Molecular, Cellular, and Tissue Engineering Commons^™

Autofluorescence Spectral Unmixing For Quantitative Metabolic Imaging Within Tissues, Lucy Woodbury

Biomedical Engineering Undergraduate Honors Theses

Label-free metabolic imaging through quantification of NADH and FAD autofluorescence has become a powerful and efficient tool for non-invasive measurements of cell metabolism. It has applications in a variety of fields including diagnostic and therapeutic monitoring. However, NADH and FAD imaging typically selects specific single excitation and emission bands where it is assumed that these fluorophores are individually isolated. In some cases of isolated cell imaging, this assumption holds true, but for samples with other intrinsic fluorophores present (i.e. collagen and elastin), it can interfere with quantitative results. Elastin and collagen autofluorescence prevent the broader application of these optical metabolic …

Stirred Suspension Bioreactor Differentiation Of Human Mesenchymal Stem Cells Into Smooth Muscle Cells, Chris Slavin

Biomedical Engineering Undergraduate Honors Theses

Human mesenchymal stem cells (hMSCs) are a promising candidate for cellular therapies due to their multipotency, self-renewal capacity, and immunomodulatory properties. However, their isolation is a difficult and potentially painful process with very low yield, and traditional static mammalian cell culture techniques are too slow and expensive for large scale growth and differentiation of stem cells to be practical. Current research is focused on improving methods for cultivating hMSCs in stirred suspension bioreactors (SSBs), but little work has been done with regard to their differentiation in dynamic conditions such as those in SSBs. Differentiation at a large scale would increase …

The Utilization Of Autofluorescence To Study The Effects Of L-Buthionine-Sulfoximine On Cellular Metabolism In Vitro, Madison Belew

Biomedical Engineering Undergraduate Honors Theses

Oxidative damage and stress impacts metabolic activity and has been implicated in a number of diseases . These medical conditions have been found to be associated with a lack of glutathione (GSH). There is a need to study the effects that the absence of glutathione has in order to expand knowledge of indicators that cause conditions like cataracts, cancer, or impaired wound healing. L-buthionine-sulfoximine (BSO) has been used in past studies to hinder GSH production and induce oxidative stress within a cell. This study explores the effects of BSO and induced oxidative stress on the metabolic activity and processes within …

Cell Line-Specific Differences In Guided Differentiation Of Adipose-Derived Mesenchymal Stem Cells Towards Smooth Muscle Cells, Sherly Makar

Biomedical Engineering Undergraduate Honors Theses

Mesenchymal stem cells (MSCs) are a population of stromal cells found traditionally in the bone marrow and adipose tissues. They can also be found in other tissues including fallopian tube, core blood, peripheral blood, fetal liver, and lungs. Mesenchymal stem cells have profound effects in regenerative engineering, tissue repair and drug discovery owing to the excellent properties such as proliferation, self-renewal, and multipotency generating multiple cell types including adipocytes, osteocytes, cardiomyocytes (CMs), pericytes (PCs), and chondrocytes. MSCs are used as immunomodulators in generating progenitor cells to be used for transplantation, creating engineered organs, and preventing graft vs. host disease (GVHD). …

Comparative Genomics Of Human Mesenchymal Stem Cells And Human Mesenchymal Stem Cells Derived Vascular Smooth Muscle Cells, Samia Ismail

Biomedical Engineering Undergraduate Honors Theses

As of 2017, vascular diseases contributed to 23.1% of all deaths in America. To address the need for more effective and sustainable treatment options for these ailments, stem cell differentiation and implantation has emerged as a viable alternative to standard bypass and graft insertions. A completely autologous treatment can be achieved by extracting adult stem cells, differentiating them into vascular smooth muscle cells (VSMCs), and then reimplanting these cells at the affected tissue site. This study aims to investigate the efficiency of the VSMC differentiation from human mesenchymal stem cells (hMSCs) by comparing 4 cell lines of untreated hMSCs with …

Generation Of A Ccl2 Knockout Using Crispr/Cas9 And Lipid Mediated Transfection In Ct-26 Murine Colon Carcinoma Cells, Emma Sullivan

Biomedical Engineering Undergraduate Honors Theses

CCL2 is an inflammatory mediator that is released by tumor cells to activate and direct immune cell species, especially macrophages, to inflammatory sites within the body. The goal of this project was to successfully generate knockout the CCL2 ligand gene using a CRISPR/Cas9 complex delivered via lipid mediated transfection. The sgRNA and Cas9 mRNA were introduced into the cells via lipid-mediated transfection. The cells were incubated for 4 days, before being analyzed using PCR and gel electrophoresis. We expected to see one band on the first gel and two bands on the second gel. Two bands appeared on the first …

Investigating Virus Clearance Via Ph Inactivation During Biomanufacturing, Wenbo Xu

Biomedical Engineering Undergraduate Honors Theses

In the processing of biopharmaceuticals, viral clearance and viral safety are important for the development of monoclonal antibodies. Murine xenotropic leukemia virus (XMuLV) is one of the retroviruses, recommended by Food and Drug Administration (FDA) as a model virus for viral clearance via inactivation from therapeutics derived from Chinese hamster ovary cells (CHO). A robust and effective method was investigated to clear or inactivate endogenous viruses by low pH inactivation. The effects of different conductivity and inactivated time on XMuLV clearance was determined. Acetate buffer was prepared with different conductivity, and 2% XMuLV was spiked into acetate buffer. XMuLV virus …

Phenotypic Characterization Of Differentiation Of Human Mesenchymal Stem Cells To Smooth Muscle Cells, Jackson Hedrick

Biomedical Engineering Undergraduate Honors Theses

Abdominal aortic aneurysms (AAAs) are expansions in the aortic wall with an increased tendency to rupture. These expansions are triggered by the breakdown of key aortic extracellular matrix (ECM) proteins—elastin and collagen. The degradation of aortic ECM proteins is the consequence of overexpressed matrix metalloproteases (MMPs)–MMP-2 and MMP-9. Adult vascular smooth muscle cells (SMCs) lack the ability to repair the elastic matrix assembly; thus, it is imperative to find a solution to both restore the imbalance of essential ECM proteins in the aorta and mitigate the effects of MMPs. Previous studies have revealed that bone marrow derived mesenchymal stem cells …

Comparing The Effects Of Fibroblast Growth Factors On Growth Rate Of Human Fibroblast Cell Lines, Tasha Repella

Biomedical Engineering Undergraduate Honors Theses

Numerous studies have demonstrated the usefulness of growth factors in medium formulations to promote proliferation and viability of human fibroblasts in laboratory settings. However, due to the differences in the source and age of fibroblasts, the ability of growth factors like hFGF1 to promote proliferation varies. Equally difficult is the ability of hFGF1 to promote proliferation of diseased fibroblasts due to complexities associated with specific mitochondrial diseases. In this context, we decided to evaluate the potential for novel hyper stable growth factors super hFGF and hFGF1 (shFGF and hFGF1 respectively), to contribute to proliferation of different normal and diseased fibroblasts. …

Effects Of Gsk101 Exposure On Astrocyte Mechanical Properties, Collagen Production, And Viability, Ethan Echols

Biomedical Engineering Undergraduate Honors Theses

Transient receptor potential vanilloid 4 (TRPV4) is a cation channel protein which is thought to facilitate extracellular matrix (ECM) modification and glial scar formation following traumatic brain injury (TBI). Activation of TRPV4 has been shown to correlate with an increase in intracellular Ca²⁺ concentration. TRPV4 is subject to both chemical and mechanical activation. This study involved chemical activation of TRPV4 in cultured mouse astrocytes using GSK101, the primary agonist of TRPV4, and subsequent evaluation of mechanical and chemical changes to the ECM. Prior to GSK101 exposure, the cells were seeded into Matrigel Matrix, a gelatinous protein mixture intended to …

Quantification Of Tumor-Associated Macrophages Following Immunomodulation Therapy In A Murine Allograft Model Of Colorectal Carcinoma, Caroline Spainhour

Biomedical Engineering Undergraduate Honors Theses

Colorectal Carcinoma (CRC) is one of the deadliest cancers in the world, with 150,000 new cases annually in the United States. Traditional treatments include chemotherapy and invasive surgery; however, research has shown that only 25% of patients that undergo traditional treatment have a positive result. Immunotherapy is an emerging form of cancer treatment that utilizes the patients’immune system to fight cancer cells by targeting inflammation, which plays a large role in the proliferation and metastasis of cancer cells.

Tumor-associated macrophages (TAMs) are immune cells that affect the inflammatory microenvironment of tumors. TAMs are M1 in the early stages of tumors, …

Analysis Of Biological Response To Ecm Hydrogel Injection, Grady Dunlap

Biomedical Engineering Undergraduate Honors Theses

Abstract

Under normal circumstances, skeletal muscle possesses the capacity to regenerate and heal via inflammatory and myogenic pathways. In cases of severe tissue loss or certain diseases, this capacity is lost, often resulting in loss of tissue function. Extracellular matrix (ECM), the protein scaffold which houses cells in physiological tissue, has been shown to have structural and chemical properties which influence cell migration and phenotype. This results in ECM’s capacity to encourage a regenerative response when implanted into severely damaged skeletal muscle. Additional advantages are apparent when an ECM scaffold is digested into a hydrogel, namely less invasive implantation via …

Effects Of Uniaxial Cyclic Strain On Endothelial Progenitor Cells, Maria Alejandra Zeballos Castro

Biomedical Engineering Undergraduate Honors Theses

Despite the high prevalence of calcific aortic valve disease (CAVD), the underlying mechanisms of pathogenesis have not been found yet. Therefore, it is extremely important to study CAVD and understand how it develops. For this matter, we decided to study the potential of endothelial progenitor cells (EPCs) for use in tissue-engineered models of heart valves. EPCs were chosen as the cell source of interest for this study due to their high neovascularization potential and use in regenerative medicine and cardiovascular tissue engineering.

In this project, we aimed to engineer the microenvironment of cells that are involved in the formation of …

Age's Effect On Regenerative Capabilities Of Myocytes Through Satellite Cell Analysis, Karam S. Sra

Biomedical Engineering Undergraduate Honors Theses

The objective of this study was to investigate potential effects aging had on muscle fiber area and satellite cell count in myocytes. This research could help elucidate the detrimental effect age has on regenerative capabilities whether in terms of satellite cell function or satellite cell number. Satellite cells are primarily responsible for generating new muscle tissue after being activated through mechanotransduction of injury. This study utilized immunofluorescence to examine the presence of the PAX7 gene expression, a unique marker of satellite cells, within a 12 month and 18 month old population of mice. The PAX7 marker was co-stained with DAPI …

Detection Of Leukocytes Stained With Acridine Orange Using Unique Spectral Features Acquired From An Image-Based Spectrometer, Courtney J. Hunter

Biomedical Engineering Undergraduate Honors Theses

A leukocyte differential count can be used to diagnosis a myriad blood disorders, such as infections, allergies, and efficacy of disease treatments. In recent years, attention has been focused on developing point-of-care (POC) systems to provide this test in global health settings. Acridine orange (AO) is an amphipathic, vital dye that intercalates leukocyte nucleic acids and acidic vesicles. It has been utilized by POC systems to identify the three main leukocyte subtypes: granulocytes, monocytes, and lymphocytes. Subtypes of leukocytes can be characterized using a fluorescence microscope, where the AO has a 450 nm excitation wavelength and has two peak emission …

Characterization Of The Response Of Trpv4 To Chemical Stimulation, Jacob V. Schluns

Biomedical Engineering Undergraduate Honors Theses

Traumatic Brain Injury (TBI) is a source of acute and chronic health issues for many patients. One of the components of the brain’s response to injury is astrogliosis, in which astrocytes that normally function to repair the brain instead form scar tissue that halts repair processes. Transient Receptor Potential Vanilloid Type 4 (TRPV4) is a trans-membrane calcium channel involved in astrogliosis. Through Fura-2AM based calcium imaging, the base activity of this channel in mouse astrocyte cells was recorded. The cells were then subjected to TRPV4 agonist and antagonist stimulation and their subsequent activity levels were recorded. The data showed that …

Role Of Trpv4 In Astrocyte Extracellular Matrix Production, Abby Terlouw

Biomedical Engineering Undergraduate Honors Theses

Traumatic Brain Injury (TBI) is an alteration of brain pathology following damage of the central nervous system (CNS) by an external force. In the CNS, glial scar formation often occurs following TBI, and astrocytes are widely believed to contribute to this scar formation. While the role of astrocytes in extracellular matrix (ECM) production is known, the exact mechanism(s) for this event remain unclear. One possible method is the activation of transient receptor potential vanilloid 4 (TRPV4). TRPV4 is a channel protein found in the astrocyte membrane which has been shown to generate intracellular calcium ions following mechanical stimulation. Previous research …

Isolation Of Metallic Single-Walled Carbon Nanotubes For Electrically Conductive Tissue Engineering Scaffolds, Jakob Hockman

Biomedical Engineering Undergraduate Honors Theses

Metallic single-walled carbon nanotubes (m-SWNTs) were separated from pristine SWNTs using affinity chromatography for use in electrically conductive tissue engineering scaffolds. Approximately one third of SWNTs have metallic properties. Separations were achieved using a protocol modified from Liu & coworkers (2011) in order to improve the method for cell culture environments. Samples enriched in m-SWNTs were isolated and characterized. However, challenges still remain for the complete separation of m-SWNTs from their semiconducting counterpart (s-SWNTs) using this protocol. Approaches to improve separation and reduce the difficulties associated with processing the nanotubes were suggested. One of the ultimate destinations of these nanotubes …

Exploring Biomarkers For Point Of Care Bladder Cancer Detection, Cassandra M. Walker

Biomedical Engineering Undergraduate Honors Theses

Bladder cancer is the 5th most common non-cutaneous human cancer in the United States. While effective methods of detecting bladder cancer are currently practiced, they are often expensive and invasive. There is a need for a noninvasive detection method that can be used in areas with few medical resources. Cell free DNA in urine is normally present only in very low concentrations. Abnormally high levels of cell free DNA in urine could be indicative of disease. This study tests the hypothesis that DNA present in urine can be used as a biomarker for bladder cancer before hematuria is seen in …