Eyk Schellenberger - Academia.edu (original) (raw)

Papers by Eyk Schellenberger

Noninvasive imaging using radioactive annexin V is an emerging strat- egy for the assessment of c... more Noninvasive imaging using radioactive annexin V is an emerging strat- egy for the assessment of cell death in vivo (F. G. Blankenberg, and H. W. Strauss. Apoptosis, 6: 117-123, 2001.). Therefore, we investigated whether annexin V labeled with the fluorophore Cy5.5 (Cy) could serve as a probe for imaging of tumor apoptosis using near infrared fluorescence (NIRF). We prepared active

Molecular imaging

Superparamagnetic iron oxide particles are used as potent contrast agents in magnetic resonance i... more Superparamagnetic iron oxide particles are used as potent contrast agents in magnetic resonance imaging. In histology, these particles are frequently visualized by Prussian blue iron staining of aldehyde-fixed, paraffin-embedded tissues. Recently, zinc salt-based fixative was shown to preserve enzyme activity in paraffin-embedded tissues. In this study, we demonstrate that zinc fixation allows combining in situ zymography with fluorescence immunohistochemistry (IHC) and iron staining for advanced biologic investigation of iron oxide particle accumulation. Very small iron oxide particles, developed for magnetic resonance angiography, were applied intravenously to BALB/c nude mice. After 3 hours, spleens were explanted and subjected to zinc fixation and paraffin embedding. Cut tissue sections were further processed to in situ zymography, IHC, and Prussian blue staining procedures. The combination of in situ zymography as well as IHC with subsequent Prussian blue iron staining on zinc-...

Cellular oncology : the official journal of the International Society for Cellular Oncology, 2005

To microscopically analyze the chemotherapeutic response of tumors using in vivo staining based o... more To microscopically analyze the chemotherapeutic response of tumors using in vivo staining based on an annexinV-Cy5.5 probe and independently asses their apoptotic count using quantitative histological analysis. Lewis Lung Carcinomas cells, that are sensitive (CS-LLC) and resistant (CR-LLC) to chemotherapy were implanted in nude mice and grown to tumours. Mice were treated with cyclophosphamide and injected with a Cy5.5-annexinV fluorescent probe. In vivo imaging was performed using Fluorescence Molecular Tomography. Subsequently tumours were excised and prepared for histology. The histological tumour sections were stained for apoptosis using a terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) assay. A minimum of ten tissue sections were analyzed per tumour for apoptosis quantification by TUNEL staining and corresponding Cy5.5 distribution. We detected higher levels of apoptosis and corresponding higher levels of Cy5.5 fluorescence in the CS-LLC vs. the CR-LLC ...

Molecular imaging

We examined the accumulation of Cy5.5-labeled annexin V in the paws of mice with and without coll... more We examined the accumulation of Cy5.5-labeled annexin V in the paws of mice with and without collagen-induced arthritis, with and without methotrexate (MTX) treatment, by near-infrared fluorescence imaging. Fluorescence reflectance imaging (FRI) of paws was performed 48 hr after MTX injection and at 10 min and 3 hr after the injection of Cy5.5-annexin V (1 nmol dye per mouse). With arthritic paws, MTX treatment caused a 7-fold increase in fluorescence intensity compared with the paws of untreated mice and a 4-fold increase compared to nonarthritic paws of MTX-treated mice (p < .001 each). Tissue samples of paws were examined histologically for Cy5.5 fluorescence and by TUNEL staining for apoptosis. Cy5.5-annexin V was seen in the hyperplastic synovia of MTX-treated mice, and TUNEL staining for apoptosis showed apoptotic cells in the hyperplastic synovia. Monitoring the uptake of Cy5.5-annexin V in arthritic paws by FRI provided a method of assessing a response to MTX, a response ...

Chembiochem : a European journal of chemical biology, Jan 5, 2004

We have developed techniques for the efficient synthesis and screening of small libraries of surf... more We have developed techniques for the efficient synthesis and screening of small libraries of surface-functionalized nanoparticles for the recognition of specific types of cells. To illustrate this concept we describe the development of a nanoparticle that preferentially recognizes apoptotic Jurkat cells in a manner similar to the apoptosis-recognizing protein annexin V. The nanoparticle, which is detectable by fluorescence or NMR relaxometry, was analyzed for the ability to recognize normal and apoptotic cells by fluorescence-activated cell sorting (FACS) analysis and fluorescence microscopy. The capability to develop nanoparticles which interact with specific target cells could be applied to the design of materials for diverse applications including quantum dots, which serve as fluorescence tracers, colloidal gold, which serves as a tracer for electron micrographs, or the crystalline forms of drugs.

Neoplasia (New York, N.Y.)

A rapid and accurate assessment of the antitumor efficacy of new therapeutic drugs could speed up... more A rapid and accurate assessment of the antitumor efficacy of new therapeutic drugs could speed up drug discovery and improve clinical decision making. Based on the hypothesis that most effective antitumor agents induce apoptosis, we developed a near-infrared fluorescent (NIRF) annexin V to be used for optical sensing of tumor environments. To demonstrate probe specificity, we developed both an active (i.e., apoptosis-recognizing) and an inactive form of annexin V with very similar properties (to account for nonspecific tumor accumulation), and tested the agents in nude mice each bearing a cyclophosphamide (CPA) chemosensitive (LLC) and a chemoresistant LLC (CR-LLC). After injection with active annexin V, the tumor-annexin V ratio (TAR; tumor NIRF/background NIRF) for untreated mice was 1.22+/-0.34 for LLC and 1.43+/-0.53 for CR-LLC (n=4). The LLC of CPA-treated mice had significant elevations of TAR (2.56+/-0.29, P=.001, n=4), but only a moderate increase was obtained for the CR-LLC...

Cancer research, Jan 15, 2003

Noninvasive imaging using radioactive annexin V is an emerging strategy for the assessment of cel... more Noninvasive imaging using radioactive annexin V is an emerging strategy for the assessment of cell death in vivo (F. G. Blankenberg, and H. W. Strauss. Apoptosis, 6: 117-123, 2001.). Therefore, we investigated whether annexin V labeled with the fluorophore Cy5.5 (Cy) could serve as a probe for imaging of tumor apoptosis using near infrared fluorescence (NIRF). We prepared active Cy-annexin (an equimolar dye:protein ratio) that bound to apoptotic Jurkat T cells and an inactive Cy-annexin probe (>2 dyes/mol protein) that did not. Active Cy annexin was used to image a 9L gliosarcoma, constitutively expressing green fluorescent protein marker, and the CR8 variant of Lewis lung carcinoma, stably transfected to express DsRed2. The expression of transfected fluorescent protein provided an indication of tumor margins and a means of defining tumor-associated NIRF signal intensity with both tumor models. Tumors were imaged with and without cyclophosphamide treatment. In both tumor models a...

Medical Laser Application, 2003

ABSTRACT

Journal of Cerebral Blood Flow & Metabolism, 2014

Focal cerebral ischemia is among the main causes of death and disability worldwide. The ischemic ... more Focal cerebral ischemia is among the main causes of death and disability worldwide. The ischemic core often progresses, invading the peri-ischemic brain; however, assessing the propensity of the peri-ischemic brain to undergo secondary damage, understanding the underlying mechanisms, and adjusting treatment accordingly remain clinically unmet challenges. A significant hallmark of the peri-ischemic brain is dysfunction of the blood-brain barrier (BBB), yet the role of disturbed vascular permeability in stroke progression is unclear. Here we describe a longitudinal in vivo fluorescence imaging approach for the evaluation of cortical perfusion, BBB dysfunction, free radical formation and cellular injury using the photothrombosis vascular occlusion model in male Sprague Dawley rats. Blood-brain barrier dysfunction propagated within the peri-ischemic brain in the first hours after photothrombosis and was associated with free radical formation and cellular injury. Inhibiting free radical signaling significantly reduced progressive cellular damage after photothrombosis, with no significant effect on blood flow and BBB permeability. Our approach allows a dynamic follow-up of cellular events and their response to therapeutics in the acutely injured cerebral cortex.

Textbook ofin vivo Imaging in Vertebrates, 2006

RöFo - Fortschritte auf dem Gebiet der Röntgenstrahlen und der bildgebenden Verfahren, 2012

To compare citrate-coated very small superparamagnetic iron oxide particles (VSOP) with gadofosve... more To compare citrate-coated very small superparamagnetic iron oxide particles (VSOP) with gadofosveset trisodium as blood pool contrast agents for cardiac magnetic resonance angiography (CMRA) in pigs. Animal experiments were approved by the responsible authority. 10 CMRA-like examinations were performed at 1.5 T after administration of VSOP (0.06 mmol Fe/kg; 5 examinations) and gadofosveset trisodium (0.03 mmol Gd/kg; 5 examinations). The CMRA protocol included ECG-gated inversion-recovery-prepared T1-weighted gradient echo imaging (IR-GRE; one slice) and ECG-gated inversion recovery prepared steady state free precession imaging (IR SSFP; one slice) before and 1, 3, 5, 10, 15, 20, 25, 30, 40, 50, and 60 min after injection. At each time point, three different inversion times (TI; 200 msec, 300 msec, and 400 msec) were applied. Contrast-to-noise ratios (CNR) between blood and myocardium were calculated and compared using mixed linear models. No significant differences of CNR were found between IR-GRE and IR SSFP. At 3 and 5 min after contrast agent administration, VSOP showed a significantly higher CNR than gadofosveset trisodium when TI of 200 msec and 300 msec were applied (TI of 200 msec at 3 min: 8.2 ± 0.7 vs. 5.4 ± 0.7; TI of 200 msec at 5 min: 7.9 ± 0.7 vs. 3.5 ± 0.8; TI of 300 msec at 3 min: 11.7 ± 0.7 vs. 8.8 ± 0.8; TI of 300 msec at 5 min: 11.4 ± 0.7 vs. 8.0 ± 0.8; p &amp;amp;amp;lt; 0.05). Moreover, significant differences in favor of VSOP were found for all time points from 10 to 40 min irrespective of TI (p &amp;amp;amp;lt; 0.05). VSOP has superior blood-pool properties compared to gadofosveset trisodium resulting in prolonged improvement of CNR on CMRA.

Radiology, 2006

To compare a superparamagnetic iron oxide (SPIO), VSOP-C184, with a gadopentetate dimeglumine wit... more To compare a superparamagnetic iron oxide (SPIO), VSOP-C184, with a gadopentetate dimeglumine with regard to signal-enhancing effects on T1-weighted dynamic magnetic resonance (MR) images and with another SPIO contrast medium with regard to signal-reducing effects on delayed T2-weighted MR images. All experiments were approved by the responsible Animal Care Committee. Twenty rabbits (five for each contrast agent and dose) implanted with VX-2 carcinoma were imaged at 1.5 T. VSOP-C184 at 0.015 and 0.025 mmol Fe/kg was compared with gadopentetate dimeglumine at 0.15 mmol Gd/kg and ferucarbotran at 0.015 mmol Fe/kg. The imaging protocol comprised a T1-weighted dynamic gradient-echo (GRE) MR before injection and at 6-second intervals for up to 42 seconds after injection and a T2-weighted turbo spin-echo MR before and 5 minutes after injection. Images were evaluated quantitatively, and contrast media were compared by using nonparametric analysis of variance. At dynamic T1-weighted GRE MR imaging with 0.015-mmol Fe/kg VSOP-C184, 0.025-mmol Fe/kg VSOP-C184, gadopentetate dimeglumine, and ferucarbotran, the median peak contrast-to-noise ratio (CNR) was 20.7 (25th percentile, 16.3; 75th percentile, 22.6), 24.2 (25th percentile, 19.3; 75th percentile, 28.5), 16.4 (25th percentile, 13.7; 75th percentile, 20.3), and 14.0 (25th percentile, 11.4; 75th percentile, 16.8), respectively. Both doses of VSOP-C184 yielded significantly higher CNR (P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; .05) than the other two agents. At T2-weighted turbo spin-echo imaging with 0.015-mmol Fe/kg VSOP-C184, 0.025-mmol Fe/kg VSOP-C184, gadopentetate dimeglumine, and ferucarbotran, the median CNR was 15.0 (25th percentile, 13.4; 75th percentile, 21.3), 15.7 (25th percentile, 14.5; 75th percentile, 19.8), 11.3 (25th percentile, 8.2; 75th percentile, 12.2), and 15.7 (25th percentile, 12.5; 75th percentile, 22.4), respectively. There was no significant difference between VSOP-C184 and ferucarbotran; both had a significantly higher CNR than did gadopentetate dimeglumine. VSOP-C184 produces higher liver-to-tumor contrast at dynamic T1-weighted imaging than does gadopentetate dimeglumine; at delayed T2-weighted imaging, the contrast is comparable to that achieved with ferucarbotran.

Proceedings of the National Academy of Sciences, 2004

In vivo imaging of treatment responses at the molecular level could have a significant impact on ... more In vivo imaging of treatment responses at the molecular level could have a significant impact on the speed of drug discovery and ultimately lead to personalized medicine. Strong interest has been shown in developing quantitative fluorescence-based technologies with good molecular specificity and sensitivity for noninvasive 3D imaging through tissues and whole animals. We show herein that tumor response to chemotherapy can be accurately resolved by fluorescence molecular tomography (FMT) with a phosphatidylserinesensing fluorescent probe based on modified annexins. We observed at least a 10-fold increase of fluorochrome concentration in cyclophosphamide-sensitive tumors and a 7-fold increase of resistant tumors compared with control studies. FMT is an optical imaging technique developed to overcome limitations of commonly used planar illumination methods and demonstrates higher quantification accuracy validated by histology. It is further shown that a 3-fold variation in background absorption heterogeneity may yield 100% errors in planar imaging but only 20% error in FMT, thus confirming tomographic imaging as a preferred tool for quantitative investigations of fluorescent probes in tissues. Tomographic approaches are found essential for small-animal optical imaging and are potentially well suited for clinical drug development and monitoring. drug discovery ͉ quantification ͉ three-dimensional T he ability to noninvasively image molecular processes in vivo is an emerging reality with different reporter and detection approaches (1, 2). Molecular imaging has been heralded to lead to earlier detection than current anatomical imaging approaches, which typically detect late-stage abnormalities. Another important prospect of molecular imaging is the ability to examine and quantify treatment responses in vivo by monitoring specific primary molecules or downstream targets. Therapeutic efficacy could then be probed dynamically on timescales of hours to days. This ability is in contrast to the mainstay of today's healthcare with traditionally late end points of drug efficacy, a practice that often impairs prompt revision and exclusion of ineffective treatment strategies with potentially lethal results.

Magnetic Resonance in Medicine, 2005

The ability to image cardiomyocyte apoptosis in vivo with high-resolution MRI could facilitate th... more The ability to image cardiomyocyte apoptosis in vivo with high-resolution MRI could facilitate the development of novel cardioprotective therapies. The sensitivity of the novel nanoparticle AnxCLIO-Cy5.5 for cardiomyocyte apoptosis was thus compared in vitro to that of annexin V-FITC and showed a high degree of colocalization. MRI was then performed, following transient coronary artery (LAD) occlusion, in five mice given AnxCLIO-Cy5.5 and in four mice given an identical dose (2 mg Fe/kg) of CLIO-Cy5.5. MR signal intensity and myocardial T2* were evaluated, in vivo, in hypokinetic regions of myocardium in the LAD distribution. Ex vivo fluorescence imaging was performed to confirm the in vivo findings. Myocardial T2* was significantly lower in the mice given AnxCLIO-Cy5.5 (8.1 versus 13.2 ms, P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.01), and fluorescence target to background ratio was significantly higher (2.1 versus 1.1, P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.01). This study thus demonstrates the feasibility of obtaining high-resolution MR images of cardiomyocyte apoptosis in vivo with the novel nanoparticle, AnxCLIO-Cy5.5.

Journal of Nuclear Medicine, 2014

This article and updated information are available at:

Journal of Magnetic Resonance Imaging, 2011

To evaluate very small superparamagnetic iron oxide particles (VSOP-C184) as blood-pool contrast ... more To evaluate very small superparamagnetic iron oxide particles (VSOP-C184) as blood-pool contrast agent for coronary MR angiography (CMRA) in humans. Six healthy volunteers and 14 patients with suspected coronary artery disease underwent CMRA after administration of VSOP-C184 at the following doses: 20 μmol Fe/kg (4 patients), 40 μmol Fe/kg (5 patients), 45 μmol Fe/kg (6 healthy volunteers), and 60 μmol Fe/kg (5 patients). In healthy volunteers, contrast-to-noise ratio (CNR), signal-to-noise ratio (SNR), and vessel edge definition (VED) of contrast-enhanced CMRA were compared with non-contrast-enhanced CMRA. In patients, a per-segment intention-to-diagnose evaluation of contrast-enhanced CMRA for detection of significant coronary stenosis (≥50%) was performed. Three healthy volunteers (45 μmol Fe/kg VSOP-C184) and two patients (60 μmol Fe/kg VSOP-C184) had adverse events of mild or moderate intensity. VSOP-C184 significantly increased CNR (15.1 ± 4.6 versus 6.9 ± 1.9; P = 0.010), SNR (21.7 ± 5.3 versus 15.4 ± 1.6; P = 0.048), and VED (2.3 ± 0.6 versus 1.2 ± 0.2; P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001) compared with non-contrast-enhanced CMRA. In patients, contrast-enhanced CMRA yielded sensitivity, specificity, and diagnostic accuracy for detection of significant coronary stenosis of 86.7%, 71.0%, 73.1%, respectively. CMRA using VSOP-C184 was feasible and yielded moderate diagnostic accuracy for detection of significant coronary stenosis within this proof-of-concept setting.

Journal of Cerebral Blood Flow & Metabolism, 2013

Intravenous administration of iron oxide nanoparticles during the acute stage of experimental str... more Intravenous administration of iron oxide nanoparticles during the acute stage of experimental stroke can produce signal intensity changes in the ischemic region. This has been attributed, albeit controversially, to the infiltration of iron-laden blood-borne macrophages. The properties of nanoparticles that render them most suitable for phagocytosis is a matter of debate, as is the most relevant timepoint for administration. Both of these questions are examined in the present study. Imaging experiments were performed in mice with 30 minutes of middle cerebral artery occlusion (MCAO). Iron oxide nanoparticles with different charges and sizes were used, and mice received 300 μmol Fe/kg intravenously: either superparamagnetic iron oxide nanoparticles (SPIOs), ultrasmall SPIOs, or very small SPIOs. The particles were administered 7 days before MCAO, at the time of reperfusion, or 72 hours after MCAO. Interestingly, there was no observable signal change in the ischemic brains that could be attributed to iron. Furthermore, no Prussian blue-positive cells were found in the brains or blood leukocytes, despite intense staining in the livers and spleens. This implies that the nanoparticles selected for this study are not phagocytosed by blood-borne leukocytes and do not enter the ischemic mouse brain.

Molecular imaging

Cellular oncology : the official journal of the International Society for Cellular Oncology, 2005

Molecular imaging

Chembiochem : a European journal of chemical biology, Jan 5, 2004

Neoplasia (New York, N.Y.)

Cancer research, Jan 15, 2003

Medical Laser Application, 2003

ABSTRACT

Journal of Cerebral Blood Flow & Metabolism, 2014

Textbook ofin vivo Imaging in Vertebrates, 2006

RöFo - Fortschritte auf dem Gebiet der Röntgenstrahlen und der bildgebenden Verfahren, 2012

Radiology, 2006

Proceedings of the National Academy of Sciences, 2004

Magnetic Resonance in Medicine, 2005

Journal of Nuclear Medicine, 2014

This article and updated information are available at:

Journal of Magnetic Resonance Imaging, 2011

Journal of Cerebral Blood Flow & Metabolism, 2013