Mark F. Lythgoe

De novo cardiomyocytes from within the activated adult heart after injury

A significant bottleneck in cardiovascular regenerative medicine is the identification of a viable source of stem/progenitor cells that could contribute new muscle after ischaemic heart disease and acute myocardial infarction. A therapeutic ideal—relative to cell transplantation—would be to stimulate a resident source, thus avoiding the caveats of limited graft survival, restricted homing to the site of injury and host immune rejection. Here we demonstrate in mice that the adult heart contains a resident stem or progenitor cell population, which has the potential to contribute bona fide terminally differentiated cardiomyocytes after myocardial infarction. We reveal a novel genetic label of the activated adult progenitors via re-expression of a key embryonic epicardial gene, Wilm’s tumour 1 (Wt1), through priming by thymosin β4, a peptide previously shown to restore vascular potential to adult epicardium-derived progenitor cells with injury. Cumulative evidence indicates an epicardial origin of the progenitor population, and embryonic reprogramming results in the mobilization of this population and concomitant differentiation to give rise to de novo cardiomyocytes. Cell transplantation confirmed a progenitor source and chromosome painting of labelled donor cells revealed transdifferentiation to a myocyte fate in the absence of cell fusion. Derived cardiomyocytes are shown here to structurally and functionally integrate with resident muscle; as such, stimulation of this adult progenitor pool represents a significant step towards resident-cell-based therapy in human ischaemic heart disease.

In vivo imaging of glucose uptake and metabolism in tumors

Tumors have a greater reliance on anaerobic glycolysis for energy production than normal tissues. We developed a noninvasive method for imaging glucose uptake in vivo that is based on magnetic resonance imaging and allows the uptake of unlabeled glucose to be measured through the chemical exchange of protons between hydroxyl groups and water. This method differs from existing molecular imaging methods because it permits detection of the delivery and uptake of a metabolically active compound in physiological quantities. We show that our technique, named glucose chemical exchange saturation transfer (glucoCEST), is sensitive to tumor glucose accumulation in colorectal tumor models and can distinguish tumor types with differing metabolic characteristics and pathophysiologies. The results of this study suggest that glucoCEST has potential as a useful and cost-effective method for characterizing disease and assessing response to therapy in the clinic.

Compartment models of the diffusion MR signal in brain white matter: A taxonomy and comparison

This paper aims to identify the minimum requirements for an accurate model of the diffusion MR signal in white matter of the brain. We construct a taxonomy of multi-compartment models of white matter from combinations of simple models for the intra- and the extra-axonal spaces. We devise a new diffusion MRI protocol that provides measurements with a wide range of imaging parameters for diffusion sensitization both parallel and perpendicular to white matter fibres. We use the protocol to acquire data from two fixed rat brains, which allows us to fit, study and compare the different models. The study examines a total of 47 analytic models, including several well-used models from the literature, which we place within the taxonomy. The results show that models that incorporate intra-axonal restriction, such as ball and stick or CHARMED, generally explain the data better than those that do not, such as the DT or the biexponential models. However, three-compartment models which account for restriction parallel to the axons and incorporate pore size explain the measurements most accurately. The best fit comes from combining a full diffusion tensor (DT) model of the extra-axonal space with a cylindrical intra-axonal component of single radius and a third spherical compartment of non-zero radius. We also measure the stability of the non-zero radius intra-axonal models and find that single radius intra-axonal models are more stable than gamma distributed radii models with similar fitting performance.

Magnetic Resonance Imaging of Mesenchymal Stem Cells Homing to Pulmonary Metastases Using Biocompatible Magnetic Nanoparticles

The ability of mesenchymal stem cells (MSC) to specifically home to tumors has suggested their potential use as a delivery vehicle for cancer therapeutics. MSC integration into tumors has been shown in animal models using histopathologic techniques after animal sacrifice. Tracking the delivery and engraftment of MSCs into human tumors will need in vivo imaging techniques. We hypothesized that labeling MSCs with iron oxide nanoparticles would enable in vivo tracking with magnetic resonance imaging (MRI). Human MSCs were labeled in vitro with superparamagnetic iron oxide nanoparticles, with no effect on differentiation potential, proliferation, survival, or migration of the cells. In initial experiments, we showed that as few as 1,000 MSCs carrying iron oxide nanoparticles can be detected by MRI one month after their coinjection with breast cancer cells that formed subcutaneous tumors. Subsequently, we show that i.v.- injected iron-labeled MSCs could be tracked in vivo to multiple lung metastases using MRI, observations that were confirmed histologically. This is the first study to use MRI to track MSCs to lung metastases in vivo. This technique has the potential to show MSC integration into human tumors, allowing early-phase clinical studies examining MSC homing in patients with metastatic tumors.

Early changes in water diffusion, perfusion, T1, and T2 during focal cerebral ischemia in the rat studied at 8.5 T †

The time evolution of water diffusion, perfusion, T1, and T2 is investigated at high magnetic field (8.5 T) following permanent middle cerebral artery occlusion in the rat. Cerebral blood flow maps were obtained using arterial spin tagging. Although the quantitative perfusion measurements in ischemic tissue still pose difficulties, the combined perfusion and diffusion data nevertheless distinguish between a “moderately affected area,” with reduced perfusion but normal diffusion; and a “severely affected area,” in which both perfusion and diffusion are significantly reduced. Two novel magnetic resonance imaging observations are reported, namely, a decrease in T2 and an increase in T1, both within the first few minutes of ischemia. The rapid initial decrease in T2 is believed to be associated with an increase in deoxyhemoglobin levels, while the initial increase in T1 may be related to several factors, such as flow effects, an alteration in tissue oxygenation, and changes in water environment. Magn Reson Med 41:479–485, 1999.

A rat decellularized small bowel scaffold that preserves villus-crypt architecture for intestinal regeneration

Management of intestinal failure remains a clinical challenge and total parenteral nutrition, intestinal elongation and/or transplantation are partial solutions. In this study, using a detergent-enzymatic treatment (DET), we optimize in rats a new protocol that creates a natural intestinal scaffold, as a base for developing functional intestinal tissue. After 1 cycle of DET, histological examination and SEM and TEM analyses showed removal of cellular elements with preservation of the native architecture and connective tissue components. Maintenance of biomechanical, adhesion and angiogenic properties were also demonstrated strengthen the idea that matrices obtained using DET may represent a valid support for intestinal regeneration.

Magnetic Tagging Increases Delivery of Circulating Progenitors in Vascular Injury

OBJECTIVES We sought to magnetically tag endothelial progenitor cells (EPCs) with a clinical agent and target them to a site of arterial injury using a magnetic device positioned outside the body. BACKGROUND Circulating EPCs are involved in physiological processes such as vascular re-endothelialization and post-ischemic neovascularization. However, the success of cell therapies depends on the ability to deliver the cells to the site of injury. METHODS Human EPCs were labeled with iron oxide superparamagnetic nanoparticles. Cell viability and differentiation were tested using flow cytometry. Following finite element modeling computer simulations and flow testing in vitro, angioplasty was performed on rat common carotid arteries to denude the endothelium and EPCs were administered with and without the presence of an external magnetic device for 12 min. RESULTS Computer simulations indicated successful external magnetic cell targeting from a vessel with flow rate similar to a rat common carotid artery; correspondingly there was a 6-fold increase in cell capture in an in vitro flow system. Targeting enhanced cell retention at the site of injury by 5-fold at 24 h after implantation in vivo. CONCLUSIONS Using an externally applied magnetic device, we have been able to enhance EPC localization at a site of common carotid artery injury. This technology could be more widely adapted to localize cells in other organs and may provide a useful tool for the systemic injection of cell therapies.

Clusters of iron-rich cells in the upper beak of pigeons are macrophages not magnetosensitive neurons

Understanding the molecular and cellular mechanisms that mediate magnetosensation in vertebrates is a formidable scientific problem. One hypothesis is that magnetic information is transduced into neuronal impulses by using a magnetite-based magnetoreceptor. Previous studies claim to have identified a magnetic sense system in the pigeon, common to avian species, which consists of magnetite-containing trigeminal afferents located at six specific loci in the rostral subepidermis of the beak. These studies have been widely accepted in the field and heavily relied upon by both behavioural biologists and physicists. Here we show that clusters of iron-rich cells in the rostro-medial upper beak of the pigeon Columbia livia are macrophages, not magnetosensitive neurons. Our systematic characterization of the pigeon upper beak identified iron-rich cells in the stratum laxum of the subepidermis, the basal region of the respiratory epithelium and the apex of feather follicles. Using a three-dimensional blueprint of the pigeon beak created by magnetic resonance imaging and computed tomography, we mapped the location of iron-rich cells, revealing unexpected variation in their distribution and number—an observation that is inconsistent with a role in magnetic sensation. Ultrastructure analysis of these cells, which are not unique to the beak, showed that their subcellular architecture includes ferritin-like granules, siderosomes, haemosiderin and filopodia, characteristics of iron-rich macrophages. Our conclusion that these cells are macrophages and not magnetosensitive neurons is supported by immunohistological studies showing co-localization with the antigen-presenting molecule major histocompatibility complex class II. Our work necessitates a renewed search for the true magnetite-dependent magnetoreceptor in birds.

The chronic vascular and haemodynamic response after permanent bilateral common carotid occlusion in newborn and adult rats

Vascular growth and redistribution of flow can compensate for arterial occlusion and possibly reduce the effects of hypoperfusion. As yet there is limited information on the age-dependent nature of vasculature remodelling. In this study, we have monitored the vascular and morphologic changes using magnetic resonance imaging and histology in a chronic bilateral common carotid artery occlusion (BCCAO) model in both newborn and adult rats. Acutely, cerebral blood flow (CBF) decreased immediately after BCCAO, producing a state of oligaemic hypoperfusion. At 6 months after BCCAO in both adult and neonatal rats, the CBF had normalised at control values. To investigate the underlying mechanism for the return of CBF to control values, intra- and extracerebral magnetic resonance angiograms (MRAs) were acquired. As expected, signal from the common carotid arteries was present in the sham-operated rats, but was absent in the BCCAO animals. India ink angiograms demonstrated more tortuous basilar arteries in the adult rats post-BCCAO and MRAs demonstrated more extracerebral midline collaterals in the neonatal rats post-BCCAO, indicating different modes of vascular adaptation dependent on the age at onset of the insult. Both groups had collateral vessels arising from the vertebral arteries, and BCCAO was also associated with increased diameter of basilar, posterior cerebral, posterior communicating, internal carotid, middle cerebral and anterior cerebral arteries. Our study suggests that the developing and mature animals exhibit different patterns of vascular remodelling and that the BCCAO hypoperfusion model will be useful for investigating age-dependent vascular events in response to vasoocclusive disease.

The measurement of diffusion and perfusion in biological systems using magnetic resonance imaging

The aim of this review is to describe two recent developments in the use of magnetic resonance imaging (MRI) in the study of biological systems: diffusion and perfusion MRI. Diffusion MRI measures the molecular mobility of water in tissue, while perfusion MRI measures the rate at which blood is delivered to tissue. Therefore, both these techniques measure quantities which have direct physiological relevance. It is shown that diffusion in biological systems is a complex phenomenon, influenced directly by tissue microstructure, and that its measurement can provide a large amount of information about the organization of this structure in normal and diseased tissue. Perfusion reflects the delivery of essential nutrients to tissue, and so is directly related to its status. The concepts behind the techniques are explained, and the theoretical models that are used to convert MRI data to quantitative physical parameters are outlined. Examples of current applications of diffusion and perfusion MRI are given. In particular, the use of the techniques to study the pathophysiology of cerebral ischaemia/stroke is described. It is hoped that the biophysical insights provided by this approach will help to define the mechanisms of cell damage and allow evaluation of therapies aimed at reducing this damage.