Surface, adhesiveness and virulence aspects of Candida haemulonii species complex.

The emerging opportunistic pathogens comprising the Candida haemulonii complex (C. haemulonii [Ch], C. duobushaemulonii [Cd] and C. haemulonii var. vulnera[Chv]) are notable for their intrinsic antifungal resistance. Different clinical manifestations are associated with these fungal infections; however, little is known about their biology and potential virulence attributes. Herein, we evaluated some surface properties of 12 clinical isolates of Ch (n = 5), Cd (n = 4) and Chv (n = 3) as well as their virulence on murine macrophages and Galleria mellonella larvae. Scanning electron microscopy demonstrated the presence of homogeneous populations among the species of the C. haemulonii complex, represented by oval yeasts with surface irregularities able to form aggregates. Cell surface hydrophobicity was isolate-specific, exhibiting high (16.7%), moderate (25.0%) and low (58.3%) hydrophobicity. The isolates had negative surface charge, except for one. Mannose/glucose- and N-acetylglucosamine-containing glycoconjugates were evidenced in considerable amounts in all isolates; however, the surface expression of sialic acid was poorly detected. Cd isolates presented significantly higher amounts of chitin than Ch and Chv. Membrane sterol and lipid bodies, containing neutral lipids, were quite similar among all fungi studied. All isolates adhered to inert surfaces in the order: polystyrene > poly-L-lysine-coated glass > glass. Likewise, they interacted with murine macrophages in a quite similar way. Regarding in vivo virulence, the C. haemulonii species complex were able to kill at least 80% of the larvae after 120 hours. Our results evidenced the ability of C. haemulonii complex to produce potential surface-related virulence attributes, key components that actively participate in the infection process described in Candida spp.

Equisetum giganteum influences the ability of Candida albicans in forming biofilms over the denture acrylic resin surface.

CONTEXT: Equisetum giganteum L. (Equisetaceae) is an endemic plant of Central and South America used in traditional medicine. Natural drugs have been frequently used in the treatment of a myriad of diseases, proving to be an alternative to synthetic chemicals, and have been intensively studied in the prevention of sicknesses, including oral diseases. OBJECTIVE: This study evaluated the in vitro antiadherent activity of E. giganteum extract against Candida albicans biofilms. MATERIALS AND METHODS: Crystal violet and colony-forming units assays were used to quantify the total biofilm biomass and biofilm living cells on a denture base acrylic resin pretreated with hydroethanolic extract of E. giganteum in different concentrations (50, 25, 16, 8, and 4 mg/mL), after 24 h of biofilm development. RESULTS: Equisetum giganteum affected biofilms by reduction of biomass and living cells per area of acrylic specimens. The results revealed reduction of 15-44% of the biofilm mass and reduction of numbers of colony-forming units (CFUs) present in biofilms (79%) compared to the untreated control (CTRL/PBS). At all concentrations, it demonstrated important antiadherent activity on Candida albicans biofilms, the main microbe in denture stomatitis. DISCUSSION AND CONCLUSION: The present findings show that E. giganteum antimicrobial effects may qualify the extract as a promising natural alternative for topical treatment or prevention of denture stomatitis. The usage of drugs made of natural products shows advantages in relation to synthetic drugs on the market, such as lower cost, lower toxicity, and in relation to the occurrence of microbial resistance.

Novel protease inhibitor-loaded Nanoparticle-in-Microparticle Delivery System leads to a dramatic improvement of the oral pharmacokinetics in dogs.

With the advent of the Highly Active Antiretroviral Therapy, the morbidity and the mortality associated to HIV have been considerably reduced. However, 35-40 million people bear the infection worldwide. One of the main causes of therapeutic failure is the frequent administration of several antiretrovirals that results in low patient compliance and treatment cessation. In this work, we have developed an innovative Nanoparticle-in-Microparticle Delivery System (NiMDS) comprised of pure drug nanocrystals of the potent protease inhibitor indinavir free base (used as poorly water-soluble model protease inhibitor) produced by nanoprecipitation that were encapsulated within mucoadhesive polymeric microparticles. Pure drug nanoparticles and microparticles were thoroughly characterized by diverse complementary techniques. NiMDSs displayed an encapsulation efficiency of approximately 100% and a drug loading capacity of up to 43% w/w. In addition, mucoadhesiveness assays ex vivo conducted with bovine gut showed that film-coated microparticles were retained for more than 6 h. Finally, pharmacokinetics studies in mongrel dogs showed a dramatic 47- and 95-fold increase of the drug oral bioavailability and half-life, respectively, with respect to the free unprocessed drug. These results support the outstanding performance of this platform to reduce the dose and the frequency of administration of protease inhibitors, a crucial step to overcome the current patient-incompliant therapy.

In vivo evaluation of a mucoadhesive polymeric caplet for intravaginal anti-HIV-1 delivery and development of a molecular mechanistic model for thermochemical characterization.

CONTEXT AND OBJECTIVE: The aim of this study was to develop, characterize and evaluate a mucoadhesive caplet resulting from a polymeric blend (polymeric caplet) for intravaginal anti-HIV-1 delivery. MATERIALS AND METHODS: Poly(lactic-co-glycolic) acid, ethylcellulose, poly(vinylalcohol), polyacrylic acid and modified polyamide 6, 10 polymers were blended and compressed to a caplet-shaped device, with and without two model drugs 3'-azido-3'-deoxythymidine (AZT) and polystyrene sulfonate (PSS). Thermal analysis, infrared spectroscopy and microscopic analysis were carried out on the caplets employing temperature-modulated DSC (TMDSC), Fourier transform infra-red (FTIR) spectrometer and scanning electron microscope, respectively. In vitro and in vivo drug release analyses as well as the histopathological toxicity studies were carried out on the drug-loaded caplets. Furthermore, molecular mechanics (MM) simulations were carried out on the drug-loaded caplets to corroborate the experimental findings. RESULTS AND DISCUSSION: There was a big deviation between the Tg of the polymeric caplet from the Tg's of the constituent polymers indicating a strong interaction between constituent polymers. FTIR spectroscopy confirmed the presence of specific ionic and non-ionic interactions within the caplet. A controlled near zero-order drug release was obtained for AZT (20 d) and PSS (28 d). In vivo results, i.e. the drug concentration in plasma ranged between 0.012-0.332 mg/mL and 0.009-0.256 mg/mL for AZT and PSS over 1-28 d. CONCLUSION: The obtained results, which were corroborated by MM simulations, attested that the developed system has the potential for effective delivery of anti-HIV-agents.

Synthesis and evaluation of antibacterial polyurethane coatings made from soybean oil functionalized with dimethylphenylammonium iodide and hydroxyl groups.

Preparation of antibacterial polyurethane coatings from novel functional soybean oil was considered in this work. First, epoxidized soybean oil (ESBO) as a low price and widely available renewable resource raw material was subjected to the reaction with aniline using an ionic liquid as a green catalyst. The intermediate phenylamine containing polyol (SAP) was then methylated by reaction with methyl iodide to produce a polyol (QAP) with pendant dimethylphenylammonium iodide groups. To regulate the physical and mechanical properties as well as biological characteristics of final coatings, QAP was mixed with different portions of a similar soybean oil-based polyol (MSP) without quaternary ammonium groups. The mixtures were reacted with isophorone diisocyanate to produce crosslinked polyurethane coatings. Evaluation of viscoelastic properties by DMA method revealed single phase structure with Tg in the range of 50-82°C. Stress-strain analysis of the prepared polyurethanes showed initial modulus, tensile strength, and elongation at break in the ranges of 13-299 MPa, 4.5-13.8 MPa, and 16-109%, respectively. Additionally, the coatings showed good adherence to aluminum and PVC substrates. The solvent extracted samples showed excellent biocompatibility as determined by monitoring L929 fibroblast cells morphology and MTT assay. Meanwhile, very promising antibacterial properties against both Gram-positive (S. aureus) and Gram-negative (E. coli) bacteria with bacterial reduction in the range of 83-100% was observed.

Development of a novel synergistic thermosensitive gel for vaginal candidiasis: an in vitro, in vivo evaluation.

The singular aim of the proposed work is the development of a synergistic thermosensitive gel for vaginal application in subjects prone to recurrent vaginal candidiasis and other microbial infections. The dual loading of Itraconazole and tea tree oil in a single formulation seems promising as it would elaborate the microbial coverage. Despite being low solubility of Itraconazole in tea tree oil, a homogeneous, transparent and stable solution of both was created by co-solvency using chloroform. Complete removal of chloroform was authenticated by GC-MS and the oil solution was used in the development of nanoemulsion which was further translated into a gel bearing thermosensitive properties. In vitro analyses (MTT assay, viscosity measurement, mucoadhesion, ex vivo permeation, etc.) and in vivo studies (bioadhesion, irritation potential and fungal clearance kinetics in rat model) of final formulation were carried out to establish its potential for further clinical evaluation.

Exploiting the synergistic effect of chitosan-EDTA conjugate with MSA for the early recovery from colitis.

The present study was aimed to exploit the antibacterial/antifungal and film coating potential of Chitosan-EDTA (CH-EDTA) conjugate in combination with mesalamine (anti-inflammatory agent) for the early recovery from TNBS induced coilitis. The results suggested CH:EDTA (1:1) spray coated mesalamine tablets has an ability to transport drug in buffer pH 6.8 with rat caecal content condition. The CH-EDTA shows high level of adhesiveness of coat with core tablet. Further, FTIR, DSC and SEM analysis suggested spray coating of CH-EDTA on tablets was beneficial as compared to ladling method as it enhances interaction density and showed resistance from pH (1.2, 6.8 and 7.4). The pharmacokinetic parameters, AUC and AUMC of spray coated tablets were respectively, 4.70 fold and 2.10 fold increased. A synergistic therapeutic effect with CH-EDTA spray coated mesalamine was observed as evaluated by colon/body weight ratio, clinical activity score and damage score. X ray image study supported that CH-EDTA conjugate successfully delivered MSA tablets to large intestine. Histopathology of colon tissues showed rapid recovery from TNBS induced colitis in rats within 4 days. The findings revealed decreased recovery period was due to combined effect of both CH-EDTA and MSA to treat TNBS induced colitis.

Mucoadhesion evaluation of polysaccharide gels for vaginal application by using rheological and indentation measurements.

The influence of hyaluronic acid (HA) and fructooligosaccharides (FOS) addition on low methyl pectin (LMP) gelation has been investigated in order to produce adhesive gel-based microparticles suitable for the development of a vaginal delivery system of pro- and prebiotics. First, dynamic rheological measurements were performed on LMP/Ca(2+) gels with or without FOS and HA in presence or not of porcine stomach mucins. This rheological method is known to translate the interactions between polymer and mucins and then simulate the polymer bioadhesion potential. Nevertheless, as this method is disputed, in vitro and ex vivo indentation test measurements were also achieved in order to correlate the results obtained. Despite some different results, the overall tendency indicates that addition of HA and FOS enhanced the mucoadhesive properties of LMP gels. Moreover, gel-based microparticles obtained according to an emulsification/gelation method and composed by LMP 3% (w/v), FOS 5% (w/v) and HA 0.5% (w/v) displayed a mucoadhesive potential adapted to vaginal delivery system.

In vivo, in vitro evaluation of linseed mucilage based buccal mucoadhesive microspheres of venlafaxine.

The purpose of the present research work was to extract linseed mucilage, use it as a mucoadhesive agent and to develop mucoadhesive microspheres for buccal delivery with an intention to avoid hepatic first-pass metabolism, by enhancing residence time in the buccal cavity. Linseed mucilage was extracted and used to prepare microspheres with varying concentrations of mucilage from formulation F1-F4 (1-2.5%) by spray-drying technique. The microspheres were evaluated for the yield, particle size, incorporation efficiency, swelling property, in vitro mucoadhesion, in vitro drug release, histological study, and stability. Microspheres were characterized by differential scanning colorimetry, scanning electron microscopy, and X-ray diffraction study. Further, the bioavailability study using the New Zealand rabbits was carried out. Formulation F4 showed the maximum mucoadhesion 89.37 ± 1.35%, 92.10 ± 1.37% incorporation efficiency, highest swelling index 0.770 ± 1.23. F4 showed a marked increase in the bioavailability after buccal administration (51.86 ± 3.95) as compared to oral route (39.60 ± 6.16). Also it took less time to reach maximum plasma concentration of 21.38 ± 1.05 ng/ml as compared to oral solution where it required 180 min to reach maximum plasma concentration of 17.98 ± 1.14. It is concluded from the results that linseed mucilage can be used in the production of the mucoadhesive microspheres.

F4H5: a novel substance for the removal of silicone oil from intraocular lenses.

AIM: Adherent silicone oil on intraocular lenses (IOLs) following retinal detachment surgery induces large and irregular refractive errors and multiple images, and gives rise to glare, distorted and often poor vision. Its removal remains challenging, often requiring mechanical wiping or explantation. F4H5 is a new semifluorinated alkane into which silicone oil is readily soluble. The aim is to establish the effectiveness of F4H5 in removing silicone oil from three different types of IOL in vitro. METHOD: Silicone lenses (Tecnis ZM900, Advanced Medical Optics, Inc.), hydrophobic acrylic lenses (MA60, Alcon Laboratories, Inc.) and PMMA lenses (Ocular Vision, Inc) were first immersed in phosphate-buffered saline, second in silicone oil, then in F4H5 (Fluoron GmbH) for 10 min and lastly vigorously agitated in F4H5 for 1 min. They were weighed at each stage using scales accurate to 0.0001 g to measure the weight of the adherent oil. Dynamic contact angle (DCA) analysis was used to assess their surface properties. RESULTS: Immersion in F4H5 alone removed 96.1% (+/-1.23) by weight of silicone oil from the hydrophobic acrylic lenses, 91.4% (+/-1.58) from the silicone and 95.6% (+/-1.44) from the PMMA IOLs. Immersion combined with 1 min of agitation increased the removal to 98.8% (+/-0.46) from the acrylic IOLs, to 93.7% (+/-0.48) from the silicone IOLs and to 100% (within +/-0.0001 g) from every PMMA IOL. After treatment with F4H5, all IOL were optically clear. DCA hysteresis curves remained permanently altered. All measurements were highly reproducible. CONCLUSION: F4H5 was highly effective at removing the bulk of the silicone oil from all three groups of IOL. The DCA measurements suggested that their surface properties were permanently modified.

In vitro inhibitory effect of Streblus asper leaf-extract on adhesion of Candida albicans to human buccal epithelial cells.

This in vitro study aimed at determining the effects of a sublethal concentration of Streblus asper Lour (Moraceae) leaf ethanolic extract on adherence of Candida albicans to human buccal epithelial cells (HBEC). The minimum concentration of Streblus asper leaf ethanolic extract (SAE) that significantly reduced adherence (P<0.05) after a 1-h exposure was 15.6 mg/ml. However, there was a significant reduction (P<0.05) of candidal adhesion to HBEC after 1-min exposure to 125 mg/ml of SAE. Pre-treatment of either Candida or HBEC, or both, with 125 mg/ml of SAE for 1h resulted in reduced adherence. SAE at concentrations of 125 and 250 mg/ml also showed 41 and 61% inhibition of germ tube formation, respectively, which might affect adherence. These findings indicate that the sublethal concentration of SAE may modulate candidal colonization of the oral mucosa thereby suppressing the invasive potential of the pathogen.

Use of phenylboronic acids to investigate boron function in plants. Possible role of boron in transvacuolar cytoplasmic strands and cell-to-wall adhesion.

The only defined physiological role of boron in plants is as a cross-linking molecule involving reversible covalent bonds with cis-diols on either side of borate. Boronic acids, which form the same reversible bonds with cis-diols but cannot cross-link two molecules, were used to selectively disrupt boron function in plants. In cultured tobacco (Nicotiana tabacum cv BY-2) cells, addition of boronic acids caused the disruption of cytoplasmic strands and cell-to-cell wall detachment. The effect of the boronic acids could be relieved by the addition of boron-complexing sugars and was proportional to the boronic acid-binding strength of the sugar. Experiments with germinating petunia (Petunia hybrida) pollen and boronate-affinity chromatography showed that boronic acids and boron compete for the same binding sites. The boronic acids appear to specifically disrupt or prevent borate-dependent cross-links important for the structural integrity of the cell, including the organization of transvacuolar cytoplasmic strands. Boron likely plays a structural role in the plant cytoskeleton. We conclude that boronic acids can be used to rapidly and reversibly induce boron deficiency-like responses and therefore are useful tools for investigating boron function in plants.

Extract from Herniaria hirsuta coats calcium oxalate monohydrate crystals and blocks their adhesion to renal epithelial cells.

PURPOSE: The interaction of calcium oxalate crystals with renal epithelial cells is a critical event in kidney stone formation. In this study we assessed the effect of aqueous extract from Herniaria hirsuta on the adhesion of calcium oxalate monohydrate (COM) crystals to cultured renal cells. MATERIALS AND METHODS: Madin Darby canine kidney cells were used as a model for studying the adhesion of radioactive COM crystals in the presence and absence of plant extract. RESULTS: COM crystal binding to cells was inhibited by extract in a concentration dependent manner. Prior exposure of crystals but not cells to extract blocked crystal binding, suggesting that plant molecules can coat and exert their effect at the crystal surface. Crystal attachment appeared related to membrane fluidity since crystal adhesion increased at higher vs lower temperatures (37C vs 0C) and Herniaria extract altered crystal adhesion only under conditions of increased fluidity (increased temperature). Extract also displaced a significant portion of prebound crystals without apparent effects on cell function or the morphology of preexisting calcium oxalate crystals. Herniaria extract exerted no adverse or toxic effect on cells, which proliferated normally in its presence even at relatively high concentrations. CONCLUSIONS: Our current data suggest a mechanism whereby Herniaria hirsuta extract used in traditional medicine might prevent and possibly eliminate preexisting kidney stones. Further characterization of the active compound(s) could identify a new candidate drug for patients with nephrolithiasis.

Nitric oxide inhalation decreases pulmonary platelet and neutrophil sequestration during extracorporeal circulation in the pig.

OBJECTIVE: The inhibiting effect of nitric oxide on the aggregation and adhesion of neutrophils and platelets has been well documented in vitro. In vivo evidence, however, is more scant. In this study, we studied the effects of inhaled nitric oxide on pulmonary cellular sequestration in our sham hemodialysis model. Accumulation of neutrophils and platelets in the lungs has been shown to be an early event in this model. DESIGN: Prospective, randomized, controlled study. SETTING: Animal laboratory at a university medical center. SUBJECTS: Twenty-six anesthetized, mechanically ventilated pigs. INTERVENTIONS: 111Indium-oxine was used to selectively label neutrophils or platelets and the activity over the lungs was followed dynamically with a gamma camera. Sham hemodialysis, using a cuprophan hollow-fiber dialyzer, was instituted via catheters in the femoral vessels. The animals were divided into two main groups: a) the nitric oxide recipient group (n = 12, with platelets labeled in seven animals and neutrophils labeled in five animals); and b) the control group (n = 14, with platelets labeled in seven animals and neutrophils labeled in seven animals). The animals in the former group were given 50 parts per million of nitric oxide in the inspiratory gas from the beginning of dialysis and for 30 mins onward. MEASUREMENTS AND MAIN RESULTS: Inhalation of nitric oxide attenuated the increase in activity over the lungs in both the neutrophil and platelet groups during sham hemodialysis. In addition, an inhibiting effect on the increase in pulmonary pressure was noted. CONCLUSION: Apart from the effects of nitric oxide on central hemodynamics in this model, the scintigraphic findings indicate an in vivo effect of nitric oxide on the accumulation of platelets and neutrophils in the lungs, probably due to inhibition of the adhesion and/or aggregation of these cells.