Johanna Rokka

Detection of Microglial Activation in an Acute Model of Neuroinflammation Using PET and Radiotracers 11C-(R)-PK11195 and 18F-GE-180

It remains unclear how different translocator protein (TSPO) ligands reflect the spatial extent of astrocyte or microglial activation in various neuroinflammatory conditions. Here, we use a reproducible lipopolysaccharide (LPS)-induced model of acute central nervous system inflammation to compare the binding performance of a new TSPO ligand 18F-GE-180 with 11C-(R)-PK11195. Using immunohistochemistry, we also explore the ability of the TSPO ligands to detect activated microglial cells and astrocytes. Methods: Lewis rats (n = 30) were microinjected with LPS (1 or 10 μg) or saline (1 μL) into the left striatum. The animals were imaged in vivo at 16 h after the injection using PET radiotracers 18F-GE-180 or 11C-(R)-PK11195 (n = 3 in each group) and were killed afterward for autoradiography of the brain. Immunohistochemical assessment of OX-42 and glial fibrillary acidic protein (GFAP) was performed to identify activated microglial cells and reactive astrocytes. Results: In vivo PET imaging revealed an increase in the ipsilateral TSPO binding, compared with binding in the contralateral hemisphere, after the microinjection of 10 μg of LPS. No increase was observed with vehicle. By autoradiography, the TSPO radiotracer binding potential in the injected hemisphere was increased after striatal injection of 1 or 10 μg of LPS. However, the significant increase was observed only when using 18F-GE-180. The area of CD11b-expressing microglial cells extended beyond that of enhanced GFAP staining and mapped more closely to the extent of 18F-GE-180 binding than to 11C-(R)-PK11195 binding. The signal from either PET ligand was significantly increased in regions of increased GFAP immunoreactivity and OX-42 colocalization, meaning that the presence of both activated microglia and astrocytes in a given area leads to increased binding of the TSPO radiotracers. Conclusion: 18F-GE-180 is able to reveal sites of activated microglia in both gray and white matter. However, the signal is increased by the presence of activated astrocytes. Therefore, 18F-GE-180 is a promising new fluorinated longer-half-life tracer that reveals the presence of activated microglia in a manner that is superior to 11C-(R)-PK11195 due to the higher binding potential observed for this ligand.

In Vivo Detection of Diffuse Inflammation in Secondary Progressive Multiple Sclerosis Using PET Imaging and the Radioligand 11C-PK11195

Patients with secondary progressive multiple sclerosis (SPMS) are lacking efficient medication to slow down the progression of their disease. PET imaging holds promise as a method to study, at the molecular level and in vivo, the central nervous system pathology of SPMS. PET might thus help to elucidate potential therapeutic targets and be useful as an imaging biomarker in future treatment trials of progressive multiple sclerosis. The objective of this study was to evaluate whether translocator protein (TSPO) imaging could be used to visualize the diffuse inflammation located in the periplaque area and in the normal-appearing white matter (NAWM) in the brains of patients with SPMS. Methods: This was an imaging study using MR imaging and PET with 11C-PK11195 binding to TSPO, which is expressed in activated, but not in resting, microglia. Ten SPMS patients with a mean expanded disability status scale score of 6.3 (SD, 1.5) and eight age-matched healthy controls were studied. The imaging was performed using High-Resolution Research Tomograph PET and 1.5-T MR imaging scanners. Microglial activation was evaluated as the distribution volume ratio (DVR) of 11C-PK11195 from dynamic PET images. DVR estimations were performed with special interest in NAWM and gray matter using region-of-interest and parametric image–based approaches. Results: The DVR of 11C-PK11195 was significantly increased in the periventricular and total NAWM (P = 0.016 and P < 0.001, respectively) and in the thalamic ROIs (P = 0.027) of SPMS patients, compared with the control group. Similarly, parametric image analysis showed widespread increases of 11C-PK11195 in the white matter of SPMS patients, compared with healthy controls. Increased perilesional TSPO uptake was present in 57% of the chronic T1 lesions in MR imaging. Conclusion: The finding of increased 11C-PK11195 binding in the NAWM of SPMS patients is in line with the neuropathologic demonstration that activated microglial cells are the source of diffuse NAWM inflammation. Evaluating microglial activation with TSPO-binding PET ligands provides a unique tool to assess diffuse brain inflammation and perilesional activity in progressive multiple sclerosis in vivo.

[11C]PIB-, [18F]FDG-PET and MRI imaging in patients with Parkinson’s disease with and without dementia

The objective of this study was to identify possible group differences between PD patients with dementia and without dementia by combining different functional and structural imaging methods in vivo, which might provide an opportunity to disentangle the pathophysiological correlates of cognitive impairment and dementia in PD. We performed a neuropsychological evaluation, structural brain MRI, [(18)F]FDG PET and [(11)C]PIB PET in 19 PD patients [eight non-demented (PD), eleven demented (PDD)] and 24 healthy elderly volunteers. [(11)C]PIB region-to-cerebellum ratios did not differ significantly between the groups in any brain region (p > 0.05). PDD patients showed impaired glucose metabolism in cortical brain regions and this reduction was associated with the degree of cognitive impairment. PDD patients had more atrophy both in the hippocampus and the frontal cortex compared with PD patients and controls, and hippocampal atrophy was associated with impaired memory. This cross-sectional data suggests that development of dementia in PD is associated with extensive spread of hypometabolism beyond the occipital cortex, and with hippocampal and frontal atrophy but not beta-amyloid deposition consistent with a unique biological process related to PD rather than co-incidental development of AD in persons with PD.

Longitudinal Amyloid Imaging in Mouse Brain with 11C-PIB: Comparison of APP23, Tg2576, and APPswe-PS1dE9 Mouse Models of Alzheimer Disease

Follow-up of β-amyloid (Aβ) deposition in transgenic mouse models of Alzheimer disease (AD) would be a valuable translational tool in the preclinical evaluation of potential antiamyloid therapies. This study aimed to evaluate the ability of the clinically used PET tracer 11C-Pittsburgh compound B (11C-PIB) to detect changes over time in Aβ deposition in the brains of living mice representing the APP23, Tg2576, and APPswe-PS1dE9 transgenic mouse models of AD. Methods: Mice from each transgenic strain were imaged with 60-min dynamic PET scans at 7−9, 12, 15, and 18−22 mo of age. Regional 11C-PIB retention was quantitated as distribution volume ratios using Logan graphical analysis with cerebellar reference input, as radioactivity uptake ratios between the frontal cortex (FC) and the cerebellum (CB) during the 60-min scan, and as bound-to-free ratios in the late washout phase (40−60 min). Ex vivo autoradiography experiments were performed after the final imaging session to validate 11C-PIB binding to Aβ deposits. Additionally, the presence of Aβ deposits was evaluated in vitro using staining with thioflavin-S and Aβ1–40, Aβ1–16, and AβN3(pE) immunohistochemistry. Results: Neocortical 11C-PIB retention was markedly increased in old APP23 mice with large thioflavin-S–positive Aβ deposits. At 12 mo, the Logan distribution volume ratio for the FC was 1.03 and 0.93 (n = 2), increasing to 1.38 ± 0.03 (n = 3) and 1.34 (n = 1) at 18 and 21 mo of age, respectively. An increase was also observed in bound-to-free ratios for the FC between young (7- to 12-mo-old) and old (15- to 22-mo-old) APP23 mice. Binding of 11C-PIB to Aβ-rich cortical regions was also evident in ex vivo autoradiograms of APP23 brain sections. In contrast, no increases in 11C-PIB retention were observed in aging Tg2576 or APPswe-PS1dE9 mice in vivo, although in the latter, extensive Aβ deposition was already observed at 9 mo of age with immunohistochemistry. Conclusion: The results suggest that 11C-PIB binding to Aβ deposits in transgenic mouse brain is highly dependent on the AD model and the structure of its Aβ plaques. Longitudinal in vivo 11C-PIB uptake studies are possible in APP23 mice.

Autoradiographic characterization of α2C‐adrenoceptors in the human striatum

Indirect experimental evidence suggests that drugs acting on the α2C‐adrenoceptor could be useful in the treatment of neuropsychiatric disorders such as depression and schizophrenia. In rodent brain, the highest levels of α2C‐adrenoceptors are found in the striatum, with lower levels in cerebral cortex and hippocampus. In human brain, because of the poor subtype‐selectivity of the available α2‐adrenoceptor ligands, the localization of α2C‐adrenoceptors has remained unknown. Recently, a selective α2C‐adrenoceptor antagonist, JP‐1302, was characterized, and to assess the presence of α2C‐adrenoceptors in human brain, we performed competition binding in vitro receptor autoradiography with JP‐1302 and the α2‐adrenoceptor subtype nonselective antagonist [ethyl‐3H]RS79948‐197 on rat and human postmortem brain sections. In striatum of both species, JP‐1302 vs. [ethyl‐3H]RS79948‐197 competition binding was biphasic, identifying high‐ and low‐affinity binding sites, whereas in cortex and cerebellum, only low‐affinity binding sites were detected. The results indicate that a significant portion of the α2‐adrenoceptors in striatum is of the α2C subtype, whereas non‐α2C‐adreocneptors predominate in cortex and cerebellum. Because the α2C‐adrenoceptor subtype distribution pattern appears to be conserved between rodents and humans, results obtained from studies on the role of the α2C‐adrenoceptor in rodent models of neuropsychiatric disorders may be relevant also for human diseases. Synapse 62:508–515, 2008.

Evaluation of the effect of fingolimod treatment on microglial activation using serial PET imaging in multiple sclerosis

Traditionally, multiple sclerosis (MS) has been considered a white matter disease with focal inflammatory lesions. It is, however, becoming clear that significant pathology, such as microglial activation, also takes place outside the plaque areas, that is, in areas of normal-appearing white matter (NAWM) and gray matter (GM). Microglial activation can be detected in vivo using 18-kDa translocator protein (TSPO)–binding radioligands and PET. It is unknown whether fingolimod affects microglial activation in MS. The aim of this study was to investigate whether serial PET can be used to evaluate the effect of fingolimod treatment on microglial activation. Methods: Ten relapsing-remitting MS patients were studied using the TSPO radioligand 11C-(R)-PK11195. Imaging was performed at baseline and after 8 and 24 wk of fingolimod treatment. Eight healthy individuals were imaged for comparison. Microglial activation was evaluated as distribution volume ratio of 11C‐(R)-PK11195. Results: The patients had MS for an average of 7.9 ± 4.3 y (mean ± SD), their total relapses averaged 4 ± 2.4, and their Expanded Disability Status Scale was 2.7 ± 0.5. The patients were switched to fingolimod because of safety reasons or therapy escalation. The mean washout period before the initiation of fingolimod was 2.3 ± 1.1 mo. The patients were clinically stable on fingolimod. At baseline, microglial activation was significantly higher in the combined NAWM and GM areas of MS patients than in healthy controls (P = 0.021). 11C‐(R)-PK11195 binding was reduced (−12.31%) within the combined T2 lesion area after 6 mo of fingolimod treatment (P = 0.040) but not in the areas of NAWM or GM. Conclusion: Fingolimod treatment reduced microglial/macrophage activation at the site of focal inflammatory lesions, presumably by preventing leukocyte trafficking from the periphery. It did not affect the widespread, diffuse microglial activation in the NAWM and GM. The study opens new vistas for designing future therapeutic studies in MS that use the evaluation of microglial activation as an imaging outcome measure.

Synthesis and evaluation of a [18F]-curcumin derivate for β-amyloid plaque imaging

INTRODUCTION Curcumin is a neuroprotective compound that inhibits the formation of amyloid oligomers and fibrils and binds to β-amyloid plaques in Alzheimers disease (AD). We aimed to synthesize an (18)F-labeled curcumin derivate ([(18)F]4) and to characterize its positron emission tomography (PET) tracer-binding properties to β-amyloid plaques in a transgenic APP23 mouse model of AD. METHODS We utilized facile one-pot synthesis of [(18)F]4 using nucleophilic (18)F-fluorination and click chemistry. Binding of [(18)F]4 to β-amyloid plaques in the transgenic APP23 mouse brain cryosections was studied in vitro using heterologous competitive binding against PIB. [(18)F]4 uptake was studied ex vivo in rodents and in vivo using PET/computed tomography of transgenic APP23 and wild-type control mice. RESULTS The radiochemical yield of [(18)F]4 was 21 ± 11%, the specific activity exceeded 1TBq/μmol, and the radiochemical purity exceeded 99.3% at the end of synthesis. In vitro studies of [(18)F]4 with the transgenic APP23 mouse revealed high β-amyloid plaque binding. In vivo and ex vivo studies demonstrated that [(18)F]4 has fast clearance from the blood, moderate metabolism but low blood-brain barrier (BBB) penetration. CONCLUSIONS [(18)F]4 was synthesized in high yield and excellent quality. In vitro studies, metabolite profile, and fast clearance from the blood indicated a promising tracer for Aβ imaging. However, [(18)F]4 has low in vivo BBB penetration and thus further studies are needed to reveal the reason for this and to possibly overcome this issue.

Cortical 11C-PIB Uptake is Associated with Age, APOE Genotype, and Gender in "Healthy Aging"

BACKGROUND/OBJECTIVE Our aim was to elucidate factors that contribute to amyloid-β (Aβ) accumulation in the brains of the seemingly healthy elderly population, and whether there is interplay between those factors. METHODS We conducted a cross-sectional positron emission tomography (PET) study with the amyloid tracer 11C-PIB, in 64 cognitively healthy subjects (54-89 years). In addition to PET, magnetic resonance imaging, neuropsychological testing, and APOE genotyping was performed. The results were assessed with a statistical general linear model as well as with Statistical Parametric Mapping (SPM). RESULTS The effects of age (p < 0.001), APOE ε4 carrier status (p = 0.003), and gender (p = 0.001) on composite cortical 11C-PIB uptake were all significant. The effect of educational level was non-significant (p = 0.37). No significant interactions were found between any of the factors. Cortical 11C-PIB uptake increased, on the average, by 0.015 cortex/cerebellar cortical ratio unit, with every year of age. APOE ε4 positive subjects exhibited higher cortical 11C-PIB uptake than APOE ε4 negative subjects (unadjusted means 1.49 ± 0.34 versus 1.29 ± 0.26) and males had higher uptake than females (1.49 ± 0.39 versus 1.29 ± 0.22), irrespective of age. The results of the voxel-based (SPM) analysis were similar. In addition, SPM analysis showed that lower CERAD score was associated with higher 11C-PIB uptake in the frontal cortex. CONCLUSIONS Age and APOE ε4 genotype were associated with higher 11C-PIB uptake. In this sample of cognitively healthy elderly individuals, men exhibited higher 11C-PIB uptake than women. Possible gender differences in Aβ accumulation have not been addressed in detail in previous studies, and deeper evaluation in the future is warranted.

Cardiac remodeling in a new pig model of chronic heart failure: Assessment of left ventricular functional, metabolic, and structural changes using PET, CT, and echocardiography

AimsLarge animal models are needed to study disease mechanisms in heart failure (HF). In the present study we characterized the functional, metabolic, and structural changes of myocardium in a novel pig model of chronic myocardial infarction (MI) by using multimodality imaging and histology.Methods and ResultsMale farm pigs underwent a two-step occlusion of the left anterior descending coronary artery with concurrent distal ligation and implantation of a proximal ameroid constrictor (HF group), or sham operation (control group). Three months after the operation, cardiac output and wall stress were measured by echocardiography. Left ventricle (LV) volumes and mass were measured by computed tomography (CT). Myocardial perfusion was evaluated by [15O]water and oxygen consumption using [11C]acetate positron emission tomography, and the efficiency of myocardial work was calculated. Histological examinations were conducted to detect MI, hypertrophy, and fibrosis. Animals in the HF group had a large anterior MI scar. CT showed larger LV diastolic volume and lower ejection fraction in HF pigs than in controls. Perfusion and oxygen consumption in the remote non-infarcted myocardium were preserved in HF pigs as compared to controls. Global LV work and efficiency were significantly lower in HF than control pigs and was associated with increased wall stress. Histology showed myocyte hypertrophy but not increased interstitial fibrosis in the remote segments in HF pigs.ConclusionsThe chronic post-infarction model of HF is suitable for studies aimed to evaluate LV remodeling and changes in oxidative metabolism and can be useful for testing new therapies for HF.

HPLC and TLC methods for analysis of [(18)F]FDG and its metabolites from biological samples.

The most used positron emission tomography (PET) tracer, 2-[18F]fluoro-2-deoxy-d-glucose ([18F]FDG), is a glucose analogue that is used to measure tissue glucose consumption. Traditionally, the Sokoloff model is the basis for [18F]FDG modeling. According to this model, [18F]FDG is expected to be trapped in a cell in the form of [18F]FDG-6-phosphate ([18F]FDG-6-P). However, several studies have shown that in tissues, [18F]FDG metabolism goes beyond [18F]FDG-6-P. Our aim was to develop radioHPLC and radioTLC methods for analysis of [18F]FDG metabolites from tissue samples. The radioHPLC method uses a sensitive on-line scintillation detector to detect radioactivity, and the radioTLC method employs digital autoradiography to detect the radioactivity distribution on a TLC plate. The HPLC and TLC methods were developed using enzymatically in vitro-produced metabolites of [18F]FDG as reference standards. For this purpose, three [18F]FDG metabolites were synthesized: [18F]FDG-6-P, [18F]FD-PGL, and [18F]FDG-1,6-P2. The two methods were evaluated by analyzing the [18F]FDG metabolic profile from rodent ex vivo tissue homogenates. The HPLC method with an on-line scintillation detector had a wide linearity in a range of 5Bq-5kBq (LOD 46Bq, LOQ 139Bq) and a good resolution (Rs ≥1.9), and separated [18F]FDG and its metabolites clearly. The TLC method combined with digital autoradiography had a high sensitivity in a wide range of radioactivity (0.1Bq-2kBq, LOD 0.24Bq, LOQ 0.31Bq), and multiple samples could be analyzed simultaneously. As our test and the method validation with ex vivo samples showed, both methods are useful, and at best they complement each other in analysis of [18F]FDG and its radioactive metabolites from biological samples.