Development of docetaxel and alendronate-loaded chitosan-conjugated polylactide-co-glycolide nanoparticles: In vitro characterization in osteosarcoma cells

Purpose: To develop docetaxel (DTX)- and alendronate (ALN)-loaded, chitosan (CS)-conjugated polylactide- co-glycolide (PLGA) nanoparticles (NPs) to increase therapeutic efficacy in osteosarcoma cells. Methods: Drug-loaded PLGA NPs were prepared by nanoprecipitation and chemically conjugated by the carboxylic group of PLGA to the amine-bearing CS polymer. The nanocarrier was characterized by dynamic light scattering, transmission electron microscopy, scanning electron microscopy, and differential scanning calorimetry as well as by in vitro drug release and cell culture studies. Results: NP size was within the tumour targeting range (~200 nm) with an effective positive charge (20 mV), thus increasing cellular uptake efficiency. Morphological analysis revealed clear spherical particles with uniform dispersion. The NPs exhibited identical sustained release kinetics for both DTX and ALN. CS-conjugated PLGA with dual-drug-loaded (DTX and AL) NPs showed typical time-dependent cellular uptake and also displayed superior cytotoxicity in MG-63 cells compared with blank NPs, which were safe and biocompatible. Conclusion: Combined loading of DTX and ALN in NPs increased the therapeutic efficacy of the formulation for osteosarcoma treatment, thus indicating the potential benefit of a combinatorial drug regimen using nanocarriers for effective treatment of osteosarcoma.

In situ epicatechin-loaded hydrogel implants for local drug delivery to spinal column for effective management of post-traumatic spinal injuries

Purpose: To prepare hydrogels loaded with epicatechin, a strong antioxidant, anti-inflammatory, and neuroprotective tea flavonoid, and characterise them in situ as a vehicle for prolonged and safer drug delivery in patients with post-traumatic spinal cord injury. Methods: Five in situ gel formulations were prepared using chitosan and evaluated in terms of their visual appearance, clarity, pH, viscosity, and in vitro drug release. In vivo anti-inflammatory activity was determined and compared with 2 % piroxicam gel as standard. Motor function activity in a rat model of spinal injury was examined comparatively with i.v. methylprednisolone as standard. Results: The N-methyl pyrrolidone solution (containing 1 % w/w epicatechin with 2 to 10 % w/w chitosan) of the in situ gel formulation had a uniform pH in the range of 4.01 ± 0.12 to 4.27 ± 0.02. High and uniform drug loading, ranging from 94.48 ± 1.28 to 98.08 ± 1.24 %, and good in vitro drug release (79.48 ± 2.84 to 96.48 ± 1.02 % after 7 days) were achieved. The in situ gel prepared from 1 % epicatechin and 2 % chitosan (E5) showed the greatest in vivo anti-inflammatory activity (60.58 % inhibition of paw oedema in standard carrageenan-induced hind rat paw oedema model, compared with 48.08 % for the standard). The gels showed significant therapeutic effectiveness against post-traumainduced spinal injury in rats. E5 elicited maximum motor activity (horizontal bar test) in the spinal injury rat model; the rats that received E5 treatment produced an activity score of 3.62 ± 0.02 at the end of 7 days, compared with 5.0 ± 0.20 following treatment with the standard. Conclusion: In situ epicatechin-loaded gel exhibits significant neuroprotective and anti-inflammatory effects, and therefore can potentially be used for prolonged and safe drug delivery in patients with traumatic spinal cord injury.

Development and evaluation of floating microspheres of curcumin in alloxan-induced diabetic rats

Purpose: To prepare and evaluate floating microspheres of curcumin for prolonged gastric residence and to study their effect on alloxan-induced diabetic rats. Methods: Floating microsphere were prepared by emulsion-solvent diffusion method, using hydroxylpropyl methylcellulose, chitosan and Eudragit S 100 polymer in varying proportions. Ethanol/dichloromethane blend was used as solvent in a ratio of 1:1. The floating microspheres were evaluated for flow properties, particle size, incorporation efficiency, as well as in-vitro floatability and drug release. The anti-diabetic activity of the floating microspheres of batch FM4 was performed on alloxaninduced diabetic rats. Result: The floating microspheres had particle size, buoyancy, drug entrapment efficiency and yield in the ranges of 255.32 - 365.65 μm, 75.58 - 89.59, 72.6 - 83.5, and 60.46 - 80.02 %, respectively. Maximum drug release after 24 h was 82.62 % for formulation FM4 and 73.879, 58.613 and 46.106 % for formulations FM1, FM2, and FM3 respectively. In-vivo data obtained over a 120-h period indicate that curcumin floating microspheres from batch FM4 showed the better glycemic control than control and a commercial brand of the drug. Conclusion: The developed floating curcumin delivery system seems economical and effective in diabetes management in rats, and enhances the bioavailability of the drug.

Development of hydroxyapatite-based nanomaterials to enhance biological response of osteoblast cells for clinical application

Purpose: To develop a novel chitosan/gelatin-hydroxyapatite (CGHaP) microspheres for evaluating the biological response of pre-osteoblast cells. Methods: The microsphere was prepared by water-in-oil emulsion method. Cell proliferation was studied using AlamarBlue colorimetric assay and DAPI staining while alkaline phosphatase assay was carried out by colorimetric assay method. Chitosan microspheres as well as chitosan-hydroxyapatite microspheres was prepared and tested for biological response from MC3T3-E1 cell line. Results: The results showed that CGHaP promotes MC3T3-E1 cell proliferation and spread on the surface of microspheres. The cells were clustered with more actin filaments and well-linked with neighbouring cells or adjacent cells when cultured in CGHaP microspheres whereas fewer cells were spread on chitosan (CH) microspheres. CGHaP microspheres significantly (p < 0.05) promoted cell attachment, proliferation and extracellular matrix mineralization. CGHaP microspheres presented significantly (p < 0.02) higher calcium deposition (0.5 ng) than CH microspheres (0.28 ng). Specifically, CGHaP microspheres exhibited high ALP activity (8 units; 2-fold) compared to CH with 3 units, after 7 days of incubation. The results suggest that CGHaP possesses a great ability to facilitate bone ingrowth formation and possibility of good osteointegration in vivo. Conclusion: The nanomaterial enhances the proliferation of pre-osteoblast cells in tissue engineering microspheres. The outcome of this study may have a major impact on the development of novel nanomaterials for bone tissue engineering.

Polymeric implant of methylprednisolone for spinal injury: preparation and characterization

Purpose: To improve the effectiveness and reduce the systemic side effects of methylprednisolone in traumatic spinal injuries, its polymeric implants were prepared using chitosan and sodium alginate as the biocompatible polymers. Methods: Implants of methylprednisolone sodium succinate (MPSS) were prepared by molding the drug-loaded polymeric mass obtained after ionotropic gelation method. The prepared implants were evaluated for drug loading, in vitro drug release and in vivo performance in traumatic spinal-injury rat model with paraplegia. Results: All the implant formulations were light pale solid matrix with smooth texture. Implants showed 86.56 ± 2.07 % drug loading. Drug release was 89.29 ± 1.25 % at the end of 7 days. Motor function was evaluated in traumatic spinal injury-induced rats in terms of its movement on the horizontal bar. At the end of 7 days, the test group showed the activity score (4.75 ± 0.02) slightly higher than that of standard (4.62 ± 0.25), but the difference was not statistically different (p > 0.05). Conclusion: MPSS-loaded implants produces good recovery in traumatic spinal-injury rats.