Date of Award

5-8-2015

Document Type

Open Access Dissertation

Department

Chemistry and Biochemistry

First Advisor

Qian Wang

Second Advisor

John H. Dawson

Abstract

Topographical cues can profoundly affect cellular behaviors. This thesis investigate how to utilize the topographical cues generated by two techniques, flow assembly and electrospinning, to regulate cellular behaviors.

First of all, a facile and robust method to align one-dimensional (1D) nanoparticles (NPs) in large scale has been developed. Using flow assembly, representative rod-like nanoparticles, including tobacco mosaic virus (TMV), gold nanorods and bacteriophage M13, have been aligned inside capillaries by controlling flow rate and substrate surface properties. The properties of 1D NPs, such as stiffness and aspect ratio, play a critical role in the alignment. Furthermore, these hierarchically organized structures can be used to support cell growth, where cell orientation and morphology are controlled. When C2C12 myoblasts were cultured on surfaces coated with aligned TMV, we found that nanoscale topographic features were critical to guide the cell orientation and myogenic differentiation. This method can be used in the fabrication of complex assemblies with 1D NPs and has wide applications in tissue engineering, sensing, electronics and optical fields.

Furthermore, we combined this flow assembly method with genetically modified TMV mutants with reported cell adhesion sequences (i.e. RGD1, RGD7, PSHRN3, P15, and DEGA) to generate biomimetic substrates with specific cell adhesion motifs and precisely controlled structural organization for guiding cell behaviors by providing desired biochemical and physical cues. We found that TMV mutants significantly promote neurite outgrowth and the resultant aligned TMV mutants substrates were able to dictate directional neurite outgrowth of N2a cells. Hence, the plant virus-based materials provide tremendous promise for neural tissue engineering in the future.

In addition, we generated the electrospun polycaprolactone (PCL) microfibers for three dimensional (3D) culture of breast cancer cell lines, MCF-7. We found that cancer stem cells (CSCs), a small group of tumor-initiating cells within tumors as the main contributors of tumor growth, metastasis, and recurrence, have significantly increased the proportion in the whole population. The expression of stem cell markers, including OCT3/4 and SOX2, and breast CSC-specific markers, SOX4 and CD49f, was significantly upregulated, and the mammosphere-forming capability in cells cultured in 3D PCL scaffolds increased. The fibrous scaffolds also induced the elongation of MCF-7 cells and extended cell proliferation. The increase of CSC properties after culturing in 3D scaffolds was further confirmed in the other two luminal-type mammary cell lines, T47D and SK-BR-3, and a basal-type cell line, MDA-MB-231. Moreover, we observed the upregulation of epithelial to mesenchymal transition and increased invasive capability in cells cultured in 3D PCL scaffolds. These data suggests that the increase of CSC proportion in 3D culture system may account for the enhanced malignancy. Therefore, our 3D PCL scaffolds can potentially be used for CSCs enrichment and anti-cancer drug screening.

Finally, by electrospinning of polycaprolactone solutions containing N-(benzoylthio)benzamide (NSHD1), a H2S donor, we fabricated fibrous scaffolds with hydrogen sulfide (H2S) releasing capability (H2S-fibers). The resultant microfibers are capable of releasing H2S upon immersion in aqueous solution containing biological thiols under physiological conditions. The H2S release peaks of H2S-fibers appeared at 2~4 hours, while the peak of donor alone showed at 45 minutes. H2S release half-lives of H2S-fibers were 10-20 times longer than that of donor alone. Furthermore, H2S-fibers can protect cells from H2O2 induced oxidative damage by significantly decreasing the production of intracellular reactive oxygen species (ROS). Given that H2S has a broad range of physiological functions, H2S-fibers hold great potential for various biomedical applications.

In summary, the emerging area of nanotechnology have been applied for biomedical application as well as the fundamental study of the interaction between cell behaviors and surrounding nanomaterials. Here, we have generated a robust and facile method that can align various 1D NPs in a large quantity. The resulting substrates in capillaries can be used to guide the directionality of cell growth. Meanwhile, we investigated the effects of electrospun microfibers on breast cancer stem cells and fabricated H2S releasing fibers, which could be used in tissue engineering.

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