Electronic Theses and Dissertations (PhDs)
Permanent URI for this collectionhttps://hdl.handle.net/10539/37985
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Item Design and development of a bioactive-loaded polymer-engineered neural device for potential application in reducing neurological deficits after spinal cord injuries and neuro-regeneration(University of the Witwatersrand, Johannesburg, 2017) Kumar, Pradeep; Choonara, Yahya Essop; Modi, Girish; Naidoo, Dinesh; Pillay, VinessTraumatic Spinal Cord Injuries (SCI), due to their devastating nature, present several interventional challenges (extensive inflammation, axonal tethering, scar formation, neuronal degeneration and functional loss) that need to be addressed before even a slight neuronal recovery can be achieved. Recent post-TSCI investigational approaches include support strategies capable of providing scaffold architecture to allow axonal growth and conformal repair. This research provided detailed insight towards the development and fabrication of six specialized Polymer-Engineered Neural Devices (PENDs): 1) poly(lactic-co-glycolic acid)-gliadin (PLGA-GLDN) nanofibrous mats, 2) polyacrylamidated chitosan (PAAm-g-HT) scaffold, 3) functionalized chitosan methoxypolyethylene glycol (CHT-mPEG) cryosponges, 4) polyacrylonitrile-elastin-collagen (PANi-EC) neurosponge, 5) methylcellulose-alginate-polyethylene glycol (MAP) thermogel, and 6) chitosan-luronic F127-β glycerophosphate (CHT-PF127-βGP) composite thermogel for potential restriction, repair, regeneration, restoration and reorganization post-SCI. The latest trends in biomaterials-based SCI intervention were reviewed, discussed and analyzed in detail throughout the thesis. The research also involved an in silico analytico mathematical interpretation of multi(biomed)material assemblies wherein quantification of energy surfaces and molecular attributes via atomistic, dynamic, and docking simulations was carried out. The in silico experimentation additionally confirmed the potential of curcumin as a bioactive of choice for SCI intervention. Curcumin and dexamethasone were incorporated into the compact scaffolds and the bioactive release was determined over a period extending up to 60 days. The PLGA-GLDN nanofibrous mats demonstrated the formation of a compatible blend among the component polymers at equal weight ratios (PG55) as confirmed by quantitative physicochemical characterizations. Image processing analysis (DiameterJ plug-in of ImageJ) was performed on the SEM images of nanofibers to quantify the size, porosity, and orientation of the samples. Nanofibers within the size range of 10nm and 250nm were obtained in case of compatible blend and the nano stack was used for in vivo implantation post-SCI. Polyacrylamidated chitosan (PAAm-g-CHT) was synthesized via a unique persulfate-mediated polymer slicing and complexation as determined by static lattice atomistic simulations. The graft copolymer so obtained was fabricated into an anisotropic neurodurable scaffold. The CHT/mPEG cryosponges showed unique morphological features such as fringe thread-like structures (CHT alone); hemispherical, pebble-like structures (CHT-mPEG); curved quartz crystal-like or crystal-flower-like structures (CHT-mPEG-CHO); and grouped, congealed, steep-sided canyon-like structures (CHT-mPEG COOH). A novel image processing protocol involving DiameterJ and ND plugins of ImageJ software was employed for analyses of the SEM micrographs in terms of % porosity, pore wall thickness and % xiiehaviorxii of the porous scaffolds. The PANi-EC interpenetrating polymer network neurosponges were synthesized employing free radical polymerization under acidic conditions wherein first-in-the-world spinomimetic scaffolds were obtained. The unique feature of the PANi-EC neurosponge was the formation of a fibrous neurotunnel architecture mimicking the native spinal cord. The physicochemical characterization revealed that the secondary structure of the peptide molecules (elastin and collagen) rearranged in the presence of PANi to their native extracellular matrix (ECM) form confirming the self-assembling nature of the polymer-peptide architecture. Furthermore, the PANi-EC neurosponge provided a perfect balance of matrix resilience and matrix hardness similar to the native collagen-elastin complex in vivo. Two very interesting tri-component thermogels were reported here viz. a simple blend thermogel comprising methylcellulose, sodium alginate and poly(ethylene glycol) and a complex thermogel incorporating chitosan, Pluronic F127 and β-glycerophosphate. Both the thermogels solidified at physiological temperature confirming their applicability in vivo. The outstanding feature of MAP thermogels was the formation of hydrogen bonded O-H…C=O which only formed in the tripolymeric blend while the bipolymeric blends showing no such interaction. We proposed that the MAP thermogel self-assembled into a repeating network structure represented by “PEG400-ALG-hydrophillicMChydrophobic}-{hydrophobicMC-hydrophilic}-ALG-PEG400” and the physical “compression” might have led to the formation of hydrogen bonded O-H…C=O among MC/alginate or PEG/alginate in the presence of PEG or MC, respectively. In case of the complex CHT/PF127/βGP thermogel, a unique triphasic gel-sol-gel transition xiiehavior was observed with the thermogel forming a gel-phase at lower temperatures (T<20°C), a sol-phase at intermediate temperatures (20°C35°C). The MTT proliferation studies indicated that all polymer engineered neural devices (PANi-alone matrix) were capable of efficiently supporting the growth of PC12 cells compared to the control over a period of 72 hours. The fundamental objective of this thesis was to test the applicability and capability of various biomaterial composites towards the repair and regeneration of neuronal tissue after traumatic spinal cord injury. Although drug-loaded scaffolds were also developed, only drug-free scaffolds (PLGA-Gliadin 5:5 electrospun nanofibers; PANi-Elastin-Collagen neurosponge; and Chitosan/Pluronic F127/β-glycerophosphate thermogel) were tested in vivo for the proof-of-concept. The 21-point scale BBB locomotor rating analysis demonstrated that PEND I (14), PEND II (19) and PEND III (18) provided significant motor recovery as compared to the lesion-control (5) group 28 days post-SCI and –implantation. The immunohistochemistry confirmed that reparative changes were accompanied by marked upregulation of iNOS, a notable influx of ED1-positive chronic inflammatory cells, the appearance of multinucleate cells characteristic of presumptive regenerative neuroblasts and near-complete loss of GFAP and NF-200 protein/structural integrity. Almost complete functional and neurostructural recovery was observed with post-SCI implantation of PEND II and III. In conclusion, the composite scaffolds tested in this research demonstrated immense potential in improving the neurological, neurochemical, and behavioral outcome after implantation post-SCI.Item Novel antiangiogenic peptide targeted therapeutic nanosystem for non-small-cell lung carcinoma(University of the Witwatersrand, Johannesburg, 2023) Ngema, Lindokuhle Malibongwe; Choonara, Yahya Essop; Marimuthu, Thashree; Adeyemi, AdebawoleLung cancer is the leading cause of cancer deaths globally, with nearly 1.8 million deaths and 2.2 million incidences recorded annually. Primarily, non-small-cell lung carcinoma (NSCLC) is the most commonly diagnosed type of lung cancer, which makes up approximately 85% of all reported lung cancer cases. Currently, the management of NSCLC is a global challenge, and although, various treatment protocols are available, such as surgery, radiotherapy, and chemotherapy, the survival outcomes remain poor. Combination chemotherapy is the current first-line treatment for NSCLC, however, it presents with a myriad of drawbacks, including non-specificity, high dosage, and detrimental side effects, resulting in patients intolerability to the regimen. Consequently, a new therapeutic approach is greatly needed and warrants the design of biocompatible targeted drug delivery nanosystems that can halt tumor proliferation and metastasis by targeting key molecules and deliver drugs directly to tumors, with limited side effects and toxicity to healthy cells. Tumor targeted drug delivery nanosystems such as magnetic nanoparticles (MNPs) modified with biomolecules and functionalized with homing peptides are of great interest for potential application as a potent nanomedicine in NSCLC management. Accordingly, the present study set to develop novel targeted paclitaxel (PTX) delivery nanosystems from the amenable superparamagnetic iron oxide nanoparticles (SPIONs) coated with trans-10,cis-12 conjugated linoleic acid (10E, 12Z) and functionalized with either a vascular endothelial growth factor receptor (VEGFR) binding or a matrix metalloproteinase 2 (MMP-2) binding peptide, for specific delivery of PTX to VEGFR and MMP-2 expressing NSCLC tumors. A preceding nanosystem without the peptides (CLA-coated PTX-SPIONs) was originally fabricated as proof of concept for the application of 10E, 12Z CLA as a surface coating and drug partitioning biomolecule. CLA-coated PTX-SPIONs exhibited a spherical shape, with an average particle size and zeta potential of 96.5 ± 0.6 nm and −27.3 ± 1.9 mV, respectively. The nanosystem had a drug loading efficiency of 98.5% and demonstrated a sustained site-specific in vitro release of PTX over 24 h (i.e., 94% at pH 6.8 mimicking the tumor microenvironment). Enhanced anti-proliferative activity was also observed with the CLA-coated PTX-SPIONs against a lung adenocarcinoma (A549) cell line after 72 h, with a recorded cell viability of 17.1%. Thereafter, the fabricated nanosystem was optimised for direct tumor-targeting by functionalization with HRH or CTT peptides, to give CLA-coated PTX-SPIONs@HRH and CLA-coated PTX-SPIONs@CTT. A new design methodology was established for the tandem surface functionalization of CLA-coated PTX-SPIONs with the antiangiogenic peptides, via coupling reactions. A series of robust nanotechnological techniques were employed for pertinent physicochemical characterization, in vitro evaluation of drug release, anti-proliferative activity, and quantification of VEGF-A and MMP-2 levels. Meanwhile, in vivo testing was carried out on a lung tumor xenograft mouse model. Both nanosystems exhibited a marked cellular uptake and internalization by A549 cells, and CLA-coated PTX-SPIONs@HRH significantly reduced secretion levels of VEGF-A in human dermal microvascular endothelial cells (HMEC-1) from 46.9 pg/mL to 35.6 pg/mL, meanwhile CLA-coated PTX-SPIONs@CTT significantly inhibited MMP-2 secretion by almost 70% , indicating specific anti-MMP-2 activity. A 76.6% and 69.7 % tumor regression was observed in a lung tumor xenograft mouse model treated with CLA-coated PTX SPIONS@HRH and CLA-coated PTX-SPIONs@CTT, respectively, demonstrating tumor targetability and angiogenesis inhibition. Lastly, the pharmacokinetics (PK) evaluation revealed that both nanosystems prolonged the half-life of PTX and circulation time in vivo. In essence, potent antiangiogenic tumor-targeted PTX delivery nanosystems were successfully fabricated, and the obtained results suggest potential application of CLA-coated PTX SPIONs@HRH and CLA-coated PTX-SPIONs@CTT for effective management of NSCLC.