Selective photodynamic effects on cervical adenocarcinoma cells provided by F127 Pluronic®-based micelles modulating hypericin delivery

Abstract Cervical cancer is a leading cause of death among women. The endocervical adenocarcinoma (ECA) represents an aggressive and metastatic type of cancer with no effective treatment options currently available. We evaluated the antitumoral and anti-migratory effects of hypericin (HYP) encapsulated on Pluronic F127 (F127/HYP) photodynamic therapy (PDT) against a human cell line derived from invasive cervical adenocarcinoma (HeLa) compared to a human epithelial cell line (HaCaT). The phototoxicity and cytotoxicity of F127/HYP were evaluated by the following assays: colorimetric assay, MTT, cellular morphological changes by microscopy and long-term cytotoxicity by clonogenic assay. In addition, we performed fluorescence microscopy to analyze cell uptake and subcellular distribution of F127/HYP, cell death pathway and reactive oxygen species (ROS) production. The PDT mechanism was determined with sodium azide and D-mannitol and cell migration by wound-healing assay. The treatment with F127/HYP promoted a phototoxic result in the HeLa cells in a dose-dependent and selective form. Internalization of F127/HYP was observed mainly in the mitochondria, causing cell death by necrosis and ROS production especially by the type II PDT mechanism. Furthermore, F127/HYP reduced the long-term proliferation and migration capacity of HeLa cells. Overall, our results indicate a potentially application of F127/HYP micelles as a novel approach for PDT with HYP delivery to more specifically treat ECA.

Enhancement of Solubility And Dissolution Profile of Clopidogrel By Various Solid Dispersion Formulations

The present research is aimed at enhancing solubility and drug dissolution of clopidogrel (CPG) used as oral antiplatelet agent by employing solid dispersion (SD) technique. Total 40 SDs formulated with drug: polymers (pluronic F127, poloxamer 407, labrafil PG, PEG 6000, gelucire 50/13), in varying ratios (1:0.5, 1:1, 1:2, 1:3, 1:4) of which CPG1 to CPG20 and CPG21 to CPG40 prepared by adopting solvent evaporation method fusion (melt) method respectively. The formulation CPG40 containing pluronic F127 as polymer showed highest solubility of 6.57±0.04 mg/ml) that is 45 folds than pure drug. Similar results reflected in the dissolution studies where CPG40 containing CPG: pluronic F127 in 1:4 ratio showed maximum % drug content, % practical yield and drug dissolution of 99.14% in 60 minutes when compared with other formulations and pure drug (32.76%) obtained by fusion melt method. From FTIR studies the optimized formulation CPG40 showed the compatibility between drug and polymers. XRD and SEM studies showed CPG40 exists in amorphous form that fetched in better drug release from the SD formulation in comparison to pure drug.

Differences in the effect of two injectable Pluronic F-127 hydrogel composites in repairing rabbit cartilage defects / 中国组织工程研究

BACKGROUND: Preliminary experiments show that bone marrow mesenchymal stem cells transfected with SOX9 gene can grow and proliferate in Pluronic F-127 hydrogel, promote the secretion of extracellular matrix, and increase the expression of cartilage matrix. OBJECTIVE: The SOX9 gene was transduced into bone marrow mesenchymal stem cells by lentivirus gene induction, and then combined with injectable Pluronic F-127 hydrogel to observe the effect of Pluronic F-127 hydrogel on repairing cartilage defects. METHODS: SOX9 gene was transfected into bone marrow mesenchymal stem cells by lentivirus gene induction. After 48 hours of transfectlon, SOX9 gene was combined with Pluronic F-127 hydrogel. Sixty New Zealand white rabbits provided by the Experimental Animal Center of Wuhan University of Science and Technology were selected to establish the models of femoral condylar cartilage defect of the right knee joint. The rabbits were randomly divided into three groups: model group without implantation of any material at the defect site, control group with implantation of non-transfected bone marrow mesenchymal stem cells and Pluronic F-127 hydrogel complex at the defect site, and experimental group with implantation of SOX9 gene-transfected bone marrow mesenchymal stem cells and Pluronic F-127 hydrogel complex at the defect site. Four and twelve weeks after operation, the defect tissues were taken for three-dimensional reconstruction of micro-CT, hematoxylin-eosin staining, Safranine O staining, type II collagen immunohistochemical staining and Wakitani soft tissue repair histological score. This study was approved by the Ethics Committee of Wuhan University of Science and Technology. RESULTS AND CONCLUSION: (1) At 12 weeks after operation, three-dimensional reconstruction of Micro-CT showed that there was no obvious repair In the defect area of the model group, and there was still a large depression In the center. In the control group, the central depression area was significantly reduced and more trabecular structures of regenerated bone were observed. In the experimental group, the defect area was basically repaired. (2) At 12 weeks after operation, hematoxylin-eosin staining showed that there was no trabecular bone structure, disordered cell distribution and no cartilage lacunae at the defect area of the model group. In the control group, more bone tissue was reconstructed, and the defect area was mainly filled with cartilage-like tissue and fibrous tissue. In the experimental group, bone tissue was reconstructed adequately, and the defect area was mainly filled with chondroid cells and chondroid extracellular matrix. Cells arranged columnariy, similar to the surrounding cartilage. (3) At 12 weeks after surgery, Safranine O staining and collagen II immunohistochemical staining results showed that a small amount of glycosamlnoglycan was observed, but no type II collagen was found in the model group. The expression of glycosaminoglycan and type II collagen was more in the control group. The expression of glycosaminoglycan and type II collagen was highest In the experimental group compared with the other two groups. (4) The histological score of Wakitani soft tissue repair in the experimental group was higher than that in the control group and model group (P < 0.05). (5) The results suggested that Pluronic F-127 hydrogel complex loaded with SOX9 gene transfected bone marrow mesenchymal stem cells can promote the repair of cartilage defects.

Development of amphotericin b Based organogels against mucocutaneous fungal infections

Amphotericin B is a broad spectrum antifungal agent used to treat fungal infections. Organogel is a semisolid preparation in which the apolar phase gets immobilized within spaces of the three-dimensional structure. The current study aimed at the formulation and comparative evaluation of sorbitan monostearate organogels and pluronic lecithin organogels (PLO). Different compositions of span 60 based organogels were prepared by varying the concentrations of the span 60 and PLO gels were prepared by varying the concentration of Pluronic F 127. The developed organogels were subjected to various characteristics such as pH, viscosity, spreadability, extrudability, and drug release studies. The optimized formulations were evaluated against Candida albicans and carried out ex vivo release study. The optimized formulation was selected from span 60 based organogels, and pluronic lecithin organogels were S1 and P1, respectively. The optimized formulation (S1) showed effective inhibition against Candida albicans. The skin irritation test was carried out on albino mice for optimized formulations and results showed that no irritation to the skin. Based on the results, organogels prepared by sorbitan monostearate showed better antifungal activity, and also all the formulations were found to be stable and safe throughout the study period.

Study on Preparation and in vitro Release Characteristic of Ursolic Acid/PF 127/TPGS-doxorubicin Mixed Nanomicelles / 中国药房

OBJECTIVE： To prepare Ursolic acid （UA）/Pluronic F127 （PF127）/TPGS-doxorubicin （DOX） mixed nanomicelles， and to characterize it and study its in vitro release behavior. METHODS： UA/PF127/TPGS nanomicelles were prepared by thin film hydration method. Using encapsulation efficiency of UA as index， combined with the results of single factor tests， L9（34） orthogonal test was used to optimize drug dosage of UA， molar ratio of PF127 to TPGS， hydration temperature and hydration volume， validation test was performed. On the basis of succinylated TPGS， TPGS-DOX was synthesized and mixed with UA/PF127/TPGS to prepare UA/PF127/TPGS-DOX mixed nanomicelles， the appearance， particle size and critical micelle concentration （PF127/TPGS） were investigated. The drug release behavior was examined by dialysis bag diffusion method. RESULTS： The optimal preparation technology of UA/PF127/TPGS nanomicelles was as follows as drug dosage of UA 8 mg， molar ratio of PF127 to TPGS 3 ∶ 7， hydration temperature 50 ℃， hydration volume 4 mL. Average encapsulation efficiency of UA in nanomicelles was 89.00％ （RSD＝0.43％， n＝3）. The prepared UA/PF127/TPGS-DOX mixed nanomicelles solution was clear with opalescence. The nanomicelles were spherical and uniform in size； average particle size was （115.00±9.42） nm； critical micelle concentration of PF127/TPGS （molecular ratio 3 ∶ 7） was 0.001 3％. The in vitro drug release of UA and DOX in the mixed nanomicelles was significantly slowed down， compared with raw materials or substance control. The drug release process of the two drugs in the nanomicelles conformed to Weibull equation. CONCLUSIONS： UA/PF127/TPGS-DOX mixed nanomicelles are successfully prepared with uniform particle size， good stability and good sustained-release effect.

Improved efficiency of adeno-associated virus mediated gene transfer to vein grafts by use of pluronic F-127-trypsin gel / 上海交通大学学报（医学版）

Objective: To investigate whether pluronic F-127-trypsin gel enhances the efficiency of adeno-associated virus (AAV) mediated gene transfer to vein grafts. Methods: Rat model of venous bridge vascular restenosis was established. The vein grafts were infected with recombinant AAV encoding an enhanced green fluorescent protein (Egfp) reporter gene, and pluronic F-127-trypsin gel was used to increase viral contact time and viral penetration. The expression of EGFP in vein grafts was determined by frozen section, immunohistochemistry staining and quantitative reverse transcriptase-mediated PCR 21 days after surgery. Structural integrity of the tissue was evaluated by measurement of tissue tensile strength. Results: Pluronic F-127-trypsin gel can significantly improve the efficiency of AAV infection in vein grafts, in which AAV serotype 6 was more efficient than AAV 1, 8, and 9 serotypes in infecting vein grafts, and tissue tensile strength was not affected. Conclusion: Pluronic F-127 trypsin gel may be a safe and effective method to improve the efficiency of AAV mediated gene transfer to vein grafts. It can be used for the prevention and treatment of venous bridge vascular restenosis.

A Review on organogel for skin aging.

Skin aging is one of the prominent problems associated with skin as each part of body ages with the time, skin is the external organ where the sign and symptoms of aging are readily evident. However cosmetics as well as pharmaceutical approaches delayed skin aging. Gel are best fitted in all these essential criteria because of their excellent appearance, smoothness, desired consistency, fast drug release, ease of manufacturing and quality assessment and admirable stability. Recently gel formulation have been modified to yield an advance drug delivery system known as ―organogels‖. Gel define as a semi-solid preparation having an external solvent phase, apolar [organogel] or polar [hydrogel] immobilized within the space available of a three dimensional network structure. Lecithin is a natural surfactant isolated from eggs or soya bean, when it combined with water and non-polar solvent, it form gels. PLO gels have gained importance in recent years as transdermal drug delivery system. It is a thermodynamically stable, visco-elastic system, which is non-irritating, odorless and biodegradable. Pluronic F127 or poloxamer is a copolymer of polyoxyethylene and polyoxypropylene which forms a thermoreversible gel in concentrations between 15-30%w/v. Water plays the role of a structure-forming agent and stabilizes the process of gel formation as it solubilizes the pluronic and other hydrophilic drugs. PLO gel system facilitates the delivery of hydrophilic as well as lipophilic drugs owing to the presence of both oil and aqueous phases within the gel system.

Preparation of Redox-responsive Pluronic F127 Micelles with High Drug-loading Efficiency / 中国药房

OBJECTIVE:To prepare redox-responsive pluronic F127 micelles with high drug-loading efficiency. METHODS:The thiol derivative F127-SH with strong hydrophobicity was synthesized by introducing tert-butyl methacrylate and cysteamine to the hydrophobic segments of F127. Paclitaxel (PTX)-loaded redox-responsive pluronic F127 micelles with high drug-loading effi-ciency(F127-SS/PTX)were prepared by film-hydration method as well as oxidation reaction of thiol groups. The diameter,encap-sulation efficiency and drug-loading amount were detected,and the change of diameter and drug release behavior of micelles were investaged under reducing environment or non-reducing environment. RESULTS:Mean diameter of F127-SS/PTX micelles was about(77.87±1.79)nm,and encapsulation efficiency and drug-loading amount were(92.73±2.35)% and(16.25±0.99)%,re-spectively. The diameter of the micelles increased rapidly and drug release rate of PTX increased significantly in the presence of 10 mmol/L dithiothreitol. CONCLUSIONS:The redox-responsive pluronic F127 micelles with high drug-loading efficiency are pre-pared successfully.

Preparation and in vitro evaluation of ampelopsin-loaded nanomicelles / 中国中药杂志

To improve the solubility and antitumor activity of ampelopsin, ampelopsin-loaded nanomicelles from the mixture of pluronic F127 and D-α-tocopheryl polyethylene glycol 1000 succinate (TPGS1000) were prepared by film-thin hydration method, in order to optimize the process conditions and physicochemical properties. The antitumor activities against MCF-7 cells between ampelopsin and nanomicelles were compared by MTT method, respectively. The results showed that the optimal nanomicelles were round with the nanometric size of (22.6±0.5) nm, encapsulation efficiency rate of (80.42±1.13)%, and drug-loading rate of (4.41±0.26)%. The solubility of ampelopsin in mixed nanomicelles significantly increased by 16 times. In different release media, the mixed nanomicelles could release more than 90% of drug in 8 h, and showed stronger cytotoxicity and inhibition against MCF-7 cells (P<0.01). The mixed nanomicelles can be used as new drug delivery system of ampelopsin.

Preparation of CI-921 mixed micelles and determination of its entrapment efficiency by method of dialysis and fitting / 中国药科大学学报

@#The aim of this research were to prepare CI-921 mixed micelles(CI-Micelles), to establish a method for determining the entrapment efficiency of CI-Micelles, to optimize the formulation and to evaluate their in vitro properties. CI-Micelles were prepared by film dispersion. Dialysis and fitting were used to calculate true entrapment efficiency(EE)and drug loading(DL)of CI-Micelles. The influences of polymer concentration, polymer radio and hydrating media on the entrapment efficiency, drug loading and particle-size were investigated. The stability at 4 °C in 6 days was evaluated. The optimal formulation of CI-921-Micelles consisting of polymer concentration of 72 mg/mL, a mass radio of Pluronic F127/Solutol HS15 at mass ratio of 1 ∶2 and 5% glucose solution as hydrating medium, showed a EE of more than 90%, and mean particle-size of 17-25 nm and PDI< 0. 210. There were no significant changes to CI-Micelles in EE and particle-size after treatment at 4 °C for 6 days. The applied method of dialysis and fitting could be used to determine EE for micelles loaded with weakly basic drug which was difficult to meet sink conditions. Adjustment of the mass radio of Pluronic F127 to Solutol HS15 had resulted in uniform particle size distribution, high entrapment efficiency and drug loading capacity and better stability of CI-Micelles.

Trimethyl chitosan nanoparticles coated with Pluronic F-127 for oral insulin delivery system / 中国药科大学学报

@#The purpose of this investigation was to develop Pluronic F-127 coated N-trimethyl chitosan nanoparticles(F-S NPs)of insulin as the model drug and asses their penetration of the mucosal barriers. Single factor screening was used to optimize the formulations of nanoparticles and the nanoparticles were characterized. Their particle size, Zeta potential, encapsulation efficiencies and drug loading were assayed to be(240. 6±6. 51)nm, (10. 42±1. 60)mV, (43. 39±2. 83)% and(3. 39±0. 57)%, respectively. The impact of PF-127 on mucin binding in vitro and nanoparticles′s transport in freshly obtained mucus were also evaluated. The mucin affinity of F-S NPs was significantly reduced when compared to that of the N-trimethyl chitosan nanoparticles(S NPs), i. e. , 28% of the latter. And F-S NPs was found to have an improved mucosal penetrating capability. Mucus-secreting HT29-MTX-E12(E12)cell monolayer was selected to investigate their cellular uptake. F-S NPs exhibited higher penetration coefficient than both free insulin and S NPs in mucus-secreting epithelium cells, i. e. , 16-fold and 1. 4-fold, respectively. Data suggest that F-S NPs be potential carriers to cross mucosal barriers and enhance the cellular uptake of insulin.

Formulation and Evaluation of Gel Forming Ocular Minitablets Containing Piroxicam.

Aims: The purpose of this in vitro and ex vivo study was to prepare and characterise ocular minitablets of piroxicam based on different polymeric matrices and to evaluate their potential to provide prolonged and controlled drug release to ocular tissues after surface administration. Study Design: Experimental study and ex-vivo study. Place and Duration of Study: School of Pharmacy, University of East Anglia, Norwich, Norfolk, UK, between July 2011 and March 2012. Methodology: A range of placebo minitablet formulations were prepared based on pharmaceutically-acceptable polymers of differing chemical and physical properties. These were evaluated using standard physical and visual imaging methods. A subset of placebo formulations was chosen to prepare medicated minitablets containing 5 %w/w piroxicam as a model drug. Three different in vitro methodologies were used to assess drug release from the minitablets. An ex vivo porcine ocular method was used to assess likely tissue distribution of the drug after surface ocular administration of the minitablets. Results: Minitablets were successfully produced from all formulations. The in vitro drug release profile was dependent on the chemistry of the polymer used, its hydration and swelling behaviour and to some extent, the methodology used for assessing the drug release profile. The ex vivo studies in porcine eyes suggested that the drug disposition was inversely related to the hydration and swelling behaviour of the polymer. Minitablets containing piroxicam based on Pluronic F127 showed the highest posterior segment ocular bioavailability of the formulations studied in the ex vivo model. Conversely, the more highly swelling minitablet formulations showed higher anterior segment bioavailability. Conclusions: Ocular minitablets containing 5 %w/w piroxicam were successfully produced from a range of polymer matrices. In vitro release was shown to be dependent on the physical and chemical properties of the polymers used as the basis of the minitablets. Posterior segment deposition in an ex vivo model was greatest in the formulation which showed limited hydration and swelling behaviour in a simulated ocular environment.

Osteogenic Activity of Cultured Human Periosteal-Derived Cells in a Three Dimensional Polydioxanone/pluronic F127 Scaffold

Experimental Transscleral Ocular Drug Delivery: Use of Pluronic F-127 Gel and Fibrin Glue as a Sustained Release System

PURPOSE: The purpose of this study was to evaluate the effect of two sustained release systems (Pluronic F-127 gel and Fibrin glue) on the diffusion of dexamethasone across the human sclera. METHODS: Scleral sections excised from moist-chamber-stored human globes were mounted in a perfusion chamber that can create transscleral pressure. In the sustained release study, sample (100 muL) of 3H-dexamethasone in Pluronic F-127 gel or Fibrin glue was added to the episcleral side of the tissue, while BSS plusR was perfused across the uveal side at an transscleral pressure of 15 mmHg. Perfusate fractions were collected and measured using scintillation spectrometry and scleral permeability was calculated. RESULTS: The apparent permeability constants of the human sclera to 3H-dexamethasone in BSS plus(R) (the control value), Pluronic F-127 gel, and Fibrin glue were 1.15+/-0.11x10(-5) cm/s (n=5), 1.49+/-0.33x10-6 cm/s (n=5), and 7.32+/-0.98x10(-6) cm/s (n=7), respectively. The permeability values of Pluronic F-127 gel and Fibrin glue were relatively lower than the control value. Pluronic F-127 gel and Fibrin glue showed a uniform sustained release characteristic during a 24-hour period. The cumulative release rates of dexamethasone through the human sclera from BSS plus(R) (the control value), Pluronic F-127 gel, and Fibrin glue were 84.0+/-1.5% (n=5), 29.3+/-5.8% (n=5), and 61.5+/-5.9% (n=4) at 20 hours, respectively. There were significant differences in the human scleral permeability constants and cumulative release rates among the three vehicles (p<0.0001). CONCLUSIONS: Pluronic F-127 gel and Fibrin glue provided a slow, uniform sustained release during a 24- hour period. This study established a strong possibility of the new transscleral drug delivery in vitro using the sustained release systems of Pluronic F-127 gel and Fibrin glue. This may be a good experimental tool in the future development of a practical drug delivery system across the sclera for the treatment of a variety of chorioretinal disorders.