micrographs. Such adequate porosity along with the presence of ZnO nanoparticles controls the fluid uptake ability, water retention and water vapor transmission rate. The fluid uptake ability in pseudo-extracellular fluid and water ranged between (554-664%) and (1281-1603%), respectively. After exposure to air for 6 h, ZnO dressings kept about 50-65% of their water content which makes them more suitable for moderate exudating wounds. Water vapor transmission rate ranged between 167 and 184 (g/(m2 h) which is sufficient to keep wound's surface moist. ZnO dressings show an efficient microbial barrier potency and profound antimicrobial activity against Staphylococcus aureus, Escherichia coli and Candida albicans. In vitro cytotoxicity and haemolytic potency evaluation showed their biocompatibility.

Effects of cyanobacteria Oscillatoria sp. lipopolysaccharide on B cell activation and Toll-like receptor 4 signaling.

Cyanobacteria ("blue-green algae"), such as Oscillatoria sp., are a ubiquitous group of bacteria found in freshwater systems worldwide that are linked to illness and in some cases, death among humans and animals. Exposure to cyanobacteria occurs via ingestion of contaminated water or food-products. Exposure of the gut to these bacteria also exposes their toxins, such as lipopolysaccharide (LPS), to B cells in the gut associated lymphoid tissue. However, the effect of Oscillatoria sp. LPS on B cell activation is unknown. To test the hypothesis that Oscillatoria sp. LPS exposure to murine B cells would result in B cell activation, murine B cells were incubated in the absence or presence of Oscillatoria sp. LPS or E. coli LPS as a positive control. The data indicate that Oscillatoria sp. LPS induces B cells to proliferate, upregulate MHC II and CD86, enhance antigen uptake and induce IgM production at low levels. Additional studies demonstrate that this low level of stimulation may be due to incomplete TLR4 signaling induced by Oscillatoria sp. LPS, since IRF-3 is not induced in B cells after stimulation with Oscillatoria sp. LPS. These findings have important implications for the mechanisms of toxicity of cyanobacteria in both humans and animals.

Cytotoxic effect of a mannogalactoglucan extracted from Agaricus bisporus on HepG2 cells.

A mannogalactoglucan (RK2-Ab; Mw 1.8×104gmol-1) composed by Man (27.3%), Gal (24.4%) and Glc (48.3%) was extracted and characterized from Agaricus bisporus, and its biological activity was evaluated on human hepatocarcinoma cells (HepG2). The partially-O-methylated alditol acetates together with the NMR data suggest the main chain to be composed of α-d-Galp (32.8%) and ß-d-Glcp (37.0%) units (1→6)-linked, with ß-d-Manp (14.6%), as non-reducing end units, substituting the side chains at O-2 (α-d-Galp units; 3.3%) and O-2 and O-4 (ß-d-Glcp units; 3.6%). (1→2)-linked ß-d-Glcp (2.7%) and ß-d-Manp (6.0%) can also be observed. RK2-Ab reduced cellular viability of HepG2 cells, by both, the MTT and lactate dehydrogenase release assays, promoted the increase of cytochrome c release and decrease of ATP content. Suggesting that the mannogalactoglucan from A. bisporus may have antitumor activity by inducing apoptosis by the mitochondrial death pathway, and could be used in cancer therapy.

Modified xanthan gum for buccal delivery-A promising approach in treating sialorrhea.

AIM: The aim of this study was to modify xanthan, a well-known gelling agent, in order to treat sialorrhea, which increases salivary flow due to an excessive stimulus of the salivary reflex. METHODS: Chemical modification occurs by covalent attachment of l-cysteine (SH) to the polymeric backbone of xanthan (X) via amide bond formation. Safety considerations, water uptake capacity, and erosion were evaluated. Furthermore, mucoadhesiveness on buccal mucosa and vapor uptake studies were performed. In vitro/in vivo correlation of reduce of salivary flow was conducted and drug release of embedded tannin was determined. RESULTS: Safety investigations ensured modified X-SH being safe to use. X-SH exposed 1.5-x higher water uptake capacity in comparison to unmodified xanthan. Then, stability of X-SH augmented 5.5-fold in the case of matrix erosion studies. Reduction of salivary flow could be obtained 1.6-fold improved in case of X-SH compared to X. Furthermore, tannin was 1.8-fold controlled released in comparison to unmodified xanthan. CONCLUSION: Taking these findings into consideration, chemical modified xanthan emerged with its distinctive properties as a promising approach in treating sialorrhea.

Nanoemulsion-based electrolyte triggered in situ gel for ocular delivery of acetazolamide.

In the present work the antiglaucoma drug, acetazolamide, was formulated as an ion induced nanoemulsion-based in situ gel for ocular delivery aiming a sustained drug release and an improved therapeutic efficacy. Different acetazolamide loaded nanoemulsion formulations were prepared using peanut oil, tween 80 and/or cremophor EL as surfactant in addition to transcutol P or propylene glycol as cosurfactant. Based on physicochemical characterization, the nanoemulsion formulation containing mixed surfactants and transcutol P was selected to be incorporated into ion induced in situ gelling systems composed of gellan gum alone and in combination with xanthan gum, HPMC or carbopol. The nanoemulsion based in situ gels showed a significantly sustained drug release in comparison to the nanoemulsion. Gellan/xanthan and gellan/HPMC possessed good stability at all studied temperatures, but gellan/carbopol showed partial drug precipitation upon storage and was therefore excluded from the study. Gellan/xanthan and gellan/HPMC showed higher therapeutic efficacy and more prolonged intraocular pressure lowering effect relative to that of commercial eye drops and oral tablet. Gellan/xanthan showed superiority over gellan/HPMC in all studied parameters and is thus considered as a promising mucoadhesive nanoemulsion-based ion induced in situ gelling formula for topical administration of acetazolamide.

Capsular Polysaccharide of Mycoplasma ovipneumoniae Induces Sheep Airway Epithelial Cell Apoptosis via ROS-Dependent JNK/P38 MAPK Pathways.

In an attempt to better understand the pathogen-host interaction between invading Mycoplasma ovipneumoniae (M. ovipneumoniae) and sheep airway epithelial cells, biological effects and possible molecular mechanism of capsular polysaccharide of M. ovipneumoniae (CPS) in the induction of cell apoptosis were explored using sheep bronchial epithelial cells cultured in air-liquid interface (ALI). The CPS of M. ovipneumoniae was first isolated and purified. Results showed that CPS had a cytotoxic effect by disrupting the integrity of mitochondrial membrane, accompanied with an increase of reactive oxygen species and decrease of mitochondrial membrane potential (ΔΨm). Of importance, the CPS exhibited an ability to induce caspase-dependent cell apoptosis via both intrinsic and extrinsic apoptotic pathways. Mechanistically, the CPS induced extrinsic cell apoptosis by upregulating FAS/FASL signaling proteins and cleaved-caspase-8 and promoted a ROS-dependent intrinsic cell apoptosis by activating a JNK and p38 signaling but not ERK1/2 signaling of mitogen-activated protein kinases (MAPK) pathways. These findings provide the first evidence that CPS of M. ovipneumoniae induces a caspase-dependent apoptosis via both intrinsic and extrinsic apoptotic pathways in sheep bronchial epithelial cells, which may be mainly attributed by a ROS-dependent JNK and p38 MAPK signaling pathways.

Anti-inflammatory and angiogenic activity of polysaccharide extract obtained from Tibetan kefir.

The search for new bioactive molecules is a driving force for research pharmaceutical industries, especially those molecules obtained from fermentation. The molecules possessing angiogenic and anti-inflammatory attributes have attracted attention and are the focus of this study. Angiogenic activity from kefir polysaccharide extract, via chorioallantoic membrane assay, exhibited a pro-angiogenic effect compared with vascular endothelial factor (pro-angiogenic) and hydrocortisone (anti-angiogenic) activity as standards with an EC50 of 192ng/mL. In terms of anti-inflammatory activity determined via hyaluronidase enzyme assay, kefir polysaccharide extract inhibited the enzyme with a minimal activity of 2.08mg/mL and a maximum activity of 2.57mg/mL. For pharmaceutical purposes, kefir polysaccharide extract is considered to be safe because it does not inhibit VERO cells in cytotoxicity assays.

Safety Assessment of Microbial Polysaccharide Gums as Used in Cosmetics.

The Cosmetic Ingredient Review Expert Panel assessed the safety of 34 microbial polysaccharide gums for use in cosmetics, finding that these ingredients are safe in cosmetic formulations in the present practices of use and concentration. The microbial polysaccharide gums named in this report have a variety of reported functions in cosmetics, including emulsion stabilizer, film former, binder, viscosity-increasing agent, and skin-conditioning agent. The Panel reviewed available animal and clinical data in making its determination of safety.