A core technology of regenerative medicine is the bioengineering of a functional tissue or organ by seeding living cells onto a biodegradable scaffold and surgically implanting the construct into a patient. This approach offers great promise to cure chronic or degenerative diseases affecting major organs, and serious battlefield traumas. Tissue regeneration involves extensive remodeling of cells and scaffolds. Despite significant advances, notably the first positive clinical tests of a bioengineered human organ – the bladder, many of the most basic problems of regenerative medicine remain unsolved. These include the prevention of inflammation and scarring, the guidance of cell migration and differentiation, and the promotion of vascularization and innervation of regenerating tissues. A major barrier to progress is our inability to monitor dynamic biological processes in a minimally invasive real-time fashion, which makes control and optimization extremely difficult. Current methods to assess tissue regeneration, such as histology or organ bath physiology, are highly invasive. Thus, there is a fundamental knowledge discrepancy between cellular processes and systems biology.
As a key component of the NIH roadmap, molecular imaging is playing an important role in revolutionizing medicine. The combination of fluorescent and bioluminescent probes with optoelectronics and computing techniques has led to the development of optical molecular imaging tools that allow the visualization of biologic interactions in complex, living systems over time. This powerful technology drives a paradigm shift from static assays of cellular function in biopsied tissue or 2D culture models towards systematic analyses of 3D systems in vitro and in vivo. This capability greatly enhances our ability to assess physiology and pathology, and improve clinical practice. However, despite the enormous potential of optical molecular imaging, it has not yet been harnessed as an enabling technology for regenerative medicine.
The overall goal of this project is to develop a first-of-its-kind multi-probe, multi-modal optical molecular tomography (OMT) system for regenerative medicine and to demonstrate its utility to visualize the development of bioengineered blood vessels in bioreactors and after implantation into living animals in real-time. We are inspired by two hypotheses: (1) optical imaging, photon transport modeling, and 3D image reconstruction will allow the non-invasive, dynamic analysis of bioengineered tissue constructs comprising scaffolds seeded with multiple types of cells; and (2) tomography of distinct fluorescent probes can remarkably improve the interrogation of the development, remodeling, and functionality of such constructs, thereby facilitating their optimization. We will test these hypotheses through the following specific aims:
Aim 1 – System Prototyping: Develop prototypes for hybrid optical molecular tomography to monitor bioengineered blood vessel constructs in bioreactors (in vitro prototype) and after implantation into sheep (in vivo prototype). We will employ fluorescent probes to label a cylindrical electrospun matrix scaffold and two cell types (vascular endothelial and smooth muscle cells). Optical fibers embedded in the scaffold will facilitate optical coherent tomography (OCT) and optical molecular tomography (OMT). The primary novelty of this multi-modal system lies in the enabling capabilities for multi-probe high-resolution analyses.
Aim 2 – Algorithm Development: Develop processing and reconstruction methods for the proposed optical molecular tomography system. Processing will be needed to minimize any excitation, measurement and reconstruction artifacts/noise. We will perform multi-spectral fluorescence tomography reconstructions based on our new phase-approximation model for photon transport. We will conduct systematic comparison in numerical simulation and tissue phantom experiments to optimize the imaging performance towards the design goals of 100μm 3D resolution in vitro and 500μm 3D resolution in vivo.
Aim 3 – In vitro Studies: Apply the imaging system to assess development of the bioengineered vessels in a pulsatile bioreactor for up to 30 days. We will investigate the endothelial coverage of the scaffold’s lumenal surface, migration of smooth muscle cells in the scaffold’s porous outer compartment, and population dynamics of the two cell types. Fluorescent reporters will be utilized to monitor cell-type-specific gene expression in real-time, and to verify physiological responses of cells in the engineered vessel. We will validate optical molecular tomography results by comparison with conventional assays, including histology and measurements of specific mRNA and proteins.
Aim 4 – In vivo Studies: Apply the imaging system to assess the further development of bioengineered blood vessel constructs after anastomosis into the carotid arteries of sheep (up to 4 months). Features of interest to be analyzed are similar to those in vitro, and include scaffold remodeling, and the survival, continued growth and migration, and tissue-appropriate gene expression of cells initially seeded in the vessels. We will validate the OMT results by comparison with traditional histological, molecular, and physiological analyses.
The VT-WFU School of Biomedical Engineering & Sciences (SBES) is a joint venture between Virginia Tech and Wake Forest University. This close collaboration between public and private institutions of national reputation seeks to improve human and animal health through interdisciplinary teamwork in engineering, science and medicine. To promote our tomographic imaging program within the SBES framework, PI Dr. Wang directs the Biomedical Imaging Division, and has the Optical Molecular Tomography Lab on both Virginia Tech and Wake Forest campuses. WFIRM is an important contributor to SBES through joint appointments, graduate courses in regenerative medicine, and research collaboration. Communication is facilitated through videoconferencing, in addition to frequent visits and seminars. WFIRM scientists were the first in the world to implant a laboratory-grown organ into humans and today are working to develop >22 different bioengineered organs and tissues. This partnership offers tremendous inter- and trans-disciplinary research opportunities as exemplified by this BRP project.
The technical work will be performed in PI Dr. Ge Wang’s Optical Molecular Tomography Laboratory and other Virginia Tech co-investigators’ research spaces on both the Wake Forest University Medical School Campus and the Virginia Tech Main Campus. The multi-probe multi-modal OMT system including the in vitro and in vivo prototypes will be first developed and validated at Virginia Tech in close collaboration with the tissue engineering team members, and then moved to Dr. Ge Wang’s Optical Molecular Tomography Laboratory on the Wake Forest University Medical School Campus. The tissue engineering work will be done by PI Dr. Shay Soker and his collaborators in the Wake Forest Institute of Regenerative Medicine in collaboration with the optical imaging team members. Over the past years, we have been successfully developing and applying other OMT systems for different projects characterized by collaboration between researchers on these two campuses in the same way, and anticipate no logistic problem in this BRP project given the geographical closeness and portability of the involved optical imaging components (we already have high-quality optical benches on both the two campuses).
Website for the NIH/BRP Project “Optical Molecular Tomography for Regenerative Medicine“
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