Using the same ultrasound detector, hybrid optoacoustic-ultrasound (OPUS) imaging provides concurrent scans of tissue slices or volumes and visualizes complementary sound- and light-based contrast at similar resolutions. In addition to the benefit of hybrid contrast, spatial co-registration enables images from one modality to be employed as prior information for improving an aspect of the performance of the other modality. We consider herein a handheld OPUS system and utilize structural information from ultrasound images to guide regional Laplacian regularization-based reconstruction of optoacoustic images. Using phantoms and data from OPUS scans of human radial and carotid arteries, we show that ultrasound-driven optoacoustic inversion reduces limited-view artefacts and improves image contrast. In phantoms, prior-integrated reconstruction leads to a 50 % higher contrast-to-noise ratio (CNR) of the image than standard reconstruction, and a 17 % higher structural similarity (SSIM) index. In clinical data, prior-integrated reconstruction detects deep-seated radial arteries with higher CNR than the standard method at three different depths. In this way, the prior-integrated method offers unique insights into atherosclerotic carotid plaques in humans (with p<0.01 between patients and healthy volunteers), potentially paving the way for new abilities in vascular imaging and more generally in optoacoustic imaging.

Multispectral optoacoustic tomography (MSOT) represents a new imaging approach revealing functional tissue information without extrinsic contrast agents. Using a clinical combined ultrasound (US)/MSOT device, we investigated the interindividual robustness and impact of intra- and interobserver variability of MSOT values in soft tissue (muscle and subcutaneous fat) of healthy volunteers. Semiquantitative MSOT values for deoxygenated (Hb), oxygenated (HbO₂) and total hemoglobin (HbT), as well as oxygen saturation (sO₂), were calculated for both forearms in transversal and longitudinal probe orientation (n = 3, 8 measurements per subject). For intraobserver reproducibility, the same examiner investigated three subjects twice. Mean values of left vs. right forearm and transversal vs. longitudinal probe orientation were compared using an unpaired Student’s t test. Bland Altmann plots with 95% limits of agreement for absolute averages and differences were calculated. Intraclass correlation coefficients (ICC 2,k) were computed for three different examiners. We obtained reproducible and consistent MSOT values with small-to-moderate deviation for muscle and subcutaneous fat tissue. Probe orientation and body side had no impact on calculated MSOT values (p > 0.05 each). Intraobserver reproducibility revealed equable mean values with small-to-moderate deviation. For muscular tissue, good ICC was obtained for sO₂. Measurements of subcutaneous tissue revealed good-to-excellent ICCs for all calculated values. Thus, in this preliminary study on healthy individuals, clinical MSOT provided consistent and reproducible functional soft tissue characterization, independent on the investigating personnel.

Purpose: As a promotor of tumor invasion and tumor microenvironment (TME) formation, the protein complex S100A8/S100A9 is associated with poor prognosis. Our aim was to further evaluate its origin and regulatory effects, and to establish an imaging biomarker for TME activity. Methods: S100A9^−/−cells (ko) were created from syngeneic murine breast cancer 4T1 (high malignancy) and 67NR (low malignancy) wildtype (wt) cell lines and implanted into either female BALB/c wildtype or S100A9^−/− mice (n = 10 each). Anti-S100A9-Cy5.5-targeted fluorescence reflectance imaging was performed at 0 h and 24 h after injection. Potential early changes of S100A9-presence under immune checkpoint inhibition (anti-PD-L1, n = 7 vs. rat IgG2b as isotype control, n = 3) were evaluated. Results: In S100A9^−/−mice contrast-to-noise-ratios were significantly reduced for wt and S100A9^−/−tumors. No significant differences were detected for 4T1 ko and 67NR ko cells as compared to wildtype cells. Under anti-PD-L1 treatment S100A9 presence significantly decreased compared with the control group. Conclusion: Our results confirm a secretion of S100A8/S100A9 by the TME, while tumor cells do not apparently release the protein. Under immune checkpoint inhibition S100A9-imaging reports an early decrease of TME activity. Therefore, S100A9-specific imaging may serve as an imaging biomarker for TME formation and activity.

Cardiovascular disease remains the most frequent cause of death worldwide. Atherosclerosis, an underlying cause of cardiovascular disease, is an inflammatory disorder associated with endothelial dysfunction. The endothelin system plays a crucial role in the pathogenesis of endothelial dysfunction and is involved in the development of atherosclerosis. We aimed to reveal the expression levels of the endothelin-A receptor (ET_AR) in the course of atherogenesis to reveal possible time frames for targeted imaging and interventions. We used the ApoE^−/− mice model and human specimens and evaluated ET_AR expression by quantitative rtPCR (qPCR), histology and fluorescence molecular imaging. We found a significant upregulation of ET_AR after 22 weeks of high-fat diet in the aortae of ApoE^−/− mice. With regard to translation to human disease, we applied the fluorescent probe to fresh explants of human carotid and femoral artery specimens. The findings were correlated with qPCR and histology. While ET_AR is upregulated during the progression of early atherosclerosis in the ApoE^−/− mouse model, we found that ET_AR expression is substantially reduced in advanced human atherosclerotic plaques. Moreover, those expression changes were clearly depicted by fluorescence imaging using our in-house designed ET_AR-Cy 5.5 probe confirming its specificity and potential use in future studies.

(1) Background: The prognosis of cancer is dependent on immune cells in the tumor microenvironment (TME). The protein S100A9 is an essential regulator of the TME, associated with poor prognosis. In this study, we evaluated early therapy effects on the TME in syngeneic murine breast cancer via S100A9-specific in vivo imaging. (2) Methods: Murine 4T1 cells were implanted orthotopically in female BALB/c mice (n = 59). Tumor size-adapted fluorescence imaging was performed before and 5 days after chemo- (Doxorubicin, n = 20), anti-angiogenic therapy (Bevacizumab, n = 20), or placebo (NaCl, n = 19). Imaging results were validated ex vivo (immunohistochemistry, flow cytometry). (3) Results: While tumor growth revealed no differences (p = 0.48), fluorescence intensities (FI) for S100A9 in Bevacizumab-treated tumors were significantly lower as compared to Doxorubicin (2.60 vs. 15.65 AU, p < 0.0001). FI for Doxorubicin were significantly higher compared to placebo (8.95 AU, p = 0.01). Flow cytometry revealed shifts in monocytic and T-cell cell infiltrates under therapy, correlating with imaging. (4) Conclusions: S100A9-specific imaging enables early detection of therapy effects visualizing immune cell activity in the TME, even before clinically detectable changes in tumor size. Therefore, it may serve as a non-invasive imaging biomarker for early therapy effects.