Imaging of translocator protein upregulation is selective for pro‐inflammatory polarized astrocytes and microglia

Abstract

Translocator protein (TSPO) expression is increased in activated glia, and has been used as a marker of neuroinflammation in PET imaging. However, the extent to which TSPO upregulation reflects a pro‐ or anti‐inflammatory phenotype remains unclear. Our aim was to determine whether TSPO upregulation in astrocytes and microglia/macrophages is limited to a specific inflammatory phenotype. TSPO upregulation was assessed by flow cytometry in cultured astrocytes, microglia, and macrophages stimulated with lipopolysaccharide (LPS), tumor necrosis factor (TNF), or interleukin‐4 (Il‐4). Subsequently, mice were injected intracerebrally with either a TNF‐inducing adenovirus (AdTNF) or IL‐4. Glial expression of TSPO and pro‐/anti‐inflammatory markers was assessed by immunohistochemistry/fluorescence and flow cytometry. Finally, AdTNF or IL‐4 injected mice underwent PET imaging with injection of the TSPO radioligand 18F‐DPA‐713, followed by ex vivo autoradiography. TSPO expression was significantly increased in pro‐inflammatory microglia/macrophages and astrocytes both in vitro, and in vivo after AdTNF injection (p < .001 vs. control hemisphere), determined both histologically and by FACS. Both PET imaging and autoradiography revealed a significant (p < .001) increase in 18F‐DPA‐713 binding in the ipsilateral hemisphere of AdTNF‐injected mice. In contrast, no increase in either TSPO expression assessed histologically and by FACS, or ligand binding by PET/autoradiography was observed after IL‐4 injection. Taken together, these results suggest that TSPO imaging specifically reveals the pro‐inflammatory population of activated glial cells in the brain in response to inflammatory stimuli. Since the inflammatory phenotype of glial cells is critical to their role in neurological disease, these findings may enhance the utility and application of TSPO imaging.

Diagnostic and Therapeutic Potential of TSPO Studies Regarding Neurodegenerative Diseases, Psychiatric Disorders, Alcohol Use Disorders, Traumatic Brain Injury, and Stroke: An Update

“…In view of these observations, much research has been carried out in order to track TSPO expression and ligand binding not just for uncovering the secrets of neuroinflammation but also in the hope of offering non-invasive techniques for locating them, thus providing a significant impact on the diagnosis of CNS diseases. From all the available in vitro studies up to this date, TSPO upregulation is indisputably confirmed in the M1 microglial phenotype [27,39] while enhanced TSPO expression in M2 has been less unequivocal [46]. Furthermore, it was also established that TSPO ligands 2-Cl-MGV-1 (full name: (2-(2-chlorophenyl) quinazolin-4-yl dimethylcarbamate)) and MGV-1 (full name: 2-phenylquinazolin-4-yl dimethylcarbamate) expressed anti-inflammatory effects by reducing IL-1β, IL-6, tumor necrosis factor alpha (TNF-α), and interferon gamma (IFNγ) levels in lipopolysaccharide (LPS) activated microglia [47].…”

“…In light of these assumptions regarding astrocytes, a recent report by Pannel et al about the significantly higher TSPO expression in proinflammatory polarized cultured murine astrocytes measured by immunohistochemistry assays, together with their increased uptake of 18F-DPA-713 PET tracer, notably enhances the potential utility of TSPO imaging in neuroinflammation [39]. This study also revealed that astrocytes stimulated by the anti-inflammatory IL-4 do not share these TSPO-overexpressing features; however, the authors noted the possible differences in the phenotype of these cells to A2 astrocytes as reported by Liddelow et al [29,39]. In summary, astrocytes can not only provide additional answers to the cellular mechanisms involved in the inflammatory response in the brain but are also worthy targets for TSPO PET ligand research.…”

PET Imaging of Peripheral Benzodiazepine Receptor Standard Uptake Value Increases After Controlled Cortical Impact, a Rodent Model of Traumatic Brain Injury.

“…TSPO/PBR, an often PETimaged protein is upregulated after injury [15,16]. Recent evidence from Pannell et al (2020) demonstrated that TSPO is upregulated in astrocytes and microglia when stimulated with lipopolysaccharide (LPS), and this is re ected with an increase in PET imaging of TSPO ligand 18 F-DPA-713 [21]. In addition, they demonstrated TSPO expression was signi cantly increased in microglia after AdTNF injections [21].…”

“…Recent evidence from Pannell et al (2020) demonstrated that TSPO is upregulated in astrocytes and microglia when stimulated with lipopolysaccharide (LPS), and this is re ected with an increase in PET imaging of TSPO ligand 18 F-DPA-713 [21]. In addition, they demonstrated TSPO expression was signi cantly increased in microglia after AdTNF injections [21]. These data indicate that TSPO is a good marker to target in order to observe increases of pro-in ammatory microglia in vivo.…”

Elevated serum chemokine CCL22 levels in first-episode psychosis: associations with symptoms, peripheral immune state and in vivo brain glial cell function

“…Complicating the issue further, different TSPO antibodies have different selectivities for TSPO in different cell lines 87 . Also, TSPO immunoreactivity can present as elevated or decreased expression in astrocytes, microglial cells and vascular endothelial cells as a response to some immunological stimuli, but not necessarily others 16,87,90 . A recent study compared TSPO PET of schizophrenia patients and controls with combined peripheral and CSF measures of IL1β, IFNγ, IL-10, IL-6 and TNF-α.…”