The Anti-Inflammatory Effects of Lipoxygenase and Cyclo-Oxygenase Inhibitors in Inflammation-Induced Human Fetal Glia Cells and the Aβ Degradation Capacity of Human Fetal Astrocytes in an Ex vivo Assay

Chronic inflammation is a common phenomenon present in the background of multiple neurodegenerative diseases, including Alzheimer's disease (AD). The arachidonic acid pathway overproduces proinflammatory eicosanoids during these states and glial cells in the brain gradually lose their vital functions of protecting and supporting neurons. In this study, the role of different key enzymes of the eicosanoid pathway mediating inflammatory responses was examined in vitro and ex vivo using human fetal glial cells. Astrocytes and microglia were exposed to proinflammatory agents i.e., cytokines interleukin 1-beta (IL-1 beta) and tumor necrosis factor (TNF-alpha). ELISA assays were used to examine the effects of inhibitors of key enzymes in the eicosanoid pathway. Inhibitors for 5-lipoxygenase (5-LOX) and cyclo-oxygenase 2 (COX-2) in both cell types and 5-, 12-, and 15-LOX-inhibitor in astrocytes reduced significantly IL-6 secretion, compared to exposed glial cells without inhibitors. The cytokine antibody array showed that especially treatments with 5, -12, and -15 LOX inhibitor in astrocytes, 5-LOX inhibitor in microglia and COX-2 inhibitor in both glial cell types significantly reduced the expression of multiple proinflammatory cytokines. Furthermore, human fetal astrocytes and microglia were cultured on top of AD-affected and control human brain sections for 30h. According to the immunochemical evaluation of the level of total A beta, astrocytes were very efficient at degrading A beta from AD-affected brain sections ex vivo; simultaneously added enzyme inhibitors did not increase their A beta degradation capabilities. Microglia were not able to reduce the level of total A beta during the 30h incubation time

Pancreatic metabolism, blood flow, and β-cell function in obese humans.

Context: Glucolipotoxicity is believed to induce pancreatic &beta;-cell dysfunction in obesity. Previously, it has not been possible to study pancreatic metabolism and blood flow in humans. Objective: The objective of the study was to investigate whether pancreatic metabolism and blood flow are altered in obesity using positron emission tomography (PET). In the preclinical part, the method was validated in animals. Design: This was a cross-sectional study. Setting: The study was conducted in a clinical research center. Participants: Human studies consisted of 52 morbidly obese and 25 healthy age-matched control subjects. Validation experiments were done with rodents and pigs. Interventions: PET and magnetic resonance imaging studies using a glucose analog ([18F]fluoro-2-deoxy-d-glucose), a palmitate analog [14(R,S)-[18F]fluoro-6-thia-heptadecanoic acid], and radiowater ([15O]H2O) were performed. In animals, a comparison between ex vivo and in vivo data was performed. Main Outcome Measures: Pancreatic glucose/fatty acid (FA) uptake, fat accumulation, and blood flow parameters of &beta;-cell function were measured. Results: PET proved to be a feasible method to measure pancreatic metabolism. Compared with healthy participants, obese participants had elevated pancreatic FA uptake (P &lt; .0001), more fat accumulation (P = .0001), lowered glucose uptake both during fasting and euglycemic hyperinsulinemia, and blunted blood flow (P &lt; .01) in the pancreas. Blood flow, FA uptake, and fat accumulation were negatively associated with multiple markers of &beta;-cell function. Conclusions: Obesity leads to changes in pancreatic energy metabolism with a substrate shift from glucose to FAs. In morbidly obese humans, impaired pancreatic blood flow may contribute to &beta;-cell dysfunction and in the pathogenesis of type 2 diabetes. &nbsp;</div

Ruthenium-Mediated 18F-Fluorination and Preclinical Evaluation of a New CB1 Receptor Imaging Agent [18F]FPATPP

Cannabinoid receptor 1 (CB1R) controls various physiological and pathological conditions, including memory, motivation, and inflammation, and is thus an interesting target for positron emission tomography (PET). Herein, we report a ruthenium-mediated radiolabeling synthesis and preclinical evaluation of a new CB1R specific radiotracer, [18F]FPATPP. [18F]FPATPP was produced with 16.7 ± 5.7% decay-corrected radiochemical yield and >95 GBq/μmol molar activity. The tracer showed high stability, low defluorination, and high specific binding to CB1Rs in mouse brain.</p

Monoacylglycerol lipase inhibitor JZL184 reduces neuroinflammatory response in APdE9 mice and in adult mouse glial cells

Conclusions: JZL184 decreased the proinflammatory reactions of microglia and reduced the total A beta burden and its precursors in the APdE9 mouse model. It also reduced the proinflammatory responses of microglia and astrocytes isolated from adult mice.</p

In vivo characterization of a novel norepinephrine transporter PET tracer [18F]NS12137 in adult and immature Sprague-Dawley rats

Norepinephrine modulates cognitive processes such as working and episodic memory. Pathological changes in norepinephrine and norepinephrine transporter (NET) function and degeneration of the locus coeruleus produce irreversible impairments within the whole norepinephrine system, disrupting cognitive processes. Monitoring these changes could enhance diagnostic accuracy and support development of novel therapeutic components for several neurodegenerative diseases. Thus, we aimed to develop a straightforward nucleophilic fluorination method with high molar activity for the novel NET radiotracer [18F]NS12137 and to demonstrate the ability of [18F]NS12137 to quantify changes in NET expression.Methods: We applied an 18F-radiolabeling method in which a brominated precursor was debrominated by nucleophilic 18F-fluorination in dimethyl sulfoxide. Radiolabeling was followed by a deprotection step, purification, and formulation of the radiotracer. The [18F]NS12137 brain uptake and distribution were studied with in vivo PET/CT and ex vivoautoradiography using both adult and immature Sprague-Dawley rats because postnatal NET expression peaks at 10-20 days post birth. The NET specificity for the tracer was demonstrated by pretreatment of the animals with nisoxetine, which is well-known to have a high affinity for NET.Results: [18F]NS12137 was successfully synthesized with radiochemical yields of 18.6±5.6%, radiochemical purity of >99%, and molar activity of >500 GBq/μmol at the end of synthesis. The in vivo [18F]NS12137 uptake showed peak standard uptake values (SUV) of over 1.5 (adult) and 2.2 (immature) in the different brain regions. Peak SUV/30 min and peak SUV/60 min ratios were calculated for the different brain regions of the adult and immature rats, with a peak SUV/60 min ratio of more than 4.5 in the striatum of adult rats. As expected, in vivo studies demonstrated uptake of the tracer in brain areas rich in NET, particularly thalamus, neocortex, and striatum, and remarkably also in the locus coeruleus, a quite small volume for imaging with PET. The uptake was significantly higher in immature rats compared to the adult animals. Ex vivo studies using autoradiography showed very strong specific binding in NET-rich areas such as the locus coeruleus and the bed nucleus of the stria terminalis, and high binding in larger grey matter areas such as the neocortex and striatum. The uptake of [18F]NS12137 was dramatically reduced both in vivo and ex vivo by pretreatment with nisoxetine, demonstrating the specificity of binding.Conclusions: [18F]NS12137 was synthesized in good yield and high molar activity and demonstrated the characteristics of a good radiotracer, such as good brain penetration, fast washout, and high specific binding to NET.Keywords: [18F]NS12137, norepinephrine transporter, NET, locus coeruleus, PET, nucleophilic fluorination</p

Basal and cold-induced fatty acid uptake of human brown adipose tissue is impaired in obesity

atty acids (FA) are important substrates for brown adipose tissue (BAT) metabolism, however, it remains unclear whether there exists a difference in FA metabolism of BAT between lean and obese healthy humans. In this study we evaluated supraclavicular BAT fatty acid uptake (FAU) along with blood perfusion in lean and obese subjects during cold exposure and at room temperature using positron emission tomography (PET)/computed tomography (CT). Additionally, tissue samples were taken from supraclavicular region (typical BAT region) from a subset of subjects to evaluate histological presence of BAT. Non-shivering cold stress elevated FAU and perfusion of BAT in lean, but not in obese subjects. Lean subjects had greater FAU in BAT compared to obese subjects during cold exposure and interestingly also at room temperature. The higher BAT FAU was related to younger age and several indicators of superior systemic metabolic health. The subjects who manifested BAT histologically had several folds higher BAT FAU compared to subjects with no such histological manifestation. Together, obese subjects have less active tissue in supraclavicular region both in basal and cold-activated state and the FA metabolism of BAT is blunted in obesity.</p

Kuopio birth cohort - design of a Finnish joint research effort for identification of environmental and lifestyle risk factors for the wellbeing of the mother and the newborn child

Background: A Finnish joint research effort Kuopio Birth Cohort (KuBiCo) seeks to evaluate the effects of genetics, epigenetics and different risk factors (medication, nutrition, lifestyle factors and environmental aspects) during pregnancy on the somatic and psychological health status of the mother and the child. Methods: KuBiCo will ultimately include information on 10,000 mother-child pairs who have given their informed consent to participate in this cohort. Identification of foetal health risk factors that can potentially later manifest as disease requires a repository of relevant biological samples and a flexible open up-to-date data handling system to register, store and analyse biological, clinical and questionnaire-based data. KuBiCo includes coded questionnaire-based maternal background data gathered before, during and after the pregnancy and bio-banking of maternal and foetal samples that will be stored in deep freezers. Data from the questionnaires and biological samples will be collected into one electronic database. KuBiCo consists of several work packages which are complementary to each other: Maternal, foetal and placental metabolism and omits; Paediatrics; Mental wellbeing; Prenatal period and delivery; Analgesics and anaesthetics during peripartum period; Environmental effects; Nutrition; and Research ethics. Discussion: This report describes the set-up of the KuBiCo and descriptive analysis from 3532 parturients on response frequencies and feedback to KuBiCo questionnaires gathered from June 2012 to April 2016. Additionally, we describe basic demographic data of the participants (n = 1172). Based on the comparison of demographic data between official national statistics and our descriptive analysis, KuBiCo represents a cross-section of Finnish pregnant women.Peer reviewe

Radiosynthesis and Preclinical Evaluation of an α2A-Adrenoceptor Tracer Candidate, 6-[18F]Fluoro-marsanidine

Purpose: The α2-adrenoceptors mediate many effects of norepinephrine and epinephrine, and participate in the regulation of neuronal, endocrine, cardiovascular, vegetative, and metabolic functions. Of the three receptor subtypes, only α2A and α2C are found in the brain in significant amounts. Subtype-selective positron emission tomography (PET) imaging of α2-adrenoceptors has been limited to the α2C subtype. Here, we report the synthesis of 6-[18F]fluoro-marsanidine, a subtype-selective PET tracer candidate for α2A-adrenoceptors, and its preclinical evaluation in rats and mice.Procedures: 6-[18F]Fluoro-marsanidine was synthesized using electrophilic F-18 fluorination with [18F]Selectfluor bis(triflate). The tracer was evaluated in Sprague Dawley rats and in α2A-knockout (KO) and wild-type (WT) mice for subtype selectivity. In vivo PET imaging and ex vivo brain autoradiography were performed to determine the tracer distribution in the brain. The specificity of the tracer for the target was determined by pretreatment with the subtype-non-selective α2-agonist medetomidine. The peripheral biodistribution and extent of metabolism of 6-[18F]fluoro-marsanidine were also analyzed.Results: 6-[18F]Fluoro-marsanidine was synthesized with [18F]Selectfluor bis(triflate) in a radiochemical yield of 6.4 ± 1.7 %. The molar activity was 3.1 to 26.6 GBq/μmol, and the radiochemical purity was > 99 %. In vivo studies in mice revealed lower uptake in the brains of α2A-KO mice compared to WT mice. The results for selectivity were confirmed by ex vivo brain autoradiography. Blocking studies revealed reduced uptake in α2A-adrenoceptor-rich brain regions in pretreated animals, demonstrating the specificity of the tracer. Metabolite analyses revealed very rapid metabolism of 6-[18F]fluoro-marsanidine with blood-brain barrier-permeable metabolites in both rats and mice.Conclusion: 6-[18F]Fluoro-marsanidine was synthesized and evaluated as a PET tracer candidate for brain α2A-adrenoceptors. However, rapid metabolism, extensive presence of labeled metabolites in the brain, and high non-specific uptake in mouse and rat brain make 6-[18F]fluoro-marsanidine unsuitable for α2A-adrenoceptor targeting in rodents in vivo.</p

Cessation of anti-VLA-4 therapy in a focal rat model of multiple sclerosis causes an increase in neuroinflammation

BackgroundPositron emission tomography (PET) can be used for in vivo evaluation of the pathology associated with multiple sclerosis. We investigated the use of longitudinal PET imaging and the 18-kDa translocator protein (TSPO) binding radioligand [F-18]GE-180 to detect changes in a chronic multiple sclerosis-like focal delayed-type hypersensitivity experimental autoimmune encephalomyelitis (fDTH-EAE) rat model during and after anti-VLA-4 monoclonal antibody (mAb) treatment. Thirty days after lesion activation, fDTH-EAE rats were treated with the anti-VLA-4 mAb (n=4) or a control mAb (n=4; 5mg/kg, every third day, subcutaneously) for 31days. Animals were imaged with [F-18]GE-180 on days 30, 44, 65, 86 and 142. Another group of animals (n=4) was used for visualisation the microglia with Iba-1 at day 44 after a 2-week treatment period.ResultsAfter a 2-week treatment period on day 44, there was a declining trend (p=0.067) in [F-18]GE-180-binding in the anti-VLA-4 mAb-treated animals versus controls. However, cessation of treatment for 4days after a 31-day treatment period increased [F-18]GE-180 binding in animals treated with anti-VLA-4 mAb compared to the control group (p=0.0003). There was no difference between the groups in TSPO binding by day 142.ConclusionsThese results demonstrated that cessation of anti-VLA-4 mAb treatment for 4days caused a transient rebound increase in neuroinflammation. This highlights the usefulness of serial TSPO imaging in the fDTH-EAE model to better understand the rebound phenomenon