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Objective: Lyme borreliosis (LB) is a tick-borne infectious disease caused by Borrelia burgdorferi spirochaetes, which are able to disseminate from the tick-bite site to distant organs. Mouse models are widely used to study LB and especially Lyme arthritis (LA), but only a few whole-animal in vivo imaging studies on the pathogenesis of B. burgdorferi infection in mice have been published so far. The existing imaging techniques have their drawbacks and, therefore, novel tools to complement the array of available LB imaging methodologies are needed.Method: The applicability of positron emission tomography combined with computed tomography (PET/CT) imaging was evaluated as a method to monitor LB and especially LA in the C3H/HeN mouse model infected with wild-type B. burgdorferi N40 bacteria. The imaging results were compared with the traditional LA analysis methods, such as tibiotarsal joint swelling and histopathological assessment of joint inflammation.Results: PET/CT imaging provided high-resolution images with quantitative information on the spatial and temporal distribution of the [18F]fluorodeoxyglucose ([18F]FDG) tracer in B. burgdorferi-infected mice. The [18F]FDG accumulated in the affected joints and activated lymph nodes of infected mice, while the tracer signal could not be visualized in these organs in uninfected control animals. Importantly, in vivo PET/CT imaging data were in agreement with the histopathological scoring of inflammation of mouse joints.Conclusion: PET/CT imaging with [18F]FDG is a reliable method to longitudinally monitor the development and progression of B. burgdorferi infection-induced inflammation in vivo in mouse joints. [ABSTRACT FROM AUTHOR]

Purpose: Positron emission tomography (PET) with ¹⁵O-labeled water (H₂ ¹⁵O) facilitates the visualization and quantification of blood flow in clinical investigations and also in small animals. The quantification of blood flow requires an input function, which is generally obtained by measuring radioactivity in arterial blood withdrawn during PET scanning. However, this approach is not always feasible, because abundant blood sampling may affect the physiological process being measured. The purpose of the present study was to develop and cross-validate two methods, namely, a blood- and an image-based method for obtaining the input function for blood flow studies from rat H₂ ¹⁵O PET.Methods: The study material consisted of two separate groups of rats. Group 1 rats were imaged twice by a high-resolution research tomograph PET camera at resting condition for a test-retest study (n = 4), and group 2 rats were imaged with and without adenosine infusion for a rest-stress study (n = 4). In group 1, radioactivity concentration in arterial blood was measured with a new flow-through detector during imaging and a blood-based input function was obtained. The image-based input function was estimated using time-activity curves from the left ventricle and myocardial regions. To validate the two input function methods, myocardial blood flow (MBF) and cerebral blood flow (CBF) were computed, and the methods were tested for reproducibility (test-retest study) and changes (rest-stress study).Results: The blood- and image-based input functions were similar, and the corresponding CBF values differed only by -6.9 ± 8.1%. In the test-retest study, both MBF and CBF showed good reproducibility, and in the rest-stress study, adenosine significantly increased both MBF (P = 0.035) and CBF (P = 0.029), compared with the resting condition.Conclusion: It is possible both to measure the input function from rat arteria femoralis during H₂ ¹⁵O PET imaging and to estimate the input function from rat H₂ ¹⁵O PET images, thereby facilitating the assessment of blood flow in organs visible in PET images. [ABSTRACT FROM AUTHOR]

Fatty acid binding protein 3 (FABP3) is expressed both in tumor cells and in the tumor vasculature, making it a potential target for medical imaging and therapy. In this study, we aimed to radiolabel a CooP peptide with a free amino and thiol group, and evaluate the radiolabeled product [ 18 F]FNA- N -CooP for imaging FABP3 expression in breast cancer brain metastases by positron emission tomography. [ 18 F]FNA- N -CooP was prepared by highly chemoselective N -acylation and characterized using different chemical approaches. We validated its binding to the target using in vitro tissue section autoradiography and performed stability tests in vitro and in vivo. [ 18 F]FNA- N -CooP was successfully synthesized in 16.8% decay-corrected radiochemical yield with high radiochemical purity (98.5%). It exhibited heterogeneous binding on brain metastasis tissue sections from a patient with breast cancer, with foci of radioactivity binding corresponding to FABP3 positivity. Furthermore, the tracer binding was reduced by 55% in the presence of nonradioactive FNA- N -CooP a blocker, indicating specific tracer binding and that FABP3 is a viable target for [ 18 F]FNA- N -CooP. Favorably, the tracer did not bind to necrotic tumor tissue. However, [ 18 F]FNA- N -CooP displayed limited stability both in vitro in mouse plasma or human serum and in vivo in mouse, therefore further studies are needed to improve the stability [ 18 F]FNA- N -CooP to be used for in vivo applications.

Purpose: Aluminum fluoride-18-labeled 1,4,7-triazacyclononane-1,4,7-triacetic acid-conjugated mannosylated dextran derivative (Al[ 18 F]F-NOTA-D10CM) is a new tracer for PET imaging. We report here on in vitro and in vivo validation of the tracer's ability to target the macrophage mannose receptor CD206., Methods: First, the uptake of intravenously (i.v.) administered Al[ 18 F]F-NOTA-D10CM was compared between wild-type (WT) and CD206 -/- knockout (KO) mice. C57BL/6N mice were injected with complete Freund's adjuvant (CFA) in the left hind leg and the uptake of Al[ 18 F]F-NOTA-D10CM after i.v. or intradermal (i.d.) injection was studied at 5 and 14 days after CFA induction of inflammation. Healthy C57BL/6N mice were studied as controls. Mice underwent PET/CT on consecutive days with [ 18 F]FDG, i.v. Al[ 18 F]F-NOTA-D10CM, and i.d. Al[ 18 F]F-NOTA-D10CM. After the last imaging, Al[ 18 F]F-NOTA-D10CM was i.v. injected for an ex vivo biodistribution study and autoradiography of inflamed tissues. Blood plasma samples were analyzed using high-performance liquid chromatography. To evaluate the specificity of Al[ 18 F]F-NOTA-D10CM binding, an in vitro competitive displacement study was performed on inflamed tissue sections using autoradiography. CD206 expression was assessed by immunohistochemical staining., Results: Compared with WT mice, the uptake of Al[ 18 F]F-NOTA-D10CM was significantly lower in several CD206 -/- KO mice tissues, including liver (SUV 8.21 ± 2.51 vs. 1.06 ± 0.16, P < 0.001) and bone marrow (SUV 1.63 ± 0.37 vs. 0.22 ± 0.05, P < 0.0001). The uptake of i.v. injected Al[ 18 F]F-NOTA-D10CM was significantly higher in inflamed ankle joint (SUV 0.48 ± 0.13 vs. 0.18 ± 0.05, P < 0.0001) and inflamed foot pad skin (SUV 0.41 ± 0.10 vs. 0.04 ± 0.01, P < 0.0001) than in the corresponding tissues in healthy mice. The i.d.-injected Al[ 18 F]F-NOTA-D10CM revealed differences between CFA-induced lymph node activation and lymph nodes in healthy mice. Ex vivo γ-counting, autoradiography, and immunohistochemistry supported the results, and a decrease of ~ 80% in the binding of Al[ 18 F]F-NOTA-D10CM in the displacement study with excess NOTA-D10CM confirmed that tracer binding was specific. At 60 min after i.v. injection, an average 96.70% of plasma radioactivity was derived from intact Al[ 18 F]F-NOTA-D10CM, indicating good in vivo stability. The uptake of Al[ 18 F]F-NOTA-D10CM into inflamed tissues was positively associated with the area percentage of CD206-positive staining., Conclusion: The uptake of mannosylated dextran derivative Al[ 18 F]F-NOTA-D10CM correlated with CD206 expression and the tracer appears promising for inflammation imaging., (© 2024. The Author(s).)

Purpose: Inflammatory bowel disease (IBD) can be imaged with positron emission tomography (PET), but existing PET radiopharmaceuticals have limited diagnostic accuracy. Vascular adhesion protein-1 (VAP-1) is an endothelial cell surface molecule that controls leukocyte extravasation into sites of inflammation. However, the role of inflammation-induced VAP-1 expression in IBD is still unclear. Therefore, this study investigated the utility of VAP-1-targeted [ 68 Ga]Ga-DOTA-Siglec-9 positron emission tomography/computed tomography (PET/CT) for assessing inflammation in two mouse models of IBD., Procedures: Studies were performed using K8 -/- mice that develop a chronic colitis-phenotype and C57Bl/6NCrl mice with acute intestinal inflammation chemically-induced using 2.5% dextran sodium sulfate (DSS) in drinking water. In both diseased and control mice, uptake of the VAP-1-targeting peptide [ 68 Ga]Ga-DOTA-Siglec-9 was assessed in intestinal regions of interest using in vivo PET/CT, after which ex vivo gamma counting, digital autoradiography, and histopathological analyses were performed. Immunofluorescence staining was performed to determine VAP-1-expression in the intestine, including in samples from patients with ulcerative colitis., Results: Intestinal inflammation could be visualized by [ 68 Ga]Ga-DOTA-Siglec-9 PET/CT in two murine models of IBD. In both models, the in vivo PET/CT and ex vivo studies of [ 68 Ga]Ga-DOTA-Siglec-9 uptake were significantly higher than in control mice. The in vivo uptake was increased on average 1.4-fold in the DSS model and 2.0-fold in the K8 -/- model. Immunofluorescence staining revealed strong expression of VAP-1 in the inflamed intestines of both mice and patients., Conclusions: This study suggests that the VAP-1-targeting [ 68 Ga]Ga-DOTA-Siglec-9 PET tracer is a promising tool for non-invasive imaging of intestinal inflammation. Future studies in patients with IBD and evaluation of the potential value of [ 68 Ga]Ga-DOTA-Siglec-9 in diagnosis and monitoring of the disease are warranted., (© 2023. The Author(s).)

Background: Vascular adhesion protein-1 (VAP-1) is an adhesion molecule and primary amine oxidase, and Gallium-68-labeled 1,4,7,10-tetraazacyclododecane-N,N',N″,N‴-tetra-acetic acid conjugated sialic acid-binding immunoglobulin-like lectin 9 motif containing peptide ([ 68 Ga]Ga-DOTA-Siglec-9) is a positron emission tomography (PET) tracer targeting VAP-1. We evaluated the feasibility of PET imaging with [ 68 Ga]Ga-DOTA-Siglec-9 for the detection of myocardial lesions in rats with autoimmune myocarditis., Methods: Rats (n = 9) were immunized twice with porcine cardiac myosin in complete Freund's adjuvant. Control rats (n = 6) were injected with Freund's adjuvant alone. On day 21, in vivo PET/computed tomography (CT) imaging with [ 68 Ga]Ga-DOTA-Siglec-9 was performed, followed by ex vivo autoradiography, histology, and immunohistochemistry of tissue sections. In addition, myocardial samples from three patients with cardiac sarcoidosis were studied., Results: [ 68 Ga]Ga-DOTA-Siglec-9 PET/CT images of immunized rats showed higher uptake in myocardial lesions than in myocardium outside lesions (SUV mean , 0.5 ± 0.1 vs 0.3 ± 0.1; P = .003) or control rats (SUV mean , 0.2 ± 0.03; P < .0001), which was confirmed by ex vivo autoradiography of tissue sections. Immunohistochemistry showed VAP-1-positive staining in lesions of rats with myocarditis and in patients with cardiac sarcoidosis., Conclusion: VAP-1-targeted [ 68 Ga]Ga-DOTA-Siglec-9 PET is a potential novel technique for the detection of myocardial lesions., (© 2023. The Author(s).)

Purpose: In addition to being expressed on liver sinusoidal endothelial cells, mannose receptors are also found on antigen-presenting cells, including macrophages, which are mainly involved in the inflammation process. Dextran derivatives of various sizes containing cysteine and mannose moieties have previously been labeled with 99m Tc and used for single-photon emission computed tomography imaging of sentinel lymph nodes. In this study, we radiolabeled 21.3-kDa D10CM with positron-emitting 18 F for initial positron emission tomography (PET) studies in rats., Procedures: D10CM was conjugated with 1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA) chelator and radiolabeled with the aluminum fluoride-18 method. The whole-body distribution kinetics and stability of the intravenously administered tracer were studied in healthy male Sprague-Dawley rats by in vivo PET/CT imaging, ex vivo gamma counting, and high-performance liquid chromatography analysis., Results: Al[ 18 F]F-NOTA-D10CM was obtained with a radiochemical purity of >99% and molar activity of 9.9 GBq/μmol. At 60 minutes after injection, an average of 84% of the intact tracer was found in the blood, indicating excellent in vivo stability. The highest radioactivity concentration was seen in the liver, spleen, and bone marrow, in which mannose receptors are highly expressed under physiological conditions. The uptake specificity was confirmed with in vivo blocking experiments., Conclusions: Our results imply that Al[ 18 F]F-NOTA-D10CM is a suitable tracer for PET imaging. Further studies in disease models with mannose receptor CD206-positive macrophages are warranted to clarify the tracer's potential for imaging of inflammation., (© 2023. The Author(s).)

Pretargeted concept in positron emission tomography (PET) together with bioorthogonal chemistry is an elegant solution to study processes with slow pharmacokinetics by utilizing radiotracers labeled with short-lived radionuclides. Namely, radiotracers based on tetrazine ligation with trans -cyclooctene (TCO) via the inverse electron demand Diels-Alder (IEDDA) reaction have become a state-of-the-art for the pretargeted PET imaging. For radiolabeling of tetrazine scaffolds, indirect radiofluorination methods are often preferred, as tetrazines are vulnerable to harsh conditions typically necessary for the direct radiofluorination. 18 F-Fluoroglycosylation is an indirect radiofluorination method, which allows the introduction of a widely accessible glucose analog 2-[ 18 F]fluoro-2-deoxy-d-glucose ([ 18 F]FDG) to aminooxy-functionalized precursors via oxime formation. Here, we report the biological evaluation of [ 18 F]FDG-Tz as a tracer for pretargeted PET imaging of TCO-functionalized molecular spherical nucleic acids (MSNA) against human epidermal growth factor receptor 2 (HER2) mRNA. The oxime ether formation between [ 18 F]FDG and tetrazine oxyamine resulted in [ 18 F]FDG-Tz with high radiochemical purity (>99%) and moderate yields (6.5 ± 3.6%, n = 5). Biological evaluation of [ 18 F]FDG-Tz in healthy mice indicated favorable pharmacokinetics with quick blood clearance, urinary excretion as the main elimination route, and the absence of GLUT1 transportation. The successful pretargeted experiments with TCO-functionalized MSNA revealed higher tumor uptake compared to preclicked MSNA in HER2-expressing human breast cancer xenograft-bearing mice., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

18 F-Labeled [60]fullerene-based molecular spherical nucleic acids (MSNAs), consisting of a human epidermal growth factor receptor 2 (HER2) mRNA antisense oligonucleotide sequence with a native phosphodiester and phosphorothioate backbone, were synthesized, site-specifically labeled with a positron emitting fluorine-18 and intravenously administrated via tail vein to HER2 expressing HCC1954 tumor-bearing mice. The biodistribution of the MSNAs was monitored in vivo by positron emission tomography/computed tomography (PET/CT) imaging. MSNA with a native phosphodiester backbone (MSNA-PO) was prone to rapid nuclease-mediated degradation, whereas the corresponding phosphorothioate analogue (MSNA-PS) with improved enzymatic stability showed an interesting biodistribution profile in vivo . One hour after the injection, majority of the radioactivity was observed in spleen and liver but also in blood with an average tumor-to-muscle ratio of 2. The prolonged radioactivity in blood circulation may open possibilities to the targeted delivery of the MSNAs.

Introduction: Non-invasive imaging techniques such as positron emission tomography (PET) are extremely important for cancer detection and characterization especially for difficult to biopsy or extremely delicate organs such as the brain. The folate analogue 1,4,7-triazacylononane-1,4,7-triacetic acid-conjugated folate radiolabeled with aluminum fluoride-18 ([ 18 F]FOL) has been previously shown to accumulate preferentially in tumor cells with an overexpression of folate receptors (FRs) and here was investigated for its ability to detect orthotopic gliomas in a rat model. In addition, we studied the expression of FRs in human glioblastoma samples to investigate if an analogous relationship may exist., Methods: Nine BDIX rats were injected with BT4C rat glioma cells into the right hemisphere of the brain. Animals were imaged with gadolinium-enhanced magnetic resonance imaging at on days prior to PET/computed tomography (CT) imaging. Animals were divided into two groups, and were PET/CT imaged with either [ 18 F]FOL or 2-deoxy-2-18F-fluoro-D-glucose ([ 18 F]FDG) on 19 and 32-days post glioma grafting. Two subjects were also PET/CT imaged with [ 18 F]FOL on day 16. Biodistribution was studied and brains were cryosectioned for autoradiography, immunofluorescence, and histological studies. Patient-derived paraffin-embedded glioblastomas were sectioned and stained with similar methods., Results: PET imaging showed an increase of [18F]FOL tumor-to-brain uptake ratio (TBR) over the study duration from day 16/19 (3.3 ± 0.9) increasing to 5.7 ± 1.0 by day 32. [ 18 F]FDG PET-imaged rats had a consistent TBR of 1.6 ± 0.1 throughout the study. Ex vivo autoradiography results revealed an exceptionally high TBR of 116.1 ± 26.9 for [ 18 F]FOL while the [18F]FDG values were significantly lower giving 2.9 ± 0.6 (P<0.0001). Immunostaining demonstrated an increased presence of FR-α in the BT4C gliomas versus the contralateral brain tissue, while FR-β was present only on glioma periphery. Human sections assayed showed similar FRs expression characteristics., Conclusion: This study shows upregulation of FR-α inside glioma regions in both human and animal tissue, providing a biochemical basis for the observed increased [ 18 F]FOL uptake in animal PET images. These results suggest that FRs targeting imaging and therapeutic compounds may possess clinically relevant translational abilities for the detection and treatment of gliomas., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Miner, Liljenbäck, Virta, Kärnä, Viitanen, Elo, Gardberg, Teuho, Saipa, Rajander, Mansour, Cleveland, Low, Li and Roivainen.)

Coronary artery disease (CAD) is a pandemic disease where up to half of the risk is explained by genetic factors. Advanced insights into the genetic basis of CAD require deeper understanding of the contributions of different cell types, molecular pathways, and genes to disease heritability. Here, we investigate the biological diversity of atherosclerosis-associated cell states and interrogate their contribution to the genetic risk of CAD by using single-cell and bulk RNA sequencing (RNA-seq) of mouse and human lesions. We identified 12 disease-associated cell states that we characterized further by gene set functional profiling, ligand-receptor prediction, and transcription factor inference. Importantly, Vcam1+ smooth muscle cell state genes contributed most to SNP-based heritability of CAD. In line with this, genetic variants near smooth muscle cell state genes and regulatory elements explained the largest fraction of CAD-risk variance between individuals. Using this information for variant prioritization, we derived a hybrid polygenic risk score (PRS) that demonstrated improved performance over a classical PRS. Our results provide insights into the biological mechanisms associated with CAD risk, which could make a promising contribution to precision medicine and tailored therapeutic interventions in the future., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Bexmarilimab is a new humanized monoclonal antibody against common lymphatic endothelial and vascular endothelial receptor-1 (CLEVER-1) and is in clinical trials for macrophage-guided cancer immunotherapy. In addition being associated with cancer, CLEVER-1 is also associated with fibrosis. To facilitate prospective human PET studies, we preclinically evaluated 89 Zr-labeled bexmarilimab in rabbits. Methods: Bexmarilimab was conjugated with desferrioxamine (DFO) and radiolabeled with 89 Zr. Retained immunoreactivity was confirmed by flow cytometry. The distribution kinetics of intravenously administered 89 Zr-DFO-bexmarilimab (0.1 mg/kg) were determined for up to 7 d in a rabbit model of renal fibrosis mediated by unilateral ureteric obstruction. The in vivo stability of 89 Zr-DFO-bexmarilimab was evaluated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis in combination with autoradiography. Additionally, we estimated the human radiation dose from data obtained in healthy rabbits. Results: Zr-DFO-bexmarilimab cleared rapidly from the blood circulation and distributed to the liver and spleen. At 24 h after injection, PET/CT, ex vivo γ-counting, and autoradiography demonstrated that there was significantly higher 89 Zr-DFO-bexmarilimab cleared rapidly from the blood circulation and distributed to the liver and spleen. At 24 h after injection, PET/CT, ex vivo γ-counting, and autoradiography demonstrated that there was significantly higher 89 Zr-DFO-bexmarilimab uptake in unilateral ureteric obstruction-operated fibrotic renal cortex, characterized by abundant CLEVER-1-positive cells, than in contralateral or healthy kidneys. The estimated effective dose for a 70-kg human was 0.70 mSv/MBq. Conclusion: The characteristics of 89 Zr-DFO-bexmarilimab support future human PET studies to, for example, stratify patients for bexmarilimab treatment, evaluate the efficacy of treatment, or monitor disease progression., (© 2023 by the Society of Nuclear Medicine and Molecular Imaging.)

Purpose: Photoperiod determines the metabolic activity of brown adipose tissue (BAT) and affects the food intake and body mass of mammals. Sympathetic innervation of the BAT controls thermogenesis and facilitates physiological adaption to seasonal changes, but the exact mechanism remains elusive. Previous studies have shown that central opioid signaling regulates BAT thermogenesis, and that the expression of the brain mu-opioid receptor (MOR) varies seasonally. Therefore, it is important to know whether MOR expression in BAT shows seasonal variation., Methods: We determined the effect of photoperiod on BAT MOR availability using [ 11 C]carfentanil positron emission tomography (PET). Adult rats (n = 9) were repeatedly imaged under various photoperiods in order to simulate seasonal changes., Results: Long photoperiod was associated with low MOR expression in BAT (β =  - 0.04, 95% confidence interval: - 0.07, - 0.01), but not in muscles. We confirmed the expression of MOR in BAT and muscle using immunofluorescence staining., Conclusion: Photoperiod affects MOR availability in BAT. Sympathetic innervation of BAT may influence thermogenesis via the peripheral MOR system. The present study supports the utility of [ 11 C]carfentanil PET to study the peripheral MOR system., (© 2022. The Author(s).)

Radiolabeled peptides have emerged as highly specific agents for targeting receptors expressed in tumors for therapeutic and diagnostic purposes. Peptides developed for positron emission tomography (PET) are typically radiolabeled using prosthetic groups or bifunctional chelators for fast "kit-like" incorporation of the radionuclide into the structure. A novel [ 18 F]alkylammoniomethyltrifluoroborate ([ 18 F]AmBF 3 ) tetrazine (Tz), [ 18 F]AmBF 3 -Tz, was developed for the [ 18 F]fluorination of trans -cyclooctene (TCO)-modified biomolecules using Tyr 3 -octreotides (TOCs) as model peptides. [ 18 F]AmBF 3 -Tz (A m = 15.4 ± 9.2 GBq/μmol, n = 14) was evaluated in healthy mice by ex vivo biodistribution and PET/computed tomography (CT), where the radiolabel in the prosthetic group was found stable in vivo , indicated by the low bone uptake in tibia (0.4 ± 0.1% ID/g, t = 270 min). TCO-TOCs tailored with polyethylene glycol (PEG) linkers were radiolabeled with [ 18 F]AmBF 3 -Tz, forming two new tracers, [ 18 F]AmBF 3 -PEG 4 -TOC (A m = 2.8 ± 1.8 GBq/μmol, n = 3) and [ 18 F]AmBF 3 -PEG 7 -TOC (A m of 6.0 ± 3.4 GBq/μmol, n = 13), which were evaluated by cell uptake studies and ex vivo biodistribution in subcutaneous AR42J rat pancreatic carcinoma tumor-bearing nude mice. The tracer demonstrating superior behavior ex vivo , the [ 18 F]AmBF 3 -PEG 7 -TOC, was further evaluated with PET/CT, where the tracer provided clear tumor visualization (SUV baseline = 1.01 ± 0.07, vs SUV blocked = 0.76 ± 0.04) at 25 min post injection. The novel AmBF 3 -Tz demonstrated that it offers potential as a prosthetic group for rapid radiolabeling of biomolecules in mild conditions using bioorthogonal chemistry.

Vascular adhesion protein-1 (VAP-1) is an inflammation-inducible adhesion molecule, which supports contact between leukocytes and inflamed endothelium. There is evidence that VAP-1 is involved in the recruitment of leukocytes to melanoma tumors. Interleukin-2 (IL-2)-based immunotherapy is an efficient therapy that promotes immune system activity against cancers but is associated with toxicity. In the present study, we evaluated the feasibility of PET/CT imaging using the radiotracer [ 68 Ga]Ga-DOTA-Siglec-9, which is targeted to VAP-1, to monitor pharmacodynamic effects of a novel FAP-IL2v immunocytokine (a genetically engineered variant of IL-2 fused with fibroblast activation protein) in the B16-FAP melanoma model. At 9 days after the inoculation of B16-FAP melanoma cells, mice were studied with [ 68 Ga]Ga-DOTA-Siglec-9 PET/CT as a baseline measurement. Immediately after baseline imaging, mice were treated with FAP-IL2v or vehicle, and treatment was repeated 3 days later. Subsequent PET/CT imaging was performed 3, 5, and 7 days after baseline imaging. In addition to in vivo PET imaging, ex vivo autoradiography, histology, and immunofluorescence staining were performed on excised tumors. B16-FAP tumors were clearly detected with [ 68 Ga]Ga-DOTA-Siglec-9 PET/CT during the follow-up period, without differences in tumor volume between FAP-IL2v-treated and vehicle-treated groups. Tumor-to-muscle uptake of [ 68 Ga]Ga-DOTA-Siglec-9 was significantly higher in the FAP-IL2v-treated group than in the vehicle-treated group 7 days after baseline imaging, and this was confirmed by tumor autoradiography analysis. FAP-IL2v treatment did not affect VAP-1 expression on the tumor vasculature. However, FAP-IL2v treatment increased the number of CD8 + T cells and natural killer cells in tumors. The present study showed that [ 68 Ga]Ga-DOTA-Siglec-9 can detect B16-FAP tumors and allows monitoring of FAP-IL2v treatment., Competing Interests: SJ owns stock in Faron Pharmaceuticals. CK, VN, and MR declare employment, patents and ownership interest with Roche, and TN was employed by Roche at the time of the study. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Viitanen, Virtanen, Liljenbäck, Moisio, Li, Nicolini, Richard, Klein, Nayak, Jalkanen and Roivainen.)

Increased glutamine metabolism by macrophages is associated with development of atherosclerotic lesions. Positron emission tomography/computed tomography (PET/CT) with a glutamine analog (2S,4 R )-4- 18 F-fluoroglutamine ( 18 F-FGln) allows quantification of glutamine consumption in vivo . Here, we investigated uptake of 18 F-FGln by atherosclerotic lesions in mice and compared the results with those obtained using the glucose analog 2-deoxy-2- 18 F-fluoro- D -glucose ( 18 F-FDG). Uptake of 18 F-FGln and 18 F-FDG by healthy control mice (C57BL/6JRj) and atherosclerotic low-density lipoprotein receptor-deficient mice expressing only apolipoprotein B100 (LDLR -/- ApoB 100/100 ) was investigated. The mice were injected intravenously with 18 F-FGln or 18 F-FDG for in vivo PET/CT imaging. After sacrifice at 70 minutes post-injection, tracer uptake was analyzed by gamma counting of excised tissues and by autoradiography of aorta cryosections, together with histological and immunohistochemical analyses. We found that myocardial uptake of 18 F-FGln was low. PET/CT detected lesions in the aortic arch, with a target-to-background ratio (SUV max , aortic arch/SUV mean , blood) of 1.95 ± 0.42 (mean ± standard deviation). Gamma counting revealed that aortic uptake of 18 F-FGln by LDLR -/- ApoB 100/100 mice (standardized uptake value [SUV], 0.35 ± 0.06) was significantly higher than that by healthy controls (0.20 ± 0.08, P = 0.03). More detailed analysis by autoradiography revealed that the plaque-to-healthy vessel wall ratio of 18 F-FGln (2.90 ± 0.42) was significantly higher than that of 18 F-FDG (1.93 ± 0.22, P = 0.004). Immunohistochemical staining confirmed that 18 F-FGln uptake in plaques co-localized with glutamine transporter SLC7A7-positive macrophages. Collectively these data show that the 18 F-FGln PET tracer detects inflamed atherosclerotic lesions. Thus, exploiting glutamine consumption using 18 F-FGln PET may have translational relevance for studying atherosclerotic inflammation., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Palani, Miner, Virta, Liljenbäck, Eskola, Örd, Ravindran, Kaikkonen, Knuuti, Li, Saraste and Roivainen.)

The 68 Gallium-labeled 1,4,7-triazacyclononane-1-glutaric acid-4,7-diacetic acid conjugated radiolabelled arginine-glycine-aspartic acid peptide ([ 68 Ga]Ga-NODAGA-RGD) is a positron emission tomography (PET) tracer binding to cell surface receptor α v β 3 integrin that is upregulated during angiogenesis and inflammation. We studied whether α v β 3 targeting PET imaging can detect myocardial inflammation in a rat model of autoimmune myocarditis. To induce myocarditis, rats ( n = 8) were immunized with porcine cardiac myosin in complete Freund's adjuvant on days 0 and 7. Control rats ( n = 8) received Freund's adjuvant alone. On day 21, in vivo PET/CT imaging with [ 68 Ga]Ga-NODAGA-RGD followed by ex vivo autoradiography and immunohistochemistry were carried out. Inflammatory lesions were detected histologically in the myocardium of 7 out of 8 immunized rats. In vivo PET images showed higher [ 68 Ga]Ga-NODAGA-RGD accumulation in the myocardium of rats with inflammation than the non-inflamed myocardium of control rats (SUV mean 0.4 ± 0.1 vs. 0.1 ± 0.02; P = 0.00006). Ex vivo autoradiography and histology confirmed that [ 68 Ga]Ga-NODAGA-RGD uptake co-localized with inflammatory lesions containing α v β 3 integrin-positive capillary-like structures. A non-specific [ 68 Ga]Ga-DOTA-(RGE) 2 tracer showed 76% lower uptake than [ 68 Ga]Ga-NODAGA-RGD in the inflamed myocardium. Our results indicate that α v β 3 integrin-targeting [ 68 Ga]Ga-NODAGA-RGD is a potential PET tracer for the specific detection of active inflammatory lesions in autoimmune myocarditis., Competing Interests: AS received consultancy or speaker fees from Amgen, Astra Zeneca, Boehringer Ingelheim, Abbott, and Bayer not related to the present study. JK received consultancy fees from GE Healthcare and AstraZeneca and speaker fees from GE Healthcare, Bayer, and Lundbeck. Boehringer-Ingelheim and Merck, outside of the submitted work. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Jahandideh, Ståhle, Virta, Li, Liljenbäck, Moisio, Knuuti, Roivainen and Saraste.)

Purpose: The three positron emission tomography (PET) imaging compounds: (2 S ,4 R )-4-[ 18 F]Fluoroglutamine ([ 18 F]FGln), L -[methyl- 11 C]Methionine ([ 11 C]Met), and 2-deoxy-2-[ 18 F]fluoro- D -glucose ([ 18 F]FDG) were investigated to contrast their ability to image orthotopic BT4C gliomas in BDIX rats. Two separate small animal imaging systems were compared for their tumor detection potential. Dynamic acquisition of [ 18 F]FGln was evaluated with multiple pharmacokinetic models for future quantitative comparison., Procedures: Up to four imaging studies were performed on each orthotopically grafted BT4C glioma-bearing BDIX rat subject (n = 16) on four consecutive days. First, a DOTAREM ® contrast enhanced MRI followed by attenuation correction CT and dynamic PET imaging with each radiopharmaceutical (20 min [ 11 C]Met, 60 min [ 18 F]FDG, and 60 min [ 18 F]FGln with either the Molecubes PET/CT (n = 5) or Inveon PET/CT cameras (n = 11). Ex vivo brain autoradiography was completed for each radiopharmaceutical and [ 18 F]FGln pharmacokinetics were studied by injecting 40 MBq into healthy BDIX rats (n = 10) and collecting blood samples between 5 and 60 min. Erythrocyte uptake, plasma protein binding and plasma parent-fraction were combined to estimate the total blood bioavailability of [ 18 F]FGln over time. The corrected PET-image blood data was then applied to multiple pharmacokinetic models., Results: Average BT4C tumor-to-healthy brain tissue uptake ratios (TBR) for PET images reached maxima of: [ 18 F]FGln TBR: 1.99 ± 0.19 (n = 13), [ 18 F]FDG TBR: 1.41 ± 0.11 (n = 6), and [ 11 C]Met TBR: 1.08 ± 0.08, (n = 12) for the dynamic PET images. Pharmacokinetic modeling in dynamic [ 18 F]FGln studies suggested both reversible and irreversible uptake play a similar role. Imaging with Inveon and Molecubes yielded similar end-result ratios with insignificant differences (p > 0.25)., Conclusions: In orthotopic BT4C gliomas, [ 18 F]FGln may offer improved imaging versus [ 11 C]Met and [ 18 F]FDG. No significant difference in normalized end-result data was found between the Inveon and Molecubes camera systems. Kinetic modelling of [ 18 F]FGln uptake suggests that both reversible and irreversible uptake play an important role in BDIX rat pharmacokinetics., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Miner, Liljenbäck, Virta, Helin, Eskola, Elo, Teuho, Seppälä, Oikonen, Yang, Kindler-Röhrborn, Minn, Li and Roivainen.)

Purpose: To evaluate fitting quality and repeatability of four mathematical models for diffusion weighted imaging (DWI) during tumor progression in mouse xenograft model of prostate cancer., Methods: Human prostate cancer cells (PC-3) were implanted subcutaneously in right hind limbs of 11 immunodeficient mice. Tumor growth was followed by weekly DWI examinations using a 7T MR scanner. Additional DWI examination was performed after repositioning following the fourth DWI examination to evaluate short term repeatability. DWI was performed using 15 and 12 b-values in the ranges of 0-500 and 0-2000 s/mm 2 , respectively. Corrected Akaike information criteria and F-ratio were used to evaluate fitting quality of each model (mono-exponential, stretched exponential, kurtosis, and bi-exponential)., Results: Significant changes were observed in DWI data during the tumor growth, indicated by ADC m , ADC s , and ADC k . Similar results were obtained using low as well as high b-values. No marked changes in model preference were present between the weeks 1-4. The parameters of the mono-exponential, stretched exponential, and kurtosis models had smaller confidence interval and coefficient of repeatability values than the parameters of the bi-exponential model., Conclusion: Stretched exponential and kurtosis models showed better fit to DWI data than the mono-exponential model and presented with good repeatability., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Merisaari, Laakso, Liljenbäck, Virtanen, Aronen, Minn, Poutanen, Roivainen, Liimatainen and Jambor.)

Cardiovascular effects of glucagon-like peptide-1 receptor (GLP-1R) agonist therapies are potentially mediated by anti-inflammatory effects on atherosclerosis. Our study demonstrates that 68 Ga-NODAGA-exendin-4, a radioligand specifically targeting GLP-1R, detects GLP-1R expression in inflamed atherosclerotic lesions in nondiabetic and diabetic hypercholesterolemic mice. Immunofluorescence staining suggests that GLP-1R is primarily localized in M2 macrophages in lesions. This study describes a new potential tool that may have translational relevance for studies of pharmacological modification of GLP-1R signaling in atherosclerosis.

Cryptochrome 2 (Cry2) is a core clock gene important for circadian regulation. It has also been associated with anxiety and depressive-like behaviors in mice, but the previous findings have been conflicting in terms of the direction of the effect. To begin to elucidate the molecular mechanisms of this association, we carried out behavioral testing, PET imaging, and gene expression analysis of Cry2 -/- and Cry2 +/+ mice. Compared to Cry2 +/+ mice, we found that Cry2 -/- mice spent less time immobile in the forced swim test, suggesting reduced despair-like behavior. Moreover, Cry2 -/- mice had lower saccharin preference, indicative of increased anhedonia. In contrast, we observed no group differences in anxiety-like behavior. The behavioral changes were accompanied by lower metabolic activity of the ventro-medial hypothalamus, suprachiasmatic nuclei, ventral tegmental area, anterior and medial striatum, substantia nigra, and habenula after cold stress as measured by PET imaging with a glucose analog. Although the expression of many depression-associated and metabolic genes was upregulated or downregulated by cold stress, we observed no differences between Cry2 -/- and Cry2 +/+ mice. These findings are consistent with other studies showing that Cry2 is required for normal emotional behavior. Our findings confirm previous roles of Cry2 in behavior and extend them by showing that the effects on behavior may be mediated by changes in brain metabolism., (© 2020 International Behavioural and Neural Genetics Society and John Wiley & Sons Ltd.)

MRI is a common method of prostate cancer diagnosis. Several MRI-derived markers, including the apparent diffusion coefficient (ADC) based on diffusion-weighted imaging, have been shown to provide values for prostate cancer detection and characterization. The hypothesis of the study was that docetaxel chemotherapy response could be picked up earlier with rotating frame relaxation times T RAFF2 and T RAFF4 than with the continuous wave T 1ρ , adiabatic T 1ρ , adiabatic T 2ρ , T 1 , T 2 or water ADC. Human PC3 prostate cancer cells expressing a red fluorescent protein were implanted in 21 male mice. Docetaxel chemotherapy was given once a week starting 1 week after cell implantation for 10 randomly selected mice, while the rest served as a control group (n = 11). The MRI consisted of relaxation along a fictitious field (RAFF) in the second (RAFF2) and fourth (RAFF4) rotating frames, T 1 and T 2 , continuous wave T 1ρ , adiabatic T 1ρ and adiabatic T 2ρ relaxation time measurements and water ADC. MRI was conducted at 7 T, once a week up to 4 weeks from cell implantation. The tumor volume was monitored using T 2 -weighted MRI and optical imaging. The histology was evaluated after the last imaging time point. Significantly reduced RAFFn, T 1ρ, T 2ρ and conventional relaxation times 4 weeks after tumor implantation were observed in the treated tumors compared with the controls. The clearest short- and long-term responses were obtained with T 1 , while no clear improvement in response to treatment was detected with novel methods compared with conventional methods or with RAFFn compared with all others. The tumor volume decreased after a two-week time point for the treated group and increased significantly in the control group, which was supported by increasing red fluorescent light emission in the control tumors. Decreased relaxation times were associated with successful chemotherapy outcomes. The results indicate altered relaxation mechanisms compared with higher dose chemotherapies previously published., (© 2021 The Authors. NMR in Biomedicine published by John Wiley & Sons Ltd.)

Seasonal rhythms influence mood and sociability. The brain μ-opioid receptor (MOR) system modulates a multitude of seasonally varying socioemotional functions, but its seasonal variation remains elusive with no previously reported in vivo evidence. Here, we first conducted a cross-sectional study with previously acquired human [ 11 C]carfentanil PET imaging data (132 male and 72 female healthy subjects) to test whether there is seasonal variation in MOR availability. We then investigated experimentally whether seasonal variation in daylength causally influences brain MOR availability in rats. Rats (six male and three female rats) underwent daylength cycle simulating seasonal changes; control animals (two male and one female rats) were kept under constant daylength. Animals were scanned repeatedly with [ 11 C]carfentanil PET imaging. Seasonally varying daylength had an inverted U-shaped functional relationship with brain MOR availability in humans. Brain regions sensitive to daylength spanned the socioemotional brain circuits, where MOR availability peaked during spring. In rats, MOR availabilities in the brain neocortex, thalamus, and striatum peaked at intermediate daylength. Varying daylength also affected the weight gain and stress hormone levels. We conclude that cerebral MOR availability in humans and rats shows significant seasonal variation, which is predominately associated with seasonal photoperiodic variation. Given the intimate links between MOR signaling and socioemotional behavior, these results suggest that the MOR system might underlie seasonal variation in human mood and social behavior. SIGNIFICANCE STATEMENT Seasonal rhythms influence emotion and sociability. The central μ-opioid receptor (MOR) system modulates numerous seasonally varying socioemotional functions, but its seasonal variation remains elusive. Here we used positron emission tomography to show that MOR levels in both human and rat brains show daylength-dependent seasonal variation. The highest MOR availability was observed at intermediate daylengths. Given the intimate links between MOR signaling and socioemotional behavior, these results suggest that the MOR system might underlie seasonal variation in human mood and social behavior., (Copyright © 2021 Sun et al.)

Background: Activated macrophages in the experimental model of multiple sclerosis (MS) express folate receptor-β (FR-β), representing a promising target for the treatment of MS. Here, we both evaluated the efficacy of a novel folate-aminopterin construct (EC2319) in a rat focal model of multiple sclerosis (MS) and investigated the utility of 68 Ga-labeled 1,4,7-triazacyclononane-1,4,7-triacetic acid-conjugated folate ( 68 Ga-FOL) for assessing inflammatory lesions. In addition, we investigated whether FR-β is expressed in the brain of patients with MS., Methods: Focal delayed-type hypersensitivity experimental autoimmune encephalomyelitis (fDTH-EAE) was induced in 40 Lewis rats; 20 healthy Lewis rats were used as controls. Rats were divided into six groups according to the duration of disease (control, acute, or chronic) and intervention (vehicle versus EC2319). 68 Ga-FOL analyses, histology, and immunofluorescence of the brain were performed to evaluate the efficacy of subcutaneously administered EC2319 on lesion development. Immunofluorescence was used to assess FR-β expression in postmortem brain samples from 5 patients with MS and 5 healthy controls., Results: Immunofluorescence and histological analyses revealed significant reductions in FR-β expression (P < 0.05) and lesion size (P < 0.01), as well as improved inducible nitric oxide synthase/mannose receptor C type 1 ratios (P < 0.01) in macrophages and microglia during the chronic but not acute phase of fDTH-EAE in EC2319-treated rats. The uptake of IV-injected 68 Ga-FOL in the brain was low and did not differ between the groups, but the in vitro binding of 68 Ga-FOL was significantly lower in EC2319-treated rats (P < 0.01). FR-β positivity was observed in chronically active lesions and in normal-appearing white matter in MS brain samples., Conclusions: EC2319 was well tolerated and attenuated inflammation and lesion development in a rat model of a chronic progressive form of MS. Human MS patients have FR-β-positive cells in chronically active plaques, which suggests that these results may have translational relevance.

Background: Activation of glucagon-like peptide-1 receptor (GLP-1R) signaling protects against cardiac dysfunction and remodeling after myocardial infarction (MI). The aim of the study was to evaluate 68 Ga-NODAGA-exendin-4 positron emission tomography (PET) for assessment of GLP-1R expression after MI in rats., Methods and Results: Rats were studied at 3 days, 1 and 12 weeks after permanent coronary ligation or a sham-operation. Rats were injected with 68 Ga-NODAGA-exendin-4 and scanned with PET and contrast-enhanced computed tomography (CT) followed by digital autoradiography and histology of left ventricle tissue sections. 68 Ga-NODAGA-exendin-4 PET/CT showed focally increased tracer uptake in the infarcted regions peaking at 3 days and continuing at 1 week after MI. Pre-treatment with an unlabeled exendin-4 peptide significantly reduced 68 Ga-NODAGA-exendin-4 uptake. By autoradiography, 68 Ga-NODAGA-exendin-4 uptake was 8.6-fold higher in the infarcted region and slightly increased also in the remote, non-infarcted myocardium at 1 week and 12 weeks post-MI compared with sham. Uptake of 68 Ga-NODAGA-exendin-4 correlated with the amount of CD68-positive macrophages in the infarcted area and alpha-smooth muscle actin staining in the remote myocardium., Conclusions: 68 Ga-NODAGA-exendin-4 PET detects up-regulation of cardiac GLP-1R expression during healing of MI in rats and may provide information on the activated repair mechanisms after ischemic myocardial injury.

Currently available imaging techniques have limited specificity for the detection of active myocardial inflammation. Aluminum 18 F-labeled 1,4,7-triazacyclononane- N,N',N″ -triacetic acid conjugated folate ( 18 F-FOL) is a PET tracer targeting folate receptor β (FR-β), which is expressed on activated macrophages at sites of inflammation. We evaluated 18 F-FOL PET for the detection of myocardial inflammation in rats with autoimmune myocarditis and studied the expression of FR-β in human cardiac sarcoidosis specimens. Methods: Myocarditis was induced by immunizing rats ( n = 18) with porcine cardiac myosin in complete Freund adjuvant. Control rats ( n = 6) were injected with Freund adjuvant alone. 18 F-FOL was intravenously injected, followed by imaging with a small-animal PET/CT scanner and autoradiography. Contrast-enhanced high-resolution CT or 18 F-FDG PET images were used for coregistration. Rat tissue sections and myocardial autopsy samples from 6 patients with cardiac sarcoidosis were studied for macrophages and FR-β. Results: The myocardium of 10 of 18 immunized rats showed focal macrophage-rich inflammatory lesions, with FR-β expression occurring mainly in M1-polarized macrophages. PET images showed focal myocardial 18 F-FOL uptake colocalizing with inflammatory lesions (SUV mean , 2.1 ± 1.1), whereas uptake in the remote myocardium of immunized rats and controls was low (SUV mean , 0.4 ± 0.2 and 0.4 ± 0.1, respectively; P < 0.01). Ex vivo autoradiography of tissue sections confirmed uptake of 18 F-FOL in myocardial inflammatory lesions. Uptake of 18 F-FOL in inflamed myocardium was efficiently blocked by a nonlabeled FR-β ligand folate glucosamine in vivo. The myocardium of patients with cardiac sarcoidosis showed many FR-β-positive macrophages in inflammatory lesions. Conclusion: In a rat model of autoimmune myocarditis, 18 F-FOL shows specific uptake in inflamed myocardium containing macrophages expressing FR-β, which were also present in human cardiac sarcoid lesions. Imaging of FR-β expression is a potential approach for the detection of active myocardial inflammation., (© 2020 by the Society of Nuclear Medicine and Molecular Imaging.)

Hydroxysteroid 17β dehydrogenase 12 (HSD17B12) is suggested to be involved in the elongation of very long chain fatty acids. Previously, we have shown a pivotal role for the enzyme during mouse development. In the present study we generated a conditional Hsd17b12 knockout (HSD17B12cKO) mouse model by breeding mice homozygous for a floxed Hsd17b12 allele with mice expressing the tamoxifen-inducible Cre recombinase at the ROSA26 locus. Gene inactivation was induced by administering tamoxifen to adult mice. The gene inactivation led to a 20% loss of body weight within 6 days, associated with drastic reduction in both white (83% males, 75% females) and brown (65% males, 60% females) fat, likely due to markedly reduced food and water intake. Furthermore, the knockout mice showed sickness behavior and signs of liver toxicity, specifically microvesicular hepatic steatosis and increased serum alanine aminotransferase (4.6-fold in males, 7.7-fold in females). The hepatic changes were more pronounced in females than males. Proinflammatory cytokines, such as interleukin-6 (IL-6), IL-17, and granulocyte colony-stimulating factor, were increased in the HSD17B12cKO mice indicating an inflammatory response. Serum lipidomics study showed an increase in the amount of dihydroceramides, despite the dramatic overall loss of lipids. In line with the proposed role for HSD17B12 in fatty acid elongation, we observed accumulation of ceramides, dihydroceramides, hexosylceramides, and lactosylceramides with shorter than 18-carbon fatty acid side chains in the serum. The results indicate that HSD17B12 is essential for proper lipid homeostasis and HSD17B12 deficiency rapidly results in fatal systemic inflammation and lipolysis in adult mice.

Folate receptor β (FR-β), a marker expressed on macrophages, is a promising target for imaging of inflammation. Here, we report the radiosynthesis and preclinical evaluation of [ 68 Ga]Ga-NOTA-folate ( 68 Ga-FOL). After determining the affinity of 68 Ga-FOL using cells expressing FR-β, we studied atherosclerotic mice with 68 Ga-FOL and 18 F-FDG PET/CT. In addition, we studied tracer distribution and co-localization with macrophages in aorta cryosections using autoradiography, histology, and immunostaining. The specificity of 68 Ga-FOL was assessed in a blocking study with folate glucosamine. As a final step, human radiation doses were extrapolated from rat PET data. We were able to produce 68 Ga-FOL with high radiochemical purity and moderate molar activity. Cell binding studies revealed that 68 Ga-FOL had 5.1 nM affinity for FR-β. Myocardial uptake of 68 Ga-FOL was 20-fold lower than that of 18 F-FDG. Autoradiography and immunohistochemistry of the aorta revealed that 68 Ga-FOL radioactivity co-localized with Mac-3-positive macrophage-rich atherosclerotic plaques. The plaque-to-healthy vessel wall ratio of 68 Ga-FOL was significantly higher than that of 18 F-FDG. Blocking studies verified that 68 Ga-FOL was specific for FR. Based on estimations from rat data, the human effective dose was 0.0105 mSv/MBq. Together, these findings show that 68 Ga-FOL represents a promising new FR-β-targeted tracer for imaging macrophage-associated inflammation.

Purpose: The glutamine analogue (2S, 4R)-4-[ 18 F]fluoroglutamine ([ 18 F]FGln) was investigated to further characterize its pharmacokinetics and acquire in vivo positron emission tomography (PET) images of separate orthotopic and subcutaneous glioma xenografts in mice., Procedures: [ 18 F]FGln was synthesized at a high radiochemical purity as analyzed by high-performance liquid chromatography. An orthotopic model was created by injecting luciferase-expressing patient-derived BT3 glioma cells into the right hemisphere of BALB/cOlaHsd-Foxn1 nu mouse brains (tumor growth monitored via in vivo bioluminescence), the subcutaneous model by injecting rat BT4C glioma cells into the flank and neck regions of Foxn1 nu/nu mice. Dynamic PET images were acquired after injecting 10-12 MBq of the tracer into mouse tail veins. Animals were sacrificed 63 min after tracer injection, and ex vivo biodistributions were measured. Tumors and whole brains (with tumors) were cryosectioned, autoradiographed, and stained with hematoxylin-eosin. All images were analyzed with CARIMAS software. Blood sampling of 6 Foxn1 nu/nu and 6 C57BL/6J mice was performed after 9-14 MBq of tracer was injected at time points between 5 and 60 min then assayed for erythrocyte uptake, plasma protein binding, and plasma parent-fraction of radioactivity to correct PET image-derived whole-blood radioactivity and apply the data to multiple pharmacokinetic models., Results: Orthotopic human glioma xenografts displayed PET image tumor-to-healthy brain region ratio of 3.6 and 4.8 while subcutaneously xenografted BT4C gliomas displayed (n = 12) a tumor-to-muscle (flank) ratio of 1.9 ± 0.7 (range 1.3-3.4). Using PET image-derived blood radioactivity corrected by population-based stability analyses, tumor uptake pharmacokinetics fit Logan and Yokoi modeling for reversible uptake., Conclusions: The results reinforce that [ 18 F]FGln has preferential uptake in glioma tissue versus that of corresponding healthy tissue and fits well with reversible uptake models.

Background and Aims: Dipeptidyl peptidase 4 (DPP-4) inhibitors have anti-inflammatory and atheroprotective effects. We evaluated the effects of the DPP-4 inhibitor linagliptin on atherosclerotic plaque and hepatic inflammation using histology and 2-deoxy-2-[ 18 F]-fluoro-d-glucose ( 18 F-FDG), a positron emission tomography tracer of inflammation, in a mouse model of hypercholesterolemia and type 2 diabetes., Methods: Igf2/Ldlr -/- Apob 100/100 mice were fed a high-fat diet (HFD) for 8 weeks and then randomly allocated to receive HFD (n = 14), or HFD with added linagliptin (n = 15) for additional 12 weeks. Five mice fed a chow diet were studied as an additional control. At the end of the study, glucose tolerance, aortic and liver uptake of 18 F-FDG, and histology were studied., Results: Mice in linagliptin and HFD groups had similar fasting glucose concentrations, but linagliptin improved glucose tolerance. Aortas of linagliptin and HFD groups showed advanced atherosclerotic plaques with no difference in the mean intima-to-media ratio or number of macrophages in the plaques. Autoradiography showed similar 18 F-FDG uptake by atherosclerotic plaques in linagliptin and HFD groups (plaque-to-wall ratio: 1.7 ± 0.25 vs. 1.6 ± 0.21; p = 0.24). In the liver, linagliptin reduced the histologic inflammation score but had no effect on 18 F-FDG uptake. Compared with chow diet, uptake of 18 F-FDG was similar in the aorta, but higher in the liver after HFD., Conclusions: Linagliptin therapy improved glucose tolerance and reduced hepatic inflammation but had no effect on plaque burden or atherosclerotic inflammation, as determined by histology and 18 F-FDG uptake, in atherosclerotic mice with type 2 diabetes., Competing Interests: Declaration of competing interest The authors declared they do not have anything to disclose regarding conflict of interest with respect to this manuscript., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: Magnetic resonance imaging (MRI) with Gadolinium 1,4,7,10-tetraazacyclododecane-N',N″,N''',N″″-tetraacetic acid (Gd-DOTA) enables assessment of myocardial perfusion during first-pass of the contrast agent, while increased retention can signify areas of myocardial infarction (MI). We studied whether Gallium-68-labeled analog, 68 Ga-DOTA, can be used to assess myocardial perfusion on positron emission tomography/computed tomography (PET/CT) in rats, comparing it with 11 C-acetate., Methods: Rats were studied with 11 C-acetate and 68 Ga-DOTA at 24 hours after permanent ligation of the left coronary artery or sham operation. One-tissue compartmental models were used to estimate myocardial perfusion in normal and infarcted myocardium. After the PET scan, hearts were sectioned for autoradiographic detection of 68 Ga-DOTA distribution., Results: 11 C-acetate PET showed perfusion defects and histology showed myocardial necrosis in all animals after coronary ligation. Kinetic modeling of 68 Ga-DOTA showed significantly higher k 1 values in normal myocardium than in infarcted areas. There was a significant correlation (r = 0.82, P = 0.001) between k 1 values obtained with 68 Ga-DOTA and 11 C-acetate. After 10 minutes of tracer distribution, the 68 Ga-DOTA concentration was significantly higher in the infarcted than normal myocardium on PET imaging and autoradiography., Conclusions: Our results indicate that acute MI can be detected as reduced perfusion, as well as increased late retention of 68 Ga-DOTA.

This study showed that treatment with a therapeutic monoclonal immunoglobulin-G1 antibody against phosphorylcholine on oxidized phospholipids preserves coronary flow reserve and attenuates atherosclerotic inflammation as determined by the uptake of 18 F-fluorodeoxyglucose in atherosclerotic mice. The noninvasive imaging techniques represent translational tools to assess the efficacy of phosphorylcholine-targeted therapy on coronary artery function and atherosclerosis in clinical studies., (© 2020 The Authors.)

Background: Folate receptor-β (FR-β) is a cell surface receptor that is significantly upregulated on activated macrophages during inflammation and provides a potential target for folate-based therapeutic and diagnostic agents. FR-β expression in central nervous system inflammation remains relatively unexplored. Therefore, we used focally induced acute and chronic phases of experimental autoimmune encephalomyelitis (EAE) to study patterns of FR-β expression and evaluated its potential as an in vivo imaging target., Methods: Focal EAE was induced in rats using heat-killed Bacillus Calmette-Guérin followed by activation with complete Freund's adjuvant supplemented with Mycobacterium tuberculosis. The rats were assessed with magnetic resonance imaging and positron emission tomography/computed tomography (PET/CT) at acute (14 days) and chronic (90 days) phases of inflammation. The animals were finally sacrificed for ex vivo autoradiography of their brains. PET studies were performed using FR-β-targeting aluminum [ 18 F]fluoride-labeled 1,4,7-triazacyclononane-1,4,7-triacetic acid conjugated folate ([ 18 F]AlF-NOTA-folate, 18 F-FOL) and 18 kDa translocator protein (TSPO)-targeting N-acetyl-N-(2-[ 11 C]methoxybenzyl)-2-phenoxy-5-pyridinamine ( 11 C-PBR28). Post-mortem immunohistochemistry was performed using anti-FR-β, anti-cluster of differentiation 68 (anti-CD68), anti-inducible nitric oxide synthase (anti-iNOS), and anti-mannose receptor C-type 1 (anti-MRC-1) antibodies. The specificity of 18 F-FOL binding was verified using in vitro brain sections with folate glucosamine used as a blocking agent., Results: Immunohistochemical evaluation of focal EAE lesions demonstrated anti-FR-β positive cells at the lesion border in both acute and chronic phases of inflammation. We found that anti-FR-β correlated with anti-CD68 and anti-MRC-1 immunohistochemistry; for MRC-1, the correlation was most prominent in the chronic phase of inflammation. Both 18 F-FOL and 11 C-PBR28 radiotracers bound to the EAE lesions. Autoradiography studies verified that this binding took place in areas of anti-FR-β positivity. A blocking assay using folate glucosamine further verified the tracer's specificity. In the chronic phase of EAE, the lesion-to-background ratio of 18 F-FOL was significantly higher than that of 11 C-PBR28 (P = 0.016)., Conclusion: Our EAE results imply that FR-β may be a useful target for in vivo imaging of multiple sclerosis-related immunopathology. FR-β-targeted PET imaging with 18 F-FOL may facilitate the monitoring of lesion development and complement the information obtained from TSPO imaging by bringing more specificity to the PET imaging armamentarium for neuroinflammation.

Mitochondrial dysfunction elicits stress responses that safeguard cellular homeostasis against metabolic insults. Mitochondrial integrated stress response (ISR mt ) is a major response to mitochondrial (mt)DNA expression stress (mtDNA maintenance, translation defects), but the knowledge of dynamics or interdependence of components is lacking. We report that in mitochondrial myopathy, ISR mt progresses in temporal stages and development from early to chronic and is regulated by autocrine and endocrine effects of FGF21, a metabolic hormone with pleiotropic effects. Initial disease signs induce transcriptional ISR mt (ATF5, mitochondrial one-carbon cycle, FGF21, and GDF15). The local progression to 2 nd metabolic ISR mt stage (ATF3, ATF4, glucose uptake, serine biosynthesis, and transsulfuration) is FGF21 dependent. Mitochondrial unfolded protein response marks the 3 rd ISR mt stage of failing tissue. Systemically, FGF21 drives weight loss and glucose preference, and modifies metabolism and respiratory chain deficiency in a specific hippocampal brain region. Our evidence indicates that FGF21 is a local and systemic messenger of mtDNA stress in mice and humans with mitochondrial disease., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Shank-associated RH domain-interacting protein (SHARPIN) is a cytosolic protein that plays a key role in activation of nuclear factor κ-light-chain enhancer of activated B cells and regulation of inflammation. Furthermore, SHARPIN controls integrin-dependent cell adhesion and migration in several normal and malignant cell types, and loss of SHARPIN correlates with increased integrin activity in mice. Arginyl-glycyl-aspartic acid (RGD), a cell adhesion tripeptide motif, is an integrin recognition sequence that facilitates PET imaging of integrin upregulation during tumor angiogenesis. We hypothesized that increased integrin activity due to loss of SHARPIN protein would affect the uptake of α v β 3 -selective cyclic, dimeric peptide 68 Ga-DOTA-E[c(RGDfK)] 2 , where E[c(RGDfk)] 2 = glutamic acid-[cyclo(arginyl-glycyl-aspartic acid-D-phenylalanine-lysine)], both in several tissue types and in the tumor microenvironment. To test this hypothesis, we used RGD-based in vivo PET imaging to evaluate wild-type (wt) and SHARPIN-deficient mice ( Sharpin cpdm , where cpdm = chronic proliferative dermatitis in mice) with and without melanoma tumor allografts. Methods: Sharpin cpdm mice with spontaneous null mutation in the Sharpin gene and their wt littermates with or without B16-F10-luc melanoma tumors were studied by in vivo imaging and ex vivo measurements with cyclic-RGD peptide 68 Ga-DOTA-E[c(RGDfK)] 2 After the last 68 Ga-DOTA-E[c(RGDfK)] 2 peptide PET/CT, tumors were cut into cryosections for autoradiography, histology, and immunohistochemistry. Results: The ex vivo uptake of 68 Ga-DOTA-E[c(RGDfK)] 2 in the mouse skin and tumor was significantly higher in Sharpin cpdm mice than in wt mice. B16-F10-luc tumors were detected 4 d after inoculation, without differences in volume or blood flow between the mouse strains. PET imaging with 68 Ga-DOTA-E[c(RGDfK)] 2 peptide at day 10 after inoculation revealed significantly higher uptake in the tumors transplanted into Sharpin cpdm mice than in wt mice. Furthermore, tumor vascularization was increased in the Sharpin cpdm mice. Conclusion: Sharpin cpdm mice demonstrated increased integrin activity and vascularization in B16-F10-luc melanoma tumors, as demonstrated by RGD-based in vivo PET imaging. These data indicate that SHARPIN, a protein previously associated with increased cancer growth and metastasis, may also have important regulatory roles in controlling the tumor microenvironment., (© 2019 by the Society of Nuclear Medicine and Molecular Imaging.)

Background: Borrelia burgdorferi sensu lato spirochetes (Borrelia) causing Lyme borreliosis are able to disseminate from the initial entry site to distant organs in the host. Outer-surface adhesins are crucial in the bacterial dissemination and adhesion to various tissues. Two well-characterized Borrelia adhesins, decorin-binding proteins A and B, have been shown to bind to 2 host receptors, decorin and biglycan. However, the role of biglycan in Borrelia infection has not been characterized in vivo., Methods: We infected biglycan knockout (KO) and wild-type (WT) C3H mice with strains representing 3 Borrelia genospecies, Borrelia burgdorferi sensu stricto, Borrelia garinii, and Borrelia afzelii. The infection was monitored by measuring joint swelling, Borrelia culture, polymerase chain reaction analysis, and serologic analysis. The host immune responses were analyzed by histological scoring of the inflammation in tissues and by cytokine profiling., Results: B. burgdorferi sensu stricto and B. garinii established long-term infection in mice of both genotypes, while B. afzelii failed to disseminate in KO mice. Further, the B. burgdorferi sensu stricto-infected KO mice had persistent inflammation in the joints., Conclusions: The dissemination and tissue colonization of Borrelia and the inflammatory response of the host differ in a mouse biglycan expression- and Borrelia genospecies-dependent manner., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)