Characterization of the exopolymer-producing Pseudoalteromonas sp. S8-8 from Antarctic sediment.

A synergistic approach using cultivation methods, chemical, and bioinformatic analyses was applied to explore the potential of Pseudoalteromonas sp. S8-8 in the production of extracellular polymeric substances (EPSs) and the possible physiological traits related to heavy metal and/or antibiotic resistance. The effects of different parameters (carbon source, carbon source concentration, temperature, pH and NaCl supplement) were tested to ensure the optimization of growth conditions for EPS production by the strain S8-8. The highest yield of EPS was obtained during growth in culture medium supplemented with glucose (final concentration 2%) and NaCl (final concentration 3%), at 15 °C and pH 7. The EPS was mainly composed of carbohydrates (35%), followed by proteins and uronic acids (2.5 and 2.77%, respectively) and showed a monosaccharidic composition of glucose: mannose: galactosamine: galactose in the relative molar proportions of 1:0.7:0.5:0.4, as showed by the HPAE-PAD analysis. The detection of specific molecular groups (sulfates and uronic acid content) supported the interesting properties of EPSs, i.e. the emulsifying and cryoprotective action, heavy metal chelation, with interesting implication in bioremediation and biomedical fields. The analysis of the genome allowed to identify a cluster of genes involved in cellulose biosynthesis, and two additional gene clusters putatively involved in EPS biosynthesis. KEY POINTS: • A cold-adapted Pseudoalteromonas strain was investigated for EPS production. • The EPS showed emulsifying, cryoprotective, and heavy metal chelation functions. • Three gene clusters putatively involved in EPS biosynthesis were evidenced by genomic insights.

Effect of gal/GalNAc regioisomerism in galactosylated liposomes on asialoglycoprotein receptor-mediated hepatocyte-selective targeting in vivo.

In this study, we describe a novel synthesis of galactosylated lipids by lipase catalysis. Lactitol (Lac), galactose (Gal), or N-acetyl galactosamine (GalNAc) was coupled with cholesterol (CHS) as target head groups by enzyme-catalyzed regioselective esterification to produce three kinds of lipids: CHS-1-Gal, CHS-6-Gal, or CHS-6-GalNAc1. The biological effects of galactosylated lipids carrying different constitutional isomers of the pendent sugar species were investigated. LP-1-Gal (liposomes containing 5.0 molar% of CHS-1-Gal) showed strong binding to tetrameric lectins of Ricinus communis agglutinin (RCA120) in vitro, while LP-6-Gal (liposomes containing 5.0 molar% of CHS-6-Gal) and LP-6-GalNAc (liposomes containing 5.0 molar% of CHS-6-GalNAc) did not. After intravenous injection, LP-6-GalNAc, LP-1-Gal and LP-6-Gal rapidly disappeared from the blood and accumulated rapidly in liver (up to 74.88 ± 4.11%, 58.67 ± 5.75%, and 47.66 ± 4.56% of injected dose/g organ within 4 h, respectively). This is significantly higher than the uptake of unmodified liposomes (Unmod-LP) (18.67 ± 6.07%). Pre-injection of asialofetuin significantly inhibits liver uptake of Gal-liposomes (P < 0.01), with the degree of inhibition appearing in the following order: LP-6-GalNAc (73.29%) > LP-1-Gal (67.06%) > LP-6-Gal (53.61%). More importantly, LP-6-GalNAc was preferentially taken up by hepatocytes and the uptake ratio by parenchymal cells (PC) and nonparenchymal cells (NPC) (PC/NPC ratio) was 11.03 higher than LP-1-Gal (7.32), LP-6-Gal (5.83) and Unmod-LP (2.39). We suggest that liposomes containing the novel galactosylated lipid CHS-6-GalNAc have potential as drug delivery carriers for hepatocyte-selective targeting.

Lactobacillus salivarius LI01 encapsulated in alginate-pectin microgels ameliorates D-galactosamine-induced acute liver injury in rats.

Acute liver failure is a clinical emergency associated with high mortality. Accumulating evidence indicates that gut microbiota participates in the progression of liver injury, and preventive therapies based on altering gut microbiota are of great interest. Previous studies demonstrated that Lactobacillus salivarius LI01 attenuates hepatic injury, though efficiency in curtailed in the harsh environment in the gastrointestinal tract. In this study, a system to encapsulate LI01 in alginate-pectin (AP) microgels was investigated. Encapsulation significantly enhances probiotic viability for long-term storage and heat treatment, and in simulated gastrointestinal fluids (SGF or SIF) and bile salt solutions. Acute liver injury was induced in Sprague-Dawley (SD) rats by D-galactosamine (D-GaIN) injection following pretreatment with probiotics. Liver and gut barrier function, cytokines, liver and gut histology, bacterial translocation, and gut microbiota were assessed. Administration of encapsulated LI01 more effectively upregulates hepatic anti-inflammatory cytokine IL-10 and TLR-3, restores expressions of gut barrier biomarkers Claudin-1 and MUC2 and attenuates destruction of mucosal ultrastructure compared with unencapsulated probiotics pretreatment. Pretreatment with AP-LI01 microgels altered the microbial community, decreasing the abundance of pathogenic taxa Ruminiclostridium, Dorea and Ruminococcaceae_UCG-004 and enriching beneficial taxa Ruminococcaceae_UCG-014, Eubacterium, and Prevotella_1 that produce short-chain fatty acids. These results suggest that AP encapsulation of LI01 boosts viability and attenuates liver injury by reducing inflammation and restoring intestinal barrier function. These beneficial effects are probably due to alternation of gut flora. These findings provide new insight into encapsulation technology and prevention of liver failure. KEY POINTS: • Alginate-pectin encapsulation enhances the viability of Lactobacillus salivarius LI01 under simulated commercial conditions and simulated gastrointestinal environment. • AP-LI01 microgel attenuates hepatic and intestinal inflammation and restores gut barrier function. • AP-LI01 microgel alters gut microbial community with increased SCFAs producers and decreased pathogenic microbes. • Beneficial improvements after administration of probiotics are highly associated with alternation of gut microbial community.

Barbaloin loaded polydopamine-polylactide-TPGS (PLA-TPGS) nanoparticles against gastric cancer as a targeted drug delivery system: Studies in vitro and in vivo.

Gastric cancer is the third leading cause of cancer-associated death worldwide. Although a decrease in its incidence is observed, gastric cancer still poses a major clinical challenge due to poor prognosis and limited treatments. Barbaloin (BBL) is a main medicinal composition of the Chinese traditional medicine aloe vera. BBL has various bioactivities, including anti-oxidant, anti-inflammatory and anti-tumor properties. Polydopamine (pD)-based surface modification is easy to functionalize polymeric nanoparticles (NPs) surfaces with ligands and/or additional polymeric layers. In the present study, BBL-loaded formulations was developed with pD-modified NPs, which was synthesized by polylactide-TPGS (PLA-TPGS) (pD-PLA-TPGS/NPs). And galactosamine (Gal) was conjugated on the prepared NPs (Gal-pD-PLA-TPGS/NPs) for targeting the gastric cancer cells. Here, we found that BBL-loaded Gal-pD-PLA-TPGS/NPs showed the highest cellular uptake efficacy in gastric cancer cells. Gal-pD-PLA-TPGS/NPs more significantly reduced the gastric cancer cell viability. Further, greater apoptosis, autophagy and ROS generation was induced by Gal-pD-PLA-TPGS/NPs in gastric cancer cells. Additionally, compared to the other two NPs, Gal-pD-PLA-TPGS/NPs most markedly decreased ATP levels in gastric cancer cells. In vivo, Gal-pD-PLA-TPGS/NPs were specifically targeted to tumor site. Moreover, Gal-pD-PLA-TPGS/NPs exhibited the most anti-tumor effects, as evidenced by the lowest tumor volume and tumor weight. Of note, there was no significant difference was observed in body and liver weight, as well as the histological changes in major organs isolated from each group of mice. Together, the findings indicated that BBL-loaded Gal-pD-PLA-TPGS/NPs could be targeted to gastric cancer cells to suppress tumor progression without toxicity.

Fast on-Site Visual Detection of Active Ricin Using a Combination of Highly Efficient Dual-Recognition Affinity Magnetic Enrichment and a Specific Gold Nanoparticle Probe.

Ricin, a highly toxic protein, is a controlled substance by both the Chemical Weapons Convention (CWC) and the Biological Weapons Convention (BWC). Therefore, fast precaution of potential ricin toxin plays an important role in national security and public safety. Herein, a simple, sensitive, and accurate visual detection of active ricin in complex samples is presented by combining magnetic affinity enrichment with a specific gold nanoparticle (AuNP) probe. In the first step, a dual-recognition magnetic absorbent was fabricated by simultaneously incorporating two different affinity ligands (concanavalin A and galactosamine) on low-foul polymer brushes grafted magnetic beads, which showed remarkable multivalent synergy binding capacity for ricin even under complex interfering environments. Subsequently, a homoadenine-constituted oligodeoxynucleotide named poly(21dA) was conjugated to AuNPs (the poly(21dA)-AuNPs), which served as a specific depurination substrate of active ricin. Coralyne can trigger the intact poly(21dA)-AuNPs aggregate by forming a non-Watson-Crick homoadenine/coralyne complex, but the poly(21dA)-AuNPs after reacting with active ricin failed to form this complex due to the loss of adenines. Based on these facts, active ricin can be detected as low as 12.5 ng mL-1 with the naked eyes. This detection strategy could be well-applied in various ricin-spiked complex matrices. The features such as ready operation, facile readout, and easy accessibility make the assay a better choice for fast on-site active ricin detection.

A combinatorial library of bi-functional polymeric vectors for siRNA delivery in vitro.

PURPOSE: To apply a combinatorial chemistry approach toward the design of polymeric vectors, and to evaluate their effectiveness as siRNA delivery systems in vitro. METHODS: Poly(acrylic acid) (pAA) was synthesized via RAFT polymerization with well-controlled molecular weights (M (n): 3 kDa, 5 kDa, 10 kDa and 21 kDa). A polymer library was generated from the pAA precursors by conjugating two distinct moieties, agmatine (Agm) and D-(+)-galactosamine (Gal), at various ratios. Biophysical and cellular characterization was evaluated in vitro for these polymeric vectors using MDA-MB-231-luc+ cells. RESULTS: A critical balance between Agm/Gal content and polymer molecular weight must be attained to achieve favorable transfection efficacies. From the library of 22 polymers, only a few had knockdown efficiencies commensurate with effective siRNA delivery, particularly those with polymer precursor M (n) of 5 kDa and 10 kDa. Highest protein knockdown of 84% was achieved by a polymer conjugate with a 5 kDa pAA backbone with a side chain composition of 55% Agm and 17% Gal. CONCLUSIONS: Effective delivery of siRNA was found to be highly dependent on the molecular structure of the polymeric vector. The combinatorial approach employed provided the tools to identify optimal structural properties leading to efficient siRNA delivery for this class of vector.

The influence of membrane molecular weight cutoff on a novel bioartificial liver.

Given the xenogeneic immune reaction relevant to the molecular weight cutoff of the membrane of a bioartificial liver (BAL) system, we investigated the influence of membrane molecular weight cutoff in our BAL system in this study. Acute liver failure in beagles was induced by d-galactosamine administration. Eight beagles were divided into two groups by the membrane molecular weight cutoff of the plasma component separator. Group 1 beagles were treated with BAL containing 200 kDa retention rating membrane. Group 2 beagles were treated with BAL containing 1200 kDa retention rating membrane. Each group underwent two 6-h BAL treatments that were performed on day 1 and day 21. The hemodynamic and hematologic response, humoral immune responses, and cytotoxic immune response to BAL therapy were studied before and after treatments. All beagles remained hemodynamically and hematologically stable during BAL treatments. BAL treatment was associated with a significant decline in levels of complement; however, a longer time of level maintenance was observed in Group 2. Group 2 beagles experienced a significant increase in levels of IgG and IgM after two BAL treatments. Significant levels of canine proteins were detected in BAL medium from Group 2; only trace levels of canine proteins were detected in BAL medium from Group 1. The posttreatment viability of co-culture cells in Group 2 was lower compared with Group 1, and the viability of co-culture cells after treatments was associated with deposition of canine proteins on the cells. Xenogeneic immune response was influenced by membrane molecular weight cutoff in the BAL.

The Pel polysaccharide is predominantly composed of a dimeric repeat of α-1,4 linked galactosamine and N-acetylgalactosamine.

The genetic capacity to synthesize the biofilm matrix exopolysaccharide Pel is widespread among Gram-negative and Gram-positive bacteria. However, its exact chemical structure has been challenging to determine. Using a Pseudomonas aeruginosa strain engineered to overproduce Pel, improvements to the isolation procedure, and selective hydrolysis with the glycoside hydrolase PelAh, we demonstrate that Pel is a partially de-N-acetylated linear polymer of α-1,4-N-acetylgalactosamine comprised predominantly of dimeric repeats of galactosamine and N-acetylgalactosamine.

Propitious maneuvering for delivery of the phytopharmaceutical "apocynin" to induced fulminant hepatitis in BALB/c mice: In vitro and in vivo assessments.

Apocynin (APO), a specific nicotinamide adenine dinucleotide phosphate-oxidase (NADPH-oxidase, NOX) inhibitor, has recently emerged as a bioactive phytochemical with eminent anti-inflammatory and anti-oxidant activities. To our knowledge, no research has been conducted to fabricate a mucoadhesive nanostructured delivery system of APO that targets the liver. Accordingly, chitosan (CS) surface decorated polymeric nanoparticulate delivery system (PNDS) was victoriously fabricated by double emulsion-solvent evaporation method. Herein, a randomized full 33 factorial design was employed to assess the impact of the independently processing parameters (IPPs) namely; (poly(d,l-lactide-co-glycolide) (PLGA) amount (A)), (polyvinyl alcohol (PVA) concentration (B)), and (CS concentration (C)), on different dependently measured attributes (DMAs). The optimal APO-loaded chitosan-coated poly(d,l-lactide-co-glycolide) nanoparticles (APO-loaded CS-coated PLGA NPs) formula (F19) would be extensively appraised through meticulous in vitro-in vivo studies. Crucially, the results revealed that oral pre-treatment with the optimal formula evoked a prodigious in vivo hepatoprotective efficacy against lipopolysaccharide (LPS)/D-(+)-galactosamine (D-GalN) induced fulminant hepatitis (FH) in BALB/c mice when compared with pure APO, uncoated F19, and plain NPs (P NPs) pretreated groups. In conclusion, APO-loaded CS-coated PLGA NPs could be considered as a promising oral mucoadhesive phytopharmaceutical PNDS to open new prospects for therapeutic intervention in inflammatory based liver diseases.

An in-depth analysis of polymer-analogous conjugation using DMTMM.

Combinatorial libraries have become increasingly popular in the field of functional biomaterials. One approach for creating diverse polymer libraries is polymer-analogous conjugation of functional groups to polymer scaffolds. In this study, we show that a water-soluble condensing agent, 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMTMM), can be employed to conjugate two disparate model ligands, d-(+)-galactosamine (Gal) and agmatine (Agm), to the side chains of either poly(methacrylic acid) (pMAA) or poly(acrylic acid) (pAA) at various substitution ratios. The degree of substitution was found to be directly influenced by media pH, polymer concentration, structure of ligands, and polymer precursor. A nearly 2-fold increase in conjugation efficiencies for both ligands to pAA was achieved as compared to pMAA under identical conditions reaching up to 56% and 78% of Gal and Agm of total content, respectively. These two structurally similar polymers showed remarkably different performances, which reveals that the selection of a polymer precursor is crucial for the optimal design of polymeric libraries, particularly when complex structural ligands are involved. The approach employed provides a basis from which larger and more diverse combinatorial libraries of functionalized polymers with multiple moieties can be generated.

Optically Manipulated Microtools to Measure Adhesion of the Nanoparticle-Targeting Ligand Glutathione to Brain Endothelial Cells.

Targeting nanoparticles as drug delivery platforms is crucial to facilitate their cellular entry. Docking of nanoparticles by targeting ligands on cell membranes is the first step for the initiation of cellular uptake. As a model system, we studied brain microvascular endothelial cells, which form the anatomical basis of the blood-brain barrier, and the tripeptide glutathione, one of the most effective targeting ligands of nanoparticles to cross the blood-brain barrier. To investigate this initial docking step between glutathione and the membrane of living brain endothelial cells, we applied our recently developed innovative optical method. We present a microtool, with a task-specific geometry used as a probe, actuated by multifocus optical tweezers to characterize the adhesion probability and strength of glutathione-coated surfaces to the cell membrane of endothelial cells. The binding probability of the glutathione-coated surface and the adhesion force between the microtool and cell membrane was measured in a novel arrangement: cells were cultured on a vertical polymer wall and the mechanical forces were generated laterally and at the same time, perpendicularly to the plasma membrane. The adhesion force values were also determined with more conventional atomic force microscopy (AFM) measurements using functionalized colloidal probes. The optical trapping-based method was found to be suitable to measure very low adhesion forces (≤ 20 pN) without a high level of noise, which is characteristic for AFM measurements in this range. The holographic optical tweezers-directed functionalized microtools may help characterize the adhesion step of nanoparticles initiating transcytosis and select ligands to target nanoparticles.

Galactosamine-modified PEG-PLA/TPGS micelles for the oral delivery of curcumin.

In this study, galactosamine-modified poly(ethylene glycol)-poly(lactide) (Gal-PEG-PLA) polymers were synthesized and Gal-PEG-PLA/D-α-tocopherol polyethylene glycol 1000 succinate (TPGS) micelles named as GPP micelles were designed to promote the oral absorption of a hydrophobic drug, curcumin (CUR). CUR-loaded Gal-PEG-PLA/TPGS micelles (CUR@GPP micelles) were fabricated using the thin-film dispersion method. CUR@GPP micelles had a size of about 100 nm, a near-neutral zeta potential, drug loading (DL) of 14.6%, and sustained release properties. GPP micelles with high Gal density (GPP3 micelles) were superior in facilitating uptake in epithelial cells and improving intestinal permeation. In situ intestinal absorption studies suggested that the jejunum and ileum were the best absorption segments in the intestinal tract. Additionally, biodistribution results revealed that GPP3 micelles could be remarkably taken up by the jejunum and ileum. Pharmacokinetics revealed that the maximum plasma concentration (Cmax) and the area under the plasma concentration-time curve from 0 to 24 h (AUC0-24) for CUR@GPP3 micelles were both significantly increased, and that the relative bioavailability of CUR@GPP3 micelles to CUR-loaded mPEG-PLA/TPGS micelles (CUR@PP micelles) was 258.8%. Furthermore, CUR-loaded micelles could reduce damage to the liver and intestinal tissues. This study highlights the importance of Gal content in the design of targeting nanocarrier Gal-modified micelles, which have broad prospects for oral delivery of hydrophobic drugs. Therefore, they could serve as a promising candidate for targeted delivery to the liver.

PEGylated lactoferrin enhances its hepatoprotective effects on acute liver injury induced by D-galactosamine and lipopolysaccharide in rats.

Polyethylene glycol (PEG) is attached to proteins in order to increase their half-life in circulation and reduce their immunogenicity in vivo. The present study was conducted to examine whether two different sizes of PEGylated bovine lactoferrin (40k- and 20k-PEG-bLf) would enhance the protective effect of native bLf on liver injury induced by D-galactosamine (GalN) and lipopolysaccharide (LPS) in rats. The treatment of PEGylated bLf more remarkably prevented the elevation of serum levels of hepatic enzyme markers and inhibited inflammatory and hemorrhagic changes and hepatic apoptosis induced by GalN/LPS than native bLf. The treatment of PEGylated bLf more significantly inhibited the increased concentration of proinflammatory cytokines (TNF-alpha and IL-6) in serum caused by GaIN/LPS, and enhanced anti-inflammatory cytokine (IL-10) production more than native bLf. PEGylated bLf decreased serum levels of nitric oxide (NO) more than native bLf. These results indicate that PEGylated bLf inhibits more significantly the induction of inflammatory mediators such as cytokines and NO than native bLf, resulting in the enhancement of its prevention of fulminant liver failure induced by GalN/LPS in rats. The present study provided evidence that PEGylated bLf may offer a novel alternative therapy for the prevention of acute hepatic failure through its anti-inflammatory and immunomodulatory properties.

In vitro imaging and in vivo liver targeting with carbohydrate capped quantum dots.

PEGylated quantum dots (QDs) capped with d-mannose, d-galactose, and d-galactosamine have been synthesized. The stable, high quantum yield fluorescence of QDs was exploited to study specific carbohydrate-protein interactions in vitro and in vivo.

From polymer to size-defined oligomers: a highly divergent and stereocontrolled construction of chondroitin sulfate A, C, D, E, K, L, and M oligomers from a single precursor: part 2.

An efficient, stereocontrolled, and highly divergent approach for the preparation of oligomers of chondroitin sulfate (CS) A, C, D, E, K, L, and M variants, starting from a single precursor easily obtained by semisynthesis from abundant natural polymer is reported for the first time. Common intermediates were designed that allowed the straightforward construction of O-sulfonated species either on the D-galactosamine unit (CS-A, -C, and -E) or on both D-glucuronic acid and D-galactosamine units (CS-D and CS-K, -L, and -M). This strategy represents a successful improvement and brings a definitive answer toward the synthesis of such complex molecules with numerous relevant biological functions.

Galactosylated PLGA nanoparticles for the oral delivery of resveratrol: enhanced bioavailability and in vitro anti-inflammatory activity.

BACKGROUND: Resveratrol (RES) is a natural anti-inflammatory and antioxidant compound with poor water solubility and oral bioavailability. The present study takes the advantages of nanocarriers combined with a ligand (galactose) anchoring to orally deliver RES in an attempt to improve its bioavailability and pharmacological activity. METHODS: RES-loaded galactosylated nanoparticles (RES-GNPs) were prepared by solvent diffusion technique using poly(lactic-co-glycolic acid), synthesized N-oleoyl-d-galactosamine and Tween 80. RES-GNPs were characterized by particle size, morphology, entrapment efficiency (EE) and in vitro release. Oral bioavailability and in vitro anti-inflammatory activity were investigated in rats and lipopolysaccharides-induced RAW 264.7 cells, respectively. RESULTS: The resulting RES-GNPs were 108.4 nm around in particle size with a polydispersity index of 0.217. Furthermore, RES-GNPs possessed a high EE and a slow drug release in water. After oral administration, RES-GNPs significantly enhanced the oral bioavailability of RES, up to 335.7% relative to RES suspensions. In situ single-pass intestinal perfusion and cellular uptake experiments showed that GNPs could improve the intestinal permeability and transcellular transport of RES. Moreover, the anti-inflammatory efficacy of RES-GNPs in RAW 264.7 cells model was superior to free RES and RES-GNPs. CONCLUSION: The results indicate that RES-GNPs can effectively promote the intestinal absorption of RES and strengthen its bioactivity, which may be a promising system for the treatment of inflammatory diseases.

Macrophage-targeting and reactive oxygen species (ROS)-responsive nanopolyplexes mediate anti-inflammatory siRNA delivery against acute liver failure (ALF).

As one of the intractable challenges in the clinic, the treatment of acute liver failure (ALF) is limited due to high mortality and resource cost. RNA interference (RNAi) provides a new modality for the anti-inflammatory therapy of ALF, while its therapeutic efficacy is greatly hampered by the lack of effective carriers to cooperatively overcome the various systemic barriers. Herein, we developed macrophage-targeting and reactive oxygen species (ROS)-responsive polyplexes to enable efficient systemic delivery of TNF-α siRNA (siTNF-α) to attenuate hepatic inflammation in mice bearing ALF. Se-PEI, obtained from the cross-linking of 600 Da polyethylenimine (PEI) via the ROS-responsive diselenide bond, was developed to condense siTNF-α, and the obtained polyplexes were further coated with carboxylated mannan (Man-COOH). Man-COOH coating allowed active targeting of polyplexes to macrophages with over-expressed mannose receptors (MRs), and it shielded the surface positive charges to enhance the serum stability of polyplexes. Se-PEI could be degraded by ROS in inflammatory macrophages to promote intracellular siRNA release to potentiate the gene knockdown efficiency, and in the meantime reduce the material cytotoxicity associated with high molecular weight. As such, i.v. injected Man-COOH/Se-PEI/siTNF-α polyplexes afforded notable TNF-α silencing by ∼80% in inflamed liver tissues at 500 µg siRNA per kg, and notably reduced serum TNF-α levels to achieve potent anti-inflammatory performance against ALF.

Concentrations of monosaccharides and their amino and alcohol derivatives in human preovulatory follicular fluid.

The study purpose was to compare sugar and polyol concentrations in preovulatory ovarian follicular fluid (FF) with those in the circulation. Samples of FF and peripheral venous blood were obtained after an overnight fast from 14 women attending an IVF program. High performance liquid chromatography measurements of seven polyols, two aminohexoses and four hexoses were the main outcome measures. Glucose concentrations in FF and plasma were 2781.26 +/- 205.64 and 4431.25 +/- 65.17 microM, respectively (P < 0.001). Mannose concentration in FF was 38.99 +/- 3.33 microM, significantly lower than plasma concentration (55.38 +/- 2.29 microM; P < 0.001). A concentration gradient from plasma to FF was also significant for glycerol (99.41 +/- 8.47 versus 74.32 +/- 6.54 microM; P < 0.002), galactose (31.69 +/- 1.58 versus 26.73 +/- 1.93 microM; P < 0.01) and galactosamine (11.49 +/- 0.69 versus 6.38 +/- 0.59 microM; P < 0.001). The plasma-to-FF concentration difference was greatest for glucose (1649.99 +/- 204.09 microM). There was a significant correlation between plasma and FF concentrations for galactose and glycerol. This study supports a substantial utilization of glucose by the oocyte/granulosa cells complex, and documents a significant concentration gradient from plasma to FF for glycerol, mannose, galactose and galactosamine. These plasma-FF differences may reflect both utilization of these carbohydrates by the cells of the preovulatory ovarian follicle and/or transport characteristics of these cells.

Low-fouling and functional poly(carboxybetaine) coating via a photo-crosslinking process.

Antifouling modification technology is developed for many biomedical applications such as blood-contact devices and biosensors. In this work, a photo-reactive polymer containing zwitterionic carboxybetaine groups was prepared by copolymerization of two kinds of methacrylic acids with carboxybetaine and azidoaniline. The carboxybetaine moiety is for low fouling and the azidophenyl moiety is for photo-crosslinking. The synthesized copolymers were coated onto polymeric substrates, and then covalently immobilized on the substrates by exposure to UV radiation. The poly(CBMA-co-AzMA) coating revealed that cell and platelet adhesion and protein adsorption to the substrates were reduced significantly compared to the untreated substrate. Furthermore, the direct immobilization of galactosamine was carried out on the polymer coating by EDC/NHS chemistry. The galactose-immobilized surface had the potential for selecting hepatocyte adhesion from the co-population of different cell types. In addition, the incorporation of photolithographic technology could make micropatterns of poly(CBMA-co-AzMA) coating for the cell co-culture of hepatocytes and fibroblasts. This work demonstrates that the reported technique is an economic and facile tool for layers of reduced adsorption of protein modification with functional groups in one step.

Nano-sized cytochrome P450 3A4 inhibitors to block hepatic metabolism of docetaxel.

Most drugs are metabolized by hepatic cytochrome P450 3A4 (CYP3A4), resulting in their reduced bioavailability. In this study, we present the design and evaluation of bio-compatible nanocarriers trapping a natural CYP3A4-inhibiting compound. Our aim in using nanocarriers was to target the natural CYP3A4-inhibiting agent to hepatic CYP3A4 and leave drug-metabolizing enzymes in other organs undisturbed. In the design of such nanocarriers, we took advantage of the nonspecific accumulation of small nanoparticles in the liver. Specific targeting functionalization was added to direct nanocarriers toward hepatocytes. Nanocarriers were evaluated in vitro for their CYP3A4 inhibition capacity and in vivo for their biodistribution, and finally injected 24 hours prior to the drug docetaxel, for their ability to improve the efficiency of the drug docetaxel. Nanoparticles of poly(lactic-co-glycolic) acid (PLGA) with a hydrodynamic diameter of 63 nm, functionalized with galactosamine, showed efficient in vitro CYP3A4 inhibition and the highest accumulation in hepatocytes. When compared to docetaxel alone, in nude mice bearing the human breast cancer, MDA-MB-231 model, they significantly improved the delay in tumor growth (treated group versus docetaxel alone, percent treated versus control ratio [%T/C] of 32%) and demonstrated a major improvement in overall survival (survival rate of 67% versus 0% at day 55).