Evaluation of oxygen sensitivity of hyperpolarized helium imaging for the detection of pulmonary ischemia.

PURPOSE: In this study, a new model of pulmonary embolism in rats was developed and tested, to examine if hyperpolarized (HP) (3) He MR images can measure impairment of the exchange of oxygen from the airspaces to the blood during pulmonary embolism. METHODS: HP (3) He MRI was used to image six treatment-group rats in which a branch of the pulmonary artery was embolized, and six control-group rats. HP (3) He MR images were used to calculate the initial partial pressure of oxygen (pO ) and the rate of oxygen depletion (R) in rat lungs. RESULTS: The pO was significantly higher in the ischemic lung than in the contralateral normal side, and pO was significantly higher in the ischemic lung than in both sides of the control lungs. Mean R in ischemic lungs was significantly lower than in the contralateral lungs, and mean R in ischemic lungs was also significantly lower than in both control lungs. CONCLUSION: These results demonstrate that pO and R, as measured by the T1 decay of HP (3) He, are sensitive to pulmonary ischemia in rats, confirming the findings in studies performed in large animal models of pulmonary ischemia.

Visualization of clot lysis in a rat embolic stroke model: application to comparative lytic efficacy.

BACKGROUND AND PURPOSE: The purpose of this study was to develop a novel MRI method for imaging clot lysis in a rat embolic stroke model and to compare tissue plasminogen activator (tPA)-based clot lysis with and without recombinant Annexin-2 (rA2). METHODS: In experiment 1 we used in vitro optimization of clot visualization using multiple MRI contrast agents in concentrations ranging from 5 to 50 µL in 250 µL blood. In experiment 2, we used in vivo characterization of the time course of clot lysis using the clot developed in the previous experiment. Diffusion, perfusion, angiography, and T1-weighted MRI for clot imaging were conducted before and during treatment with vehicle (n=6), tPA (n=8), or rA2 plus tPA (n=8) at multiple time points. Brains were removed for ex vivo clot localization. RESULTS: Clots created with 25 µL Magnevist were the most stable and provided the highest contrast-to-noise ratio. In the vehicle group, clot length as assessed by T1-weighted imaging correlated with histology (r=0.93). Clot length and cerebral blood flow-derived ischemic lesion volume were significantly smaller than vehicle at 15 minutes after treatment initiation in the rA2 plus tPA group, whereas in the tPA group no significant reduction from vehicle was observed until 30 minutes after treatment initiation. The rA2 plus tPA group had a significantly shorter clot length than the tPA group at 60 and 90 minutes after treatment initiation and significantly smaller cerebral blood flow deficit than the tPA group at 90 minutes after treatment initiation. CONCLUSIONS: We introduce a novel MRI-based clot imaging method for in vivo monitoring of clot lysis. Lytic efficacy of tPA was enhanced by rA2.

Using hyperpolarized 3He MRI to evaluate treatment efficacy in cystic fibrosis patients.

PURPOSE: To use hyperpolarized (HP) (3)He MR imaging to assess functional lung ventilation in subjects with cystic fibrosis (CF) before and after treatment. MATERIALS AND METHODS: We performed HP (3)He static ventilation MRI scans on three subjects, using a Philips 3.0 Tesla (T) Achieva MRI scanner, before and after 11 days of in-patient treatment with combined intravenous and inhaled therapies for pulmonary exacerbations of CF. We also collected spirometry data. We quantified pulmonary ventilation volume measured with HP (3)He MRI using an advanced semi-automated analysis technique. RESULTS: Following 11 days of treatment with intravenous antibiotics, hypertonic saline, and rhDNase, HP (3)He MR images in one subject displayed a 25% increase in total ventilation volume. Total ventilation volume in the other two subjects slightly decreased. All three subjects showed increases in FEV(1) and FVC following treatment. CONCLUSION: In all subjects, the HP (3)He MR images provided detailed information on precisely where in the lungs gas was reaching. These data provide additional support for the conclusion that HP noble gas MRI can be a powerful tool for evaluating lung ventilation in patients with cystic fibrosis, but also raise important questions about the correlation between spirometry and HP gas MRI measurements.

In vitro and In vivo imaging of antivasculogenesis induced by Noggin protein expression in human venous endothelial cells.

Noggin protein is a potent bone morphogenetic protein (BMP) antagonist capable of inhibiting vasculogenesis even in the presence of provasculogenic VEGF and FGF-2. We found that human umbilical vein endothelial cells (HUVECs) do not express Noggin in culture and used these cells for modeling of antivasculogenesis. We hypothesized that high-efficiency transduction of HUVECs with bicistronic lentiviral vector encoding Noggin and enhanced green fluorescent protein (EGFP) enables direct visualization of Noggin effects in homogenous primary cell populations in vitro and in vivo. By comparing HUVECs transduced with a control GFP and GFP/Noggin expression cassettes, we showed that constitutive and orthotopic Noggin protein expression did not influence cell proliferation, down-regulated BMP-2 expression, and showed no effect on BMP receptor transcripts. We demonstrated that in contrast to GFP-only control, Noggin expression in endothelial cells abrogated endothelial migration in response to monolayer injury, blocked endothelial transmigration, and caused abrogation of cord formation in vitro. Adding exogenous BMP-4 restored the formation of cords. Imaging experiments in vivo investigated vessel formation in Matrigel implants in athymic mice by utilizing GFP imaging or magnetic resonance imaging of perfusion in the implants. Both approaches demonstrated the lack of functional vessel formation after the adoptive transfer of GFP/Noggin-expressing human endothelial cells in mice.

Estimation and correction of cardiac respiratory motion in SPECT in the presence of limited-angle effects due to irregular respiration.

PURPOSE: One issue with amplitude binning list-mode studies in SPECT for respiratory motion correction is that variation in the patient's respiratory pattern will result in binned motion states with little or no counts at various projection angles. The reduced counts result in limited-angle reconstruction artifacts which can impact the accuracy of the necessary motion estimation needed to correct the images. In this work, the authors investigate a method to overcome the effect of limited-angle reconstruction artifacts in SPECT when estimating respiratory motion. METHODS: In the first pass of the reconstruction method, only the projection angles with significant counts in common between the binned respiratory states are used in order to better estimate the motion between them. After motion estimation, the estimates are used to correct for motion within iterative reconstruction using all of the acquired projection data. RESULTS: Using simulated SPECT studies based on the NCAT phantom, the authors demonstrate the problem caused by having data available for only a limited number of angles when estimating motion and the utility of the proposed method in diminishing this error. For NCAT data sets with a clinically appropriate level of Poisson noise, the average registration error for motion with the proposed method was always less with the use of their algorithm, the reduction being statistically significant (p<0.05) in the majority of cases. The authors illustrate the ability of their method to correct the degradations caused by respiratory motion in short-axis slices and polar maps of the NCAT phantom for cases with 1 and 2 cm amplitudes of respiratory motion. In four cardiac-perfusion patients acquired on the same day, the authors demonstrate the large variability of the number of counts in the amplitude-binned projections. Finally, the authors demonstrate a visual improvement in the slices and polar maps of patient studies with the algorithm for respiratory motion correction. CONCLUSIONS: The authors' method shows promise in reducing errors in respiratory motion estimation despite the presence of limited-angle reconstruction effects due to irregularity in respiration. Improvements in image quality were observed in both simulated and clinical studies.

A method for estimating intracellular sodium concentration and extracellular volume fraction in brain in vivo using sodium magnetic resonance imaging.

In this feasibility study we propose a method based on sodium magnetic resonance imaging (MRI) for estimating simultaneously the intracellular sodium concentration (C1, in mM) and the extracellular volume fraction (α) in grey and white matters (GM, WM) in brain in vivo. Mean C1 over five healthy volunteers was measured ~11 mM in both GM and WM, mean α was measured ~0.22 in GM and ~0.18 in WM, which are in close agreement with standard values for healthy brain tissue (C1 ~ 10-15 mM, α ~ 0.2). Simulation of 'fluid' and 'solid' inclusions were accurately detected on both the C1 and α 3D maps and in the C1 and α distributions over whole GM and WM. This non-invasive and quantitative method could provide new biochemical information for assessing ion homeostasis and cell integrity in brain and help the diagnosis of early signs of neuropathologies such as multiple sclerosis, Alzheimer's disease, brain tumors or stroke.

Assessment of repeatability of hyperpolarized gas MR ventilation functional imaging in cystic fibrosis.

RATIONALE AND OBJECTIVES: Hyperpolarized (HP) gas magnetic resonance imaging (MRI) is an advanced imaging technique that provides high-resolution regional information on lung function without using ionizing radiation. Before this modality can be considered for assessing clinical or investigational interventions, baseline repeatability needs to be established. We assessed repeatability of lung function measurement using HP helium-3 MRI (HP (3)He MRI) in a small cohort of patients with cystic fibrosis (CF). MATERIALS AND METHODS: We examined repeatability of HP (3)He MR images of five patients with CF in four scanning sessions over a 4-week period. We acquired images on a Philips 3.0 Tesla Achieva MRI scanner using a quadrature, flexible, wrap-around, (3)He radiofrequency coil with a fast gradient-echo pulse sequence. We determined ventilation volume and ventilation defect volume using an advanced semiautomatic segmentation algorithm and also quantified ventilation heterogeneity. RESULTS: There were no significant differences in total ventilation volume, ventilation defect volume, ventilation defect percentage, or mean ventilation heterogeneity (repeated-measures analysis of variance, P = .2116, P = .2825, P = .2871, and P = .7265, respectively) in the patients across the four scanning sessions. CONCLUSIONS: Our results indicate that total ventilation volume, ventilation defect volume, ventilation defect percentage, and mean ventilation heterogeneity as assessed by HP gas MRI in CF patients with stable health are reproducible over time. This repeatability and the technique's capability to provide noninvasive high-resolution data on regional lung function without ionizing radiation make (3)He MRI a potentially useful outcome measure for CF-related clinical trials.

Distribution of hyperpolarized xenon in the brain following sensory stimulation: preliminary MRI findings.

In hyperpolarized xenon magnetic resonance imaging (HP (129)Xe MRI), the inhaled spin-1/2 isotope of xenon gas is used to generate the MR signal. Because hyperpolarized xenon is an MR signal source with properties very different from those generated from water-protons, HP (129)Xe MRI may yield structural and functional information not detectable by conventional proton-based MRI methods. Here we demonstrate the differential distribution of HP (129)Xe in the cerebral cortex of the rat following a pain stimulus evoked in the animal's forepaw. Areas of higher HP (129)Xe signal corresponded to those areas previously demonstrated by conventional functional MRI (fMRI) methods as being activated by a forepaw pain stimulus. The percent increase in HP (129)Xe signal over baseline was 13-28%, and was detectable with a single set of pre and post stimulus images. Recent innovations in the production of highly polarized (129)Xe should make feasible the emergence of HP (129)Xe MRI as a viable adjunct method to conventional MRI for the study of brain function and disease.

Characterization of gadolinium-based dynamic susceptibility contrast perfusion measurements in permanent and transient MCAO models with volumetric based validation by CASL.

Perfusion imaging is crucial in imaging of ischemic stroke to determine 'tissue at risk' for infarction. In this study we compared the volumetric quantification of the perfusion deficit in two rat middle-cerebral-artery occlusion (MCAO) models using two gadolinium-based contrast agents (P1152 (Guerbet) and Magnevist (Bayer-Schering, Pittsburgh, PA, USA)) as compared with our well established continuous arterial spin labeling (CASL) perfusion imaging technique. Animals underwent either permanent MCAO or transient MCAO with 80-min reperfusion. Imaging was performed at four different time points after MCAO. A region-of-interest (ROI) analysis of the subregions of the ischemic zone (core, penumbra, transient reversal (TR), and sustained reversal (SR)) using P1152 showed significant reduction in blood flow in the core and TR subregions relative to the penumbral and SR subregions while occluded. After reperfusion, a significant increase in blood flow was recorded at all time points after reperfusion in all regions except TR. From the ROI analysis the threshold for the penumbra was determined to be -62+/-11% and this value was subsequently used for quantification of the volumetric deficit. The ischemic volume as defined by dynamic susceptibility contrast (DSC), was only statistically different from the CASL-derived ischemic volume when using Magnevist at post-reperfusion time points.

Marked pericardial inhomogeneity of specific ventilation at total lung capacity and beyond.

We measured regional ventilation at 1l above functional residual capacity (FRC+1L) and total lung capacity (TLC) in three normal subjects and four elite breath-hold divers, and above TLC after glossopharyngeal insufflation (TLC+GI) in the divers. Hyperpolarized (3)He MRI was used to map the local ventilation per unit volume over the entire lung. At TLC and above, there was markedly increased regional ventilation of the lungs in the pericardial region compared with the relatively uniform ventilation throughout the rest of the lung. The distribution of fractional ventilation regionally was relatively uniform at FRC+1L, with a small non-gravitational cephalocaudal gradient of specific ventilation in the supine posture. Our observations at high lung volumes are consistent with the effect of high pleural tension in the concave pericardial region, which promotes expansion of the subjacent lung, leading to a higher local effective compliance and a higher specific ventilation.

Membrane order and molecular dynamics associated with IgE receptor cross-linking in mast cells.

Cholesterol-rich microdomains (or "lipid rafts") within the plasma membrane have been hypothesized to exist in a liquid-ordered phase and play functionally important roles in cell signaling; however, these microdomains defy detection using conventional imaging. To visualize domains and relate their nanostructure and dynamics to mast cell signaling, we use two-photon (760 nm and 960 nm) fluorescence lifetime imaging microscopy and fluorescence polarization anisotropy imaging, with comparative one-photon anisotropy imaging and single-point lifetime and anisotropy decay measurements. The inherent sensitivity of ultrafast excited-state dynamics and rotational diffusion to the immediate surroundings of a fluorophore allows for real-time monitoring of membrane structure and organization. When the high affinity receptor for IgE (FcepsilonRI) is extensively cross-linked with anti-IgE, molecules associated with cholesterol-rich microdomains (e.g., saturated lipids (the lipid analog diI-C(18) or glycosphingolipids)) and lipid-anchored proteins coredistribute with cross-linked IgE-FcepsilonRI. We find an enhancement in fluorescence lifetime and anisotropy of diI-C(18) and Alexa 488-labeled IgE-FcepsilonRI in the domains where these molecules colocalize. Our results suggest that fluorescence lifetime and, particularly, anisotropy permit us to correlate the recruitment of lipid molecules into more ordered domains that serve as platforms for IgE-mediated signaling.

Dynamics imaging of lipid phases and lipid-marker interactions in model biomembranes.

Biomembranes are complex systems that regulate numerous biological processes. Lipid phases that constitute these membranes influence their properties and transport characteristics. Here, we demonstrate the potential of short-range dynamics imaging (excited-state lifetime, rotational diffusion, and order parameter) as a sensitive probe of lipid phases in giant unilamellar vesicles (GUVs). Liquid-disordered and gel phases were labeled with Bodipy-PC at room temperature. Two-photon fluorescence lifetime imaging microscopy of single-phase GUVs reveals more heterogeneity in fluorescence lifetimes of Bodipy in the gel phase (DPPC: 3.8+/-0.6 ns) as compared with the fluid phase (DOPC: 5.2+/-0.2 ns). The phase-specificity of excited-state lifetime of Bodipy-PC is attributed to the stacking of ordered lipid molecules that possibly enhances homo-FRET. Fluorescence polarization anisotropy imaging also reveals distinctive molecular order that is phase specific. The results are compared with DiI-C12-labeled fluid GUVs to investigate the sensitivity of our fluorescence dynamics assay to different lipid-marker interactions. Our results provide a molecular perspective of lipid phase dynamics and the nature of their microenvironments that will ultimately help our understanding of the structure-function relationship of biomembranes in vivo. Furthermore, these ultrafast excited-state dynamics will be used for molecular dynamics simulation of lipid-lipid, lipid-marker and lipid-protein interactions.