EMIM 2019
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New Methods in Neuroimaging

Session chair: Jan Klohs (Zurich, Switzerland); Bastian Zinnhardt (Münster, Germany)
 
Shortcut: PS 06
Date: Wednesday, 20 March, 2019, 2:15 p.m.
Room: BOISDALE | level 0
Session type: Parallel Session

Contents

Click on an contribution to preview the abstract content.

2:15 p.m. PS 06-01

Introductory Lecture

Fabien Chauveau

France

This talk provides an overview of state-of-the-art research and refers to the following presentations selected from abstract submissions.

2:33 p.m. PS 06-02

Magnetic resonance spectroscopic imaging of hyperpolarized [1-13C]pyruvate in glioblastoma: a promising tool for investigating tumor metabolism and heterogeneity. (#423)

Fulvio Zaccagna1, James T. Grist1, Mary A. McLean2, Charlotte J. Daniels1, Frank Riemer1, Joshua Kaggie1, Surrin Deen1, Ramona Woitek1, Rolf F. Schulte3, Kieren Allinson4, Anita Chhabra5, Marie-Christine Laurent1, Amy Frary1, Tomasz Matys1, Ilse Patterson6, Bruno Do Carmo6, Stephan Urpsrung1, Ian Wilkinson7, Bristi Basu8, Colin Watts9, Stephen J. Price9, Sarah Jefferies8, Jonathan H. Gillard1, Kevin M. Brindle2, Ferdia A. Gallagher1

1 University of Cambridge, Department of Radiology, Cambridge, United Kingdom
2 University of Cambridge, Cancer Research UK Cambridge Institute, Cambridge, United Kingdom
3 GE Global Research, Munich, Germany
4 Cambridge University Hospitals NHS Foundation Trust, Department of Pathology, Cambridge, United Kingdom
5 Cambridge University Hospitals NHS Foundation Trust, Cambridge Cancer Centre, Cambridge, United Kingdom
6 Cambridge University Hospitals NHS Foundation Trust, Department of Radiology, Cambridge, United Kingdom
7 University of Cambridge, Department of Medicine, Cambridge, United Kingdom
8 Cambridge University Hospitals NHS Foundation Trust, Department of Oncology, Cambridge, United Kingdom
9 University of Cambridge, Department of Clinical Neurosciences, Cambridge, United Kingdom

Introduction

Metabolic reprogramming is one of the major driving forces in determining Glioblastoma (GBM) heterogeneity, therefore the in vivo characterization of metabolic heterogeneity could help development of novel therapeutic strategies targeting cancer-specific metabolic pathways.1 13C magnetic resonance imaging using dynamic nuclear polarization (DNP) allows for non-invasive assessment of the metabolism of hyperpolarized (HP) 13C-labelled molecules in vivo. The purpose of this study was to explore metabolic reprogramming in GBM and the surrounding microenvironment using HP [1-13C]pyruvate.

Methods

Three treatment-naïve patients (1 male, 58±13 yrs) were imaged on a 3T scanner (MR750 GE Healthcare, WI) using a dual-tuned 13C/1H quadrature transmit/receive head coil (Rapid Biomedical, Rimpar, Germany). [1-13C]pyruvate was hyperpolarized using a clinical hyperpolarizer (Research Circle Technology, Albany NY). 13C MRSI acquisition was performed using a dynamic IDEAL spiral sequence2. Apparent kinetic rate constant maps for the conversion of pyruvate to lactate (kPL) and pyruvate to bicarbonate (kPB) were derived using a two-site model.3 Regions of interest (ROIs) were obtained for tumour sub regions and normal appearing brain parenchyma (NABP) on the unenhanced T1W FSPGR using the post gadolinium sequence as guidance. Non-parametric and unpaired t-test were used as appropriate (p<0.05).

Results/Discussion

Dynamic signal from hyperpolarized pyruvate, lactate, and bicarbonate was observed in the brain following intravenous injection of [1-13C]pyruvate (representative case in Figure 1). Kinetic analysis (Table 1) demonstrated a significantly lower kPL in the whole tumor compared to the contralateral NABP (p = 0.03) but not compared to the ipsilateral NABP (p = 0.05). The kPB in glioblastoma was significantly lower compared to both the contralateral NABP (p < 0.0001) and the ipsilateral NABP (p < 0.0001). In the peritumoral region, kPL was not significantly different from the contralateral NABP (p = 0.22), however the peritumoral kPB was lower than in the contralateral NABP (p = 0.017). Regional analysis demonstrated intralesional heterogeneity showing a trend of higher kPL in the lateral aspect of the region. /however, only in one patient was the difference statistically significant (p = 0.03).

No adverse events were observed in patients following injection of [1-13C]pyruvate.

Conclusions

This study demonstrates the conversion of hyperpolarized pyruvate to lactate and bicarbonate in the human brain. GBMs demonstrate marked intratumoral and intertumoral heterogeneity. kPL was lower in GBM compared to the NABP. The unidirectional formation of bicarbonate was also consistently reduced within GBM compared to NABP. Moreover, intralesional heterogeneity in the labelling exchange highlighted the presence of different metabolic habitats.

References

1. Corbin Z, Spielman D, Recht L. A Metabolic Therapy for Malignant Glioma Requires a Clinical Measure. Curr. Oncol. Rep. 2017;19(12).

2. Wiesinger F, Weidl E, Menzel MI, et al. IDEAL spiral CSI for dynamic metabolic MR imaging of hyperpolarized [1-13C]pyruvate. Magn. Reson. Med. 2012;68(1):8–16.

3. Khegai O, Schulte RF, Janich MA, et al. Apparent rate constant mapping using hyperpolarized [1-13C]pyruvate. NMR Biomed. 2014;27(10):1256–1265.

Acknowledgement

This work has been funded by a Wellcome Trust Strategic Award, Cancer Research UK (CRUK; C19212/A16628) and the CRUK & Engineering and Physical Sciences Research Council (EPSRC) Cancer Imaging Centre in Cambridge and Manchester (C197/A16465). Additional support has been provided by the CRUK Cambridge Centre, the National Institute of Health Research (NIHR) Cambridge Biomedical Research Centre and Addenbrooke’s Charitable Trust.

The views expressed are those of the author(s) and not necessarily those of the NHS, the NIHR or the Department of Health and Social Care.

Figure 1

73-year old female with a left frontal IDH-wildtype GBM: T1-weighted image (a), summed pyruvate (b) and lactate (c) over the time course, post-Gd T1W image (d), kPL (e) and kPB (f). Pyruvate, lactate, kPL and kPB maps are overlaid on the T1WI. The area corresponding to the GBM is highlighted in green on b, c, e and f.

Table 1
Apparent kinetic rate constant maps for the conversion of pyruvate to lactate (kPL) and pyruvate to bicarbonate (kPB).
Keywords: hyperpolarized MRI, hyperpolarized [1-13C]pyruvate, [1-13C]pyruvate, glioblastoma, cancer metabolism
2:45 p.m. PS 06-03

Monitoring of calcium changes with bioresponsive MRI probes during ischemic stroke (#280)

Tanja Savic1, Giuseppe Gambino1, Vahid Bokharaie2, Hamid R. Noori2, Nikos K. Logothetis3, 4, Goran Angelovski1

1 Max Planck Institute for Biological Cybernetics, MR Neuroimaging Agents, Tuebingen, Germany
2 Max Planck Institute for Biological Cybernetics, Neuronal Convergence, Tuebingen, Germany
3 Max Planck Institute for Biological Cybernetics, Physiology of Cognitive Processes, Tuebingen, Germany
4 Centre for Imaging Sciences, University of Manchester, Manchester, United Kingdom

Introduction

Real-time monitoring of biological processes under physiological and pathological conditions is still great challenge for magnetic resonance imaging (MRI), despite its extensive clinical applications. Calcium is involved in an immense number of signaling events in the brain, hence it is an ideal target for functional MRI purposes. For instance, its extracellular concentration substantially varies during the ischemic stroke [1, 2]. Thus, possibility to track Ca2+ noninvasively would deepen the understanding of numerous physiological processes and allow direct monitoring of neuronal activity.

Methods

To monitor Ca2+ changes in vivo, an ischemic stroke model (transient remote MCAo-middle cerebral artery occlusion) was used. Gd2L1 (responsive probe) and Gd2L2 (control probe) were continuously infused intracranially in Wistar rats using osmotic pump, and the MCAo was induced remotely. Functional MRI measurements were divided in three segments: pre-ischemia, ischemia, and reperfusion and consisted of T1w image acquisition every 2 min. Existence of ischemia was confirmed with standard diffusion weighted imaging. Control MRI experiments included intracranial injection of Gd2L1 and Gd2L2, without causing MCAo. Data analysis was based on K-means clustering applied to the raw signals, while the data detrending was done using 4th order spline fitted to the pre-ischemia and reperfusion segments.

Results/Discussion

T1w images and corresponding cluster maps show that cluster 1 clearly corresponds to the center of injection (Figure 1). Furthermore, T1w MRI signal of Gd2L1 varied noticeably in response to the MCAo, declining through the ischemia segment. Albeit, MRI signal enhanced by Gd2L2 is merely a consequence of the infusion of the contrast agent, and did not show any alterations due to MCAo induction or tissue reperfusion. This observation can be explained by the decrease of [Ca2+] during the ischemia and accordingly longitudinal relaxivity (r1) reduction of Gd2L1. Upon reperfusion and restoration of [Ca2+], r1 recovers and so do the initial MRI signal slope. Detrended signals, freed from the transient behavior, show even greater differentiation between MCAo experiments with Gd2L1 on one hand and the ones with Gd2L2 and controls on the other hand (Figure 2), confirming that changes in MRI signal occur only during MCAo induction with Gd2L1.

Conclusions

Here we report the successful use of calcium-responsive MRI probes in vivo for early detection and monitoring of the ischemic stroke. The potential of this molecular fMRI technique is tremendous. It could allow the visualization and mapping of neural signaling using Ca2+ as its direct indicator, supplementing the use of conventional fMRI based on BOLD signal and enabling assessment of neuronal activity in direct fashion.

References

1.         Li, P.-A., et al., The influence of insulin-induced hypoglycemia on the calcium transients accompanying reversible forebrain ischemia in the rat. Experimental Brain Research, 1990. 105(3): p. 363-369.

2.         Siemkowicz, E. and A.J. Hansen, Brain extracellular ion composition and EEG activity following 10 minutes ischemia in normo- and hyperglycemic rats. Stroke, 1981. 12(2): p. 236.

Acknowledgement

The financial support of the German Exchange Academic Service (DAAD, PhD fellowship to T.S.) is gratefully acknowledged.

T1w MRI images and corresponding cluster maps

T1w MRI (7T) images and corresponding maps obtained with K-means clustering; (upper left) corresponding cluster centroids upon ischemia with continuous injection of Gd2L1 or Gd2L2.

Detrended signals
a) Gd2L1 during ischemia and control experiment b) Gd2L2 during ischemia and control experiment c) Mean values of detrended signals for Gd2L1 and Gd2L2 recordings in the three segments: pre-ischemic, ischemic, and post-ischemic period (n=5)
Keywords: calcium, fMRI, ischemia, MRI contrast agents
2:57 p.m. PS 06-04

CEST MRI of Implanted Hydrogel Cell Scaffolds (#195)

Wei Zhu1, Chengyan Chu1, Shreyas Kuddannaya1, Yue Yuan1, Piotr Walczak1, Anirudha Singh2, Xiaolei Song1, Jeff Bulte1

1 Johns Hopkins University School of Medicine, Dept. of Radiology, Baltimore, Maryland, United States of America
2 Johns Hopkins University School of Medicine, Dept. of Urology, BALTIMORE, Maryland, United States of America

Introduction

For cell therapy, scaffolding cells with hydrogels is a promising strategy to overcome initial cell loss and manipulate cell function post-transplantation. Matrix degradation directly influences cell release, with major effects on tissue regeneration or repair.1 Monitoring such degradation is essential for developing scaffolded cell-based therapies. We applied chemical exchange saturation transfer (CEST) MRI to monitor hydrogel degradation in vivo in a label-free fashion, using a commonly used hyaluronic (HA)-gelatin composite hydrogel.

Methods

A covalently cross-linked hydrogel composing of 20 mg/mL thiol-modified gelatin (Gel-SH), 20 mg/mL thiol-modified HA (HA-SH), and 20 mg/mL polyethylene (glycol) diacrylate (PEGDA) (crosslinker) was prepared (Fig. 1a). In vitro CEST MRI was carried out at 37 °C. For in vivo visualization of degradation, 3 μL hydrogel was injected into mouse striatum. CEST MRI was performed using a horizontal bore 11.7 T Bruker scanner up until 42 days. To validate the in vivo CEST MRI findings and identify the main decomposing component in the hydrogel, gelatin and HA were labeled with green and red near-infrared (NIR) dyes, respectively. The labeled hydrogel was then visualized using a LI-COR optical in vivo imaging system.

Results/Discussion

In vitro CEST MRI demonstrated a stable peak at 3.6 ppm for B1 = 1.2-7.2 μT (Fig. 1b). Amongst the three individual components of the hydrogel, gelatin was the major contributor to the CEST signal at 3.6 ppm (Fig. 1c), in agreement with earlier studies.2 An increase of the gelation proportion increased the CEST signal (Fig. 1d-e). When the hydrogel was injected into the brain of mice, it could be clearly distinguished from the surrounding brain tissue (Fig. 1f-g). When using a 4:1 hydrogel ratio, a decrease in CEST MRI signal was observed over time (Fig. 2a-b). The NIR signal of gelatin decreased gradually over 42 days, while the HA signal remained relatively stable (Fig. 2c). This suggests that gelatin, the main source of CEST signal at 3.6 ppm, is the major decomposing component in the hydrogel. An excellent correlation was found between the decay of CEST signal and gelatin NIR fluorescence signal (R2=0.94, Fig. 2d).

Conclusions

Hydrogel degradation can be monitored using CEST MRI. Gelatin was found to be the major contributor of CEST contrast and also the main degradation component in the hydrogel. This imaging approach may be used further to develop hydrogels with optimal biodegradation properties for delivering cells.

References

1. Madl CM, Lesavage BL, Dewi RE et al.: Maintenance of neural progenitor cell stemness in 3D hydrogels requires matrix remodelling. Nature Materials. 2017;16:233-1242.

2. Liang Y, Bar-Shir A, Song X, Gilad AA, Walczak P, Bulte JWM: Label-free imaging of gelatin-containing hydrogel scaffolds. Biomaterials. 2016;42:144-150.

Figure 1
(a) Chemical scheme. (b) Hydrogel MTRasym for different saturation powers. (c) MTRasym of individual hydrogel components at 3.6 μT. (d) MTRasym for different Gel-SH:HA-SH ratios. (e) Quantitative MTRasym and CEST maps at 3.6 ppm. (f) MRI of hydrogel scaffolds injected into mouse brain striatum (arrow: needle track). (g) MTRasym measurements for the implanted hydrogel scaffold Region-of-Interest.
Figure 2
(a) Serial MRI (arrow = hydrogel implantation site). (b) Decay of CEST MTRasym over time. (c) NIR imaging of the same mice shown in (a), with gelatin labeled NIR-green and HA labeled NIR-red. Quantification shows gelatin signal intensity decreasing with time, while the HA intensity remains relatively unchanged. (d) In vivo correlation of CEST MRI signal and fluorescence intensity.
Keywords: CEST MRI, Hydrogel, Cell therapy, Neuroimaging
3:09 p.m. PS 06-05

Multimodal and Multiscale Optical Imaging of Sonopermeation-induced Nanocarrier Translocation across the Blood-Brain Barrier (#117)

Twan Lammers1, Susanne Golombek1, Jan-Niklas May1, Anshuman Dasgupta1, Felix Gremse1, Fabian Kiessling1

1 RWTH Aachen University Clinic, Experimental Molecular Imaging, Aachen, North Rhine-Westphalia, Germany

Introduction

Efficient and safe drug delivery across the blood-brain barrier is a major challenge in biomedical and (nano-) pharmaceutical research. The combination of ultrasound (US) and microbubbles (MB) can induce a temporarily and spatially controlled opening of the BBB [1]. This phenomenon is termed sonopermeation and has already been successfully employed to improve drug delivery to tumors [2], even in patients [3]. Here, we employed multimodal and multiscale optical imaging to systematically study the ability of sonopermeation to shuttle nanocarriers of different sizes across the BBB (see Fig. 1).

Methods

CD-1 nude mice were intravenously co-injected with MB and with pHPMA polymers (10 nm) or PEGylated liposomes (100 nm). Both nanocarriers were labeled with Alexa488 and Cy7. Upon US treatment (16 MHz, 50% power, MI 0.45, 10 min), the accumulation of the two drug delivery systems was longitudinally monitored using hybrid computed tomography-fluorescence molecular tomography (CT-FMT [4]). Prior to sacrifice, rhodamine-labeled lectin was injected to stain perfused blood vessels in the brain. Ex vivo analysis included fluorescence reflectance imaging (FRI), confocal microscopy (CM), multiphoton microscopy (MPM) and stimulated emission depletion (STED) nanoscopy. Possible side effects and the overall extent of BBB opening were also investigated, using H&E and anti-IgG immunofluorescence. 

Results/Discussion

In all experimental groups, the efficiency of sonopermeation-induced BBB opening was confirmed via a significant increase in the number of blood vessels positive for IgG extravasation. H&E stainings showed that sonopermeation, at the US times and settings employed, did not induce obvious brain damage. While the extravasation and penetration of the 10 nm-sized polymeric drug carriers upon sonopermeation was clearly visible with all optical imaging modalities, clear-cut extravasation of 100 nm liposomes could only be observed using highly sensitive high-resolution techniques, such as  multiphoton microscopy and STED nanoscopy. Using in-house developed software tools to quantify the penetration depth of fluorescent drugs and drug delivery systems out of the blood vessels into the brain  [5], we found that 10 nm polymers penetrated significantly more efficiently and much deeper into the CNS upon sonopermeation than 100 nm liposomes.

Conclusions

Sonopermeation can be used to efficiently and safely open up the BBB, allowing drug delivery systems to translocate across the endothelium and penetrate into the brain. Multimodal and multiscale optical imaging shows that small nanocarriers accumulate more efficiently and penetrate deeper than large nanocarriers. These findings contribute to development of novel theranostic strategies to improve the treatment of brain tumors and CNS disorders.

References

1: Dasgupta A, Liu M, Ojha T, Storm G, Kiessling F, Lammers T. Ultrasound-mediated drug delivery to the brain: principles, progress and prospects. Drug Discov Today Tech 20: 41-48 (2016)

2: Theek B, Baues M, Ojha T, Möckel D, Steitz J, van Bloois L, Storm G, Kiessling F, Lammers T. Sonoporation enhances liposome accumulation and penetration in tumors with low EPR. J Control Release 231: 77-85 (2016)

3: Dimcevski G, Kotopoulis S, Bjanes T, Hoem D, Schjott J, Gjertsen B, Biermann M, Molven A, Sorbye H, McCormack E, Postema M, Gilja O. A human clinical trial using ultrasound and microbubbles to enhance gemcitabine treatment of inoperable pancreatic cancer. J Control Release 243: 172-181 (2016)

4: Kunjachan S, Gremse F, Theek B, Koczera P, Pola R, Pechar M, Etrych T, Ulbrich K, Storm G, Kiessling F, Lammers T. Non-invasive optical imaging of nanomedicine biodistribution. ACS Nano 7: 252-262 (2013)

5: Lammers T, Koczera P, Fokong S, Gremse F, Storm G, Van Zandvoort M, Kiessling F. Theranostic USPIO-loaded microbubbles for mediating and monitoring blood-brain barrier permeation. Adv Funct Mater 25: 36-43 (2015)

Acknowledgement

The authors gratefully acknowledge financial support by the European Research Council (ERC-StG 309495: NeoNaNo), the European Commission (EU-ERANET-EuroNanoMed-III: NSC4DIPG), and the German Research Foundation (DFG: GRK 2375).

Figure 1: BBB sonopermeation - Study setup
Figure 2: BBB sonopermeation - Results
Keywords: drug delivery, blood-brain barrier, ultrasound, microbubbles, optical imaging
3:21 p.m. PS 06-06

Contrast-enhanced MR microangiography of cerebral collateralization after chronic hypoperfusion in the mouse (#50)

Philipp Boehm-Sturm1, 2, Till de Bortoli3, 4, Stefan Paul Koch1, 2, Melina Nieminen3, Susanne Mueller1, 2, Giovanna Diletta Ielacqua5, Jan Klohs5, Ulrich Dirnagl1, Peter Vajkoczy3, Nils Hecht3

1 Charité – Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Department of Experimental Neurology and Center for Stroke Research Berlin, Berlin, Berlin, Germany
2 Charité - Universitätsmedizin Berlin, Cluster of Excellence NeuroCure and Charité Core Facility 7T Experimental MRIs, Berlin, Berlin, Germany
3 Charité - Universitätsmedizin Berlin, Department of Neurosurgery and Center for Stroke Research Berlin, Berlin, Berlin, Germany
4 Charité – Universitätsmedizin Berlin, Charité Comprehensive Cancer Center, Berlin, Berlin, Germany
5 University of Zurich & ETH Zurich, Institute for Biomedical Engineering, Zurich, Zürich, Switzerland

Introduction

Collateral outgrowth with flow augmentation remains the most important mechanism for prevention of hemodynamic ischemic stroke. The hemodynamic rescue is mainly provided by new growth of collateral arterioles in the range of ~20-50 µm diameter.1 However, the underlying mechanisms remain poorly understood and tools for in vivo assessment of the degree, morphology and dynamics of the process are lacking.2,3 The goal of this study was to develop contrast-enhanced MR microangiography-based methods4 to quantitatively characterize the collateral vasculature during cerebral arteriogenesis.

Methods

Male C57/Bl6 mice were randomized to undergo right side internal carotid artery occlusion (ICAO, n=8) or a sham (n=8) procedure.2 21 d after surgery, animals underwent MRI at 7T with a cryoprobe (Bruker, Ettlingen, Germany). The ~1.5 h protocol comprised T2w MRI, contrast-enhanced MR microangiography ((65 µm)3 resolution) with Ferumoxytol (300 µmol Fe/kg i.v.), and DSC MRI. Vasculature was stained (100 µL FITC-lectin i.v.) for either confocal fluorescence microscopy or serial two photon tomography (TissueVision, Somerville, MA/USA). Cortical vessel densities were calculated from angiograms4 and relative cerebral blood volume (rCBV) was mapped.5 MR and histology images were registered to the Allen brain atlas.6 All data analyses were performed blind to the group assignment of animals. 

Results/Discussion

One animal (sham) was excluded due to unsuccessful i.v. injection of the contrast agent, one (ICAO) died during MRI leading to a final n=7 in each group. Higher contrast was seen in MR microangiograms of ICAO animals but our pre-study hypothesis that the vessel tracing algorithm would show an increase in vessel density was rejected independent of parameter settings (Fig.1). However, in an exploratory data analysis, rCBV-based markers e.g. number of voxels with blood volume fraction >1% represented the increase in vasculature seen in previous studies. Registration of MRI on fluorescence microscopy images of FITC-lectin stained tissue sections showed that vessels as small as ~25 µm were directly detectable by MR microangiography (Fig. 2a). 3D serial two photon angiograms and MR microangiograms were mapped on the Allen brain atlas (Fig. 2b). A biophysical model to quantitatively compare the two modalities is currently investigated.

Conclusions

Contrast-enhanced MR microangiography visualizes changes in the collateral vasculature. rCBV seems to be a more appropriate marker of arteriogenesis but this remains to be confirmed by voxel-wise comparison to 3D histology. In the future, the quantitative tools developed in this study will help to investigate mechanisms and strategies for therapeutic stimulation of cerebral arteriogenesis in experimental models.

References

1         Faber JE, Chilian WM, Deindl E, van Royen N, Simons M. A Brief Etymology of the Collateral Circulation. Arterioscler Thromb Vasc Biol 2014; 34: 1854–1859.

2         Hecht N, He J, Kremenetskaia I, Nieminen M, Vajkoczy P, Woitzik J. Cerebral Hemodynamic Reserve and Vascular Remodeling in C57/BL6 Mice Are Influenced by Age. Stroke 2012; 43: 3052–3062.

3         Hecht N, Marushima A, Nieminen M, Kremenetskaia I, von Degenfeld G, Woitzik J et al. Myoblast-Mediated Gene Therapy Improves Functional Collateralization in Chronic Cerebral Hypoperfusion. Stroke 2015; 46: 203–211.

4         Klohs J, Baltes C, Princz-Kranz F, Ratering D, Nitsch RM, Knuesel I et al. Contrast-Enhanced Magnetic Resonance Microangiography Reveals Remodeling of the Cerebral Microvasculature in Transgenic ArcA Mice. J Neurosci 2012; 32: 1705–1713.

5         Lemasson B, Bouchet A, Maisin C, Christen T, Le Duc G, Rémy C et al. Multiparametric MRI as an early biomarker of individual therapy effects during concomitant treatment of brain tumours. NMR Biomed 2015; 28: 1163–1173.

6         Koch S, Mueller S, Foddis M, Bienert T, von Elverfeldt D, Knab F et al. Atlas registration for edema-corrected MRI lesion volume in mouse stroke models. J Cereb Blood Flow Metab 2017; : 0271678X17726635.

Acknowledgement

Work was supported by the Deutsche Forschungsgemeinschaft Cluster of Excellence NeuroCure (Exc 257) and the German Federal Ministry of Education and Research (BMBF; 01EO0801, Center for Stroke Research Berlin).

Fig. 1: MRI-based vessel density measurement after chronic hypoperfusion

Vessel density (left+right hemisphere) calculated from MR microangiograms decreased in ICAO animals for three different settings (connectivity threshold 3, 6, 9 voxels) of the previously published vessel tracing algorithm. Comparsion using two sample t-test (*p<0.05).

Fig. 2: Comparison of MR microangiograms with histology using image registration

A) Example of cortical MR microangiogram (red, Frangi filtered) registered to FITC-lectin stained vasculature on a single coronal tissue slice (green). Registration artefacts and false negatives are still visible on high resolution microscopy images but circles denote areas with a qualitative match of histology and MRI. Zoomed overlay (left) shows MR detection of a single penetrating cortical vessel of 20-30 µm diameter. B) Allen atlas registration of a cortical MR microangiogram and FITC-lectin-based whole brain serial two photon tomography angiogram (downsampled to 25 µm isotropic resolution) for 3D histological validation.

Keywords: mri, collateralization, chronic hypoperfusion, angiography, mouse
3:33 p.m. PS 06-07

Statistical parametric mapping of TSPO PET reveals divergent signal changes after anti-inflammatory treatment in a mouse model of epileptogenesis (#259)

Bettina Wolf1, 2, Pablo Bascunana1, Ina Leiter1, 2, Laura Langer1, Tobias L. Ross1, Frank M. Bengel1, Marion Bankstahl2, Jens P. Bankstahl1

1 Hannover Medical School, Nuclear Medicine, Hannover, Germany
2 University of Veterinary Medicine Hannover, Pharmacology, Hannover, Germany

Introduction

Neuroinflammation is considered as target for anti-epileptogenic treatment in insult-induced epileptogenesis. The sphingosine-1-phosphate analog fingolimod (FTY720), clinically used for anti-inflammatory treatment in multiple sclerosis, has shown promising results in preclinical epilepsy models (Gao et al. 2012). Here, we used molecular positron emission tomography (PET) imaging targeting the translocator protein (TSPO) to quantify effects of FTY720 during epileptogenesis.

Methods

Status epilepticus (SE) was induced by unilaterally injecting kainate into the dorsal hippocampus of male NMRI mice resulting in epileptogenesis thereafter. Starting six hours after SE induction, mice received either FTY720 (0.3 mg/kg, n=8) or vehicle injections (n=18) once daily for 5 days. Static [18F]GE180 TSPO PET scans (40-60 min after tracer injection) were performed 1 and 2 weeks after SE. For analysis, statistical parametric mapping (SPM) and standard MRI atlas-based regional quantification was used. Subsequently, imaging results were validated by autoradiography and histological analysis of brain slices.

Results/Discussion

[18F]GE180 PET identified divergent FTY720 treatment effects. Contrary to our expectations, FTY720 treatment resulted in increased TSPO expression in the major epileptogenic focus. SPM analysis at 1 and 2 weeks as well as autoradiography at 2 weeks post SE revealed a focal increase of TSPO expression in FTY720-treated mice close to the kainate injection area of up to 30% compared to vehicle-treated animals. Conversely, SPM analysis and autoradiography 2 weeks post SE showed a decrease of up to 42% mainly in the contralateral hemisphere of FTY720-treated mice. In contrast to the SPM analysis, atlas-based regional analysis did reveal only very limited signal changes. This might be caused by dilution effects due to changes only in parts of the pre-defined regions. Immunohistochemical analyses at 2 weeks post SE revealed reduced astrocyte activation in the contralateral hippocampus after FTY720 treatment, but no neuroprotection.

Conclusions

Sole use of atlas-based regional analysis might mask changes in mouse brain PET studies. Both activation and reduction of local inflammation by FTY720 may support reported anti-epileptogenic properties of the treatment, as TSPO expression is not totally specific to different pro- or anti-inflammatory microglia subtypes. It is currently investigated, if this scheme of changes will be observed after other anti-inflammatory treatment approaches.

References

Gao, F., Y. Liu, X. Li, Y. Wang, D. Wei and W. Jiang (2012). "Fingolimod (FTY720) inhibits neuroinflammation and attenuates spontaneous convulsions in lithium-pilocarpine induced status epilepticus in rat model." Pharmacology, Biochemistry and Behavior 103(2): 187-196.

Acknowledgement

B. Wolf is supported by a scholarship from Studienstiftung des Deutschen Volkes. I. Leiter was supported by a scholarship from the Konrad-Adenauer-Stiftung e.V. This study was funded by the European Seventh’s Framework Program (FP7/2007-2013) under grant agreement No. 602102 (EPITARGET).

TSPO PET after FTY720 treatment during epileptogenesis

(a) Average coronal brain images of [18F]GE180 uptake (%ID/cc) 7 and 14 days after SE of vehicle (Veh.) and FTY720 treated animals. (b) Statistical parametric mapping analysis of [18F]GE180 uptake. Coronal and horizontal t-maps resulting from comparisons between FTY720 and vehicle treated animals 7 and 14 days after SE (Student’s t-test, p<0.05, minimum cluster size 50 voxels).

Keywords: epilepsy, PET, TSPO, treatement, neuroinflammation