Doctor of Philosophy, Case Western Reserve University, 0, Neurosciences

Neural circuits integrate experience and store information by the formation and remodeling of synapses. Defining the signaling pathways that underlie such plasticity has been a major goal of modern neuroscience. BMP signaling critically regulates both morphological plasticity and neurotransmission in motorneurons at the Drosophila neuromuscular junction (NMJ). However the mechanisms that establish BMP pathway directionality and diversify pathway action in motorneurons remain elusive. In this thesis, I describe a novel regulator of the BMP signaling pathway in Drosophila motorneurons that I name Crimpy (Cmpy). Cmpy acts at the interface between morphological plasticity and synaptic transmission to specify a role for motorneuronal BMP ligand in promoting synaptic transmission, as opposed to morphological growth, at the NMJ. Using the Drosophila NMJ as a model system, I examined the role of Cmpy in motorneuron development. BMP signaling scales growth of the presynaptic axon arbor to growth of the postsynaptic muscle during larval development in order to preserve synaptic input onto the muscle cell. Genetic analyses suggest that the relevant BMP ligand at the NMJ, Glass bottom boat (Gbb), acts in a retrograde fashion, secreted from the postsynaptic muscle to activate presynaptic BMP receptors and promote motoraxon growth. However establishment of this directionality is unclear since Gbb is also produced by and active within motorneurons, albeit to regulate synaptic transmission as opposed to morphological growth. I found that loss of cmpy results in excessive presynaptic growth due to ectopic motorneuron-derived, autocrine pro-growth BMP signaling at the NMJ. Biochemical analyses demonstrated that Cmpy binds to precursor BMP. Expressing a novel Gbb-HA transgene within larval motorneurons that preserves protein processing and secretion, and functions like a wild-type Gbb transgene, I detected presynaptic Gbb-HA localization at the NMJ. Cmpy is necessary and suffic (open full item for complete abstract)

Committee: Heather Broihier (Advisor); Ben Strowbridge (Committee Chair); Christopher Wilson (Committee Member); Stephen Maricich (Committee Member); Jocelyn McDonald (Committee Member) Subjects: Developmental Biology; Neurosciences

PhD, University of Cincinnati, 2022, Medicine: Molecular Genetics, Biochemistry, & Microbiology

Bone Morphogenetic Proteins (BMPs) are the largest subgroup of the Transforming Growth Factor ß (TGFß) superfamily, one of the fundamental protein signaling pathways in biology. BMPs are involved in regulating numerous biological functions, with a particular focus on development, immune modulation, cell homeostasis and wound healing. When dysregulated, aberrant BMP signaling can indue a host of developmental and autoimmune disorders, as well as many different cancers. Given the wide array of biological functions BMPs regulate, precise regulation of signaling is a key component of their biology. Mechanistically, these secreted dimeric signaling proteins function by forming complex with two type 1 and two type 2 serine/threonine kinase receptors on the cell surface to drive signaling by intracellular SMAD proteins. Accordingly, the regulation of these potent signaling molecules in the extracellular space is a vital area of study. The purpose of the work outlined in this thesis is to explore certain understudied mechanisms of extracellular modulation of BMP signaling. We particularly focused on studying these mechanisms not in isolation, but rather as they actually exist in nature, as part of a complex environment with many competing biomolecules. We present studies contrasting related protein antagonists with different function in an attempt to gain insight into the key components needed for BMP inhibition. In addition, we explore a newly discovered interaction between the BMP and Wnt signaling pathways, where BMP ligands may directly antagonize canonical Wnt signaling. Lastly, we describe a procedure for the production of artificial BMP heterodimeric signaling molecules using chains with differential activity with respect to receptor preference, antagonist targeting, and affinity to the extracellular matrix. These asymmetrical signaling molecules were then used to isolate the key components of biological function across multiple different experimental systems. Overal (open full item for complete abstract)

Committee: Thomas Thompson Ph.D. (Committee Member); Rhett Kovall Ph.D. (Committee Member); Aaron Zorn Ph.D. (Committee Member); James Wells Ph.D. (Committee Member); Michael Tranter Ph.D. (Committee Member) Subjects: Biochemistry

Master of Science (MS), Bowling Green State University, 2020, Geology

Conventional tillage, a soil preparation practice to produce a fine seedbed, can disturb the soil profile by promoting soil compaction and soil organic matter (SOM) degradation. In contrast, conservation tillage, such as no-till and minimal tillage (30% or more crop residue) have the potential to sustain or increase soil organic carbon (SOC). Additional benefits of conservation tillage include; improvement to soil structure, reducing soil erosion, greater water retention, buffering soil temperatures, and greater crop residue retention. Conservation tillage practices promote nutrient retention in soils. Furthermore; Fe- and Mn-(oxy)hydroxide minerals play an important role in SOC stabilization and sequestration, which also promotes nutrient adsorption. This study aimed to 1) quantify SOC under varying agricultural managements, 2) qualitatively describe the degree of aromaticity and recalcitrance of SOC using fluorescence spectroscopy, 3) correlate SOC quantity with nitrogen and phosphorous retention in soils, and 4) understand the mineral phases responsible for the stabilization and sequestration of SOC, as well as phosphate and nitrate using a four-step chemical sequential extraction. Results showed that no till and minimal tillage sites consistently had greater SOC and fluorescence intensity in the humic-like acids region, when compared to conventional tilled fields. The SOC quality was obtained using relatively quick and cost-effective methods. No till and minimal tillage enhanced SOC stabilization. In addition, conservation tillage practices retained the largest total nitrogen and total phosphorous concentrations at all studied depths (0-30 cm), when compared to conventional tilled fields. Sequential extraction results showed that SOC was stabilized in the following order: crystalline Fe-oxides > amorphous Fe-oxides > Mn-oxides. Fe- and Mn-(oxy)hydroxide minerals can promote the stabilization and long-term sequestration of SOC via the formation of inner sphere (open full item for complete abstract)

Committee: Angélica Vázquez-Ortega Dr. (Advisor); Andrew Gregory Dr. (Committee Member); Ganming Liu Dr. (Committee Member) Subjects: Geology

Doctor of Philosophy, University of Toledo, 0, Biomedical Engineering

Cellulose and chitosan (CS) are two of the most abundant biopolymers in nature. CS is highly explored in tissue engineering applications mainly because of its unique chemical structure of possessing cationic amine groups. This bestows it with the ability to have specific interaction with extracellular matrix components such as glycosaminoglycan and proteoglycans making it even more attractive in bone regeneration. This study focused on the application of these biopolymers towards the development of microparticles and hence robust injectable bone scaffolds. The current strategy of applying MPs for bone regeneration involves their direct injection to the defect site. This technique most of the time, however, leads to the improper localization of those MPs. This becomes more problematic when these MPs are loaded with therapeutic agents or growth factors as the intended localized delivery of those agents could not be achieved. Another strategy that has been implemented for the use of MPs in bone regeneration involves the development of preformed solid scaffolds created mostly through chemical agglomeration or sintering techniques. This technique, however, takes away the injectable property of MPs making them like the conventional solid scaffolds. To address this issue related to the applications of the MPs and to develop effective therapeutics releasing scaffolds, the concept of embedding microparticles into a thermosensitive gel was used. This MPs-gel composite was developed to stay as a liquid at a lower temperature enabling an easy injection, but it undergoes a temperature-induced transition to gel at physiological temperature (37 oC). This gel enables the proper localization of MPs at the target defect site enabling efficient therapeutic effects. The blend of methylcellulose (MC) and alginate (Alg) was used to develop thermosensitive gel. The gelation occurs primarily due to the physical interaction among the hydrophobic chains of MC and is further strengthened by (open full item for complete abstract)

Committee: A. Champa Jayasuriya (Committee Chair); Arunan Nadarajah (Committee Member); Sarit Bhaduri (Committee Member); Ahalapitiya H. Jayatissa (Committee Member); Jiayong Liu (Committee Member) Subjects: Biomedical Engineering

Master of Sciences, Case Western Reserve University, 2018, Biology

The Drosophila blastoderm embryo is patterned by morphogens, molecules that inform cells to adopt specific fates in a threshold-dependent fashion. The morphogen Bicoid subdivides the antero-posterior axis into head, thorax and abdomen, whereas the morphogens Dorsal/NFk-B and Decapentaplegic/BMP-4 regulate cell fates along the dorso-ventral axis and subdivide the embryo into three germ layers (mesoderm, neuroectoderm and ectoderm). Together, the concentration gradients produced by these morphogens form a coordinate system along the main embryonic axes and cell fate is influenced depending on where a cell falls inside this system. For several years, the blastoderm was considered to be a static stage due to the lack of mitotic divisions or any major gastrulation movements. However, recent work demonstrated that a stereotyped movement of cells towards the dorsal midline from the lateral regions and the poles of the embryo takes place. As a result, by the end of the blastoderm stage, there is a higher density of cells in the dorsal region of the embryo and a lower density of cells in the ventral region. Results from our lab and others suggest that Decapentaplegic and Dorsal are required for these stereotyped cell movements. In this work, we tested whether the candidate gene frazzled (fra) we identified in a bioinformatics screening is a downstream regulator responsible for directing cells towards the dorsal region of the embryo. fra encodes a transmembrane receptor previously implicated in glial cell migration during late embryonic stages. The function of fra in the early blastoderm is unknown. In this work, we show that in embryos mutant for fra, cells lose their migration direction towards the dorsal midline and display an increased density within the ventral side compared to wild type embryos. We have also observed that FRA protein is located on the dorsal side of the Drosophila embryo and is localized on the apical surface of the cell vertices. Our findings suggest (open full item for complete abstract)

Committee: Claudia Mizutani (Advisor); Sarah Diamond (Committee Chair); Dianne Kube (Committee Member); Emmitt Jolly (Committee Member) Subjects: Biology; Developmental Biology; Genetics

Master of Science, University of Toledo, 2018, Geology

The Maumee Watershed, the largest of the Great Lakes drainages, is located in NW Ohio, and is comprised of 70% agricultural land. In an effort to increase crop yields, fertilizer containing phosphorous (P) and nitrogen (N) are applied to the agricultural land. During large rain events or melt off periods these nutrients are mobilized, ultimately making their way to Lake Erie. Anthropogenic nutrient loading and increased water temperature allow for cyanobacterial harmful algal blooms (HABs) to prosper. Climate change is expected to increase both average temperature and precipitation events, potentially changing nutrient loading from runoff and influencing an increase in HABs. Best management practices (BMPs) have been utilized on agricultural land to help reduce and slow nutrient runoff. As HABs are detrimental to human health, understanding how climate change and BMPs interact to reduce nutrient loading is essential. A target reduction of 40% dissolved reactive phosphorus (DRP) nine years out of ten was established by the Ohio Phosphorus Task Force, in an effort to reduce HAB occurrences. To investigate how climate change and BMPs interact, three climate models, MIROC, MRI, and CNRM, and a baseline scenario and two BMPs are run in a Soil and Water Assessment Tool (SWAT) model of the Maumee Watershed. MIROC predicts the warmest and driest climate amongst the three climate models, while MRI foresees the coolest and wettest climate. MIROC and MRI both anticipate a decrease in P loading on average, with some decreases in P offset by climate change for MRI. Finally, CNRM anticipates a mild increase in temperature and precipitation, but increases P loading overall. Based on similar hindcast results to NOAA's measured data it is likely that CNRM is the most realistic climate model, but is inconclusive. BMPs are an effective practice in reducing P loading as scenario 2 and scenario 3 reduce P in reference to baseline management. More aggressive management will need to be im (open full item for complete abstract)

Committee: Richard Becker (Committee Chair); Kevin Czajkowski (Committee Member); James Martin-Hayden (Committee Member) Subjects: Climate Change; Environmental Geology

Master of Science, The Ohio State University, 2018, Dentistry

Guided bone regeneration (GBR) has been used to promote osteogenesis in a bone defect surrounding a dental implant. While bone quantity at the implant interface has been investigated to evaluate stability of the implant system, bone quality is also responsible for determining mechanical response of bone around the implant. The objective of this study is to examine the nanoindentation based elastic modulus (E) and plastic hardness (H) at different bone regions adjacent to titanium dental implants with GBR treated with demineralized bone matrix (DBM) and bone morphogenetic protein (BMP) during different post-implantation periods. Six adult male beagle dogs were used to create circumferential defects (Ø6.3 mm x 4 mm) with buccal bone removal at each implantation site of mandibles following 3 months post-extraction healing periods. Four titanium dental implants were bilaterally placed in each dog mandible. The implant systems were randomly assigned to only GBR (control), GBR with DBM (DBM), and GBR with DBM+rhBMP-2 (BMP) groups. Three animals were sacrificed at each 4 and 8 weeks of post-implantation healing periods. The bone-implant constructs were buccolingually dissected for nanoindentation. The E and H values were assessed by 980 indentations at the defects (Defect), interfacial bone tissue adjacent to the implant (Interface), and pre-existing bone tissue away from the implant (Pre-existing). The E and H values of BMP group had significantly higher than control and DBM groups for interface and defect regions at 4 weeks of post-implantation period and for the defect region at 8 weeks (p<0.043). The DBM group had higher E and H values than control group only for the defect region at 4 weeks (p<0.001). The current results indicate that treatment of rhBMP-2 with GBR accelerates bone tissue mineralization, which enhances bone quality of the defect region during early post-implantation healing and maintain it for longer healing period. The GBR likely facilitate a microenv (open full item for complete abstract)

Committee: Do-Gyoon Kim (Advisor); Michael Beck (Committee Member); Toru Deguchi (Committee Member) Subjects: Dentistry; Mechanics

Master of Engineering, Case Western Reserve University, 2017, Biomedical Engineering

Lower-limb fracture exhibits a limited capacity to heal critically sized defects and biomimetic tissue engineering is a promising approach for addressing this clinical need. Here, scaffold-free tubular mesenchymal condensations featuring temporally controlled TGF-Β1 and BMP-2 morphogen release from incorporated microparticles were engineered to form self-assembled rings and tubes. In vitro culture, subcutaneous, and femoral defect implantation was performed to establish bone forming capacity with the treatment groups: 1) TGF-Β1, 2) BMP-2, or 3) BMP-2+TGF-Β1. Bone formation was evaluated by biochemical, μCT, and histological analyses. Dual-delivery enhanced bone volume versus single morphogens at 6wks, and histology revealed bone within tubular geometries with enhanced cartilage, mimetic of endochondral ossification. Overall: 1) geometry of scaffold-free mesenchymal condensations guided 3D ossification, 2) BMP-2+TGF-Β1 presentation augmented ectopic bone formation, and 3) tubular architecture promoted femoral defect bridging over random organization.

Committee: Eben Alsberg Dr. (Advisor); Steven Eppell Dr. (Committee Member); Ozan Akkus Dr. (Committee Member) Subjects: Biology; Biomedical Engineering; Biomedical Research; Cellular Biology; Engineering; Health Sciences; Histology; Medical Imaging; Microbiology; Morphology

PhD, University of Cincinnati, 2017, Medicine: Molecular and Developmental Biology

The digestive system forms during early fetal development, through intricate events coordinated by growth factors. The embryonic endoderm and mesoderm layers coordinate the formation of the gut progenitors that will give rise to foregut-derived organs such as the liver, pancreas and stomach, and hindgut derived intestine. Signaling pathways like Bone Morphogenetic Protein (BMP) and Wnt are reiteratively used during development. Their downstream targets are variable upon time and target tissue, and the specificity of the outcome is achieved through crosstalk with other factors. Although we know what growth factors act when and where during development, it is still largely unknown how multiple signaling pathways integrate their signals at the genomic level for correct embryonic patterning. In this study we aimed to unveil the transcriptional program of foregut and hindgut progenitors and how BMP/Smad1 and Wnt/β-catenin coordinate their signals to pattern these progenitors. We used genome wide analysis to determine the foregut and hindgut programs in early Xenopus laevis embryos. We found that these transcriptional programs are highly conserved with mammals. Upon signaling manipulation, we found that a subset of foregut endoderm and mesoderm genes depend on BMP signaling. Moreover, we found that BMP/Smad1 directly activates ventral genes while repressing dorsal genes. Meanwhile we showed that the Wnt signaling pathway acts on anterior-posterior patterning, directly activating the hindgut while repressing the foregut programs. Detailed analysis identified several factors, other than Tcf, to be involved with β-catenin direct repression of foregut genes. We further reported that hundreds of key foregut and hindgut genes presented Smad1 and β-catenin integration on the same CRMs. Functional analysis showed some of these cis-regulatory modules (CRMs) to be functional and responsive to BMP and Wnt manipulations. Overall, this part of our study revealed a new layer of comp (open full item for complete abstract)

Committee: Aaron Zorn Ph.D. (Committee Chair); Daniel Buchholz Ph.D. (Committee Member); Brian Gebelein Ph.D. (Committee Member); Joo-Seop Park Ph.D. (Committee Member); James Wells Ph.D. (Committee Member); Katherine Yutzey Ph.D. (Committee Member) Subjects: Developmental Biology

Master of Science, Miami University, 2016, Computational Science and Engineering

Recombinant human bone morphogenetic protein-2 (rhBMP-2) can be used clinically to promote bone healing as an alternative to bone grafting treatment. The rhBMP-2 can stimulate cellular differentiation of osteoprogenitor cells to promote bone healing. However, delivery of rhBMP-2 is a challenge since rhBMP-2 has a short half-life and has therefore been delivered from collagen sponges implanted at the injury site. While this has led to effective bone regeneration, ectopic bone growth associated with the rapid degradation of collagen and subsequent rhBMP-2 release are clinical problems. We are investigating keratins as alternative rhBMP-2 carriers. Keratins are structural intermediate proteins and can be extracted from human hair. Oxidatively extracted keratin (KOS) cannot achieve disulfide crosslinks whereas reductively extracted keratin (KTN) can form disulfide crosslinks. The rate of degradation of keratin can be tuned by mixing keratose and kerateine in varying ratios. The hypothesis guiding this thesis is that keratin can be formulated with varying ratios of KOS and KTN to modulate the rate of scaffold degradation and thereby control the releasing rate of rhBMP-2. The in vivo release kinetics of rhBMP-2 was assessed by a critically-sized rat femur defect model. The biodistribution of rhBMP-2 after implantation in the critically-sized femur model was assessed in the vital organs.

Committee: Justin Saul (Advisor); Lei Kerr (Committee Member); Michael Robinson (Committee Member) Subjects: Academic Guidance Counseling; Biomedical Engineering

Master of Science, The Ohio State University, 2016, Dentistry

Objectives: Bone marrow mesenchymal stem cells (BM-MSCs) from limb bones have demonstrated promises in regenerating craniofacial bones; yet little is known about the potential of BM-MSCs from craniofacial bones. This study compared BM-MSCs isolated from limb and craniofacial bones in a commonly used preclinical large animal model, the pig. Methods: Bone marrow was aspirated from the tibia and mandible (symphysis) of 4-month-old pigs (n=4). Subsequently, BM-MSCs were isolated, expanded and confirmed by flow cytometry. To assess cell proliferation, cell doubling times were calculated from serial cell number counts over 2 weeks. Total mRNA was extracted from freshly isolated BM-MSCs and analyzed for gene expression using an Affymetrix GeneChip porcine genome array, followed by real-time RT-PCR for validation. Osteogenic capacity was assessed by quantifying alkaline phosphatase activity. Using temperature-responsive culturing plates, the abilities of BM-MSCs to form multi-layer cell sheets (for future in-vivo transplantation), along with cell viability and morphology, were evaluated by fluorescent labeling and histological staining. Results: BM-MSCs from both locations expressed MSC markers but not hematopoietic markers. Mandibular BM-MSCs proliferated significantly faster than tibial BM-MSCs (median cell doubling times: 2.25 vs. 2.80 days, Mann-Whitney U test, p<0.01). Without any osteogenic induction, mandibular BM-MSC alkaline phosphatase activities were 3.3-fold (factorial ANOVA, p<0.001) to those of tibial BM-MSCs. Microarray analysis on one hand confirmed that overall mandibular and tibial BM-MSC gene expressions are highly correlated with each other and genes related to osteogenesis and angiogenesis were strongly expressed, and on the other hand revealed that there were several dozen genes which were indeed differentially expressed between mandibular and tibial BM-MSCs. They include cranial neural crest-related genes nestin (1.23-fold) and BMP-4 (1.79-f (open full item for complete abstract)

Committee: Zongyang Sun DDS/PhD/MS/MSD (Advisor) Subjects: Dentistry

PhD, University of Cincinnati, 2015, Medicine: Molecular and Developmental Biology

The endoderm germ layer gives rise to the digestive and respiratory tracts and their associated organs such as liver, pancreas, and lungs. Thanks to previous studies on different vertebrate model organisms, including frog, fish, chick, and mouse, we now have a general idea on how endoderm is formed during gastrulation, patterned along anterior/posterior (A/P) axis, induced by multiple signaling pathways and finally gives rise to the endodermal organ buds that will eventually serve critical functions in the digestive, respiratory and endocrine systems. What is more, utilizing the knowledge that was gathered from studying endodermal development in multiple model organisms, huge and rapid advancement has been made in the area of human embryonic stem cells and induced pluripotent stem cells to direct the differentiation of these pluripotent cell types to form multiple endoderm cell types and even endodermal organs with biological functions (Cai et al., 2007; Kroon et al., 2008; Basma et al., 2009; Zhang et al., 2009; Green et al., 2011; Nostro et al., 2011). Despite these advances, there are still many unanswered questions, especially in the area of the molecular basis for early endoderm development, mainly due to the fact that molecular signaling is highly dynamic and the same factor can have a dramatically different impact on organ development with changes of just hours in development or fractions of millimeters in the embryonic gut tube (McLin et al., 2007; Wandzioch and Zaret, 2009; Kenny et al., 2012). It's critical to understand the mechanisms that regulate these signaling dynamics, as developmental defects resulting in disrupted endodermal organ function are the underlying cause of many congenital diseases that affect millions of people, especially small children every year. In addition there are still many inefficient steps in the directed differentiation of stem cells that could benefit a lot from a better understanding of normal organogenesis. 3 (open full item for complete abstract)

Committee: Aaron Zorn Ph.D. (Committee Chair); Samantha Brugmann Ph.D. (Committee Member); Robert Brackenbury Ph.D. (Committee Member); Richard Lang Ph.D. (Committee Member); James Wells Ph.D. (Committee Member) Subjects: Biology

Doctor of Philosophy, Case Western Reserve University, 2014, Molecular Biology and Microbiology

Glioblastoma Multiforme (GBM) and other cancers are challenging to treat due to their intertumoral and intratumoral heterogeneity. The hierarchical model of intratumoral heterogeneity describes the existence of a cellular hierarchy in glioblastoma with a cancer stem cell (CSC) population at the apex. These CSCs can initiate tumors and are resistant to therapies, suggesting that they are responsible for tumor recurrence. CSCs give rise to differentiated cells, which have limited tumor initiation abilities but provide a supportive CSC niche. Here, I examine the cellular cues that maintain both CSCs and the niche by tuning the relative levels of differentiation and self-renewal in GBM. I suggest Bone Morphogenetic Proteins (BMPs) as a driver of differentiation, as BMPs have been shown to promote CSC differentiation and are strongly expressed in GBM. Extrapolating from existing paradigms in development and in cancer, I then hypothesize that extracellular BMP antagonists could be the parallel drivers of CSC self-renewal.I subsequently demonstrate that a BMP antagonist, Gremlin1, is secreted specifically by CSCs, supporting CSCs in the context of the endogenous differentiation signals provided by BMP. I promote self-renewal and increase growth of non-CSCs by overexpression of Gremlin1, and drive differentiation and slow growth of CSCs by Gremlin1 knockdown. Finally, I examine the mechanisms downstream of Gremlin1 that drive its effects on growth and self-renewal, as well as the mechanisms upstream of Gremlin1 that promote its differential secretion by CSCs. Downstream of Gremlin1, cell proliferation effects are mediated by p21 inhibition, and self-renewal is partly mediated by activation of Wnt signaling. Upstream of Gremlin1, I identify XBP1, a pro-survival factor in the unfolded protein response, as a CSC-specific signal that might be promoting Gremlin1 expression. In the course of this thesis, I identify a novel molecular target, Gremlin1, as well as severa (open full item for complete abstract)

Committee: Jeremy Rich (Advisor); Thomas Egelhoff (Committee Chair); Eric Arts (Committee Member); William Schiemann (Committee Member); Paul Tesar (Committee Chair) Subjects: Biology; Biomedical Research; Cellular Biology

Doctor of Philosophy, University of Toledo, 2014, Bioengineering

The objective of this study is to fabricate chitosan microparticles under mild environmental conditions. These microparticles will serve as a better carrier for relevant growth factors and facilitate the repair and regeneration of critical-sized defects in bones. Chitosan is structurally similar with glucosaminoglycans, and this makes it particularly attractive as a biomaterial for bone defects, as glucosaminoglycans are usually the interacting molecules with osteoblasts. Chitosan and tri-ppolyphosphate (TPP) microparticles were employed because this system can be engineered quantitatively to obtain a scaffold with desired physical and biological properties. Coacervation technique was employed to cross-link amine groups in chitosan with phosphate groups in TPP. Temperatures below room temperature were maintained during the cross-linking process to ensure maximum encapsulation efficiency of the growth factors. Physical and morphological characteristics of the microparticles were determined using scanning electron microscopy; the microparticles have a diameter in the range of 400-700 µm, and the surface was found to be groovy and rough. Degradation study conducted in vitro indicated that the microparticles remained unaltered for 30 weeks when suspended in phosphate buffered saline (PBS) of pH 7.4 containing lysozyme (10 mg/L), but when the microparticles were suspended in PBS solution of pH 5.1 containing lysozyme enzyme, the microparticles degraded within 15 weeks. In order to evaluate the harms of organic solvents like hexane and acetone on growth factor encapsulation, cumulative release profiles of insulin-like growth factor 1 (IGF-1) was compared between particles prepared using the emulsification and coacervation techniques. These results indicated a significant decrease (p<0.05) in the encapsulation efficiency, which in turn also decreased the cumulative release from the emulsification microparticles in comparison with coacervation technique during the two-week (open full item for complete abstract)

Committee: Ambalangodage Jayasuriya (Committee Chair); Beata Lecka Czernik (Committee Member); Mark Wooten (Committee Member); Patricia Relue (Committee Member); Stephen Callaway (Committee Member) Subjects: Biomedical Engineering; Biomedical Research

MS, University of Cincinnati, 2013, Medicine: Molecular and Developmental Biology

The cloaca is a transient embryonic structure that develops from the posterior hindgut that gives rise to the endodermal lining of the rectum, anus, and the urogenital system, including the bladder and urethra through a process known as septation. When this does not occur properly it can lead to anorectal and urogenital malformations, varying in severity from a simple fistula to complex anomalies like cloaca malformation. The developmental process of septation is poorly understood and the complex nature of these defects leaves much room for therapeutic treatment improvement. An overall lack of early cloacal markers makes identifying progenitor cells as well as following the identity of regions of the cloaca during development difficult. After identifying useful cloacal markers, K8 and Sox2, looking at the expression pattern during early development in mouse from e8.5 to e15.5, we mapped early cloacal domains that give rise to bladder, urethra, and parts of the anorectal system. We used these newly defined markers to establish a model of human cloaca development using a recently published method to derive gut tube structures (spheroids) from human embryonic stem cells. Given that the cloaca develops from the posterior hindgut, we hypothesized that posteriorizing factors could direct three dimensional spheroids into organoids containing cloacal progenitors that would form differentiated cells of the urogenital and anorectal tracts. Our data suggests that timing and dose of Wnt and BMP signaling posteriorized gut tube spheroids and that this promoted the subsequent formation of urothelial lineages. The ability to generate cloacal progenitors and its derivates will allow study of anorectal and urogenital malformations in a human in vitro system, which could lead to new therapeutic treatments.

Committee: James Wells Ph.D. (Committee Chair); Geraldine Guasch Ph.D. (Committee Member); Aaron Zorn Ph.D. (Committee Member) Subjects: Developmental Biology

PhD, University of Cincinnati, 2006, Medicine : Molecular and Developmental Biology

Mesoderm induction and endoderm formation are controlled by localized maternal determinants during early Xenopus embryogenesis. TGF-βs, including activins, BMPs, and nodal-related proteins are essential regulators of these processes. They are secreted growth factors, synthesized as proproteins and cleaved by proprotein convertases. One such convertase, PACE4, is important for processing of Nodal protein in mouse. The primary goal of this dissertation study was to determine whether Xenopus PACE4 homolog, XPACE4, regulates mesoderm induction through specific processing of different TGF-β proteins. We showed that XPACE4 was a vegetally localized maternal mRNA. Loss of maternal XPACE4 function resulted in gastrulation abnormalities and mesoderm induction defect. XPACE4 was required for processing of a subset of TGF-β proteins, Xnr1, 2, 3, and Vg1 (Chapter 2). The second goal of this study was to resolve the function of a localized maternal TGF-β, Vg1, in Xenopus development. We showed that Vg1 mRNA and protein were enriched on the dorsal side of 32-cell stage embryos and Vg1 was required for the early expression of dorso-anterior antagonists chordin and cerberus (Chapter 3). Additionally, we described the serine allele of Vg1 protein which was secreted and could partially rescue the Vg1 depletion phenotype (Chapter 2 and 3). Overall, these results suggest that proper regulation of TGF-β processing by XPACE4 is critical for gastrulation and mesoderm induction and Vg1 is an essential component of these developmental processes.

Committee: Dr. Janet Heasman (Advisor) Subjects: Biology, Molecular

Master of Science (MS), Ohio University, 2012, Civil Engineering (Engineering and Technology)

The exfiltration trench (ExT) is one of the BMPs that the Ohio department of Transportation (ODOT) is using to remove pollutants from stormwater runoff. A 1-year study was conducted on an ExT located in Reno, OH. Samples from the stormwater runoff and trench's effluent were collected. These samples were tested for water quality parameters (COD, turbidity, oil and grease). The results showed that the ExT was able to reduce COD in the runoff by 24.5%, turbidity by 33.9%, while oil and grease removal was indeterminable. Maintenance was necessary to remove accumulated sediments which caused clogging of the porous concrete surface. However, pollutants removal after maintenance did not improve considerably. Using total suspended solids and COD data, the first flush (FF) volume was estimated. The required length of the ExT to capture the FF was estimated by numerical solution. The solution showed that the required length was much larger than the length estimated using ODOT's equations.

Committee: Gayle Mitchell (Advisor); Guy Riefler (Committee Member); Teruhisa Masada (Committee Member); Dina Lopez (Committee Member); Shad Sargand (Committee Chair) Subjects: Civil Engineering

Master of Science (MS), Ohio University, 2012, Civil Engineering (Engineering and Technology)

Highway storm water runoff contributes to the degradation of surface water quality in the United States. The most important pollutant in highway runoff is the total suspended solids (TSS). The United States Environmental Protection Agency (U.S.EPA) requires the runoff to be treated before it is conveyed to surface water. The exfiltration trench is one of the best management practices (BMP's) utilized by the Ohio Department of Transportation (ODOT) to treat highway runoff in-situ and consists of three layers: pervious concrete, type 3 backfill material (aggregate) and type 2 backfill material (sand). In this study, the exfiltration trench located on SR7, OH, is evaluated based on the pollutants' removal efficiency, particularly, TSS. TSS was removed at an efficiency of 41% in the originally constructed trench while the reconstructed trench achieved 69% TSS removal. Other water quality parameters investigated in this study include particle size analysis, turbidity and pH which were performed on both influent (highway runoff) and effluent (treated influent) samples.

Committee: Gayle Mitchell F (Advisor) Subjects: Civil Engineering

Doctor of Philosophy (PhD), Ohio University, 2007, Civil Engineering (Engineering)

A new hydrodynamic treatment device is developed and tested in this study. The device contains two vertical concentric cylinders designed to intercept and retain sediments, oil and grease, and floatables from storm water runoff. This dissertation presents the findings on the performance of the device using simulated storm water. Removal characteristics are analyzed. This design provides good removal performance for suspended solids and oil and grease. Other advantages of this device are its compactness and flexibility for configuring in different modes. Analyzing cost-benefits can provide a reference for engineers and researchers to estimate performance of best management practices (BMPs), conduct storm water management plans, and develop discharge regulations, etc. Two popular mathematical expressions are formulated and then utilized to illustrate the cost-benefit relationships. Cost-benefit relations for storm water BMPs are conceptually illustrated in a graph and discussed. Suspended solids removals via an ideal settling tank and via the hydrodynamic treatment device are used to illustrate the relationship and effectiveness calculation. Calculated effectiveness coefficients are higher for the hydrodynamic separator compared to an ideal settling tank. A first order wash-off model is used to evaluate the significance of first flush. Assuming an exponentially decreasing concentration during a storm event, concentration is modeled as a function of a first flush coefficient (Cff), cumulative runoff depth, and initial pollutant concentration. Pollutant mitigation data from a wetland receiving runoff from a highway section in Ohio were obtained for the first flush analysis. It shows that the first order model and Cff provide a means to quantitatively evaluate the significance of first flush. Moreover, element models are established to analyze the first flush characteristics under complicated runoff situations, such as runoff from a large area, via a detention/retention (open full item for complete abstract)

Committee: Gayle Mitchell (Advisor) Subjects:

Master of Science, Miami University, 2008, Zoology

The embryonic chick eye is an ideal model to study retina regeneration since it has the ability to regenerate via two different modes: via the activation of stem/progenitor cells located in the anterior region of the eye or via the transdifferentiation of the retinal pigmented epithelium. Both modes give rise to a laminated retina with all major retinal cell types present. In this study we analyze the role of the Bone Morphogenetic Protein (BMP) pathway during the process of chick retina regeneration. The BMP pathway has been well documented in being involved in stem cell maintenance, cell differentiation and apoptosis. We demonstrate that activating the BMP pathway is sufficient to induce retina regeneration from stem/progenitor cells. During the early stages of retina regeneration, the BMP pathway promotes Fibroblast Growth Factor (FGF) signaling through the activation of its canonical pathway to promote proliferation, leading to the formation of a neuroepithelium. In later stages of retina regeneration, the BMP pathway inhibits the FGF pathway and signals through a non-canonical pathway, promoting cell death.

Committee: Katia Del Rio-Tsonis PhD (Advisor); Paul James PhD (Committee Member); Paul Harding PhD (Committee Member) Subjects: Biology; Cellular Biology; Molecular Biology