Molecular Imaging and Biology

To image inflammation and monitor therapeutic response to anti-inflammatory intervention using 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) positron emission tomography (PET) in a preclinical model of acute lung injury (ALI). Mice were intratracheally administered lipopolysaccharide (LPS, 2.5 mg/kg) to induce ALI or phosphate-buffered saline as the vehicle control. A subset of mice in the ALI group received two intraperitoneal doses of dexamethasone 1 and 24 h after LPS. [18F]FDG PET/CT was performed 2 days after the induction of ALI. [18F]FDG uptake in the lungs was quantified by PET (%ID/mLmean and standardized uptake value (SUVmean)) and ex vivo γ-counting (%ID/g). The severity of lung inflammation was determined by quantifying the protein level of inflammatory cytokines/chemokines and the activity of neutrophil elastase and glycolytic enzymes. In separate groups of mice, flow cytometry was performed to estimate the contribution of individual immune cell types to the total pulmonary inflammatory cell burden under different treatment conditions. Lung uptake of [18F]FDG was significantly increased during LPS-induced ALI, and a decreased [18F]FDG uptake was observed following dexamethasone treatment to an intermediate level between that of LPS-treated and control mice. Protein expression of inflammatory biomarkers and the activity of neutrophil elastase and glycolytic enzymes were increased in the lungs of LPS-treated mice versus those of control mice, and correlated with [18F]FDG uptake. Furthermore, dexamethasone-induced decreases in cytokine/chemokine protein levels and enzyme activities correlated with [18F]FDG uptake. Neutrophils were the most abundant cells in LPS-induced ALI, and the pattern of total cell burden during ALI with or without dexamethasone therapy mirrored that of [18F]FDG uptake. [18F]FDG PET noninvasively detects lung inflammation in ALI and its response to anti-inflammatory therapy in a preclinical model. However, high [18F]FDG uptake by bone, brown fat, and myocardium remains a technical limitation for quantification of [18F]FDG in the lungs.

Prenatal infection during pregnancy is a risk factor for schizophrenia, as well as for other developmental psychiatric disorders, such as autism and bipolar disorder. Schizophrenia patients were reported to have altered brain metabolism and neuroinflammation. However, the link between prenatal infection, altered brain inflammation and metabolism, and schizophrenia remains unclear. In this project, we aimed to evaluate whether there are changes in brain glucose consumption and microglia activation in the offspring of pregnant rats exposed to maternal immune activation (MIA), and if so, whether these changes occur before or after the initiation of schizophrenia-like behaviour. Pregnant rats were treated with the viral mimic polyinosinic-polycytidylic acid (MIA group) or saline (control group) on gestational day 15. Static PET scans of the male offspring were acquired on postnatal day (PND) 21, 60, and 90, using [11C]-PK11195 and deoxy-2-[18F]fluoro-D-glucose ([18F]-FDG) as tracers to measure TSPO expression in activated microglia and brain glucose consumption, respectively. On PND60 and PND90, anxiety-like behaviour, recognition memory, and sensorimotor gating were measured using the open field test (OFT), novel object recognition test (NOR), and prepulse inhibition test (PPI). [18F]-FDG PET demonstrated that MIA offspring displayed higher brain glucose consumption in the whole brain after weaning (p = 0.017), and in the frontal cortex during late adolescence (p = 0.001) and adulthood (p = 0.037) than control rats. [11C]-PK11195 PET did not reveal any changes in TSPO expression in MIA offspring. Prenatal infection induced age-related behavioural alterations. Adolescent MIA offspring displayed a more anxious state in the OFT than controls (p = 0.042). Adult MIA offspring showed recognition memory deficits in the NOR (p = 0.003). Our study did not show any PPI deficits. Our results suggest that prenatal immune activation changed neurodevelopment, resulting in increased brain glucose consumption, but not in microglia activation. The increased brain glucose consumption in the frontal cortex of MIA offspring remained until adulthood and was associated with increased anxiety-like behaviour during adolescence and recognition memory deficits in adulthood.

[18F]AZD4694 (2-(2-18F-fluoro-6-(methylamino)-3-pyridyl)benzofuran-5-ol) is a radioligand suitable for imaging of amyloid beta deposits in the living human brain using positron emission tomography (PET). Here, we report the preparation and pharmacokinetic profile of its carbon-11 (t 1/2  = 20.4 min) labeled isotopolog [11C]AZD4694 and compare [11C]AZD4694 with the hitherto most widely applied amyloid PET radioligand [11C]Pittsburgh Compound B (PiB). The immediate unlabeled precursor to [11C]AZD4694 was prepared in a four-step convergent synthesis. Subsequent N-11C-methylation of this precursor with [11C]methyl iodide yielded [11C]AZD4694, which after isolation and formulation was injected into cynomolgus monkeys. The radioactivity in nonhuman primate brain following injection of [11C]AZD4694 and [11C]PiB was measured using PET. [11C]AZD4694 was prepared in a 60 % incorporation yield. In a head to head comparison with [11C]PiB, it appeared that [11C]AZD4694 displayed slightly lower nonspecific binding in white matter than [11C]PiB as well as more rapid pharmacokinetics in the brain. The advantageous pharmacokinetic profile and low nonspecific binding render [11C]AZD4694 a promising PET radioligand for imaging of amyloid beta in the human brain with PET.

The objective of this study was to evaluate the use of molecular ultrasound (US) imaging for monitoring the early inflammatory effects following acute kidney injury. A population of rats underwent 30 min of renal ischemia (acute kidney injury, N = 6) or sham injury (N = 4) using established surgical methods. Animals were divided and molecular US imaging was performed during the bolus injection of a targeted microbubble (MB) contrast agent to either P-selectin or vascular cell adhesion molecule 1 (VCAM-1). Imaging was performed before surgery and 4 and 24 h thereafter. After manual segmentation of renal tissue space, the molecular US signal was calculated as the difference between time-intensity curve data before MB injection and after reaching steady-state US image enhancement. All animals were terminated after the 24 h imaging time point and kidneys excised for immunohistochemical (IHC) analysis. Renal inflammation was analyzed using molecular US imaging. While results using the P-selectin and VCAM-1 targeted MBs were comparable, it appears that the former was more sensitive to biomarker expression. All molecular US imaging measures had a positive correlation with IHC findings. Acute kidney injury is a serious disease in need of improved noninvasive methods to help diagnose the extent of injury and monitor the tissue throughout disease progression. Molecular US imaging appears well suited to address this challenge and more research is warranted.

Noninvasive positron emission tomography (PET) imaging of reporter gene is combined with quantitative real-time polymerase reverse transcription (RT-PCR) method to study the time course of death and proliferation of stem cells transplanted in the myocardium. Male murine embryonic stem cells (ESCs) were stably transfected with a mutant version of herpes simplex virus type 1 thymidine kinase (HSV1-sr39tk) reporter gene; 5 × 106 such cells were injected into the myocardium of female athymic rats. While the transplanted cells was monitored by in vivo 9-(4-[F-18]fluoro-3-hydroxymethylbutyl)guanine ([F-18]FHBG) PET imaging of the heart, their absolute number was estimated by RT-PCR from hearts harvested at 3–5 h, 24 h, days 4, 7, and 14 after transplantation. (1) Forty percent of injected cells were retained in the heart while majority of injected cells were lost within a few hours after injection. Cell death was peaked at 24 h when 18% of donor cells retained in the heart were dead. (2) The substantial cell loss was reversed by significant proliferation of ESCs. This led to the recovery of cell number to 3.4 million (70% of injected dose) at day 4 and first visual observation of in vivo [F-18] signal in the heart. (3) A robust correlation (R 2 = 0.9) between percent of injected dose per gram of tissue derived from in vivo PET signal and the number of donor cells estimated by RT-PCR was revealed. The time course of transplanted stem cells surviving in the heart reveals a process of substantial cell loss within 24 h of injection and subsequent recovery of cell number through proliferation. Such proliferation can be noninvasively monitored by reporter gene imaging.

Ultrasound-induced cavitation facilitates cellular uptake of drugs via increased membrane permeability. Here, the purpose was to evaluate the duration of enhanced membrane permeability following ultrasound treatment in cell culture. Optical chromophores with fluorescence intensity increasing 100–1,000-fold upon intercalation with nucleic acids served as smart agents for reporting cellular uptake. Opticell chambers with a monolayer of C6 cells were subjected to ultrasound in the presence of microbubbles followed by varying delays between 0 and 24 h before addition of Sytox Green optical contrast agent. Micro- and macroscopic fluorescence were used for qualitative and quantitative analysis. Up to 25% of viable cells showed uptake of contrast agent with a half time of 8 h, with cellular uptake persisting even at 24 h. Only cells exposed to ultrasound showed the effect. The temporal window of increased membrane permeability is much longer in these studies than previously suggested. This may have important repercussions for in vivo studies in which membrane permeability may be temporally separated from drug delivery.

SimPET/M7 system is a small-animal dedicated simultaneous positron emission tomography and magnetic resonance imaging (PET/MRI) scanner. The SimPET insert has been upgraded from its prototype with a focus on count rate performance and sensitivity. The M7 scanner is a 1-T permanent magnet-based compact MRI system without any cryogens. Here, we present performance evaluation results of SimPET along with the results of mutual interference evaluation and simultaneously acquired PET/MR imaging. Following NEMA NU 4-2008 standard, we evaluated the performance of the SimPET system. The M7 MRI compatibility of SimPET was also assessed by analyzing MRI images of a uniform phantom under different PET conditions and PET count rates with different MRI pulse sequences. Mouse imaging was performed including a whole-body 18F-NaF PET scan and a simultaneous PET/MRI scan with 64Cu-NOTA-ironoxide. The spatial resolution at center based on 3D OSEM without and with warm background was 0.7 mm and 1.45 mm, respectively. Peak sensitivity was 4.21 % (energy window = 250–750 keV). The peak noise equivalent count rate with the same energy window was 151 kcps at 38.4 MBq. The uniformity was 4.42 %, and the spillover ratios in water- and air-filled chambers were 14.6 % and 12.7 %, respectively. In the hot rod phantom image, 0.75-mm-diameter rods were distinguishable. There were no remarkable differences in the SNR and uniformity of MRI images and PET count rates with different PET conditions and MRI pulse sequences. In the whole-body 18F-NaF PET images, fine skeletal structures were well resolved. In the simultaneous PET/MRI study with 64Cu-NOTA-ironoxide, both PET and MRI signals changed before and after injection of the dual-modal imaging probe, which was evident with the exact spatiotemporal correlation. We demonstrated that the SimPET scanner has a high count rate performance and excellent spatial resolution. The combined SimPET/M7 enabled simultaneous PET/MR imaging studies with no remarkable mutual interference between the two imaging modalities.