Stefan Polei

One Year Follow-Up Risk Assessment in SKH-1 Mice and Wounds Treated with an Argon Plasma Jet

Multiple evidence in animal models and in humans suggest a beneficial role of cold physical plasma in wound treatment. Yet, risk assessment studies are important to further foster therapeutic advancement and acceptance of cold plasma in clinics. Accordingly, we investigated the long-term side effects of repetitive plasma treatment over 14 consecutive days in a rodent full-thickness ear wound model. Subsequently, animals were housed for 350 days and sacrificed thereafter. In blood, systemic changes of the pro-inflammatory cytokines interleukin 1β and tumor necrosis factor α were absent. Similarly, tumor marker levels of α-fetoprotein and calcitonin remained unchanged. Using quantitative PCR, the expression levels of several cytokines and tumor markers in liver, lung, and skin were found to be similar in the control and treatment group as well. Likewise, histological and immunohistochemical analysis failed to detect abnormal morphological changes and the presence of tumor markers such as carcinoembryonic antigen, α-fetoprotein, or the neighbor of Punc 11. Absence of neoplastic lesions was confirmed by non-invasive imaging methods such as anatomical magnetic resonance imaging and positron emission tomography-computed tomography. Our results suggest that the beneficial effects of cold plasma in wound healing come without apparent side effects including tumor formation or chronic inflammation.

[18F]fallypride-PET/CT Analysis of the Dopamine D2/D3 Receptor in the Hemiparkinsonian Rat Brain Following Intrastriatal Botulinum Neurotoxin A Injection

Intrastriatal injection of botulinum neurotoxin A (BoNT-A) results in improved motor behavior of hemiparkinsonian (hemi-PD) rats, an animal model for Parkinson’s disease. The caudate–putamen (CPu), as the main input nucleus of the basal ganglia loop, is fundamentally involved in motor function and directly interacts with the dopaminergic system. To determine receptor-mediated explanations for the BoNT-A effect, we analyzed the dopamine D2/D3 receptor (D2/D3R) in the CPu of 6-hydroxydopamine (6-OHDA)-induced hemi-PD rats by [18F]fallypride-PET/CT scans one, three, and six months post-BoNT-A or -sham-BoNT-A injection. Male Wistar rats were assigned to three different groups: controls, sham-injected hemi-PD rats, and BoNT-A-injected hemi-PD rats. Disease-specific motor impairment was verified by apomorphine and amphetamine rotation testing. Animal-specific magnetic resonance imaging was performed for co-registration and anatomical reference. PET quantification was achieved using PMOD software with the simplified reference tissue model 2. Hemi-PD rats exhibited a constant increase of 23% in D2/D3R availability in the CPu, which was almost normalized by intrastriatal application of BoNT-A. Importantly, the BoNT-A effect on striatal D2/D3R significantly correlated with behavioral results in the apomorphine rotation test. Our results suggest a therapeutic effect of BoNT-A on the impaired motor behavior of hemi-PD rats by reducing interhemispheric changes of striatal D2/D3R.

APPswe/PS1dE9 mice with cortical amyloid pathology show a reduced NAA/Cr ratio without apparent brain atrophy: A MRS and MRI study

Transgenic animal models of Aβ pathology provide mechanistic insight into some aspects of Alzheimer disease (AD) pathology related to Aβ accumulation. Quantitative neuroimaging is a possible aid to improve translation of mechanistic findings in transgenic models to human end phenotypes of brain morphology or function. Therefore, we combined MRI-based morphometry, MRS-based NAA-assessment and quantitative histology of neurons and amyloid plaque load in the APPswe/PS1dE9 mouse model to determine the interrelationship between morphological changes, changes in neuron numbers and amyloid plaque load with reductions of NAA levels as marker of neuronal functional viability. The APPswe/PS1dE9 mouse showed an increase of Aβ plaques, loss of neurons and an impairment of NAA/Cr ratio, which however was not accompanied with brain atrophy. As brain atrophy is one main characteristic in human AD, conclusions from murine to human AD pathology should be drawn with caution.

[ 68 Ga]pentixafor for CXCR4 imaging in a PC-3 prostate cancer xenograft model – comparison with [ 18 F]FDG PET/CT, MRI and ex vivo receptor expression

Purpose The aim was to characterize the properties of [68Ga]Pentixafor as tracer for prostate cancer imaging in a PC-3 prostate cancer xenograft mouse model and to investigate its correlation with [18F]FDG PET/CT, magnetic resonance imaging (MRI) and ex vivo analyses. Methods Static [68Ga]Pentixafor and [18F]FDG PET as well as morphological/ diffusion weighted MRI and 1H MR spectroscopy was performed. Imaging data were correlated with ex vivo biodistribution and CXCR4 expression in PC-3 tumors (immunohistochemistry (IHC), mRNA analysis). Flow cytometry was performed for evaluation of localization of CXCR4 receptors (in vitro PC-3 cell experiments). Results Tumor uptake of [68Ga]Pentixafor was significantly lower compared to [18F]FDG. Ex vivo CXCR4 mRNA expression of tumors was shown by PCR. Only faint tumor CXCR4 expression was shown by IHC (immuno reactive score of 3). Accordingly, flow cytometry of PC-3 cells revealed only a faint signal, cell membrane permeabilisation showed a slight signal increase. There was no significant correlation of [68Ga]Pentixafor tumor uptake and ex vivo receptor expression. Spectroscopy showed typical spectra of prostate cancer. Conclusion PC-3 tumor uptake of [68Ga]Pentixafor was existent but lower compared to [18F]FDG. No significant correlation of ex vivo tumor CXCR4 receptor expression and [68Ga]Pentixafor tumor uptake was shown. CXCR4 receptor expression on the surface of PC-3 cells was existent but rather low possibly explaining the limited [68Ga]Pentixafor tumor uptake; receptor localization in the interior of PC-3 cells is presumable as shown by cell membrane permeabilisation. Further studies are necessary to define the role of [68Ga]Pentixafor in prostate cancer imaging.

Application of in vivo imaging techniques to monitor therapeutic efficiency of PLX4720 in an experimental model of microsatellite instable colorectal cancer

Objectives Patient-derived tumor cell lines are a powerful tool to analyze the sensitivity of individual tumors to specific therapies in mice. An essential prerequisite for such an approach are reliable quantitative techniques to monitor tumor progression in vivo. Methods We have employed HROC24 cells, grown heterotopically in NMRI Foxn1nu mice, as a model of microsatellite instable colorectal cancer to investigate the therapeutic efficiencies of 5’-fluorouracil (5’-FU) and the mutant BRAF inhibitor PLX4720, a vemurafenib analogue, by three independent methods: external measurement by caliper, magnetic resonance imaging (MRI) and positron emission tomography/computed tomography (PET/CT) with 2-deoxy-2-(18F)fluoro-D-glucose (18F-FDG). Results Repeated measure ANOVA by a general linear model revealed that time-dependent changes of anatomic tumor volumes measured by MRI differed significantly from those of anatomic volumes assessed by caliper and metabolic volumes determined by PET/CT. Over the investigation period of three weeks, neither 5’-FU, PLX4720 nor a combination of both drugs affected the tumor volumes. Also, there was no drug effect on the apparent diffusion constant (ADC) value as detected by MRI. Interestingly, however, PET/CT imaging showed that PLX4720-containing therapies transiently reduced the standardized uptake value (SUV), indicating a temporary response to treatment. Conclusions 5’-FU and PLX4720 were largely ineffective with respect to HROC24 tumor growth. Tumoral uptake of 18F-FDG, as expressed by the SUV, proved as a sensitive indicator of small therapeutic effects. Metabolic imaging by 18F-FDG PET/CT is a suitable approach to detect effects of tumor-directed therapies early and even in the absence of morphological changes.

Current-Dependent Periodicities of Si(553)-Au

We investigate quasi one-dimensional atomic chains on Si(553)-Au with a scanning tunneling microscope (STM). The observed periodicity at the Si step edge can be altered by the STM and depends on the magnitude of the tunneling current. In a recent report this reversible structural transition was attributed to transient doping with a characteristic time scale of a few milliseconds. Here we explore the evolution of the STM topography as a function of the magnitude of the tunneling current for a wide temperature range. Based on a decomposition of topographic line profiles and a detailed Fourier analysis we conclude that all observed currentdependent STM topographies can be explained by a time-averaged linear combination of two fluctuating step-edge structures. These data also reveal the precise relative alignment of the characteristic STM features for both phases along the step edges. A simple diagram is developed, presenting the relative contribution of these phases to the STM topography as a function of tunneling current and temperature. Time- and current-dependent measurements of fluctuations in the tunneling current reveal two different transition regimes that are related to two specific current injection locations within the surface unit cell. A method based on spatially resolved I(z)curves is presented that enables a quantitative analysis of contributing phases.

Young and healthy C57BL/6 J mice performing sprint interval training reveal gender- and site-specific changes to the cortical bone

Physical exercise is considered to impede the bone loss associated with physiological ageing however, a training program that efficiently leads to bone accrual in the healthy does not yet exist. We turned to the C57BL/6 J mouse and designed a sprint interval training for treadmill that was tailored to the individual performance limits. It consisted of four weeks with five training sessions each, followed by another four weeks with three. After completion of the training period, mice were sacrificed and the hind legs were analyzed via µCT and MRI for changes in bone parameters and muscle volume, respectively. Increased performance limits in both sexes confirmed an effect of the treadmill training. However, while male tibiae after eight weeks revealed a significant reduction of cortical bone mass at the distal metaphysis, the cross sectional analysis of female tibiae showed a transient decrease of cortical bone mass after four weeks that was reversed into a significant accrual after eight weeks of training and occurred over the entire length of the tibia. The observed net reduction of female bone mass after four weeks of training is suggestive of a remodelling process which may be delayed in the males.

[18F]-florbetaben PET/CT imaging in the Alzheimer`s disease mouse model APPswe/PS1dE9

BACKGROUND Positron-emission-tomography (PET) using 18F labeled florbetaben allows noninvasive in vivo-assessment of amyloid-beta (Aβ), a pathological hallmark of Alzheimers disease (AD). In preclinical research, [18F]-florbetaben-PET has already been used to test the amyloid-lowering potential of new drugs, both in humans and in transgenic models of cerebral amyloidosis. The aim of this study was to characterize the spatial pattern of cerebral uptake of [18F]-florbetaben in the APPswe/ PS1dE9 mouse model of AD in comparison to histologically determined number and size of cerebral Aβ plaques. METHODS Both, APPswe/PS1dE9 and wild type mice at an age of 12 months were investigated by smallanimal PET/CT after intravenous injection of [18F]-florbetaben. High-resolution magnetic resonance imaging data were used for quantification of the PET data by volume of interest analysis. The standardized uptake values (SUVs) of [18F]-florbetaben in vivo as well as post mortem cerebral Aβ plaque load in cortex, hippocampus and cerebellum were analyzed. RESULTS Visual inspection and SUVs revealed an increased cerebral uptake of [18F]-florbetaben in APPswe/ PS1dE9 mice compared with wild type mice especially in the cortex, the hippocampus and the cerebellum. However, SUV ratios (SUVRs) relative to cerebellum revealed only significant differences in the hippocampus between the APPswe/PS1dE9 and wild type mice but not in cortex; this differential effect may reflect the lower plaque area in the cortex than in the hippocampus as found in the histological analysis. CONCLUSION The findings suggest that histopathological characteristics of Aβ plaque size and spatial distribution can be depicted in vivo using [18F]-florbetaben in the APPswe/PS1dE9 mouse model.

Ultrahigh field MR imaging of a subconjunctival anti-glaucoma drug delivery system in a rabbit model

Local drug delivery systems (DDS) have become a favourable approach for the treatment of numerous diseases. Biomedical imaging techniques such as ultrahigh field magnetic resonance imaging (UHF-MRI) offer unique insight into DDS biodegradation in vivo. We describe the establishment of a 7 Tesla MRI routine for longitudinal in vivo examinations of a subconjunctival DDS for the treatment of glaucoma in a rabbit model. In initial in vitro examinations the T2-relaxation times of the polymeric DDS components were assessed. Imaging of enzymatically degraded depot samples in vitro did not reveal changes in sample morphology or T2-relaxation time. Ex vivo investigations with an enucleated porcine eye showed good correlation of anatomical MRI and histological data. In longitudinal in vivo studies in rabbits, we repeatedly scanned the depot in the same animal over the course of 5 months with an in-plane resolution of 130 µm at scan times of less than 30 minutes. The degradation was quantified using volumetric analysis showing a volume reduction of 82% between 3 and 21 weeks after depot implantation. We have thereby demonstrated the feasibility of our UHF-MRI protocol as a non-invasive imaging routine for qualitative and quantitative, longitudinal evaluation of biodegradable subconjunctival DDS.