Biomarkers capable of diagnosing and tracking the progression of Parkinson’s disease (PD) are urgently needed. Saliva has been explored as an alternative to a more traditional sample source such as cerebral spinal fluid (CSF), which involves painful and invasive collection processes, and blood. Indeed, PD patients typically have dysfunction of salivary production, indicating that the disease involves this organ system. In order to increase sample availability, many protocols use cotton or plastic to stimulate saliva production. However, this raises the question of how these materials affect salivary protein composition, specifically in regards to proteins of interest as biomarkers. In order to explore these effects I incubated various materials potentially capable of stimulating increased saliva production in saliva samples and measured the concentration of alpha-synuclein (α-syn), a protein believed to be important in PD pathogenesis and a possible biomarker, and total protein. By comparing the concentrations of α-syn, total protein, and the α-syn:total protein ratio between the various material groups and control (no material) groups I am able to determine whether the materials increased, decreased, or had no effect on protein concentration. It was hypothesized that all tested materials would decrease both α-syn and total protein concentrations by some degree due to protein attraction to the materials. The results of this study contribute to a clearer basis for salivary biomarker studies involving induction techniques, facilitating collection techniques in a clinical setting, particularly in older patients with decreased saliva production.

Chitosan and alginate are widely used natural polymeric biomaterials that are biocompatible, biodegradable, and non-toxic. Chitosan is cationic as well as hydrophobic in nature whereas alginate is anionic and hydrophilic in nature. Ionic interaction between the amine and carboxyl group causes the formation of polyelectrolyte complex in the scaffold and thus enhancing the mechanical properties of the scaffold. Mechanical properties of the scaffolds are dependent on the fabrication process, cross-linking process, and sterilization process. Similarly, porosity plays a crucial role in determining the mechanical strength of the scaffold which relates to the lyophilization process and molecular weight of the polymer used. Chitosan-Alginate (CA) scaffolds are prepared by mixing equal wt% amounts of each polymer and due to chitosan's hydrophobic nature, acetic acid was added. The solution was mixed using a Thinky mixer at 2000 rpm for five minutes. After obtaining a homogeneous mixture, pH was tested and solution  casted into desired well plates. The solution was then refrigerated at 4 degree Celsius for 12 h at 20 degree Celsius overnight, and lyophilized for 24 hours. Scaffolds were then cross-linked with 70% CaCl2, and sterilized with Phosphate-buffered saline (PBS). Scanning electron microscope (SEM) is used in to image pore morphology of the scaffold. High mechanical strength and porous structure is presented in pore size ranges of 100-500 micrometers which is required for bone tissue engineering. Mechanical strength of the scaffold is obtained by performing compressive test. Samples are compressed at 4.00 mm/min using micro-tensile tester with 1N load cell. Dry samples were found to have a young's modulus (YM) of 1-2 MPa, whereas wet samples were found to have a YM of 1-2 KPa.  CA scaffolds were found to be mechanically strong with good porosity which was directly impacted by fabrication process. Mechanically strong CA scaffolds with proper porosity were achieved with the fabrication process used. Further work will allow an optimization of CA scaffolds for specific applications and cell lines.  

Tissue engineering is a field of study where porous scaffolds made of materials such as polymers and/or ceramics are used to promote tissue regeneration. This is important for treatment of tissue defects resulting from trauma or disease where repair does not occur via normal, physiological processes. In this work we fabricated 3D, porous scaffolds made of naturally-derived polymers, chitosan and alginate, using a phase separation and lyophilization technique. This chitosan-alginate (CA) scaffold is promising for tissue engineering because it is made of sustainable materials and is biocompatible, biodegradable, and cost-effective. We characterized the porosity of the scaffolds using scanning electron microscopy and observed interconnected pores on the order of 100 microns in diameter. As a first step in determining the potential of this CA scaffold for muscle tissue engineering, we cultured C2C12 mouse myoblasts and evaluated cell viability and proliferation using the Alamar Blue assay. The scaffolds support cell attachment and viability and we expect to observe cell proliferation.

The goal of my research is to  find an efficient drug delivery system to kill brain cancer cells by considering that some hydrogels change their viscosity dramatically at different temperatures (called thermosensitive). The whole project is regarded as “rocket”system, with a drug-loaded device that delivers content into the right place as a rocket and the drug  inside as an astronaut. The innovative idea of the project is that the drug-loaded system has the capability to be liquid-like at low temperature while becoming gel at body temperature, making it easy to produce and inject into tumor tissue to concentrate the effect at the tumor site. To find a stable, biocompatible and biodegradable material to be drug-loaded, we tested the in vitro stability and drug-release during 1hr, 24hr and 96hr incubation of various polymer gels including PEG-chitosan, alginate, and methyl cellulose. We then studied the drug delivery system with brain cancer cells in vitro and monitored the migration of the cancer cells towards the gel.

Some symptoms of Parkinson’s disease, a movement disorder characterized by neuronal death in the basal ganglia, may result from or be exacerbated by low-level long-term exposure to manganese. Manganese-mediated impairment of normal glutamate uptake and metabolism may also contribute to abnormalities in the neuroprotective functioning of astrocytes, including manganese and other toxicant storage. Misfolding and aggregation of the protein α-synuclein, a product of the SNCA gene, has been implicated in neuronal degeneration in Parkinson’s disease, and various studies have proposed links between manganese exposure or defects in manganese metabolic systems and synuclein toxicity. Preliminary data from our laboratory suggest that knockout (KO) of SNCA in astrocytes attenuates some physiological effects of manganese exposure. The present study seeks to compare toxicant attenuation in SNCA KO and wild type (WT) astrocytes after increasing low-concentration 24-hour manganese pre-treatments. Imaging with fluo-4 dye will be used to observe glutamate-induced calcium waves in astrocyte cultures exposed to 1, 5 and 10 μM Mn2+. We predict significant deviations from a control waveform profile in both SNCA KO and WT cultures treated with higher concentrations of manganese. We further predict WT cultures to demonstrate significantly altered waveform profiles compared to SNCA KO cultures treated with equal concentrations of manganese.