Doctor of Philosophy, The Ohio State University, 2016, Biophysics

Collagen type 1 is the most abundant extracellular matrix protein in vertebrates. Collagen fibrils provide vital scaffolding, support, and strength to tissues as well as modulate cell-matrix interactions. The quantity and quality of collagen fibrils are important for a variety of physiological and pathological processes. Much is known about how collagen quantity impacts health and disease, but little is known about the impact of collagen quality, i.e., the collagen fibril structure. A large number of factors can cause alterations in collagen fibril structure including mutations in collagen genes, dysregulation of collagen processing enzymes, or altered expression of collagen binding proteins (CBP). Previous studies have established that the CBP, discoidin domain receptor 1 (DDR1) alters collagen fibril structure in vitro. The first aim of this thesis was to determine if DDR1 impacts collagen fibril structure in vivo. To address this question, collagen fibrils in the adventitia of DDR1 knockout (KO) mice aortas were imaged using high resolution microscopy techniques, namely transmission electron microscopy (TEM), scanning electron microscopy (SEM), and atomic force microscopy (AFM). The aortic adventitia of DDR1 deficient mice exhibited increased collagen deposition, a higher percentage of large diameter fibrils, and increased D-periodic depth compared to their wild-type (WT) littermates. The second aim was to determine if the structural changes in DDR1-deficient collagen fibrils have a functional consequence. Platelet-collagen adhesion has been shown to exhibit some sensitivity to the structure of collagen fibrils. Given this, we tested whether DDR1-deficient collagen fibrils impact platelet-collagen adhesion. To investigate this question, we examined the binding of human platelets and of the CBPs primarily involved in platelet-collagen interactions, i.e., glycoprotein VI (GPVI) and von Willebrand factor (VWF) to the collagen fibrils in aortic sections from D (open full item for complete abstract)

Committee: Gunjan Agarwal (Advisor); Alevriadou Barbara (Committee Member); Gooch Keith (Committee Member) Subjects: Biophysics; Histology; Pathology; Physiology

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

Purpose: The cornea stroma consists of collagen fibrils that exist in a highly ordered structure, however the mechanisms by which collagen expression and its complex architecture are regulated are not well understood. Insight into the mechanisms that underlie collagen expression and fibril architecture would shed light onto the etiologies of corneal diseases. Thus, identifying specific pathways or proteins that are in involved in stromal collagen regulation could help better explain how collagen is regulated in health, and what goes wrong in cases of pathology. A protein recently shown to induce collagen expression is the cytoplasmic protein called Shroom3. This F-actin and Rho-kinase binding protein is best known for regulating epithelial cell shape during embryonic morphogenesis, but the mechanisms by which it regulates collagen expression have not been widely studied. Previous research has suggested that abnormal SHROOM3 disrupts cornea development in mouse embryos and that a homozygous missense mutation in this gene has been associated with the stromal thinning disease known as keratoconus. Therefore, the purpose of this research was to further characterize the corneal phenotype in Shroom3-deficient embryos and discern what effect the keratoconus-associated mutation has on Shroom3 function. Methods: Embryonic corneal Shroom3 expression was observed using X-gal labeling of Shroom3+/- embryos that express X-galactosidase under the control of the endogenous Shroom3 promoter. To determine the function of Shroom3 during cornea development, control and Shroom3-deficient mouse embryos were analyzed using histological staining, immunofluorescent imaging, and electron microscopy to ascertain the effect on collagen expression, fibril size, keratocyte number, stromal thickness and corneal epithelial shape. The consequences of mutations on Shroom3 function were analyzed in kidney- (MDCK) and cornea-derived (SIRC) cells following transfection with plasmids expressing wild (open full item for complete abstract)

Committee: Timothy Plageman PhD (Advisor); Heather Chandler PhD (Committee Member); Andrew Hartwick OD, PhD (Committee Member) Subjects: Developmental Biology; Genetics; Ophthalmology

Master of Science, The Ohio State University, 2024, Dentistry

Background and objective: Socket preservation is a bone regeneration procedure indicated immediately following tooth extraction to preserve existing alveolar ridge height and to control alveolar ridge width. Primary wound closure following socket preservation is not indicated since it is a flapless procedure and wound edges cannot be proximated to close the bone graft through suturing. Instead, a collagen plug and/or a resorbable membrane is generally used to stabilize bone graft and seal the orifice of the socket. The purpose of this approach is to allocate time for soft tissue granulation and complete epithelization. To our knowledge the indications for these two types of biomaterials are not well-established and there are no clinical studies in the literature comparing clinical outcomes in relation to the use of these wound management approaches. Thus, the purpose of this study was to evaluate and compare the early clinical outcomes following socket preservation procedures performed by using collagen plug (CP) or collagen membrane (CM) as a sealing material. In addition, soft tissue phenotype was evaluated as a variable possibly affecting clinical healing outcomes. Material and Methods: Patients who need single tooth extraction (surrounded with mesial and distal intact teeth) and socket preservation procedures for a future implant placement procedure were recruited (IRB protocol #2022H0277). Impressions (digital) were obtained prior to surgery and at 3-6 months to study ridge dimensional changes. Soft tissue phenotype was determined prior to extraction by probing and buccal tissue thickness was determined by using a surgical caliper. Buccal bone integrity was determined by probing immediately after extraction. FDBA was used as bone graft material. Decision to use a collagen plug or collagen membrane was clinician's choice based on the case and site anatomy. The membrane and plug were stabilized via simple interrupted and cross matrix suture using a resorbab (open full item for complete abstract)

Committee: Binnaz Leblebicioglu (Advisor); Guo-Liang Cheng (Committee Member); Hanin Hammoudeh (Committee Member); Luiz Meirelles (Committee Member) Subjects: Dentistry

Doctor of Philosophy, The Ohio State University, 2021, Biomedical Engineering

Collagen fibrils are the most abundant component of the extracellular matrix (ECM) present in vertebrate tissues. While the quantity of collagen deposition is routinely evaluated in diseases, much less is understood about the quality of collagen, in particular perturbations in its fibril structure. This dissertation aims to evaluate the quality of collagen fibrils in the vascular disease, abdominal aortic aneurysm (AAA) and its functional consequences. Studies are conducted on human AAA tissue extracted at the time of vascular surgery and on aorta extracted from a mouse model of AAA and on accompanying control samples. The first aim of this study was to characterize collagen fibrils at the single-fibril, ultrastructural level. Two high-resolution microscopy techniques, namely transmission electron microscopy (TEM) and atomic force microscopy (AFM) were employed to ascertain quantifiable parameters on the collagen fibril structure. We have identified the presence of structurally abnormal collagen fibrils in AAA, which consisted of compromised D-periodic banding, reduced diameter and increased fibril curvature. Our second aim was to correlate ultrastructural abnormalities with biochemical markers for a high-throughput evaluation of structure-function relationship of the collagen fibril. Towards this goal we optimized staining the tissues with collagen hybridizing peptide (CHP), a newly developed marker for degraded collagen. Multimodal approaches coupling CHP staining with AFM imaging or second harmonic generation microscopy (SHG) were employed. We found that in AAA tissue abnormal fibrils (ascertained using AFM) co-localized with CHP signal whereas SHG signals (corresponding to normal fibrils) were spatially distinct from CHP. We also ascertained a functional consequence of the abnormal collagen fibrils, namely how they impact platelet adhesion. Towards this goal AAA tissue sections were incubated with human platelets, and binding of platelets to normal or ab (open full item for complete abstract)

Committee: Gunjan Agarwal (Advisor); Arunark Kolipaka (Committee Member); Jonathan Song (Committee Member); Michael Go (Committee Member) Subjects: Biomedical Engineering; Biomedical Research

Doctor of Engineering, Cleveland State University, 2018, Washkewicz College of Engineering

The intrinsic repair mechanism of human heart is not sufficient to overcome the impact placed by adverse pathological conditions, such as myocardial infarction (MI). Current clinical approaches have played significant role in reducing the mortality rate; however, these approaches do not replace the lost cells and tissues of the myocardium. Human bone marrow-derived mesenchymal stem cells (BM-MSCs) are gaining attention in cardiac therapy due to their ability to differentiate into cardiomyocyte like cells and release a wide repertoire of paracrine factors. However, clinical trials of longer duration have shown mixed results in improving cardiac functions. In addition, in-depth studies on the secretome profile of MSCs/ differentiated MSCs and optimal approaches to modulate their outcomes are still lacking. Hence, in the first project of this study, we investigated the role of cell-cell interactions (MSC spheroids), cell-matrix interactions (collagen concentration, topography) and cell-signaling cues [5-azacytidine (aza)] on the cardiac differentiation of BM-MSCs. We found that collagen hydrogel (2 mg/ml), in the presence of 10 µM of aza, offered higher cell survival and caused time-dependent cardiomyogenic evolution of MSC spheroids. We also identified that canonical Wnt/ß-catenin signaling pathway primarily mediated the observed benefits of aza on cardiac differentiation of MSC spheroids. In the second project, we quantified the secretome profile and matrix synthesis by collagen gel-laden BM-MSC spheroids, under optimized culture conditions from the first project, and extended our investigation to MSC spheroids within human fibroblasts-derived collagen. Human collagen promoted higher matrix synthesis over time but severely impacted cell proliferation. The release of inflammatory cytokines was drastically reduced with spheroid formation and collagen cultures, specifically within human collagen. In the last project, we examined the influence of adult human cardiomyocyt (open full item for complete abstract)

Committee: Chandra Kothapalli (Committee Chair); Moo-Yeal Lee (Committee Member); Nolan B. Holland (Committee Member); Mekki Bayachou (Committee Member); Anand Ramamurthi (Committee Member) Subjects: Biomedical Engineering

Master of Science, University of Akron, 2016, Polymer Science

Collagen is a protein that is a major component of the human body and makes up thirty percent of the total protein in the human body. Collagen mimetic peptides (CMPs) have been used to study the formation of the triple helix of collagen and its higher-ordered structures. This article discusses how CMPs have been used to learn about the physical properties of the triple helix and how the amino acid sequence affects the physical properties, formation, and stability of the triple helix. CMPs have also been designed to form higher-ordered structures for various applications. This work reviews the design principles and strategies employed with various interactions to form higher-ordered structures from CMPs. These strategies include the use of electrostatic interactions, the incorporation of cysteine knots, and metal-induced self-assembly, among others. Many of these higher-ordered structures have applications such as biomolecule attraction, cellular adhesion, nanowire fabrication, and hydrogel formation.

Committee: Nita Sahai PhD (Advisor) Subjects: Biochemistry; Biology

MS, University of Cincinnati, 2003, Engineering : Engineering Mechanics

The objective of this study was to examine the cell-extracellular matrix interactions in mesenchymal stem cell-seeded collagen gel implants under varied levels of cell-to-collagen ratio. Higher cell-to-collagen ratios were hypothesized to increase total collagen content and implant contraction, and improve the collagen fiber architecture. These tendon implants must introduce active repair-cells to the injury site under the protection of the collagen matrix, until newly synthesized collagen fibrils can bear the everyday tensile load. Rabbit mesenchymal stem cells were mixed with type I collagen gel and allowed to contract onto two vertical posts in a custom made deformable silicone dish. Implants with four different cell-to-collagen ratios (0.04, 0.08, 0.40, 0.80 million cells/mg collagen) were evaluated after 14 and 28 days. The constructs were evaluated for hydroxyproline content, planar area, and matrix fiber architecture using the small angle light scattering (SALS) technique. While the two higher cell-to-collagen ratio implants (0.40 and 0.80 million cells/mg collagen) did not show changes in collagen content over 28 days in culture, the two lower cell-to-collagen ratio implants (0.04 and 0.08 million cells/mg collagen) significantly declined in collagen content. The lowest cell-to-collagen ratio implants (0.04) recorded a significant change in implant contraction between 14 and 28 days in culture. Differences in planar area among the four treatments were not significant after 28 days. During the second two weeks in culture, the highest cell-to-collagen ratio implants (0.80) significantly improved its matrix architecture by aligning collagen fibers in a longitudinal direction between the posts.

Committee: Dr. David Butler (Advisor) Subjects: Engineering, Biomedical

Master of Science, The Ohio State University, 1967, Graduate School

Committee: Not Provided (Other) Subjects:

Doctor of Philosophy, Case Western Reserve University, 2024, Macromolecular Science and Engineering

Helices are abundant and crucial examples of rigidity throughout biology from protein lever arms in molecular motors to collagen type II that assembles into fibrils in the extracellular matrix (ECM). The origin of the collagen fibril radial length scales is not fully understood but is hypothesized to be related to the flexibility of the protofibril and environmental effects. In this work, we systemically investigate both the elasticity of biomacromolecules and their surrounding elastic environments using simple polyacrylamide gels. We determine the persistence length (lp), a measure of elasticity, of model polypeptide single helices and collagen type II triple helices by using static and dynamic light scattering. Using circular dichroism, we observe that the model polypeptide transitions from a random coil to a helix with increasing pH, and lp increases from ~1 - 2 nm to ~20 nm. In addition, we crosslink the model polypeptide to utilize its increase in lp and produce hydrogels with stain stiffening behavior at low crosslink densities. In various pH and ionic strength environments, triple helical lp varies from 60 - 90 nm but has an intrinsic lp of 90 nm when backbone interactions are neutralized. We correlate the triple helical lp to the fibril diameter as determined by transmission electron microscopy (TEM) in various ionic strength solutions and determine that the values are of similar magnitude unless in high ionic strength solutions. We then investigate the environmental elasticity effects on self-assemblies of complex coacervates using light microscopy and collagen type II fibrils using cryogenic TEM. The volume of the complex coacervate droplets is inversely proportional to the modulus of the gel that the complex coacervates are formed in and have a non-monotonic salt resistance as a function of gel moduli. Collagen fibrils in 100 mM PBS solution are ~50 nm in diameter, and the fibril diameter drops to ~30 nm in gels across 63-8700 Pa moduli. Collagen's lp, s (open full item for complete abstract)

Committee: Svetlana Morozova (Committee Chair); Lydia Kisley (Committee Member); Valentin Rodionov (Committee Member); Michael Hore (Committee Member) Subjects: Biophysics; Materials Science; Physics

Doctor of Philosophy, The Ohio State University, 2024, Oral Biology

Periodontal diseases affect the supporting tissues of teeth and lead to tooth mobility, tooth loss, and loss of masticatory function and quality of life. Approximately 56% of adults present with periodontitis, which manifests due to dysregulation between oral bacteria and host immune response. Discoidin Domain Receptor 1 (DDR1) is a transmembrane non-integrin collagen receptor that binds fibrillar collagens and collagen IV. It is widely expressed in epithelial, immune and bone cells. Binding to collagen and receptor activation leads to extracellular matrix remodeling and changes in cell behavior. Previous studies in our lab have shown that genetically modified DDR1-null mice (Ddr1-/-) present with progressive periodontal attachment and alveolar bone (AB) loss by 6 months of age. Thus, we hypothesized that DDR1 is necessary for maintenance of periodontal tissues and wound healing. Our first series of experiments (Aim 1) concluded that Ddr1-/- mice present with decreased junctional epithelium (the gingival epithelium mediating attachment of gingiva to the tooth surface) permeability compared to Ddr1+/+ mice. Additionally, epithelial migration over dorsal cutaneous wounds progressed faster in Ddr1-/- versus Ddr1+/+ mice. For our second aim, we used the ligature-induced periodontitis (LIP) model to predictably induce periodontitis in Ddr1+/+ and Ddr1-/- mice. At 3 days post-ligature (dpl), Ddr1-/- presented with higher bone loss than Ddr1+/+; the difference in bone loss disappeared at 5 and 8 dpl. Analysis of immune cells via flow cytometry showed that neutrophil and macrophage numbers were higher in gingiva and cardiac blood of Ddr1-/- mice at 3dpl. Finally, in aim 3, we employed Raman spectroscopy and showed that femurs of Ddr1-/- mice presented with decreased mineral-matrix ratio and increased pyridinoline cross-linking versus Ddr1+/+ mice at 50 days postnatal (dpn). We then proceeded to examine effects of DDR1 on AB resorption by employing the traumatic occlusion (T (open full item for complete abstract)

Committee: Brian Foster (Advisor); Gunjan Agarwal (Committee Member); Parvathi Ranganathan (Committee Member); Dimitris Tatakis (Committee Member) Subjects: Dentistry

Master of Science, University of Toledo, 2023, Bioengineering

Extracellular vesicles have emerged as a promising tool for cell-free therapies in tissue regeneration and disease treatment. Although regenerative properties and therapeutic effects of extracellular vesicles have already been started to be investigated, most of the EV research is associated with stem cell lineage. As both fibroblasts and macrophages are ubiquitous in the human body and are highly active in tissue repair and remodeling, the effect of extracellular vesicles secreted by fibroblasts on macrophages towards immunomodulation is yet to be known. In this regard, the objective of this study is to first isolate extracellular vesicles successfully from fibroblasts, and then to observe whether the fibroblast-derived extracellular vesicles promote macrophage polarization from pro-inflammatory to anti-inflammatory phenotype within ECM like 3D matrix, leading that EVs are enough to create anti-inflammatory response in case of treating diseases with cell-free therapies towards tissue regeneration.

Committee: Dr. Eda Yildirim-Ayan (Committee Chair); Dr. Yuan Tang (Committee Member); Dr. Brent Cameron (Committee Member) Subjects: Biomedical Engineering; Engineering

Bachelor of Science (BS), Ohio University, 2023, Biological Sciences

Fibrosis, a pathological process characterized by excess extracellular matrix (ECM) deposition, can occur in many internal organs and tissues in response to various stimuli. As fibrosis progresses, scarring occurs, which ultimately leads to tissue dysfunction and organ failure. Patients with acromegaly, a rare disease usually caused by a benign, GH-producing pituitary tumor, have been reported to have prominent ECM deposition and scarring in certain tissues, which is indicative of fibrosis. In bGH transgenic mice, which express high levels of bovine growth hormone, several tissues [white adipose tissue (WAT), heart, intestine, and kidney] demonstrate a fibrotic phenotype. However, there is no previous research that investigates various bGH tissues – particularly from mice derived from a single cohort – for fibrosis. Additionally, WAT fibrosis is associated with obesity and lipodystrophy, and seems to be particularly associated with excess GH. This study aims to investigate the role of different cell types and genes involved in the development and progression of WAT fibrosis and determine if fibrosis is increased in BAT, liver, quad, kidney, lung, and spleen of aged bGH mice. Results of this thesis included a striking observation of increased fibrosis in all bGH tissues examined. For WAT, decreases in fibrosis-associated RNA expression in 3-month-old bGH mice via qPCR analysis was only observed in the perigonadal depot and not the subcutaneous depot that has more prominent collagen deposition. Interestingly, we observed an intriguing increase in fibrosis-associated RNA expression in a population of adipose stem and progenitor cells in 6-month-old mice within subcutaneous bGH WAT. These results indicate a potential common GH-induced mechanism of fibrosis across bGH tissues and pave the way for future research into WAT fibrosis.

Committee: Darlene Berryman (Advisor) Subjects: Biology; Biomedical Research

Master of Science (MS), Ohio University, 2023, Biomedical Engineering (Engineering and Technology)

A study on a solution to repair osteochondral defects was investigated. This work contained the use of a novel collagen-based biomaterial that was structured to mimic the composition and structure of osteochondral tissue. Collagen extraction from the bovine achilles was optimized in terms of atelocollagen yield and stability. It was found that collagen enzymatically digested at a 1.25:10 pepsin to tendon weight ratio in the superior tendon region, gave optimal results in terms of atelocollagen quantity and hydrogel formation. Mineralized collagen scaffolds were fabricated to reflect the composition of subchondral bone. Controlled freezing was applied, which successfully oriented collagen fibers mimicking those in each native zonal tissue. Multiple approaches were attempted to replicate the collagen orientations of osteochondral tissue, ultimately a T-shaped mold was designed to guide directional freezing, resulting in an anisotropic scaffold structure. Native composition of bone hydroxyapatite and cartilage hyaluronic acid were also taken into consideration when fabricating such multizonal scaffolds.

Committee: Mei Wei (Advisor); Shouan Zhu (Committee Member); Doug Goetz (Committee Member); Andrew Weems (Committee Member) Subjects: Biomedical Engineering; Biomedical Research

Master of Science, The Ohio State University, 2023, Biomedical Sciences

Injury to the superficial digital flexor tendon (SDFT) accounts for up to half of the musculoskeletal injuries in Thoroughbred racehorses and is a common reason for retirement. There is growing evidence that SDFT injuries are the result of cumulative microdamage from racehorses working at high speeds that place the tendon under maximal tensile capacity, rather than a single traumatic event. Current therapies consisting of prolonged rest, rehabilitation, and intralesional biologics aim to improve healing of the SDFT, but reduced elasticity of the ‘healed' tendon results in re-injury rates of up to 50%. Therefore, documenting flexor tendon adaptation to athletic training and delineating the mechanisms leading to SDFT injury in racing Thoroughbreds is crucial for reducing tendon injury incidence and associated loss. The objectives of this research are two-fold. First, to assess changes in SDFT hierarchical structure and elastin in mid-metacarpal SDFTs obtained from 2-, 3-, and 4year-old training/racing Thoroughbred cadaver distal limbs. Second, to determine if the biomechanical properties of SDFT are altered by athletic training. Distal forelimb SDFTs were collected from 2-, 3-, and 4-year-old Thoroughbred racehorses (N=50) within 48 hours of death or euthanasia for reasons other than SDFT injuries and that were necropsied through the California Animal Health and Food Safety Laboratory System (CAHFS). A randomly chosen forelimb SDFT was processed for histology and whole tendon biochemical quantification. The contralateral forelimb SDFT was stored at -20°C for whole tendon biomechanical testing. For histological assessment, cryopreserved sections from 1-cm mid-metacarpal SDFT segments were used for elastin immunostaining, prior to concurrent immunofluorescence and second harmonic generation (SHG) confocal microscopy. Stained, paraffin-embedded, histological sections were used for fascicle cross-sectional area (CSA) and interfascicular matrix (IFM) measurements using (open full item for complete abstract)

Committee: Sushmitha Durgam BVSc, MS, PhD (Advisor); Rebecca Urion (Committee Member); Hilary Rice DVM, MS (Committee Member) Subjects: Animal Sciences; Biomechanics; Biomedical Research

Doctor of Philosophy, The Ohio State University, 2022, Biomedical Engineering

Microscale flows are central to many different physiological and biological systems. As such, microfluidic technology, which provides the means for efficient flow manipulation at the microscale, has emerged as an important tool for numerous biomedical applications, such as perfusion-based cell culture and control of complex cell microenvironments. Yet, one research area ripe for further exploration is the application of microfluidic technology for characterizing the physical properties of biomaterials-based matrices. Biomaterials focused on recapitulating the native three-dimensional (3-D) extracellular matrix (ECM) and the interstitial space surrounding tissue provide controlled conditions for studying physiological function. Careful control of ECM properties, changing parameters such as mechanical stiffness enables researchers to investigate different developmental processes alongside distinct physiological and pathological environments. After a brief introduction, this dissertation describes three major parts that define the application of microscale flow phenomena to unravel biomaterial and biological properties of reconstituted tissue. Part 1 (Chapter 2-4) emphasizes the need to describe the initial conditions by studying the kinetics of ECM hydrogels and connecting these to their physical properties. Here, I expand on a characterization framework that probes the biophysical properties of ECM based materials, measuring mechanical stiffness, convective transport via microfluidics, matrix architecture via confocal reflectance, and ECM polymerization kinetics. While these properties are commonly characterized individually, a combined approach is necessary to comprehensively understand how ECM composition influences the biophysical properties that direct cellular responses and fates. For collagen-based ECM hydrogels, the material properties can start being adjusted in the pre-polymerization stage. Therefore, we can quantify how small changes in the prepolymer sta (open full item for complete abstract)

Committee: Jonathan Song (Advisor); Gunjan Agarwal (Committee Member); Daniel Gallego Perez (Committee Member); Gina Sizemore (Committee Member) Subjects: Biomedical Engineering

Doctor of Philosophy, The Ohio State University, 2022, Mechanical Engineering

Bone is a rigid connective tissue providing a load-bearing function along with the skeletal system in mammals. The material and structure of bone are optimally designed and adjusted to fulfill functional demands. From an engineering point of view, bone is a smart material that can modulate its material composition and properties in response to mechanical loadings applied to the bone. This unique characteristic of bone shows the potential to influence vast gains in the safety, effectiveness, and affordability of designs in diverse fields. To maintain a mechanically ideal design, bone actively remodels its structure and composition by replacing its old part with the new one. This newly formed bone mainly consists of a soft organic matrix (i.e., Type I collagen) that is subsequently mineralized to enhance the modulus and toughness of bone tissues. Thus, bone mineralization plays a key role in determining bone material integrity. During bone mineralization, the collagen matrix recruits ionized mineral precursors on the outer and into the inner space of collagen. In previous works, many driving forces, such as non-collagenous proteins, electrostatic interaction, osmotic pressure, and streaming potentials, have been suggested to explain the attraction of mineral precursors to collagen space. However, it seems not sufficient to fully address bone's ability to adapt itself to external loadings. Particularly, the infiltration of amorphous mineral precursors through the gap regions of collagen for intrafibrillar mineralization has still not been fully clarified. In this dissertation, I have focused on the mechanoelectrical response of collagen (i.e., piezoelectricity). The piezoelectricity is an inherent function of a collagen fibril, by which an electric charge generates on the collagen surface in response to mechanical stresses. Considering that ionized mineral clusters are electrically charged, and bone has an ability to adapt to mechanical stresses, I have hypothesized (open full item for complete abstract)

Committee: Hanna Cho (Advisor); Carlos Castro (Committee Member); Soheil Soghrati (Committee Member); Gunjan Agarwal (Committee Member) Subjects: Mechanical Engineering

Master of Science, The Ohio State University, 2022, Vision Science

Shroom3 is a cytoskeletal protein that is known to trigger the contraction of the cytoskeleton and alter epithelial cell shape. It is comprised of three major domains that are conserved among the Shroom gene family. The SD1 and SD2 domains are well conserved regions among family members and function to localize the protein to cellular junctions, bind actin and Rho-kinase, and induce actomyosin contractile activity. Only the longest isoforms of Shroom3 exons encode a PDZ domain within the translated Shroom proteins. Using whole exome sequencing, a homozygous missense mutation of the SHROOM3 gene was identified in a patient with keratoconus and hypothesized to be the underlying pathogenetic factor [Tariq et al., 2011]. Furthermore, evidence exists that Shroom3 plays a role in regulating collagen expression. This study is intended to expand our understanding about how Shroom3 may regulate collagen expression in the cornea. Specifically, it investigates the role of the N-terminal domain found solely in the longest isoform of Shroom3 and a specific amino acid change within this domain that has been associated with keratoconus. Methods: To evaluate the role of the Shroom3 PDZ domain two distinct mouse lines were generated. A homozygous mutation was generated in the mouse Shroom3 coding sequence (G59V) that is homologous to the homozygous polymorphism associated with keratoconus (SHROOM3G60V). The second mouse line was missing a single nucleotide in iii an alternative exon that encodes the N-terminal PDZ domain that was predicted to cause a frameshift and premature protein termination (Shroom3iso1). Embryonic neuroepithelial tissue collected from Shroom3Δiso1/Δiso1 and wild type mice were analyzed by immunofluorescent labeling using Shroom3 and β-catenin antibodies to evaluate the presence of the longest isoform of Shroom3 containing the PDZ domain. Wild type, heterozygous, and homozygous animals of the iso1 line were inves (open full item for complete abstract)

Committee: Timothy Plageman (Advisor); Andrew Hartwick (Committee Member); Heather Chandler (Committee Member) Subjects: Biochemistry; Biology; Ophthalmology

Master of Science in Biological Sciences, Youngstown State University, 2021, Department of Biological Sciences and Chemistry

The calcaneal (Achilles) tendon is capable of handling tremendous tensile loads during locomotion. However, cases of Achilles tendon ruptures have increased in recent years, requiring long healing times. Repaired tendons are more prone to re-rupture after healing, which may negatively impact patient quality of life. Thus, there exists a need for new methods of treatment aimed to improve and accelerate tendon healing. We studied the effect a combination of collagen, platelet-rich plasma (PRP), and mesenchymal stromal cells (MSC) on healing a complete Achilles tendon rupture in a Lewis rat model. The PRP was produced from rat blood collected during exsanguination procedures. MSCs from rat bone marrow met the criteria to be considered stem cells in a rat model, as they were seen to be plastic adherent and capable of tri-lineage differentiation. Rupture was surgically simulated by a full-thickness transection of the tendon, followed by surgical repair. All treatments included a strip of CollaTapeTM wrapped around the repair, acting as a vehicle for the biologics prior to closure of the wound. A single, 100µL subcutaneous injection of MSCs, PRP, or both were administered adjacent to the incision and assigned 1- or 2-week recovery periods before harvesting the operated and unoperated tendons. We observed promising trends which show an increase in gene expression activity in the treated tendons and differences in the expression of Col1a1 and Col3a1 which align with our predicted response to the treatments. However, due to contamination of the GAPDH RT-PCR results, the collagen analysis results remain inconclusive. The biomechanical properties of the tendons were determined using force-extension analysis. When normalized as a percent of the unoperated tendon, a significant improvement was seen in the strain at failure and in ultimate tensile strength after only one week of recovery in the rats who received any biological treatments used in this study, when compared to a sur (open full item for complete abstract)

Committee: Diana Fagan PhD (Advisor); Gary Walker PhD (Committee Member); Carmen Panaitof PhD (Committee Member) Subjects: Biology; Biomechanics; Biomedical Research; Physiology; Surgery

Doctor of Philosophy (PhD), Wright State University, 2020, Engineering PhD

This research focuses on the hierarchical structure of bone and associated mechanical properties at different scales to assess damage accumulation leading to premature failure, with or without instrumentation. In this work, an attempt was made to develop a framework of macro, micro, and nano damage accumulation models and implementing them to derive mechanical behavior of the bone. At macrolevel, retrospective evaluation of 313 subjects was conducted, and the damage of bone tissue was investigated with respect to subject demography including age, gender, race, body mass index (BMI), height and weight, and their role in initiating fracture. Experimental data utilized 28 human femoral bones implanted with cephalomedullary nails were used to develop damage prediction models. Investigation of three real life medical device failures identified the mechanical and clinical bases of bone failure. At the micro level, microdamage accumulation of the bone was investigated in 307 subjects and new effective morphological parameters at microscale were proposed. At the nano level, molecular dynamics simulation was performed to investigate the effect of interaction, orientation, and hydration on the atomic models of the bone composed of collagen helix and hydroxyapatite crystal. The results showed that bone density and maximum von Mises stress decreased drastically in elderly patients, implying fixation devices and implants used by the young cannot be used. The results also showed that the two-dimensional representation of the morphological parameters of the bone at microscale does not provide a realistic description of bone structure. Therefore, in this work, three-dimensional representations at microscale indicated that bone interconnectivity is higher in female patients than in male patients. Gender has a significant effect on microdamage distribution in the bone. More precautions should be taken into consideration for older female patients. Race should also be considered during (open full item for complete abstract)

Committee: Tarun Goswami D.Sc. (Advisor); Caroline GL Cao Ph.D. (Committee Member); Arnab K. Shaw Ph.D. (Committee Member); Partha P. Banerjee Ph.D. (Committee Member); Richard T. Laughlin M.D. (Committee Member); Jennie J. Gallimore Ph.D. (Committee Member) Subjects: Biomechanics; Biomedical Engineering; Biomedical Research; Engineering; Industrial Engineering; Molecular Biology; Nanoscience; Nanotechnology

Doctor of Philosophy, The Ohio State University, 2020, Biomedical Engineering

The extracellular matrix is extensively reorganized throughout breast cancer progression. This reorganization contributes to cancer cell invasion and intravasation and is an independent prognostic factor for breast cancer patients. Cancer-associated fibroblasts appear to play a major role in this reorganization but the cellular signaling pathways contributing to this reorganization remain unclear. We show here that loss of the tumor suppressor phosphatase and tensin homolog (Pten) in fibroblasts promotes extracellular matrix alignment both in vitro and in vivo through increasing cell traction forces. Furthermore, low stromal PTEN expression correlated with high mammographic density, one of the major risk factors for breast cancer development. Matrix reorganization was concomitant with a marked increase in collagen deposition within the mammary gland. We therefore investigated the mechanism of collagen deposition and showed that loss of PTEN upregulated SPARC, which mediated both collagen and fibronectin assembly without modulating cell traction force. To further determine how Pten loss connected to matrix alignment, we designed a novel screening platform to examine matrix alignment in vitro using fibroblast-derived matrices, automated microscopy, and automated image analysis through MATLAB. We discovered a number of novel matrix alignment modulators, including protein kinase C (PKC), dual specificity tyrosine regulated kinase 1b (DYRK1b), platelet-derived growth factor receptor β (PDGFRβ), and Janus kinase (JAK), among others. The effect of these markers on patient survival was examined using publicly available patient datasets. Finally, we examined the effects of hypoxia and matrix metalloproteinase activity on matrix organization.

Committee: Jennifer Leight (Advisor); Samir Ghadiali (Committee Member); Jonathan Song (Committee Member) Subjects: Biomedical Engineering