Kenneth A. Barbee - US grants

9984276
Barbee
The promise of tissue engineering lies in the prospect of replacing tissue that has become dysfunctional due to trauma or disease with new tissue capable of responding and adapting to environmental stimuli. Of primary importance in tissues that serve a mechanical or structural function is the ability to sense and respond to mechanical forces in order to adapt to the changing physical demands on the tissue. Previous in vivo and in vitro studies suggest that the structure and mechanical properties of blood vessel walls develop in response to the stress history of the tissue. The endothelium mediates vascular tone and structural remodeling in response to changes in blood flow while the vascular smooth muscle (VSM) cells sense and respond to changes in stress within the vessel wall. These responses are essential to the maintenance of structural integrity and the regulation of blood flow.

The central hypothesis of this research is that in normal development, structural relationships in vascular tissue are optimized for efficient sensing and transduction of the mechanical environment by the cells of the vessel wall. To engineer a tissue structure intended to acquire the property of adaptability present in normal tissues, we must first understand the salient features of the cells' interaction with their surrounding structures that allow appropriate mechanotransduction to occur. The structure and biochemistry of engineered matrices as well as pre-conditioning with physiological loading regimes will be analyzed and optimized based on initial functional properties and the acquisition of adaptive behaviors that will allow long-term replacement of tissue.

The objective of this career plan is to develop a program of research and education in the area of cellular and molecular mechanics. The research focus will be on the cellular response to mechanical stimuli with a special emphasis on the altered stress environments created by tissue-implant interfaces and engineered tissue constructs. The educational emphasis will be on the integration of engineering mechanics with current developments in the biological sciences. This will require not only a merging of course content but also a blending of the disparate course formats and teaching styles from the physical and biological sciences. Graduate courses that integrate theoretical and experimental foundations of cell mechanics with current concepts in the biological literature will be developed. In addition, the introduction of clinical rotations into the graduate curriculum will expose students to the state of the art in medical practice and patient care. This experience is intended to broaden their perspective and to focus their research objectives. At the undergraduate level, emphasis will be placed on providing research opportunities to stimulate students' interest in research-oriented careers either in industrial R&D or in further academic training.

0216343
Lowman
The acquisition of an Environmental Scanning Electron Microscope (ESEM),is requested. To date, no Philadelphia area universities have ESEM capabilities, so this project would also be of significant interest to local universities and industries through partnerships established through the Nanotechnology Center. Additionally, the proposed work will have a strong impact on Drexel University's educational initiatives with respect to community outreach, mentorship activity and graduate/undergraduate education. This will also include high students through summer courses designed for underrepresented minorities from Philadelphia schools as well as students from Math and Science academies. Additionally, new graduate and undergraduate curricula are being established that will make use of ESEM as a training tool.

This award supports construction and testing of a biosensor intended to measure the strength of adhesion of cells to surfaces through use of acoustic shear waves generated by a piezoelectric device. The sensor, to be called the Acoustic Shear Wave Biosensor for the Analysis of Cell Adhesion and Structure (ABACAS), is expected to measure mass accumulation in the nanogram range, chemical species concentration on the order of 1 ppb, and the complex elastic modulus for a large range of materials ranging from purely viscous to purely elastic media. Several model systems employing polystyrene microspheres as cell mimics, polymer gels simulating extracellular matrix proteins, and controlled spreading of cells will be used to calibrate the ABACAS measurements. Standard tests for cell adhesion strength, endothelial cell adhesion, spreading, and realignment in response to flow will be used to correlate ABACAS measurements with biological function.

Currently, characterization of the dynamics of interactions between biomaterials and cells relies on techniques that require many samples for temporal definition of any property. The new sensor is expected to meet the need for reliable, multipurpose and low cost miniature instrumentation for characterization of such interactions, allowing real-time measurements using a single sample.

Developments from the ABACAS project in both sensor design and cellular mechanics will also be incorporated into the institution's curriculum at the undergraduate and graduate levels. An undergraduate sequence on Biosensors taught by the PI will use ABACAS as both an example to illustrate lecture topics and a laboratory exercise. ABACAS will also be used as part of a a graduate course in Cellular Biomechanics to emphasize the relationship between theoretical modeling and experimental design in characterizing the mechanics of cell behavior. In addition, the project will be incorporated into the Biosensors Collaboratory, a unique on-line environment that gives access to the laboratory to collaborators and educational partners. This Collaboratory facilitates outreach efforts to 2-year colleges and high schools in the Philadelphia area.

0301446
Jaron
Nitric oxide (NO) plays an important role in regulation of blood flow and metabolism, however, its transport properties and mechanisms of action in the microcirculation are not well understood. The general goal of the proposed research is to use an integrated engineering approach combining mathematical modeling and in vitro and in vivo experiments to improve our understanding of the interacting mechanisms between NO, oxygen, carbon dioxide, hemoglobin, calcium, oxygen free radicals, and thiols in blood and tissue. A comprehensive state-of-the-art computational model coupled with appropriate experiments will be used to evaluate and quantify hypothesized mechanisms of NO transport, evaluate interactions between mechanisms, and to assess the relative contributions of each. The mathematical model will utilize dynamic mass transport and fluid mechanics calculations in conjunction with multiple chemical reactions to simulate production, transport, scavenging, feedback regulation, and other mechanisms of action of NO and associated chemical species in the microcirculation and tissue. Experimental studies have been designed to provide key parameters for mathematical modeling and will be used to test model validity. Quantitative data obtained from the validated model will be used to predict parameters that cannot be measured in vivo, analyze the hypotheses and further the understanding of NO production and transport mechanisms, and shape future studies.

0730547
Jaron

This proposal concerns the development a multi-scale approach to modeling nitric oxide production, transport and metabolism. A total of 5 objectives are stated for this proposal: (1) Expand an existing cell-scale model to include intracellular signaling and extra-cellular transport, (2) Couple a vascular-scale and cellular-scale model, (3) Develop a vascular network-scale model of NO transport, (4) Extend the model to a tissue level scale, and (5) Validate the theoretical simulations with in vitro and in vivo studies. The transport and metabolism of NO plays an important role in many normal and pathological tissue functions. As a result, this study could make important contributions to medicine and vascular physiology in general. The coupling of hemodynamics, multiple chemical reactions, and biophysical processes in an integrated multi-scale model is central to a vast number of important physiological and pathophysiological processes.

The coupling of the proposed research with existing courses and other education components is a strength of this proposal. In particular, including this work in cellular biomechanics, tissue engineering, and cardiovascular engineering course work will give students an opportunity to learn some cutting edge research. The PIs have a track record of including undergraduates in their research programs. Also, multi-scale and multi-temporal methods are becoming more important in biology and physiology. As a result, this work is likely to impact a number of other areas besides NO metabolism.

This grant provides funding for research and development of a scientific and engineering knowledge base for studying the effect of bio-deposition process on living cells. The project activities involve: 1) studying the feasibility of depositing living cells and constructing cell-embedded structures; 2) developing an engineering model as well as a multi-scale modeling approach to predict the deposition process-induced mechanical forces and the effect of the process and the mechanical forces on the biological behavior of cells; and 3) quantifying the biological responses of cells, its viability, recovery, proliferation, and damage under varying processing conditions. This project can lead to developing new techniques and tools to help the advancement of emerging cell-based bio-fabrication for broad applications in therapeutic products, biochips and biosensors, diagnostic arrays, microfluidic systems, and pharmaceutical methods.

If successful, this research will foster the convergence of engineering and life sciences, and provide viable manufacturing tools to biology and life sciences community. This project will actively engage industry, government and medical institution. The outcome of research can be applied to help the mission of safe exploration of space, the development of biofabrication product standardizations, and the prioritization of the future cell-based products for tissue engineering manufacturer, thus have broad impacts on the technological development, economic growth, and improvement of the quality of life. This project will also develop a new interdisciplinary course spanning Design/Manufacturing, Biomaterials, and Bio/Cell Mechanics to educate the next generation of scientists and engineers.

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5)

0853733
Marcolongo


Intellectual Merit: One of the critical challenges of modern reproductive biology is to develop viable clinical approaches to treat infertility. Embryo implantation represents a crucial step of the reproductive process and its success relies on the culmination of a well-orchestrated series of spatially and temporally controlled events. The attachment of the blastocyst to the endometrium is mediated, at least in part, by the L-selectin adhesion system. Interactions between L-selectin expressed on the peripheral cells of the human blastocyst, the trophoblasts, and its oligosaccharide ligands on the surface of the human endometrium are thought to provide a "braking" mechanism for the embryo as it enters the endometrial cavity, similar to its role in other biological systems. While the L-selectin mediated interaction is likely not strong enough to sustain attachment (which is further enhanced by secondary integrin-ligand interactions), it is thought to provide strength adequate to allow the blastocyst to be initially captured by the uterine luminal epithelium. However, there is a lack of understanding of the attachment mechanisms associated with primary and secondary adhesion. A principle cause of the 30% failure rates of in vitro fertilization is associated with a defect in the attachment process during implantation; without further understanding of the embryo attachment mechanisms, little can be done to address this critical clinical issue.

There are two objectives of this application: 1) to utilize model cell lines for trophoblasts (Jeg-3) and uterine epithelial cells (Ishikawa) and cell mechanics techniques to examine the dependence of molecular expression (controlled by environmental factors: hormonal stimulation and fluid flow) on primary and secondary attachment strength of the blastocyst/uterine epithelium interactions and 2) to develop and characterize a 3D Jeg-3 trophosphere to better mimic the architecture of the blastocyst and examine biochemical and mechanical interactions of the trophosphere with a 3D tissue engineered structure of human endometrial epithelial cells obtained from biopsies (IRB 15822) to more fully mimic the in vivo environment. The central hypothesis is that the expression and retention of a critical level of L-selectin and L-selectin ligands are necessary to facilitate adequate initial attachment of the blastocyst to the uterine epithelium and similarly the expression of molecules in the integrin-mediated attachment will control the strength of attachment for secondary bonding. The preliminary work has shown that the research team can manipulate L-selectin and L-selectin ligand expression with hormonal conditioning in trophoblast and endometrial epithelial cell models, respectively. For the first time, the attachment strength of a model L-selectin mediated implantation system has been quantified using a parallel plate flow chamber customized with a quartz crystal microbalance (QCM) sensor and functional attachment dependence based on L-selectin expression has been demonstrated. Further, the secondary attachment strength of the model integrin/ligand system has been quantified using a spinning disc apparatus. In addition to our supportive preliminary data, we are particularly poised to undertake this research because an inter-disciplinary team of bioengineers, biochemists and cell biologists and a fertility clinician with the range of skills necessary to answer these critical questions has been assembled.

This project is innovative because it will be the first time that the primary and secondary attachment strengths for blastocyst implantation have been investigated. An additional innovation comes in the use of the 3D trophosphere model to examine the spatial effects that may contribute to the attachment mechanisms. This work also realizes an engineering approach toward challenges of maternal fetal medicine and therefore serves as a milestone at the intersection of these disciplines.

Broader Impacts: Development of the model blastocyst/uterine endometrial epithelial cell system to examine the effect of microenvironment (hormonal and physical) on the primary and secondary attachment mechanisms will allow further research to examine the effects of endogenous factors and possible clinically relevant treatments on the functional behavior of the cells. The 3D models of the trophosphere and the hEEC enable the advancement of the understanding of the effect of the spatial distribution of molecules on attachment. These insights may inspire new treatments for implantation-related fertility, one of the greatest impediments to successful reproductive outcomes in IVF, where, despite fertilization rates of up to 60%, pregnancy rates per transfer of embryos to the uterus are only 42.5%. This program lays the groundwork for exploration of further questions related to infertility that could greatly benefit from an engineering approach. These methods and discoveries will be disseminated to the scientific community through publications and presentations at conferences. The proposed research at the intersection of disciplines will offer excellent training opportunities to students as they study this clinically relevant and important problem.

DESCRIPTION (provided by applicant): The goal of the proposed work is to determine the mechanisms of the early structural consequences of neural trauma and to develop strategies for acute intervention in traumatic brain injury (TBI) that address the primary mechanisms of axonal injury. The underlying hypothesis for this research is that initial mechanical trauma to the cell membranes leads to cytoskeletal disruptions and alterations of axonal transport and that acute intervention to restore membrane integrity and preserve axonal cytoskeleton and transport processes can dramatically reduce secondary degeneration and cell death. Specific Hypotheses to be tested are 1) Axonal cytoskeletal disruption and impaired axonal transport are causally related to membrane disruption, and acute repair of the axolemma by poloxamer P188 can prevent these effects;2) JNK-3 activation is causally related to membrane damage, and axonal transport is, in part, impaired by the actions of JNK-3;and 3) Mild injury will be manifest in axonal pathology, the time course of which is modulated by injury severity. We have developed and in vitro model of focal axonal injury in primary chick forebrain (CFB) neurons that mimics many features observed in vivo. In particular, focal swelling, or axonal beads, appeared within one hour following the mechanical insult. Co-localized with the beads were focal disruptions of microtubules and the accumulation of membrane bound organelles indicating a disruption of axonal transport. We characterized the membrane damage as a result of the mechanical trauma and showed that treatment with Poloxamer 188 (P188), a water soluble, non-ionic surfactant restored membrane integrity and significantly inhibited axonal beading in CFB neurons. We also have an in vivo model that produces axonal injury in the deep white matter. In this model, we have demonstrated membrane damage, focal accumulation of amyloid precursor protein (APP), and focal activation of JNK-3. Significantly, injured JNK-3 deficient mice do not exhibit the severe cognitive deficits seen in age-matched WT littermates. The following Specific Aims were developed to test these hypotheses: Aim 1: To determine the causal relationship between mechanically-induced membrane damage and subsequent alterations of cytoskeletal structure and axonal transport and to test whether acute treatment with an agent that promotes membrane repair can preserve axonal structure and function and thereby prevent secondary degeneration. Aim 2: To determine the mechanism of focal activation of the MAP kinase, JNK-3, in injured axons and its role in axonal pathology and to test the effect of acute membrane repair on JNK activation. Aim 3: To determine the window of opportunity for therapeutic intervention for both membrane repair and inhibition of JNK-3. PUBLIC HEALTH RELEVANCE: Currently, acute care for trauma victims deals mainly with preserving or restoring basic life support systems, e.g., cardiac and respiratory function, and managing mass lesions in the brain to prevent death and/or further brain damage. This research, if successful, will provide the basis for a new approach to the treatment of traumatic brain injury in which early intervention to preserve the structural integrity of neurons will stave off the secondary degenerative processes that result in persistent neurological deficits. Successful completion of the proposed research will hopefully emphasize the importance of the early treatment of neuronal injury as an important therapeutic consideration in addition to the current focus on delayed treatments aimed at halting secondary degeneration.

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5).

This project involves the renovation of the Drexel University Microfabrication Facility (MFF). Parts of the facility will be renovated to the standards of Class 100 and Class 1000 clean rooms. While the MFF houses a number of micro-fabrication and nano-fabrication instruments acquired through recent Major Research Instrumentation awards and other sources, the existing MFF is a low-dust environment rather than a true cleanroom and this limits the types of research that can be done there.

The renovated facility will be used for research in the areas of micro- and nanofluidics, Micro-Electro-Mechanical Systems and Nano-Electro-Mechanical Systems, bioreactors, tissue engineering, biomechanics and biophysics, biosensors, lab-on-a-chip technology, surgical engineering, neuroengineering, optoelectronics, solar cells and alternative energy, colloidal suspension dynamics, and novel nanoelectronic devices, all of which require micro- and nanofabrication.

The facility provides research infrastructure. The facility will be used by researchers from both Drexel and the wider Philadelphia region. The project will support the research of a number of young faculty members from several different engineering departments. The users of the renovated facility will include undergraduates, graduate students and post-doctoral research associates. The MFF will be also be used in a number of outreach programs, including training and mentoring K-12 teachers and students from the Delaware Valley region.

DESCRIPTION (provided by applicant): The overall goal of this Project is to enhance the senior design program at Drexel by sharpening the focus on translational design solutions and facilitating the achievement of completed designs with working prototypes. The proposed enhancements leverage the existence of an active program for the support of translational research supported by the Coulter Translational Partners program. The goal is to transform senior design projects from academic exercises to the development of design solutions to unmet clinical needs by matching students with clinical and engineering investigators who are actively developing and commercializing biomedical technologies. Three Specific Aims were developed to meet these objectives. Specific Aim 1 is to enhance the design program through vertical integration of Junior and Senior level design teams. Junior design teams will be matched with a senior design team and interact with them throughout the Junior Design course. The Senior Design team will make periodic presentations and reports, and the Junior Design group will critique them and provide feedback. The junior team will be responsible for completing a competitive technology analysis through patent and business database searches, and drafting a report. The involvement of students in the junior year with real, ongoing projects will enhance their understanding of the design process in ways that cannot be simulated by the didactic components of the current course. Specific Aim 2 is to create a new 'Translational Design Clinical Immersion Co-op. Selected participants will, in their junior year, complete a 10 week clinical rotation with the Clinical co-PI of a Coulter translational project. They will get fist-hand experience of the clinical environment and how it affects the design solution, identify clinical needs for new design solutions, and develop an understanding of the design constraints imposed by the environment in which the design will be used. Following this clinical experience, the students will complete a 10 week design lab co- op in the lab of the Biomedical Engineering co-PI of the Coulter project. They will develop the initial designs for the newly identified applications or participate in the design validation and refinement of an existing solution and recruit a multidisciplinary team to carry out the design. The Clinical co-PI will be involved in advising the team throughout this two-year process. Specific Aim 3 Is to improve Senior Design outcomes by providing resources that facilitate prototyping and testing and, thus, allowing more iterations in the design process. We will provide computational tools for design, funding for materials and supplies and funding and training for advanced prototyping techniques. We have developed a system to evaluate student learning outcomes and provide instructors with feedback on the content and its delivery from the students' perspective. This system assigns objectives (selected by the instructor in accordance with ABET criteria) to a class and evaluates students on these objectives before the beginning of each class and at its conclusion. It tracks changes in response to comments and success of implementing any changes.