PET imaging of lung inflammation with [18F]FEDAC, a radioligand for translocator protein (18 kDa)

Abstract

Purpose: The translocator protein (18 kDa) (TSPO) is highly expressed on the bronchial and bronchiole epithelium, submucosal glands in intrapulmonary bronchi, pneumocytes and alveolar macrophages in human lung. This study aimed to perform positron emission tomography (PET) imaging of lung inflammation with [(18)F]FEDAC, a specific TSPO radioligand, and to determine cellular sources enriching TSPO expression in the lung.

Methods: An acute lung injury model was prepared by intratracheal administration of lipopolysaccharide (LPS) to rat. Uptake of radioactivity in the rat lungs was measured with small-animal PET after injection of [(18)F]FEDAC. Presence of TSPO was examined in the lung tissue using Western blot and immunohistochemical assays.

Results: The uptake of [(18)F]FEDAC increased in the lung with the progress of inflammation by treatment with LPS. Pretreatment with a TSPO-selective ligand PK11195 showed a significant decrease in the lung uptake of [(18)F]FEDAC due to competitive binding to TSPO. TSPO expression was elevated in the inflamed lung section and its level responded to the [(18)F]FEDAC uptake and severity of inflammation. Increase of TSPO expression was mainly found in the neutrophils and macrophages of inflamed lungs.

Conclusion: From this study we conclude that PET with [(18)F]FEDAC may be a useful tool for imaging TSPO expression and evaluating progress of lung inflammation. Study on human lung using [(18)F]FEDAC-PET is promising.

Figure 1. Representative coronal PET lung images acquired between 0 and 30 min after injection of radioligand.

[¹⁸F]FEDAC: (A) control; (B) LPS-2 h; (C) LPS-6 h; (D) LPS-24 h inducement; (E) Pretreatment with PK11195 for control; (F) PK11195 for LPS-6 h; (G) PK11195 for LPS-24 h. [¹¹C](R)-PK11195: (H) control; (I) LPS-24 h; (J) Pretreatment with PK11195 for control; (K) PK11195 for LPS-24 h. The lung became visible over time after the LPS inducement. Contrast of the [¹⁸F]FEDAC images between the control and LPS-24 h was greater than that for the [¹¹C](R)-PK11195 images. Inhibitory experiment with PK11195 diminished the difference in radioactivity among all lung images. [¹⁸F]FEDAC showed higher lung uptake than [¹¹C](R)-PK11195.

Figure 2. Radioactivity accumulating in the lungs of the control and inflammatory rats.

The values of areas under time-activity curves (AUC_{0–30 min}; SUV×min, mean ± SEM., n = 4 for each group) were calculated from the time-activity curves between 0 and 30 min after injection of [¹⁸F]FEDAC (white bars) and [¹¹C](R)-PK11195 (black bars). The lung uptake became higher with time after LPS inducement. Unlabelled PK11195 treatment largely reduced the lung uptake. A significant difference (P<0.05) was seen on the following comparisons, *: control vs. LPS-2 h, 6 h, or 24 h, PK11195 treatment for control, LPS-6 h or 24 h; #: LPS-6 h or 24 h vs. PK11195 treatment for LPS-6 h or 24 h; ##: [¹⁸F]FEDAC vs. [¹¹C](R)-PK11195 for LPS-24 h.

Figure 3. TSPO expression of the control and LPS-induced lungs by Western blot assay.

(A) Expression of TSPO and b-actin in lungs. (B) Relative levels of TSPO in the lung sections (n = 4 for each group). The relative intensity of LPS-2 h, 6 h and 24 h vs. control was compared using each corrected band intensity (TSPO/beta-actin). A low level of TSPO was noted on the band of the control lung. Increase in the TSPO expression was observed over time after LPS inducement. The relative intensities on the bands of LPS-2 h, 6 h, and 24 h, showing a stepwise increase of TSPO expression compared to the control band.

Figure 4. TSPO expression in the lung tissues determined by immunohistochemical assay.

(A) TSPO was seen in the intrapulmonary bronchial and bronchiole epithelium, and bronchus-associated lymphoid tissue of the control. (D) Histological alteration was shown on the H&E staining image of the LPS-2 h section. (B, D) TSPO over the alveoli in LPS-2 h increased higher than that in the control, with few over the blood in the artery. (C, D) Many neutrophils over the alveoli and blood in the artery were detected in LPS-2 h. Abbreviations: bronchial and bronchiole epithelium (Ep), bronchus-associated lymphoid tissue (Balt), arterial wall (Ar w), alveoli (Al), blood in artery (Bl). Scale bar: 1 mm.

Figure 5. Comparison of the immunofluorescence labeling of TSPO and chloroacetate esterase staining for neutrophils.

Arrows indicate examples of cells doubly positive for TSPO (green) and chloroacetate esterase (red spots) staining. (A–D) No neutrophils were seen in the control. (E–H) Many neutrophils showing TSPO immunoreactivity were observed in the alveoli walls of LPS-2 h. (I–L) Some alveoli were collapsed in LPS-6 h, and a considerable number of neutrophils with TSPO infiltrated into the alveolar spaces. (M–P) Further alveoli collapse and neutrophil infiltration were seen in LPS-24 h. Scale bar: 20 µm.

Figure 6. Double immunofluorescence labeling of ED1 (red) and TSPO (green) displayed as a two-channel image.

The images demonstrated temporal alterations of TSPO expression in macrophages at 2 h (D–F), 6 h (G–I) and 24 h (J–L) after LPS inducement, and in the control (A–C). Arrows indicate examples of cells doubly positive for ED1 and TSPO. (A–C) TSPO expression was seen in a few macrophages. (D–F) Several macrophages showing TSPO immunoreactivity were observed in the alveolar spaces. (G–I, J–L) Numbers of macrophages with TSPO increased gradually over time. Scale bar: 20 µm.