PDADMAC/Alginate-Coated Gold Nanorod For Eradication of Staphylococcus Aureus Biofilms.

Introduction: Over 75% of clinical microbiological infections are caused by bacterial biofilms that grow on wounds or implantable medical devices. This work describes the development of a new poly(diallyldimethylammonium chloride) (PDADMAC)/alginate-coated gold nanorod (GNR/Alg/PDADMAC) that effectively disintegrates the biofilms of Staphylococcus aureus (S. aureus), a prominent pathogen responsible for hospital-acquired infections. Methods: GNR was synthesised via seed-mediated growth method, and the resulting nanoparticles were coated first with Alg and then PDADMAC. FTIR, zeta potential, transmission electron microscopy, and UV-Vis spectrophotometry analysis were performed to characterise the nanoparticles. The efficacy and speed of the non-coated GNR and GNR/Alg/PDADMAC in disintegrating S. aureus-preformed biofilms, as well as their in vitro biocompatibility (L929 murine fibroblast) were then studied. Results: The synthesised GNR/Alg/PDADMAC (mean length: 55.71 ± 1.15 nm, mean width: 23.70 ± 1.13 nm, aspect ratio: 2.35) was biocompatible and potent in eradicating preformed biofilms of methicillin-resistant (MRSA) and methicillin-susceptible S. aureus (MSSA) when compared to triclosan, an antiseptic used for disinfecting S. aureus colonisation on abiotic surfaces in the hospital. The minimum biofilm eradication concentrations of GNR/Alg/PDADMAC (MBEC50 for MRSA biofilm = 0.029 nM; MBEC50 for MSSA biofilm = 0.032 nM) were significantly lower than those of triclosan (MBEC50 for MRSA biofilm = 10,784 nM; MBEC50 for MRSA biofilm 5967 nM). Moreover, GNR/Alg/PDADMAC was effective in eradicating 50% of MRSA and MSSA biofilms within 17 min when used at a low concentration (0.15 nM), similar to triclosan at a much higher concentration (50 µM). Disintegration of MRSA and MSSA biofilms was confirmed by field emission scanning electron microscopy and confocal laser scanning microscopy. Conclusion: These findings support the potential application of GNR/Alg/PDADMAC as an alternative agent to conventional antiseptics and antibiotics for the eradication of medically important MRSA and MSSA biofilms.

Development of Nanostructured Lipid Carrier-Loaded Flavonoid-Enriched Zingiber officinale.

Flavonoids, which are bioactive molecules found in Zingiber officinale, have been widely used as antioxidant and anti-inflammatory drugs. The presence of nanostructured lipid carriers (NLCs) as sophisticated delivery systems for bioactive compounds, such as flavonoids, can increase their bioavailability and stability, thus potentially producing better therapeutic effects. This study aimed to develop an anti-inflammatory topical gel using NLC-containing flavonoids derived from Zingiber officinale. The NLC formulation was prepared using stearic acid, a mixture of medium-chain triglycerides and isopropyl myristate, Tween 20, and Span 20 by using a hot homogenization method. The total flavonoid content obtained through sequential maceration stages was 4.04 mg of QUE/g of dry extract. The highest encapsulation efficiency of flavonoid-loaded NLC was observed at a flavonoid, Zingiber officinale extract (ZOE) concentration of 2%. It was found that a ZOE concentration of 0.4% provided excellent stability with a particle size of 302-344 nm and a polydispersity index of 0.14-0.23 after 28 days of observation. Morphological analysis of the ZOE-loaded NLC revealed a stable and well-developed formulation with a fairly uniform distribution. The presence of distinctive and uniformly distributed single particles suggests a promising alternative drug delivery system for conventional topical preparations. ZOE-loaded NLC gel showed solid-like properties and higher quality stability than the gel.

Physicochemical Properties and Release Study of Antimetabolite-Incorporated Stearoyl Chitosan.

Stearoyl chitosan (SC), derived from the acylation of chitosan, contributes to the efficiency of drug delivery systems because of its structure, which accommodates the drug in a particle. Nonetheless, its role in chemotherapy has been largely unexplored. The present study involves the synthesis of stearoyl chitosan through the reaction of depolymerized chitosan with stearoyl chloride under mild reaction conditions. The resulting compound was subjected to structural analysis utilizing Fourier-transform infrared (FTIR) spectroscopy, 1H NMR, and X-ray diffraction (XRD) spectroscopy. The dispersion of SC molecules in phosphate-buffered saline (PBS) forms SC nanoparticles. The best dispersion of SC in the solution was achieved at a 1:60 chitosan-to-stearoyl chloride weight ratio. Three antimetabolite drugs, methotrexate, pemetrexed, and raltitrexed, were selected to examine the loading efficacy of SC. Pemetrexed had the highest drug-loading value of 36.8% among the three antimetabolites incorporated into SC, along with an encapsulation efficiency of 85.1%. The size of SC loaded with antimetabolites ranged from 225 to 369 nm, and their spherical form was verified via a transmission electron microscope. The in vitro release study showed that SC demonstrated controlled drug release, suggesting that SC nanoparticles have significant promise as a delivery strategy for chemotherapy.

Cholesterol-linoleic acid liposomes induced extracellular vesicles secretion from immortalized adipose-derived mesenchymal stem cells for in vitro cell migration.

Extracellular vesicles (EVs) are small vesicles that are naturally released by cells and play a crucial role in cell-to-cell communication, tissue repair and regeneration. As naturally secreted EVs are limited, liposomes with different physicochemical properties, such as 1,2-dioleoyl-3-trimethylammonium propane (DOTAP) and linoleic acid (LA) with modifications have been formulated to improve EVs secretion for in vitro wound healing. Various analyses, including dynamic light scattering (DLS) and transmission electron microscopy (TEM) were performed to monitor the successful preparation of different types of liposomes. The results showed that cholesterol-LA liposomes significantly improved the secretion of EVs from immortalized adipose-derived mesenchymal stem cells (AD-MSCs) by 1.5-fold. Based on the cell migration effects obtained from scratch assay, both LA liposomal-induced EVs and cholesterol-LA liposomal-induced EVs significantly enhanced the migration of human keratinocytes (HaCaT) cell line. These findings suggested that LA and cholesterol-LA liposomes that enhance EVs secretion are potentially useful and can be extended for various tissue regeneration applications.

ROS-generating alginate-coated gold nanorods as biocompatible nanosonosensitisers for effective sonodynamic therapy of cancer.

Sonodynamic therapy (SDT) emerges as a promising non-invasive alternative for eradicating malignant tumours. However, its therapeutic efficacy remains limited due to the lack of sonosensitisers with high potency and biosafety. Previously, gold nanorods (AuNRs) have been extensively studied for their applications in photodynamic or photothermal cancer therapy, but their sonosensitising properties are largely unexplored. Here, we reported the applicability of alginate-coated AuNRs (AuNRsALG) with improved biocompatibility profiles as promising nanosonosensitisers for SDT for the first time. AuNRsALG were found stable under ultrasound irradiation (1.0 W/cm2, 5 min) and maintained structural integrity for 3 cycles of irradiation. The exposure of the AuNRsALG to ultrasound irradiation (1.0 W/cm2, 5 min) was shown to enhance the cavitation effect significantly and generate a 3 to 8-fold higher amount of singlet oxygen (1O2) than other reported commercial titanium dioxide nanosonosensitisers. AuNRsALG exerted dose-dependent sonotoxicity on human MDA-MB-231 breast cancer cells in vitro, with âˆ¼ 81% cancer cell killing efficacy at a sub-nanomolar level (IC50 was 0.68 nM) predominantly through apoptosis. The protein expression analysis showed significant DNA damage and downregulation of anti-apoptotic Bcl-2, suggesting AuNRsALG induced cell death through the mitochondrial pathway. The addition of mannitol, a reactive oxygen species (ROS) scavenger, inhibited cancer-killing effect of AuNRsALG-mediated SDT, further verifying that the sonotoxicity of AuNRsALG is driven by the production of ROS. Overall, these results highlight the potential application of AuNRsALG as an effective nanosonosensitising agent in clinical settings.

Bispecific antibodies for targeted delivery of anti-cancer therapeutic agents: A review.

Monospecific antibodies have been utilised increasingly for anti-cancer drug targeting owing to their ability to minimise off-target toxicity by binding specifically to a tumour epitope, hence selectively delivering drugs to the tumour cells. Nevertheless, the monospecific antibodies only engage a single cell surface epitope to deliver their drug payload. Hence, their performance is often unsatisfactory in cancers where multiple epitopes need to be engaged for optimal cellular internalisation. In this context, bispecific antibodies (bsAbs) that simultaneously target two distinct antigens or two distinct epitopes of the same antigen offer a promising alternative in antibody-based drug delivery. This review describes the recent advances in developing bsAb-based drug delivery strategies, encompassing the direct conjugation of drug to bsAbs to form bispecific antibody-drug conjugates (bsADCs) and the surface functionalisation of nanoconstructs with bsAbs to form bsAb-coupled nanoconstructs. The article first details the roles of bsAbs in enhancing the internalisation and intracellular trafficking of bsADCs with subsequent release of chemotherapeutic drugs for an augmented therapeutic efficacy, particularly among heterogeneous tumour cell populations. Then, the article discusses the roles of bsAbs in facilitating the delivery of drug-encapsulating nanoconstructs, including organic/inorganic nanoparticles and large bacteria-derived minicells, that provide a larger drug loading capacity and better stability in blood circulation than bsADCs. The limitations of each type of bsAb-based drug delivery strategy and the future prospects of more versatile strategies (e.g., trispecific antibodies, autonomous drug delivery systems, theranostics) are also elaborated.

Synthesis of N-acyl glycine surfactant from palm oil as green repellent and toxicant to termite (Microcerotermes diversus).

This study described for the first time, the synthesis of a greener, safer, and more effective termiticide using a bio-based surfactant, N-acyl glycine derived from palm oil for the control of Microcerotermes diversus. Laboratory findings showed that the highest repellent activity was observed in N-acyl glycine surfactant (83.33%) at 50 ppm. In addition, N-acyl glycine surfactant also exhibited substantial time and concentration-dependent anti-termiticidal activity in which the highest termite mortality was observed after 3 days of exposure at 50 ppm of the surfactant (100%). Furthermore, 32.49 ppm concentration of N-acyl glycine surfactant (LC50 = 32.49 ppm) attained 50% of termite lethality. The current innovated termiticide with the use of N-acyl glycine surfactant offers a better efficacy, lower cost, and prevents the use of dangerous termiticides that are critical in creating a more sustainable environment, and controls Microcerotermes diversus at the same time.

Nanoscaled PAMAM Dendrimer Spacer Improved the Photothermal-Photodynamic Treatment Efficiency of Photosensitizer-Decorated Confeito-Like Gold Nanoparticles for Cancer Therapy.

A critical factor in developing an efficient photosensitizer-gold nanoparticle (PS-AuNP) hybrid system with improved plasmonic photosensitization is to allocate a suitable space between AuNPs and PS. Poly(amidoamine) (PAMAM) dendrimer is selected as a spacer between the PS and confeito-like gold nanoparticles (confeito-AuNPs), providing the required distance (≈2.5-22.5 nm) for plasmon-enhanced singlet oxygen generation and heat production upon 638-nm laser irradiation and increase the cellular internalization of the nanoconjugates. The loading of the PS, tetrakis(4-carboxyphenyl) porphyrin (TCPP), and modified zinc phthalocyanine (ZnPc1) onto PAMAM-confeito-AuNPs demonstrate better in vitro cancer cell-killing efficacy, as the combined photothermal-photodynamic therapies (PTT-PDTs) outperforms the single treatment modalities (PTT or PDT alone). These PS-PAMAM-confeito-AuNPs also demonstrate higher phototoxicity than photosensitizers directly conjugated to confeito-AuNPs (TCPP-confeito-AuNPs and ZnPc1-confeito-AuNPs) against all breast cancer cell lines tested (MDA-MB-231, MCF7, and 4T1). In the in vivo studies, TCPP-PAMAM-confeito-AuNPs are biocompatible and exhibit a selective tumor accumulation effect, resulting in higher antitumor efficacy than free TCPP, PAMAM-confeito-AuNPs, and TCPP-confeito-AuNPs. In vitro and in vivo evaluations confirm PAMAM effectiveness in facilitating cellular uptake, plasmon-enhanced singlet oxygen and heat generation. In summary, this study highlights the potential of integrating a PAMAM spacer in enhancing the plasmon effect-based photothermal-photodynamic anticancer treatment efficiency of PS-decorated confeito-AuNPs.

Chitosan-Coated-PLGA Nanoparticles Enhance the Antitumor and Antimigration Activity of Stattic - A STAT3 Dimerization Blocker.

PURPOSE: The use of nanocarriers to improve the delivery and efficacy of antimetastatic agents is less explored when compared to cytotoxic agents. This study reports the entrapment of an antimetastatic Signal Transducer and Activator of Transcription 3 (STAT3) dimerization blocker, Stattic (S) into a chitosan-coated-poly(lactic-co-glycolic acid) (C-PLGA) nanocarrier and the improvement on the drug's physicochemical, in vitro and in vivo antimetastatic properties post entrapment. METHODS: In vitro, physicochemical properties of the Stattic-entrapped C-PLGA nanoparticles (S@C-PLGA) and Stattic-entrapped PLGA nanoparticles (S@PLGA, control) in terms of size, zeta potential, polydispersity index, drug loading, entrapment efficiency, Stattic release in different medium and cytotoxicity were firstly evaluated. The in vitro antimigration properties of the nanoparticles on breast cancer cell lines were then studied by Scratch assay and Transwell assay. Study on the in vivo antitumor efficacy and antimetastatic properties of S@C-PLGA compared to Stattic were then performed on 4T1 tumor bearing mice. RESULTS: The S@C-PLGA nanoparticles (141.8 ± 2.3 nm) was hemocompatible and exhibited low Stattic release (12%) in plasma. S@C-PLGA also exhibited enhanced in vitro anti-cell migration potency (by >10-fold in MDA-MB-231 and 5-fold in 4T1 cells) and in vivo tumor growth suppression (by 33.6%) in 4T1 murine metastatic mammary tumor bearing mice when compared to that of the Stattic-treated group. Interestingly, the number of lung and liver metastatic foci was found to reduce by 50% and 56.6%, respectively, and the average size of the lung metastatic foci was reduced by 75.4% in 4T1 tumor-bearing mice treated with S@C-PLGA compared to Stattic-treated group (p < 0.001). CONCLUSION: These findings suggest the usage of C-PLGA nanocarrier to improve the delivery and efficacy of antimetastatic agents, such as Stattic, in cancer therapy.

Isolation of exosome from the culture medium of Nasopharyngeal cancer (NPC) C666-1 cells using inertial based Microfluidic channel.

Isolation of exosome from culture medium in an effective way is desired for a less time consuming, cost saving technology in running the diagnostic test on cancer. In this study, we aim to develop an inertial microfluidic channel to separate the nano-size exosome from C666-1 cell culture medium as a selective sample. Simulation was carried out to obtain the optimum flow rate for determining the dimension of the channels for the exosome separation from the medium. The optimal dimension was then brought forward for the actual microfluidic channel fabrication, which consisted of the stages of mask printing, SU8 mould fabrication and ended with PDMS microchannel curing process. The prototype was then used to verify the optimum flow rate with polystyrene particles for its capabilities in actual task on particle separation as a control outcome. Next, the microchip was employed to separate the selected samples, exosome from the culture medium and compared the outcome from the conventional exosome extraction kit to study the level of effectiveness of the prototype. The exosome outcome from both the prototype and extraction kits were characterized through zetasizer, western blot and Transmission electron microscopy (TEM). The microfluidic chip designed in this study obtained a successful separation of exosome from the culture medium. Besides, the extra benefit from this microfluidic channels in particle separation brought an evenly distributed exosome upon collection while the exosomes separated through extraction kit was found clustered together. Therefore, this work has shown the microfluidic channel is suitable for continuous separation of exosome from the culture medium for a clinical study in the future.

Development and Evaluation of 1'-Acetoxychavicol Acetate (ACA)-Loaded Nanostructured Lipid Carriers for Prostate Cancer Therapy.

1'-acetoxychavicol acetate (ACA) extracted from the rhizomes of Alpinia conchigera Griff (Zingiberaceae) has been shown to deregulate the NF-ĸB signaling pathway and induce apoptosis-mediated cell death in many cancer types. However, ACA is a hydrophobic ester, with poor solubility in an aqueous medium, limited bioavailability, and nonspecific targeting in vivo. To address these problems, ACA was encapsulated in a nanostructured lipid carrier (NLC) anchored with plerixafor octahydrochloride (AMD3100) to promote targeted delivery towards C-X-C chemokine receptor type 4 (CXCR4)-expressing prostate cancer cells. The NLC was prepared using the melt and high sheer homogenization method, and it exhibited ideal physico-chemical properties, successful encapsulation and modification, and sustained rate of drug release. Furthermore, it demonstrated time-based and improved cellular uptake, and improved cytotoxic and anti-metastatic properties on PC-3 cells in vitro. Additionally, the in vivo animal tumor model revealed significant anti-tumor efficacy and reduction in pro-tumorigenic markers in comparison to the placebo, without affecting the weight and physiological states of the nude mice. Overall, ACA-loaded NLC with AMD3100 surface modification was successfully prepared with evidence of substantial anti-cancer efficacy. These results suggest the potential use of AMD3100-modified NLCs as a targeting carrier for cytotoxic drugs towards CXCR4-expressing cancer cells.

Development of nanostructured lipid carrier (NLC) assisted with polysorbate nonionic surfactants as a carrier for l-ascorbic acid and Gold Tri.E 30.

Lipid nanocarrier displays the advantages over conventional drug carriers as they are formulated with biodegradable and non-irritant lipids. However, the main drawbacks are the agglomeration of lipid particles, instability over storage, low drug loading, and the burst release of active ingredients. In this study, we investigated the effects of various polysorbate nonionic surfactants namely Tween 20, 40, 60, or 80 on the nanostructured lipid carrier (NLC). NLC incorporated with polysorbate nonionic surfactant was prepared by using high-pressure homogenization technique. The average size was reduced to 139.9 ± 15.8 nm in the presence of Tween 80 and remained stable in nano-size even incubated for 28 days. Encapsulation of l-ascorbic acid or Gold Tri.E 30 showed a high encapsulation efficiency of more than 75%, where the highest was Gold Tri.E in the presence of Tween 60 at 99.7%. In vitro release study showed that the release of both l-ascorbic acid and Gold Tri.E was significantly reduced in NLC with Tween as compared to bare active ingredients and NLC without Tween. In conclusion, the incorporation of Tween successfully produced a lipid nanocarrier that has the potential to be developed as a carrier of various active ingredients such as nutrients, extracts, and drugs.

Mixed Oleic Acid-Erucic Acid Liposomes as a Carrier for Anticancer Drugs.

BACKGROUND: Liposomes are mostly known to be prepared from phospholipids and lipids and have a remarkable capacity to encapsulate both lipophobic and lipophilic molecules. However, there is little research on developing fatty acid liposomes for chemotherapy. OBJECTIVE: We have successfully prepared mixed fatty acid liposomes from two monounsaturated fatty acids, namely oleic acid and erucic acid, which stabilised by DOPEPEG2000. The Critical Vesicular Concentration (CVC) of liposomes was found to be within 0.09 to 0.21 mmol dm-3, with an average particle size of 400 nm. METHODS: Encapsulation of various anticancer drugs such as folinic acid, methotrexate, doxorubicin, or irinotecan resulted in Encapsulation Efficiency (%EE) of up to 90%. Using a 3-(4, 5-dimethylthiazol-2- yl)-2,5-diphenyltetrazolium bromide (MTT) assay, the median Inhibitory Concentration (IC50) values of mixed oleic acid-erucic acid encapsulating hydrophilic drugs was remarkably reduced at the end of 24 hours of incubation with the human lung carcinoma cell line A549. RESULTS: The results suggest that mixed oleic acid-erucic acid liposomes are a potential new approach to further develop as an alternative vehicle of various drugs for cancer treatment.

Large-scale structure formation in ionic solution and its role in electrolysis and conductivity.

That water may not be an inert medium was indicated by the presence at water's interfaces a negatively charged solute free zone of several hundred microns in thickness called the exclusion zone (EZ). Further evidence was demonstrated by Ovchinnikova's experiments (2009) showing that water can store and release substantial amount of charge. We demonstrate that the charge storage capacity of water arises from highly stable large-scale ionic structures with measurable charge imbalances and discrete levels of charge density. We also show evidence that the charge zones formation requires ionic solutes, and their formation correlate to large change in conductivity, by as much as 250%. Our experiments indicate that large-scale structuring plays a pivotal role in electrolysis and conductivity in ionic solution. We propose that water is an electrochemically active medium and present a new model of electrolysis and conductivity in ionic solution.

Interaction between C18 fatty acids and DOPE PEG2000 in Langmuir monolayers: effect of degree of unsaturation.

In this study, we address the effect of the cis-double bond in 1,2-dioleoyl-sn-glycero-3-phosphoethanolamide-N-[methoxy(polyethylene glycol)-2000, DOPE PEG2000 (DP), on the Langmuir monolayer of C18 fatty acids-namely, stearic acid (SA), oleic acid (L1), linoleic acid (L2), and linolenic acid (L3)-with the same head group but different degrees of saturation on their hydrocarbon chains. Negative values of Gibbs free energy of mixing (ΔG mix) were obtained throughout the investigated ranges of the unsaturated C18 fatty-acid (L1, L2 and L3) mixed systems, indicating that very strong attractions occurred between molecules in the monolayers. The bend and kink effects from the cis-double bond(s) in the hydrocarbon chain affected the membrane fluidity and molecular packing in the monolayers, which resulted in a greater interaction between unsaturated C18 fatty acids and DP. The most thermodynamically stable mole composition of unsaturated C18 fatty acids to DP was observed at 50:1; this ratio is suggested to be the best mole ratio and will be subsequently used to prepare DP-C18 fatty-acid nanoliposomes. The presence of cis-double bonds in both hydrocarbon chains of DOPE in DP also created an imperfection in the membrane structure of lipid-drug delivery systems, which is expected to enhance lipid-based systems for antibody conjugation and drug encapsulation.

Local application of osteoprotegerin-chitosan gel in critical-sized defects in a rabbit model.

BACKGROUND: Osteoprotegerin (OPG) is used for the systemic treatment of bone diseases, although it has many side effects. The aim of this study was to investigate a newly formulated OPG-chitosan gel for local application to repair bone defects. Recent studies have reported that immunodetection of osteopontin (OPN) and osteocalcin (OC) can be used to characterise osteogenesis and new bone formation. METHODS: The osteogenic potential of the OPG-chitosan gel was evaluated in rabbits. Critical-sized defects were created in the calvarial bone, which were either left unfilled (control; group I), or filled with chitosan gel (group II) or OPG-chitosan gel (group III), with rabbits sacrificed at 6 and 12 weeks. Bone samples from the surgical area were decalcified and treated with routine histological and immunohistochemical protocols using OC, OPN, and cathepsin K (osteoclast marker) antibodies. The toxicity of the OPG-chitosan gel was evaluated by biochemical assays (liver and kidney function tests). RESULTS: The mean bone growth in defects filled with the OPG-chitosan gel was significantly higher than those filled with the chitosan gel or the unfilled group (p < 0.05). At 6 and 12 weeks, the highest levels of OC and OPN markers were found in the OPG-chitosan gel group, followed by the chitosan gel group. The number of osteoclasts in the OPG-chitosan gel group was lower than the other groups. The results of the liver and kidney functional tests indicated no signs of harmful systemic effects of treatment. In conclusion, the OPG-chitosan gel has many characteristics that make it suitable for bone repair and regeneration, highlighting its potential benefits for tissue engineering applications.

Formulation and in vitro and in vivo evaluation of a new osteoprotegerin-chitosan gel for bone tissue regeneration.

The osteoprotegerin (OPG) system plays a critical role in bone remodelling by regulating osteoclast formation and activity. The study aimed to determine the physicochemical properties and biocompatibility of a newly formulated OPG-chitosan gel. The OPG-chitosan gel was formulated using human OPG protein and water-soluble chitosan. The physicochemical properties were determined using Fourier transform infrared (FTIR) spectroscopy, thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC). Gel morphology was determined using scanning electron microscopy (SEM) and then it was subjected to a protein release assay and biodegradability test. An in vitro cytotoxicity test on normal human periodontal ligament (NHPL) fibroblasts and normal human (NH) osteoblasts was carried out using the AlamarBlue assay. In vivo evaluation in a rabbit model involved creating critical-sized defects in calvarial bone, filling with the OPG-chitosan gel and sacrificing at 12 weeks. In vitro results demonstrated that the 25 kDa OPG-chitosan gel had the highest rate of protein release and achieved 90% degradation in 28 days. At 12 weeks, the defects filled with 25 kDa OPG-chitosan gel showed significant (p < 0.05) new bone formation and the highest expression of osteocalcin and osteopontin compared to controls. Thus, the 25 kDa OPG-chitosan gel could be a promising new biomaterial for tissue engineering. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 398-407, 2017.

In vitro evaluation of osteoprotegerin in chitosan for potential bone defect applications.

BACKGROUND: The receptor activator of nuclear factor kappa-B (RANK)/RANK ligand/osteoprotegerin (OPG) system plays a critical role in bone remodelling by regulating osteoclast formation and activity. OPG has been used systemically in the treatment of bone diseases. In searching for more effective and safer treatment for bone diseases, we investigated newly formulated OPG-chitosan complexes, which is prepared as a local application for its osteogenic potential to remediate bone defects. METHODS: We examined high, medium and low molecular weights of chitosan combined with OPG. The cytotoxicity of OPG in chitosan and its proliferation in vitro was evaluated using normal, human periodontal ligament (NHPL) fibroblasts in 2D and 3D cell culture. The cytotoxicity of these combinations was compared by measuring cell survival with a tetrazolium salt reduction (MTT) assay and AlamarBlue assay. The cellular morphological changes were observed under an inverted microscope. A propidium iodide and acridine orange double-staining assay was used to evaluate the morphology and quantify the viable and nonviable cells. The expression level of osteopontin and osteocalcin protein in treated normal human osteoblast cells was evaluated by using Western blot. RESULTS: The results demonstrated that OPG in combination with chitosan was non-toxic, and OPG combined with low molecular weight chitosan has the most significant effect on NHPL fibroblasts and stimulates proliferation of cells over the period of treatment.

Chitosan-Coated Poly(lactic-co-glycolic acid)-Diiodinated boron-Dipyrromethene Nanoparticles Improve Tumor Selectivity and Stealth Properties in Photodynamic Cancer Therapy.

Their limited solubility and lack of tumor selectivity limit the clinical usefulness of photosensitizers. Various nanostructures have been evaluated as delivery agents for photosensitizers in an attempt to overcome these obstacles, but these have typically been limited by premature clearance by the reticuloendothelial system (RES) and non-specific interactions with normal cells that result from their hydrophobic surfaces. In this study, we report our attempt to circumvent these problems by applying a low molecular weight chitosan (25 kDa) coating to a poly(lactic-co-glycolic acid)-diiodinated boron dipyrromethene (PLGA-I2BODIPY) nanoparticle-photosensitizer construct. This chitosan coating increased the hydrophilicity and decreased the charge of PLGA-I2 BODIPY nanoparticle surfaces without changing their size (average diameter 147 nm) or morphology. In comparison to the uncoated controls, the coated nanoparticles reduced the burst release of I2BODIPY, increased its predominantly lysosomal cellular uptake, and enhanced its photocytotoxicity in 4T1 murine and MDA-MB-231 human breast cancer cells. PLGA-Chitosan-I2BODIPY nanoparticles also showed reduced serum protein adsorption and macrophage uptake compared to the uncoated controls. In 4T1 tumor-bearing mice, the PLGA-Chitosan-I2BODIPY nanoparticles exhibited better tumor-targeting selectivity and significantly reduced accumulation in RES tissues, including the lymph nodes, spleen and liver (by 10.2-, 2.1- and 1.3-fold, respectively), and in non-tumorous organs, such as the skin and eyes (by 22.7- and 4-fold, respectively). The PLGA-Chitosan-I2BODIPY and PLGA-I2BODIPY nanoparticles also showed increased anticancer efficacy compared to free I2BODIPY. These results suggest that the low molecular weight chitosan (25 kDa) is a promising nanoparticle "stealth coating" that improves tumor selectivity.

Experimental investigation on the use of highly charged nanoparticles to improve the stability of weakly charged colloidal system.

The present work highlighted on the implementation of a unique concept for stabilizing colloids at their incipiently low charge potential. A highly charged nanoparticle was introduced within a coagulated prone colloidal system, serving as stabilizer to resist otherwise rapid flocculation and sedimentation process. A low size asymmetry of nanoparticle/colloid serves as the new topic of investigation in addition to the well-established large size ratio nanoparticle/microparticle study. Highly charged Al2O3 nanoparticles were used within the present research context to stabilize TiO2 and Fe3O4 based colloids via the formation of composite structures. It was believed, based on the experimental evidence, that Al2O3 nanoparticle interact with the weakly charged TiO2 and Fe3O4 colloids within the binary system via absorption and/or haloing modes to increase the overall charge potential of the respective colloids, thus preventing further surface contact via van der Waal's attraction. Series of experimental results strongly suggest the presence of weakly charged colloids in the studied bimodal system where, in the absence of highly charged nanoparticle, experience rapid instability. Absorbance measurement indicated that the colloidal stability drops in accordance to the highly charged nanoparticle sedimentation rate, suggesting the dominant influence of nanoparticles to attain a well-dispersed binary system. Further, it was found that the level of colloidal stability was enhanced with increasing nanoparticle fraction within the mixture. Rheological observation revealed that each hybrid complexes demonstrated behavior reminiscence to water with negligible increase in viscosity which serves as highly favorable condition particularly in thermal transport applications.