Employing Gas Challenges and Breathing Manoeuvres to Assess Microvascular Functional Dynamics

Proper blood vessel function entails the transport of blood and modulation of perfusion in response to stimuli. This serves to maintain adequate blood pressure and nutrient delivery. The ability of blood vessels to modulate perfusion in response to demand is a key feature of microvascular integrity (MI). Loss of MI is a characteristic feature of endothelial dysfunction (ED) and is found in several conditions, including hypertension, chronic kidney disease (CKD) and diabetes. To date, most of the research on ED diagnosis has focused on the brain and although there is a clear link between cerebrovascular dysfunction and microvascular pathologies, little work has been done to diagnose extra-cranial ED. Assessment of MI necessitates two steps: a vasoactive stimulus to generate a response (i.e. vasoconstriction/ vasodilatation) and an imaging method to continuously monitor the response. The works presented in this thesis aim to further our understanding of microvascular reactivity with the aim of creating a non-invasive Magnetic Resonance Imaging (MRI) based technique to diagnose extra-cranial MD with the aid of a vasoactive gas challenge stimuli. To achieve this aim, we used a series of hypercapnic and hypoxic gas challenges to induce maximal hemodynamic responses so as navigate the spectrum of vessel calibre. We also employed a long life blood pool agent (Gadofosveset) to allow us to track the changes in vessel calibre. Using this agent, we could navigate the spectrum of vessel calibre in the high perfusion organs such as the kidney and low perfusion skeletal muscle. Furthermore, by employing a specific sequence of vasoactive gases we were able to assess the degree of ED in a hind limb ischemic rat model. Though still its infancy the works presented in this thesis offer us an introduction to the utility of using a gas challenge algorithm in conjunction with MRI to be used for the assessment of vessel health in a number of pathologies.Ph.D

Concurrent Dual Contrast for Cellular Magnetic Resonance Imaging Using Gadolinium Oxide and Iron Oxide Nanoparticles

Rationale and Objectives. Concurrent visualization of differential targets in cellular and molecular imaging is valuable for resolving processes spatially and temporally, as in monitoring different cell subtypes. The purpose of this study was to demonstrate concurrent, dual (positive and negative) contrast visualization on magnetic resonance imaging (MRI) of two colocalized cell populations labeled with Gadolinium “Gd” oxide and iron “Fe” oxide nanoparticles. Materials and Methods. Human aortic endothelial cells (EC) and smooth muscle cells (SMC) were labeled with various concentrations of Gd oxide and Fe oxide, respectively. MRI on single- or mixed-cell samples was performed at 7 tesla. Proper cell phenotype expressions, cell uptake of contrast agents, and the effect of labeling on cell viability and proliferation were also determined. Results. Both contrast agents were efficiently taken up by cells, with viability and proliferation largely unaffected. On MRI, the positive contrast associated with Gd oxide-labeled EC and negative contrast associated with Fe oxide-labeled SMC discriminated the presence of each cell type, whether it existed alone or colocalized in a mixed-cell sample. Conclusion. It is feasible to use Gd oxide and Fe oxide for dual contrast and concurrent discrimination of two colocalized cell populations on MRI at 7 tesla.Peer Reviewe

A novel MRI analysis for assessment of microvascular vasomodulation in low-perfusion skeletal muscle

Compromised microvascular reactivity underlies many conditions and injuries, but its assessment remains difficult, particularly in low perfusion tissues. In this paper, we develop a new mathematical model for the assessment of vasomodulation in low perfusion settings. A first-order model was developed to approximate changes in T1 relaxation times as a result of vasomodulation. Healthy adult rats (N = 6) were imaged on a 3-Tesla clinical MRI scanner, and vasoactive response was probed on gadofosveset using hypercapnic gases at 20% and 5% CO2 to induce vasoconstriction and vasodilation, respectively. MRI included dynamic 3D T1 mapping and T1-weighted images during gas challenge; heart rate was continuously monitored. Laser Doppler perfusion measurements were performed to corroborate MRI findings. The model was able to identify hypercapnia-mediated vasoconstriction and vasodilation through the partial derivative [Formula: see text]. MRI on animals revealed gradual vasoconstriction in the skeletal muscle bed in response to 20% CO2 followed by gradual vasodilation on transitioning to 5% CO2. These trends were confirmed on laser Doppler perfusion measurements. Our new mathematical model has the potential for detecting microvascular dysfunction that manifests in the early stages across multiple metabolic and ischemic pathologies.T.G. is supported by an NSERC doctoral scholarship (PGS D2). H-L.M.C. is funded by the Natural Sciences and Engineering Research Council of Canada (#355795) and the Canada Foundation for Innovation/Ontario Research Fund (#34038)

Concurrent Dual Contrast for Cellular Magnetic Resonance Imaging Using Gadolinium Oxide and Iron Oxide Nanoparticles

Rationale and Objectives. Concurrent visualization of differential targets in cellular and molecular imaging is valuable for resolving processes spatially and temporally, as in monitoring different cell subtypes. The purpose of this study was to demonstrate concurrent, dual (positive and negative) contrast visualization on magnetic resonance imaging (MRI) of two colocalized cell populations labeled with Gadolinium "Gd" oxide and iron "Fe" oxide nanoparticles. Materials and Methods. Human aortic endothelial cells (EC) and smooth muscle cells (SMC) were labeled with various concentrations of Gd oxide and Fe oxide, respectively. MRI on single-or mixed-cell samples was performed at 7 tesla. Proper cell phenotype expressions, cell uptake of contrast agents, and the effect of labeling on cell viability and proliferation were also determined. Results. Both contrast agents were efficiently taken up by cells, with viability and proliferation largely unaffected. On MRI, the positive contrast associated with Gd oxide-labeled EC and negative contrast associated with Fe oxide-labeled SMC discriminated the presence of each cell type, whether it existed alone or colocalized in a mixed-cell sample. Conclusion. It is feasible to use Gd oxide and Fe oxide for dual contrast and concurrent discrimination of two colocalized cell populations on MRI at 7 tesla

T2* and T1 assessment of abdominal tissue response to graded hypoxia and hypercapnia using a controlled gas mixing circuit for small animals

Purpose To characterize urn:x-wiley:10531807:media:jmri25169:jmri25169-math-0003 and T1 relaxation time response to a wide spectrum of gas challenges in extracranial tissues of healthy rats. Materials and Methods A range of graded gas mixtures (hyperoxia, hypercapnia, hypoxia, and hypercapnic hypoxia) were delivered through a controlled gas‐mixing circuit to mechanically ventilated and intubated rats. Quantitative magnetic resonance imaging (MRI) was performed on a 3T clinical scanner; urn:x-wiley:10531807:media:jmri25169:jmri25169-math-0004 and T1 maps were computed to determine tissue response in the liver, kidney cortex, and paraspinal muscles. Heart rate and blood oxygen saturation (SaO2) were measured through a rodent oximeter and physiological monitor. Results urn:x-wiley:10531807:media:jmri25169:jmri25169-math-0005 decreases consistent with lowered SaO2 measurements were observed for hypercapnia and hypoxia, but decreases were significant only in liver and kidney cortex (P 10% CO2 and 90% O2 and >5% CO2. Conclusion urn:x-wiley:10531807:media:jmri25169:jmri25169-math-0008 and T1 provide complementary roles for evaluating extracranial tissue response to a broad range of gas challenges. Based on both measured and known physiological responses, our results are consistent with urn:x-wiley:10531807:media:jmri25169:jmri25169-math-0009 as a sensitive marker of blood oxygen saturation and T1 as a weak marker of blood volume changes. J. Magn. Reson. Imaging 2016;44:305–316.Tameshwar Ganesh is the recipient of the Queen Elizabeth II Graduate Scholarship in Science & Technology (QEII‐GSST)

A non-invasive magnetic resonance imaging approach for assessment of real-time microcirculation dynamics

We present a novel, non-invasive magnetic resonance imaging (MRI) technique to assess real-time dynamic vasomodulation of the microvascular bed. Unlike existing perfusion imaging techniques, our method is sensitive only to blood volume and not flow velocity. Using graded gas challenges and a long-life, blood-pool T 1-reducing agent gadofosveset, we can sensitively assess microvascular volume response in the liver, kidney cortex, and paraspinal muscle to vasoactive stimuli (i.e. hypercapnia, hypoxia, and hypercapnic hypoxia). Healthy adult rats were imaged on a 3 Tesla scanner and cycled through 10-minute gas intervals to elicit vasoconstriction followed by vasodilatation. Quantitative T 1 relaxation time mapping was performed dynamically; heart rate and blood oxygen saturation were continuously monitored. Laser Doppler perfusion measurements confirmed MRI findings: dynamic changes in T 1 corresponded with perfusion changes to graded gas challenges. Our new technique uncovered differential microvascular response to gas stimuli in different organs: for example, mild hypercapnia vasodilates the kidney cortex but constricts muscle vasculature. Finally, we present a gas challenge protocol that produces a consistent vasoactive response and can be used to assess vasomodulatory capacity. Our imaging approach to monitor real-time vasomodulation may be extended to other imaging modalities and is valuable for investigating diseases where microvascular health is compromised.T.G. is supported by an NSERC doctoral scholarship (PGS D2). H.-L.M.C. is funded by the Heart and Stroke Foundation of Canada (#000223), the Natural Sciences and Engineering Research Council of Canada (#355795), and the Canada Foundation for Innovation/Ontario Research Fund (#34038)

Manganese-Enhanced Magnetic Resonance Imaging for Early Detection and Characterization of Breast Cancers

Very early cancer detection is the key to improving cure. Our objective was to investigate manganese (Mn)-enhanced magnetic resonance imaging (MRI) for very early detection and characterization of breast cancers. Eighteen NOD scid gamma mice were inoculated with MCF7, MDA, and LM2 breast cancer cells and imaged periodically on a 3 T scanner beginning on day 6. T1-weighted imaging and T1 measurements were performed before and 24 hours after administering MnCl2. At the last imaging session, Gd-DTPA was administered and tumors were excised for histology (hematoxylin-eosin and CD34 staining). All mice, except for two inoculated with MCF7 cells, developed tumors. Tumors enhanced uniformly on Mn and showed clear borders. Early small tumors (# 5 mm3) demonstrated the greatest enhancement with a relative R1 (1/T1) change of 1.57 6 0.13. R1 increases correlated with tumor size (r 5 2.34, p 5 .04). Differences in R1 increases among the three tumor subtypes were most evident in early tumors. Histology confirmed uniform cancer cell distribution within tumor masses and vasculature in the periphery, which was consistent with rim-like enhancement on Gd-DTPA. Mn-enhanced MRI is a promising approach for detecting very small breast cancers in vivo and may be valuable for very early cancer detection.This study was supported by the Natural Sciences and Engineering Research Council of Canada (#355795) and the Garron Family Cancer Centre Grant through the SickKids Foundation

A manganese porphyrin-based T1 contrast agent for cellular MR imaging of human embryonic stem cells

MRI for non-invasive cell tracking is recognized for enabling pre-clinical research on stem cell therapy. Yet, adoption of cellular imaging in stem cell research has been restricted to sites with experience in MR contrast agent synthesis and to small animal models that do not require scaled-up synthesis. In this study, we demonstrate the use of a gadolinium-free T1 contrast agent for tracking human embryonic stem cells. The agent, MnPNH2, is an easily synthesized manganese porphyrin that can be scaled for large cell numbers. MRI was performed on a 3 T clinical scanner. Cell pellets labeled at different MnPNH2 concentrations for 24 hours demonstrated a decrease in T1 relaxation time of nearly two-fold (P < 0.05), and cellular contrast was maintained for 24 hours (P < 0.05). Cell viability (Trypan blue) and differentiation (embryoid body formation) were unaffected. Cell uptake of Mn on inductively coupled plasma atomic emission spectroscopy corroborated MRI findings, and fluorescence microscopy revealed the agent localized mainly in cell-cell boundaries and cell nuclei. Labeled cells transplanted in rats demonstrated the superior sensitivity of MnPNH2 for in-vivo cell tracking.We thank Dr. Michael Laflamme and Ms. Tamilla Valdman Sadikov for guidance on maintaining human embryonic stem cells. A.V. is supported by an NSERC scholarship (CGS-M) and Ontario Graduate Scholarships (OGS). H.-L.M.C. is funded by the Natural Sciences and Engineering Research Council of Canada (#355795), the Heart and Stroke Foundation of Canada (#000223), funds from the Ted Rogers Centre for Heart Research, the Canada First Research Excellence Fund/Medicine by Design Cycle 1 Team Project Award, and the Canada Foundation for Innovation/Ontario Research Fund (#34038)

Ultrashort echo time magnetic resonance imaging of the lung using a high-relaxivity t1 blood-pool contrast agent

The lung remains one of the most challenging organs to image using magnetic resonance imaging (MRI) due to intrinsic rapid signal decay. However, unlike conventional modalities such as computed tomography, MRI does not involve radiation and can provide functional and morphologic information on a regional basis. Here we demonstrate proof of concept for a new MRI approach to achieve substantial gains in a signal to noise ratio (SNR) in the lung parenchyma: contrast-enhanced ultrashort echo time (UTE) imaging following intravenous injection of a high-relaxivity blood-pool manganese porphyrin T1 contrast agent. The new contrast agent increased relative enhancement of the lung parenchyma by over 10-fold compared to gadolinium diethylene triamine pentaacetic acid (Gd-DTPA), and the use of UTE boosted the SNR by a factor of 4 over conventional T1-weighted gradient echo acquisitions. The new agent also maintains steady enhancement over at least 60 minutes, thus providing a long time window for obtaining high-resolution, high-quality images and the ability to measure a number of physiologic parameters.This study was supported by the Heart and Stroke Foundation (#000223 awarded to H.- L.M.C.) and the Natural Sciences and Engineering Research Council of Canada (#355795 awarded to H.-L.M.C. and #489075 awarded to X.-a.Z.). A patent application has been filed for the contrast agent MnP2. Zhang XA, Cheng WR, Haedicke I, Cheng HL, Gd-free MRI contrast agents (US patent appl. #2013101159, Canadian patent appl. #2013101162)

Manganese-Enhanced Magnetic Resonance Imaging for Early Detection and Characterization of Breast Cancers

Very early cancer detection is the key to improving cure. Our objective was to investigate manganese (Mn)-enhanced magnetic resonance imaging (MRI) for very early detection and characterization of breast cancers. Eighteen NOD scid gamma mice were inoculated with MCF7, MDA, and LM2 breast cancer cells and imaged periodically on a 3 T scanner beginning on day 6. T 1 -weighted imaging and T 1 measurements were performed before and 24 hours after administering MnCl 2 . At the last imaging session, Gd-DTPA was administered and tumors were excised for histology (hematoxylin-eosin and CD34 staining). All mice, except for two inoculated with MCF7 cells, developed tumors. Tumors enhanced uniformly on Mn and showed clear borders. Early small tumors (# 5 mm 3 ) demonstrated the greatest enhancement with a relative R 1 (1/T 1 ) change of 1.57 ± 0.13. R 1 increases correlated with tumor size ( r = −.34, p − .04). Differences in R 1 increases among the three tumor subtypes were most evident in early tumors. Histology confirmed uniform cancer cell distribution within tumor masses and vasculature in the periphery, which was consistent with rim-like enhancement on Gd-DTPA. Mn-enhanced MRI is a promising approach for detecting very small breast cancers in vivo and may be valuable for very early cancer detection