Trichomonas vaginalis Motility Is Blocked by Drug-Free Thermosensitive Hydrogel.

Trichomonas vaginalis motility in biological fluids plays a prominent, but understudied, role in parasite infectivity. In this study, the ability of a thermosensitive hydrogel (pluronic F127) to physically immobilize T. vaginalis was investigated. Blocking parasite motility could prevent its attachment to the mucosa, thus reducing the acquisition of the infection. The trajectory of individual parasites was monitored by multiple particle tracking. Mean square displacement, diffusivity, and velocity were calculated from x, y coordinates during time. Major results are that T. vaginalis exhibited different types of trajectories in a diluted solution composed of lactate buffer similar to "run-and-tumble" motion reported for flagellated bacteria. The fastest T. vaginalis specimen moves with a velocity of 19 µm/s. Observation of T. vaginalis movements showed that the cell body remains rigid during swimming and that the propulsive forces necessary to generate the movement are the result of flagellar beating. Parasite motility was partially slowed down using hydroxyethylcellulose hydrogel, used as a reference for the development of vaginal microbicides, while 100% of T. vaginalis were immobile in F127 hydrogel. Once completed by biological investigations on mice, this report suggests using drug-free formulation composed of F127 as a new strategy to prevent T. vaginalis attachment to the mucosa. The concept will be extended to other flagellated organisms where the motility is driven by cilia and flagella.

Topically Applied Chitosan-Coated Poly(isobutylcyanoacrylate) Nanoparticles Are Active Against Cutaneous Leishmaniasis by Accelerating Lesion Healing and Reducing the Parasitic Load.

Parenteral administration of amphotericin B-deoxycholate (AmB-DOC) or pentavalent antimonials to cure cutaneous leishmaniasis (CL) results in severe adverse reactions, while topically applied antileishmanial drugs are ineffective despite their good tolerance. This work is aimed to investigate whether poly(isobutylcyanoacrylate) nanoparticles coated with chitosan (Cs-NPs) could provide intrinsic antileishmanial activity after topical application. In vitro evaluations revealed that nanoparticles were active against the promastigote, axenic amastigote, and intramacrophage forms of Leishmania major. In vivo evaluations after repetitive topical applications on the skin of mice infected with L. major showed that Cs-NPs combined or not with AmB-DOC allowed partial healing of the lesion characterized by histological analyses. The parasitic load of skin specimens collected from mice was significantly reduced compared with that from nontreated mice, as analyzed by quantitative polymerase chain reaction (q-PCR). Ultrastructure characterizations by electron microscopy of L. major promastigotes after incubation with Cs-NPs showed morphological alterations, including aberrant shape and swelling of mitochondria and parasitic vacuoles.

Surface-dependent endocytosis of poly(isobutylcyanoacrylate) nanoparticles by Trichomonas vaginalis.

Previous data from our research group showed that chitosan-coated poly(isobutylcyanoacrylate) nanoparticles (NPs) (denoted PIBCA/Chito20) exhibited intrinsic anti-Trichomonas vaginalis activity, while PIBCA/pluronic® F68 without chitosan (PIBCA/F68) were inactive. However, the mechanism of anti-T. vaginalis activity of chitosan-coated PIBCA NPs is still unknown. Our hypothesis is that chitosan-coated NPs are internalized by the parasite, contrarily to PIBCA/F68. In this investigation, the impact of NP surface on their internalization by the protozoan was studied using flow cytometry and parasite morphological changes after different incubation times with PIBCA/Chito20 NPs were monitored by electron microscopy. Flow-cytometry revealed that PIBCA/Chito20 NPs were uptaken by T. vaginalis as early as 10-min-incubation. Drastic cell morphological transformations were observed from scanning electron microscopy and transmission electron microscopy after incubation with PIBCA/Chito20 NPs. Numerous pits were seen on cell membrane since 10 min. Gradual increase in contact time increased NP endocytosis and induced proportional damages to T. vaginalis membrane. Then, investigation of whether PIBCA/Chito20 NPs can improve MTZ anti-T. vaginalis activity was studied using checkerboard experiment. Calculation of fractional inhibitory concentration index (FICI = 3.53) showed an additive effect between NPs and MTZ.

New insights on the structure of hexagonally faceted platelets from hydrophobically modified chitosan and α-cyclodextrin.

A new class of non-spherical particles was recently designed in our research group by mixing a polysaccharide grafted with fatty acids and α-cyclodextrin in water. Because their flat surfaces, and according to their size, particles are called micro- or nano-platelets. Here, we varied the composition of fatty acids grafted on chitosan (oleic acid, palmitic acid or stearic acid) and characterized platelet morphology. Transmission electron microscopy (TEM), cryogenic TEM, scanning electron microscopy, atomic force microscopy (AFM) and confocal laser scanning microscopy experiments showed that the platelets have a preferentially hexagonal shape with sharp edges, independently on alkyl chain grafted on chitosan. Furthermore, AFM topographic analysis of platelet surface showed parallel thin terraces with 12-14-nm height, suggesting a multi-layered structure alternating chitosan and fatty-acid/α-cyclodextrin inclusion complexes. We also revealed for the first time that a simple magnetic mixing of fatty acids with α-cyclodextrin in water results from solid inclusion complexes with a crystalline structural organization characterized by powder X-ray diffraction. Our results demonstrate that fatty acid/α-cyclodextrin interaction is the driving force for platelet formation.

Phase solubility studies and anti-Trichomonas vaginalis activity evaluations of metronidazole and methylated ß-cyclodextrin complexes: Comparison of CRYSMEB and RAMEB.

Metronidazole (MTZ) is a 5-nitroimidazole drug used for the treatment of Trichomonas vaginalis parasitic infection. Aqueous formulations containing MTZ are restricted because apparent solubility in water of this drug is low. In this context, two methylated-ß-cyclodextrins (CRYSMEB and RAMEB) were used as a tool to increase apparent solubility of MTZ in water. CRYSMEB was limited by its own solubility in water (15% w/w, 12.59 mM), while RAMEB at a concentration of 40% w/w (300.44 mM) allowed a maximal increase of apparent solubility of MTZ (3.426% w/w, 200.19 mM). From our knowledge, this corresponds to the highest enhancement of MTZ apparent aqueous solubility ever reported in the literature using methylated cyclodextrins. In vitro evaluations showed that anti-T. vaginalis activity of MTZ formulated with CRYSMEB and RAMEB was preserved.

In situ forming pluronic® F127/chitosan hydrogel limits metronidazole transmucosal absorption.

The objective of this work is to design topically-applied thermosensitive and mucoadhesive hydrogel containing metronidazole (MTZ) for the treatment of Trichomonas vaginalis infections. Hydrogel composed of pluronic® F127 (20wt%), chitosan (1wt%) and metronidazole MTZ (0.7wt%) mixture showed its ability to decrease by a factor 4 MTZ flux and apparent permeability absorption through vaginal mucosa. The impact of hydrogel on transmucosal penetration of MTZ was evaluated ex vivo on excised porcine vaginal mucosa mounted on Franz diffusion cell. The anti-T. vaginalis activity of MTZ formulated into F127/chitosan hydrogel was preserved since the viability curve evaluated in vitro was similar to MTZ solution.