Wijit Banlunara

Mucoadhesive curcumin nanospheres: Biological activity, adhesion to stomach mucosa and release of curcumin into the circulation

Although mucoadhesive drug carriers for the gastro-intestinal tract (GIT) have been reported, the mucoadhesive property and drug release characteristics have never been evaluated separately, whilst the adherence of the carriers to the surface of GIT has not been directly visualized. Here, a monopolymeric carrier made from ethylcellulose (EC) and a dipolymeric carrier made from a blend of methylcellulose (MC) and EC (ECMC) were easily fabricated through a self-assembling process and yielded the highest reported curcumin loading of ~48-49%. Both curcumin loaded ECMC (C-ECMC) and curcumin loaded EC (C-EC) particles showed an in vitro free radical scavenging activity and a dose-dependent in vitro cytotoxic effect towards MCF-7 human breast adenocarcinoma and HepG2 hepatoblastoma cells in tissue culture. The in vivo evaluation of their adherence to stomach mucosa and their ability to release curcumin into the circulation were carried out through quantification of curcumin levels in the stomach tissue and in blood of mice orally administered with the two spheres. Direct evidence of the adherence of the C-EC and C-ECMC particles along the mucosal epithelia of the stomach is also presented for the first time through SEM images. The mucoadhesive property of self-assembled C-EC nanoparticles is discussed.

Acemannan sponges stimulate alveolar bone, cementum and periodontal ligament regeneration in a canine class II furcation defect model

BACKGROUND AND OBJECTIVE Periodontal disease is a common infectious disease, found worldwide, causing the destruction of the periodontium. The periodontium is a complex structure composed of both soft and hard tissues, thus an agent applied to regenerate the periodontium must be able to stimulate periodontal ligament, cementum and alveolar bone regeneration. Recent studies demonstrated that acemannan, a polysaccharide extracted from Aloe vera gel, stimulated both soft and hard tissue healing. This study investigated effect of acemannan as a bioactive molecule and scaffold for periodontal tissue regeneration. MATERIAL AND METHODS Primary human periodontal ligament cells were treated with acemannan in vitro. New DNA synthesis, expression of growth/differentiation factor 5 and runt-related transcription factor 2, expression of vascular endothelial growth factor, bone morphogenetic protein-2 and type I collagen, alkaline phosphatase activity, and mineralized nodule formation were determined using [(3)H]-thymidine incorporation, reverse transcription-polymerase chain reaction, enzyme-linked immunoabsorbent assay, biochemical assay and alizarin red staining, respectively. In our in vivo study, premolar class II furcation defects were made in four mongrel dogs. Acemannan sponges were applied into the defects. Untreated defects were used as a negative control group. The amount of new bone, cementum and periodontal ligament formation were evaluated 30 and 60 d after the operation. RESULTS Acemannan significantly increased periodontal ligament cell proliferation, upregulation of growth/differentiation factor 5, runt-related transcription factor 2, vascular endothelial growth factor, bone morphogenetic protein 2, type I collagen and alkaline phosphatase activity, and mineral deposition as compared with the untreated control group in vitro. Moreover, acemannan significantly accelerated new alveolar bone, cementum and periodontal ligament formation in class II furcation defects. CONCLUSION Our data suggest that acemannan could be a candidate biomolecule for periodontal tissue regeneration.

Molecular Epidemiology Reveals Genetic Diversity amongst Isolates of the Cryptococcus neoformans/C. gattii Species Complex in Thailand

To gain a more detailed picture of cryptococcosis in Thailand, a retrospective study of 498 C. neoformans and C. gattii isolates has been conducted. Among these, 386, 83 and 29 strains were from clinical, environmental and veterinary sources, respectively. A total of 485 C. neoformans and 13 C. gattii strains were studied. The majority of the strains (68.9%) were isolated from males (mean age of 37.97 years), 88.5% of C. neoformans and only 37.5% of C. gattii strains were from HIV patients. URA5-RFLP and/or M13 PCR-fingerprinting analysis revealed that the majority of the isolates were C. neoformans molecular type VNI regardless of their sources (94.8%; 94.6% of the clinical, 98.8% of the environmental and 86.2% of the veterinary isolates). In addition, the molecular types VNII (2.4%; 66.7% of the clinical and 33.3% of the veterinary isolates), VNIV (0.2%; 100% environmental isolate), VGI (0.2%; 100% clinical isolate) and VGII (2.4%; 100% clinical isolates) were found less frequently. Multilocus Sequence Type (MLST) analysis using the ISHAM consensus MLST scheme for the C. neoformans/C. gattii species complex identified a total of 20 sequence types (ST) in Thailand combining current and previous data. The Thai isolates are an integrated part of the global cryptococcal population genetic structure, with ST30 for C. gattii and ST82, ST83, ST137, ST141, ST172 and ST173 for C. neoformans being unique to Thailand. Most of the C. gattii isolates were ST7 = VGIIb, which is identical to the less virulent minor Vancouver island outbreak genotype, indicating Thailand as a stepping stone in the global spread of this outbreak strain. The current study revealed a greater genetic diversity and a wider range of major molecular types being present amongst Thai cryptococcal isolates than previously reported.

Bringing Macromolecules into Cells and Evading Endosomes by Oxidized Carbon Nanoparticles

A great challenge exists in finding safe, simple, and effective delivery strategies to bring matters across cell membrane. Popular methods such as viral vectors, positively charged particles and cell penetrating peptides possess some of the following drawbacks: safety issues, lysosome trapping, limited loading capacity, and toxicity, whereas electroporation produces severe damages on both cargoes and cells. Here, we show that a serendipitously discovered, relatively nontoxic, water dispersible, stable, negatively charged, oxidized carbon nanoparticle, prepared from graphite, could deliver macromolecules into cells, without getting trapped in a lysosome. The ability of the particles to induce transient pores on lipid bilayer membranes of cell-sized liposomes was demonstrated. Delivering 12-base-long pyrrolidinyl peptide nucleic acids with d-prolyl-(1S,2S)-2-aminocyclopentanecarboxylic acid backbone (acpcPNA) complementary to the antisense strand of the NF-κB binding site in the promoter region of the Il6 gene into the macrophage cell line, RAW 264.7, by our particles resulted in an obvious accumulation of the acpcPNAs in the nucleus and decreased Il6 mRNA and IL-6 protein levels upon stimulation. We anticipate this work to be a starting point in a new drug delivery strategy, which involves the nanoparticle that can induce a transient pore on the lipid bilayer membrane.

Silicone surface with drug nanodepots for medical devices.

An ideal surface of poly(dimethylsiloxane) (PDMS) medical devices requires sustained drug release to combat various tissue responses and infection. At present, a noncovalent surface coating with drug molecules using binders possesses a detachment problem, while covalently linking drug molecules to the surface provides no releasable drug. Here, a platform that allows the deposition of diverse drugs onto the PDMS surface in an adequate quantity with reliable attachment and a sustained-release character is demonstrated. First, a PDMS surface with carboxyl functionality (PDMS-COOH) is generated by subjecting a PDMS piece to an oxygen plasma treatment to obtain silanol moieties on its surface, then condensing the silanols with (3-aminopropyl)triethoxysilane molecules to generate amino groups, and finally reacting the amino groups with succinic anhydride. The drug-loaded carriers with hydroxyl groups on their surface can then be esterified to PDMS-COOH, resulting in a PDMS surface covalently grafted with drug-filled nanocarriers so that the drugs inside the securely grafted carriers can be released. Demonstrated here is the covalent linking of the surface of a PDMS endotracheal tube with budesonide-loaded ethylcellulose nanoparticles. A secure and high drug accumulation at the surface of the tubes (0.025 mg/cm2) can be achieved without changes in its bulk property such as hardness (Shore-A), and sustained release of budesonide with a high release flux during the first week followed by a reduced release flux over the subsequent 3 weeks can be obtained. In addition, the grafted tube possesses more hydrophilic surface and thus is more tissue-compatible. The grafted PDMS pieces show a reduced in vitro inflammation in cell culture and a lower level of in vivo tissue responses, including a reduced level of inflammation, compared to the unmodified PDMS pieces, when implanted in rats. Although demonstrated with budesonide and a PDMS endotracheal tube, this platform of grafting a PDMS surface with drug-loaded particles can be applied to other drugs and other devices.

Apoptosis and gene expression in the developing mouse brain of fusarenon-X-treated pregnant mice.

Fusarenon-X (FX), a type B trichothecene mycotoxin, is mainly produced by Fusarium crookwellense, which occurs naturally in agricultural commodities, such as wheat and barley. FX has been shown to exert a variety of toxic effects on multiple targets in vitro. However, the embryonic toxicity of FX in vivo remains unclear. In the present study, we investigated FX-induced apoptosis and the relationship between the genetic regulatory mechanisms and FX-induced apoptosis in the developing mouse brain of FX-treated pregnant mice. Pregnant mice were orally administered FX (3.5 mg/kg b.w.) and were assessed at 0, 12, 24 and 48 h after treatment (HAT). Apoptosis in the fetal brain was determined using hematoxylin and eosin staining, the TUNEL method, immunohistochemistry for PCNA and electron microscopy. Gene expressions were evaluated using microarray and real time-reverse transcription polymerase chain reaction (qRT-PCR). Histopathological changes showed that the number of apoptotic cells in the telencephalon of the mouse fetus peaked at 12 HAT and decreased at 24 and 48 HAT. FX induced the up-regulation of Bax, Trp53 and Casp9 and down-regulated Bcl2 but the expression levels of Fas and Casp8 mRNA remained unchanged. These data suggested that FX induces apoptosis in the developing mouse brain in FX-treated dams. Moreover, the genetic regulatory mechanisms of FX-induced apoptosis are regulated by Bax, Bcl2, Trp53 and Casp9 or can be defined via an intrinsic apoptotic pathway.

Combating Helicobacter pylori infections with mucoadhesive nanoparticles loaded with Garcinia mangostana extract

AIM To combat the resistance of Helicobacter pylori to antibiotics through the use of Garcinia mangostana extract (GME) in the form that can be localized at stomach mucosa. MATERIALS & METHODS GME and its major active component, α-mangostin, are encapsulated into the moderately acid stable mucoadhesive nanocarriers, and tested for anti-H. pylori and antiadhesion activities in vitro and their ability to eradicate H. pylori in infected mice. RESULTS The two in vitro activities are observed and are enhanced when the materials are encapsulated into nanocarriers. Preliminary in vivo tests revealed the ability to combat H. pylori in mice following oral administration of the encapsulated GME, but not the unencapsulated GME. CONCLUSION Nanoencapsulated GME is a potential anti-H. pylori agent.

Ethyl cellulose nanoparticles: clarithomycin encapsulation and eradication of H. pylori.

The extreme acidic environment of the stomach, its regular voidance of contents and the restricted access to the mucus covered habitat combined with the antibiotic resistance of the bacteria, all contribute to the poor success in the treatment of Helicobacter pylori gastric infections. Here, we demonstrate that by encapsulating clarithromycin into ethyl cellulose (EC) nanoparticles, the efficiency of H. pylori clearance in C57BL/6 mice infected with these bacteria was significantly improved. Clarithomycin-loaded EC nanoparticles were prepared via a simple yet effective anti-solvent particle induction method, to yield sub-micron sized particles with 22.3 ± 0.17% (w/w) clarithromycin loading at 86 ± 0.5% (w/w) encapsulation efficiency. The particles dispersed well in water and simulated gastric fluid and gave a minimum inhibitory concentration of 0.09-0.18 μg/ml against four strains of H. pylori. Encapsulation into EC particles not only enhanced the anti-adhesion activity of clarithromycin when tested with H. pylori and Hep-2 cells, but also gave significant enhancement of H. pylori clearance in the stomach of C57BL/6 mice infected with the bacteria.

Urinary bladder wall substitution using autologous tunica vaginalis in male dogs

Fresh autologous tunica vaginalis was experimentally used for partial substitution of the excised urinary bladder wall in ten male mongrel dogs. The substituted areas of two dogs were examined macroscopically and histologically at 2, 4, 6, 8, and 10 weeks after surgery. Two control dogs underwent partial cystectomy and primary wall closure without substitution. The regenerated transitional epithelium completely covered the substituted portion and smooth muscle regeneration was present at 6 weeks. The bladder walls at the closure area of one control dog and at the substituted portions of two dogs at 10 weeks were indistinguishable macroscopically from the native bladder with all layers of the bladder wall present histologically. According to the macroscopic and histological findings and simplicity of the technique, tunica vaginalis can be used as an alternative graft for bladder wall substitution. Calcification and bone metaplasia observed were similar to those found after using other tissue grafts.

Acrylate-Tethering Drug Carrier: Covalently Linking Carrier to Biological Surface and Application in the Treatment of Helicobacter pylori Infection

The development of carriers to sustain drugs at stomach surface is an attractive strategy to increase drug bioavailability locally and systematically. So far, the only reported carrier that can form a covalent bond with mucus, the thiolated carrier, relies on a reversible disulfide exchange reaction between thiols on the carrier and disulfide bridges on the mucus. Here we show the design and fabrication of a cellulose carrier with tethering acrylate groups (denoted here as clickable carrier) that, under a nontoxic condition, can efficiently react with thiols on biomaterials in situ through the thermodynamically driven and kinetically probable Michael thiol-ene click reaction. Here we show the attachments of the clickable carriers to a mucin protein, a surface of human laryngeal carcinoma cells, and a surface of a fresh porcine stomach. We also show that the required thiol moieties can be generated in situ by reducing existing cystine disulfide bridges with either the edible vitamin C or the relatively nontoxic tris(2-carboxyethyl) phosphine. Comparing to a control carrier, the clickable carrier can increase some drug concentrations in an ex vivo stomach tissue, and improve the Helicobacter pylori treatment in infected C57BL/6 mice.