Matching Items (15)

In Vitro Osteogenic Study of hMSCs Under Diabetic Conditions

Description

Patients with type 2 diabetes mellitus experience a slower healing process and poor osteointegration, making it difficult for them to heal properly after a bone fracture. This study aims to

Patients with type 2 diabetes mellitus experience a slower healing process and poor osteointegration, making it difficult for them to heal properly after a bone fracture. This study aims to compare the proliferation and differentiation of human mesenchymal stromal cells at different glucose concentrations, as well as with an advanced glycated end-product (AGE) concentration, to mimic a healthy, prediabetic, and diabetic environment in an in vitro model over several experiments. Each experiment was composed of treatment groups in either growth or osteogenic media, with varying levels of glucose concentration or an advanced glycated end-product concentration. The treatment groups were cultured in 24 well plates over 28 days with staining of FITC-maleimide, DAPI, or alkaline phosphatase conducted at varying time points. The plates were imaged, then analyzed in ImageJ and GraphPad Prism. The study supports that at 28 days in culture, the more glucose added to osteogenic media treatment groups, the lower the nuclear count. At 14 days the same is true of growth media treatment groups, though the trend does not persist until 28 days. It does not seem that cell surface area of osteogenic groups, and growth media treatment groups was affected by glucose level. At 14 days, the alkaline phosphatase expression was unaffected by glucose level. However, at the 28 day time point the higher the glucose level of osteogenic treatment groups, the less expression of alkaline phosphatase. The effect of the added AGE concentration on hMSC osteogenesis was inconclusive. Overall, this study enhanced understanding of the role that glucose and AGEs play in the bone healing process for diabetic patients, allowing for future improvements of biomaterials and engineered tissue.

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  • 2019-05

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Controlling Calcium Binding on NorHA Scaffolds using a Biomineralization Peptide

Description

The tendon-bone junction is essential for allowing humans to transfer mechanical loads during activities. When injury does occur to this important area, current surgical techniques improperly bypass important physical and

The tendon-bone junction is essential for allowing humans to transfer mechanical loads during activities. When injury does occur to this important area, current surgical techniques improperly bypass important physical and chemical gradients and do not restore proper function. It is essential to create tissue engineered scaffolds that create proper models for the region and induce healing responses for repair. To advance research into these scaffolds, electrospinning fibers and hydrogels made of norbornene functionalized hyaluronic acid (NorHA) were used to promote bone growth by adhering calcium to the material. To further improve calcium adherence, which is indicative of bone regions, a mineralization peptide was allowed to soak through the fibers. NorHA proved to be a suitable material for biomineralization experiments, showing slow calcium adherence within the first hour before accelerating in adherence over 24 hours in both fibers and hydrogels. When the mineralization peptide was implemented calcium adherence on fibers increased nearly eight times within the first 15 minutes of experimentation.

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Date Created
  • 2020-05

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Fabrication and Characterization of a 3D Printed and Electrospun Hybrid Scaffold for Regenerative Medicine

Description

Tissue engineering scaffold fabrication methods often have tradeoffs associated with them that prevent one method from fulfilling all design requirements of a desired scaffold. This undergraduate thesis seeks to combine

Tissue engineering scaffold fabrication methods often have tradeoffs associated with them that prevent one method from fulfilling all design requirements of a desired scaffold. This undergraduate thesis seeks to combine 3D printing and electrospinning tissue engineering fabrication methods into a hybrid fabrication method that can potentially fulfill more design requirements than each method alone. The hybrid scaffolds were made by inserting electrospun scaffolds between layers of 3D printed scaffolds of increasing print temperature and effects on adhesion and mechanical properties were characterized. The fabrication method proved to be feasible and print temperature affected both adhesion and mechanical properties of the scaffolds. A positive, non-linear relationship was seen between print temperature and adhesion and resulting force. Insertion of electrospun mats led to increased damping of scaffolds. Evidence from characterization indicated factors other than print temperature were likely contributing to adhesion and mechanical properties. If studied further, this fabrication method could potentially be used to improve overall structure and regenerative potential of tissue engineering scaffolds.

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Date Created
  • 2020-05

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Utilizing Magnetic Electrospinning to Create Gradients in Fiber Alignment for Interfacial Tissue Engineering

Description

Heterogeneous tissues are composed of chemical and physical gradients responsible for transferring load from one tissue type to another, through the thickness or the length of the tissue. Musculoskeletal tissues

Heterogeneous tissues are composed of chemical and physical gradients responsible for transferring load from one tissue type to another, through the thickness or the length of the tissue. Musculoskeletal tissues include these junctions, such as the tendon-bone and ligament-bone, which consist of an alignment gradient through the length of the interfacial regions. These junctions are imperative for transferring mechanical loadings between dissimilar tissues. Engineering a proper scaffold that mimics the native architecture of these tissues to prompt proper repair after an interfacial injury has been difficult to fabricate within tissue engineering. Electrospinning is a common technique for fabricating nanofibrous scaffolds that can mimic the structure of the native extracellular matrix (ECM). However, current electrospinning techniques do not easily allow for the replication of the chemical and physical gradients present in musculoskeletal interfacial tissues. In this work, a novel magnetic electrospinning technique was developed to fabricate polycaprolactone (PCL) nanofibrous scaffolds that recapitulate the gradient alignment structure of the tendon-bone junction. When exposed to the natural magnetic field from a permanent magnet, PCL fibers innately aligned near the magnet with unalignment at distances further away from the magnetic field.

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Date Created
  • 2018-05

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Controlling the Electrospun Scaffold Profile at the Interface between Varying Fiber Alignment

Description

Musculoskeletal heterogenous tissues are crucial for dissipating mechanical load during physical activity. Modern procedures to repair these tissues have proven inadequate to restore full functionality, thus there is a need

Musculoskeletal heterogenous tissues are crucial for dissipating mechanical load during physical activity. Modern procedures to repair these tissues have proven inadequate to restore full functionality, thus there is a need for alternative reconstructive methods. Consequently, tissue engineered scaffolds can mimic the native structure of tissues and trigger a healing response. Heterogenous tissues like the tendon-bone junction consist of an interdigitated fiber alignment gradient from the tendon to the bone. It has been shown that electrospun fiber alignment gradients can be fabricated from the incorporation of magnetic fields. In this study, manipulating electrostatic and magnetic interactions from various electrospinning collector arrangements were investigated for creating an interdigitated fiber alignment gradient. The collector arrangement consisting of a magnet overlaid with razor cut aluminum foil proved to provide increased control over the interfacial shape. The rapid transition at the interfacial region was verified with brightfield microscopy revealing an interdigitated gradient from highly aligned fibers to unaligned fibers.

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Date Created
  • 2020-05

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Electrically Conductive Hydrogel-Based Topographies for Development of Three Dimensional (3D) Cardiac Tissues

Description

Cardiac tissue engineering is an emerging field that has the potential to regenerate and repair damaged cardiac tissues after myocardial infarction. Numerous studies have introduced hydrogel-based cardiac tissue constructs featuring

Cardiac tissue engineering is an emerging field that has the potential to regenerate and repair damaged cardiac tissues after myocardial infarction. Numerous studies have introduced hydrogel-based cardiac tissue constructs featuring suitable microenvironments for cell growth along with precise surface topographies for directed cell organization. Despite significant progress, previously developed cardiac tissue constructs have suffered from electrically insulated matrices and low cell retention. To address these drawbacks, we fabricated micropatterned hybrid hydrogel constructs (uniaxial microgrooves with 50 µm with) using a photocrosslinkable gelatin methacrylate (GelMA) hydrogel incorporated with gold nanorods (GNRs). The electrical impedance results revealed a lower impedance in the GelMA-GNR constructs versus the pure GelMA constructs. Superior electrical conductivity of GelMA-GNR hydrogels (due to incorporation of GNRs) enabled the hybrid tissue constructs to be externally stimulated using a pulse generator. Furthermore, GelMA-GNR tissue hydrogels were tested to investigate the biological characteristics of cultured cardiomyocytes. The F-actin fiber analysis results (area coverage and alignment indices) revealed higher directed (uniaxial) cytoskeleton organization of cardiac cells cultured on the GelMA-GNR hydrogel constructs in comparison to pure GelMA. Considerable increase in the coverage area of cardiac-specific markers (sarcomeric α-actinin and connexin 43) were observed on the GelMA-GNR hybrid constructs compared to pure GelMA hydrogels. Despite substantial dissimilarities in cell organization, both pure GelMA and hybrid GelMA-GNR hydrogel constructs provided a suitable microenvironment for synchronous beating of cardiomyocytes.

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Date Created
  • 2016-05

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Edible Microfluidics: 3D Printing Hydrogels Using a Co-Flow Nozzle Extruder

Description

3D printing has recently become a popular manufacturing process and the goal of the project was to take that process to the kitchen. This was done by utilizing existing knowledge

3D printing has recently become a popular manufacturing process and the goal of the project was to take that process to the kitchen. This was done by utilizing existing knowledge of the culinary process of "spherification", by which a liquid is encapsulated in an edible shell, and combining it with the hydrogel research advancements in tissue engineering to make robust fibers. A co-flow nozzle was constructed and the two fluids needed for spherification were flowed in various configurations to create different fibers. By outlining a stability regime and measuring the outer diameters for both regular and reverse spherification, the optimal method of production and fibers that would be suitable for 3D printing were discovered. The results of the experiments can be used to begin 3D printing edible 2D patterns and eventually 3D structures.

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Date Created
  • 2015-05

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Creating Biochemical Gradients via Photoconjugation and an In-House Designed Sliding Photomask

Description

Tissue engineering is an emerging field focused on the repair, replacement, and regeneration of damaged tissue. Engineered tissue consists of three factors: cells, biomolecular signals, and a scaffold. Cell-free scaffolds

Tissue engineering is an emerging field focused on the repair, replacement, and regeneration of damaged tissue. Engineered tissue consists of three factors: cells, biomolecular signals, and a scaffold. Cell-free scaffolds present a unique opportunity to develop highly specific microenvironments with tunable properties. Norbornene-functionalized hyaluronic acid (NorHA) hydrogels provide spatial control over biomolecule binding through a photopolymerization process. With this, biomimetic gradients can be produced to model a variety of tissue interfaces. To produce these patterns, a gradient mechanism was developed to function in tandem with a syringe pump. A conversion equation was derived to calculate a panel speed from the volumetric flow rate setting on the pump. Seven speeds were used to produce fluorophore gradients on the surface of NorHA hydrogels to assess changes in the length and slope of the gradient. The results indicated a strong positive linear correlation between the speed of the panel and the length of the gradient as well as a strong negative correlation between the speed of the panel and the slope of the gradient. Additionally, the mechanism was able to successfully produce several other types of gradients including multiregional, dual, and triregional.

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Date Created
  • 2019-05

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Improving Offset Electrospinning for the Tendon-Bone Junction

Description

The tendon-bone junction, also known as the enthesis, is crucial for properly transferring mechanical loadings during physical activity. During injury, current restoration procedures are insufficient for properly restoring tissue function.

The tendon-bone junction, also known as the enthesis, is crucial for properly transferring mechanical loadings during physical activity. During injury, current restoration procedures are insufficient for properly restoring tissue function. Thus, it is paramount to design alternative tissue engineered scaffolds to act as a template to the injured region and a regenerative response for tendon-bone repair. Thus, we utilized an offset electrospinning technique to fabricate a scaffold that mimics the native biochemical gradients present within the tendon-bone junction. To improve chemical gradient resolution, we implemented both insulating and conductive shields during offset electrospinning. Polycaprolactone fibers with either rhodamine or fluorescein were used to measure the scaffold fluorescent strength with distance. Without shields, at an offset of 4 cm, the chemical gradient resolution for rhodamine and fluorescein were 2.5 cm and 6.0 cm, respectively. During implementation of insulating shields, the gradient resolution for rhodamine and fluorescein improved to 2 cm and 0.5 cm, respectively. Lastly, grounded conductive shields improved gradient resolution for rhodamine and fluorescein to 1.0 cm and 1.5 cm, respectively.

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Date Created
  • 2019-05

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Modulating chemokine receptor expression in neural stem cell transplants to promote migration after traumatic brain injury

Description

Traumatic brain injury (TBI) is a significant public health concern in the U.S., where approximately 1.7 million Americans sustain a TBI annually, an estimated 52,000 of which lead to death.

Traumatic brain injury (TBI) is a significant public health concern in the U.S., where approximately 1.7 million Americans sustain a TBI annually, an estimated 52,000 of which lead to death. Almost half (43%) of all TBI patients report experiencing long-term cognitive and/or motor dysfunction. These long-term deficits are largely due to the expansive biochemical injury that underlies the mechanical injury traditionally associated with TBI. Despite this, there are currently no clinically available therapies that directly address these underlying pathologies. Preclinical studies have looked at stem cell transplantation as a means to mitigate the effects of the biochemical injury with moderate success; however, transplants suffer very low retention and engraftment rates (2-4%). Therefore, transplants need better tools to dynamically respond to the injury microenvironment.

One approach to develop new tools for stem cell transplants may be to look towards the endogenous repair response for inspiration. Specifically, activated cell types surrounding the injury secrete the chemokine stromal cell-derived factor-1α (SDF-1α), which has been shown to play a critical role in recruiting endogenous neural progenitor/stem cells (NPSCs) to the site of injury. Therefore, it was hypothesized that improving NPSC response to SDF-1α may be a viable mechanism for improving NPSC transplant retention and migration into the surrounding host tissue. To this end, work presented here has 1. identified critical extracellular signals that mediate the NPSC response to SDF-1α, 2. incorporated these findings into the development of a transplantation platform that increases NPSC responsiveness to SDF-1α and 3. observed increased NPSC responsiveness to local exogenous SDF-1α signaling following transplantation within our novel system. Future work will include studies investigating NSPC response to endogenous, injury-induced SDF-1α and the application of this work to understanding differences between stem cell sources and their implications in cell therapies.

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Date Created
  • 2015