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Oxygen is the single most important molecule for sustaining life and, therefore, an important variable in tissue engineering and regenerative medicine. It has been shown that the change in oxygen concentration in an artificial or tissue-engineered graft affects cell survival, differentiation, and tissue growth in profound ways. However, at present, there are no reliable methods to map partial oxygen pressure (pO2) in growing artificial tissues. Here, we adapt and test the suitability of electron paramagnetic resonance oxygen imaging (EPROI) in assessing tissue graft oxygenation in vitro. EPROI is an established method to assess absolute pO2and has been widely applied to study tumor hypoxia in small animals. In this study, we demonstrate the feasibility of EPROI in evaluating oxygen dynamics in tissue grafts. We measured oxygen concentration in mesenchymal stem cell (MSC)-seeded polylactic-co-glycolic acid (PLGA) scaffolds with variable porosity. The pO2maps of these scaffolds showed that the mean pO2inside the scaffolds was smaller than the ambient air pO2(21% oxygen, 160 torr) and was gradually increased with increasing pore size. We assessed the local oxygen dynamics of the MSC-seeded osteogenic scaffold made from collagen–chitosan hydrogels in a partially sealed Eppendorf tube. The change in pO2values as a function of time inside the graft showed that the cells had used available oxygen within first 2 h of the experiment and then went to a dormant low oxygen consumption state until the oxygen supply was reestablished. Collectively, these data suggest that EPROI could be successfully used for mapping pO2in tissue-engineered grafts. The knowledge of tissue graft oxygenation may be used to improve scaffold design and to assess the tissue viability and growth.

A key technical challenge in cartilage tissue engineering is the development of a noninvasive method for monitoring the composition, structure, and function of the tissue at different growth stages. Due to its noninvasive, three-dimensional imaging capabilities and the breadth of available contrast mechanisms, magnetic resonance imaging (MRI) techniques can be expected to play a leading role in assessing engineered cartilage. In this review, we describe the new MR-based tools (spectroscopy, imaging, and elastography) that can provide quantitative biomarkers for cartilage tissue development both in vitroand in vivo. Magnetic resonance spectroscopy can identify the changing molecular structure and alternations in the conformation of major macromolecules (collagen and proteoglycans) using parameters such as chemical shift, relaxation rates, and magnetic spin couplings. MRI provides high-resolution images whose contrast reflects developing tissue microstructure and porosity through changes in local relaxation times and the apparent diffusion coefficient. Magnetic resonance elastography uses low-frequency mechanical vibrations in conjunction with MRI to measure soft tissue mechanical properties (shear modulus and viscosity). When combined, these three techniques provide a noninvasive, multiscale window for characterizing cartilage tissue growth at all stages of tissue development, from the initial cell seeding of scaffolds to the development of the extracellular matrix during construct incubation, and finally, to the postimplantation assessment of tissue integration in animals and patients.

The purpose of this study was to assess the natural pO2 of subcutaneous (SC) and intraperitoneal (IP) sites in mice to determine their relative suitability as sites for placement of implants. The pO2 measurements were performed using oxygen imaging of solid probes using lithium phthalocyanine (LiPc) as the oxygen sensitive material. LiPc is a water-insoluble crystalline probe whose spin-lattice and spin-spin relaxation rates (R1 and R2) are sensitive to the local oxygen concentration. To facilitate direct in vivo oxygen imaging, we prepared a solid probe containing encapsulated LiPc crystals in polydimethylsiloxane (PDMS), an oxygen-permeable and bioinert polymer. While LiPc-PDMS or similar probes have been used in repeated spectroscopic or average oxygen measurements using continuous wave (CW) electron paramagnetic resonance (EPR) since the late 1990s and now have advanced to clinical applications, they have not been used for pulse EPR oxygen imaging. One LiPc-PDMS probe of 2 mm diameter and 10 mm length was implanted in subcutaneous or intraperitoneal sites (left or right side) in each animal. The pO2 imaging of implanted LiPc-PDMS probes was performed weekly for six weeks using O2M’s preclinical 25 mT oxygen imager, JIVA-25, using the pulse inversion recovery electron spin echo (IRESE) method. At week 6, the probes were recovered, and histological examinations were performed. We report here, first-ever solid probe oxygen imaging of implanted devices and pO2 assessment of subcutaneous and intraperitoneal sites.

Engineering biomaterials mimicking the biofunctionality of the extracellular matrix (ECM) is important in instructing and eliciting cell response. The native ECM is highly dynamic and has been shown to support cellular attachment, migration, and differentiation. The advantage of synthesizing an ECM-based biomaterial is that it mimics the native cellular environment. However, the ECM has tissue-specific composition and patterned arrangement. In this study, we have employed biomimetic strategies to develop a novel collagen/chitosan template that is embedded with the native ECM of differentiating human marrow stromal cells (HMSCs) to facilitate osteoblast differentiation. The scaffold was characterized for substrate stiffness by magnetic resonance imaging and nanoindentation and by immunohistochemical analysis for the presence of key ECM proteins. Gene expression analysis showed that the ECM scaffold supported osteogenic differentiation of undifferentiated HMSCs as significant changes were observed in the expression levels of growth factors, transcription factors, proteases, receptors, and ECM proteins. Finally, we demonstrate that the scaffold had the ability to nucleate calcium phosphate polymorphs to form a mineralized matrix. The results from this study suggest that the three-dimensional native ECM scaffold directly controls cell behavior and supports the osteogenic differentiation of mesenchymal stem cells.

Electric field gradients (efg) at the sites of sodium in the ferroelectric NaNO2 are calculated for various local motions. It is found that the values of efg obtained from 23Na NMR data are best represented when the NO2 groups are considered to be executing flipping motion by 170° about an axis whose direction cosines are 0.99, 0.11, 0.06 with respect to the crystal axes (c, a, b). Such a motion is in conformation with the not-so-widely known fact that the electric polarization in ferroelectric NaNO2 has a spiraling orientation as one moves from one domain to the other.