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Optoacoustic imaging (OAI), or photoacoustic imaging (PAI), has fundamentally influenced basic science by providing high-resolution visualization of biological mechanisms. With the introduction of multispectral optoacoustic tomography (MSOT), these technologies have now moved closer to clinical applications. MSOT utilizes short-pulsed near-infrared laser light to induce thermoelastic expansion in targeted tissues. This results in acoustic pressure waves, which are used to resolve specific endo- and exogenous chromophores. Especially in the pediatric population, this non-invasive imaging approach might hold fundamental advantages compared to conventional cross-sectional imaging modalities. As this technology allows the visualization of quantitative molecular tissue composition at high spatial resolution non-invasively in sufficient penetration depth, it paves the way to personalized medicine in pediatric diseases.

This study aimed at evaluating hybrid multispectral optoacoustic tomography/ultrasound for imaging of thyroid disorders, including Graves' disease and thyroid nodules. Methods: The functional biomarkers and tissue parameters deoxygenated hemoglobin, oxygenated hemoglobin, total hemoglobin, saturation of hemoglobin, fat content, and water content were analyzed in thyroid lobes affected by Graves' disease (n = 6), thyroid lobes with healthy tissue (n = 8), benign thyroid nodules (n = 13), and malignant thyroid nodules (n = 3). Results: In Graves' disease, significantly higher deoxygenated hemoglobin (3.18 ± 0.52 vs. 2.13 ± 0.62; P = 0.0055) and total hemoglobin (8.34 ± 0.88 vs. 6.59 ± 1.16; P = 0.0084) and significantly lower fat content (0.64 ± 0.37 vs. 1.69 ± 1.25; P = 0.0293) were found than in healthy controls. Malignant thyroid nodules showed significantly lower saturation of hemoglobin (55.4% ± 2.6% vs. 60.8% ± 7.2%; P = 0.0393) and lower fat content (0.62 ± 0.19 vs. 1.46 ± 0.87; P = 0.1295) than benign nodules. Conclusion: This pilot study showed the applicability and the potential of hybrid multispectral optoacoustic tomography/ultrasound to semiquantitatively provide tissue characterization and functional parameters in thyroid disorders for improved noninvasive diagnostics of thyroid diseases.

The emerging research field of functional micro devices and microbots aims at developing remote-controlled microsensors and actuators that can operate inside living organisms for biomedical applications. However, a significant gap remains in controlling with high accuracy such microdevices in complex environments. This is primarily due to the lack of high spatial and temporal resolution techniques. Here we demonstrate the detection of synthetic microstructures with sizes between 10 and 50 µm size, in deep tissue-mimicking phantoms, using multispectral optoacoustic tomography (MSOT). When referring to guidance/propulsion, cargo-delivery, and imaging capabilities, these microstructures represent key components of the physical, chemical and biohybrid microbots. They are conceived for diverse in vivo medical applications, including non-invasive diagnosis and therapy.

A preliminary study suggests that intestinal-wall assessment by means of noninvasive multispectral optoacoustic tomography may distinguish remission from active disease in patients with Crohn’s disease.

Medical imaging plays an important role in diagnosis and treatment of multiple diseases. It is a field under continuous development which seeks for improved sensitivity and spatiotemporal resolution to allow the dynamic monitoring of diverse biological processes that occur at the micro- and nanoscale. Emerging technologies for targeted diagnosis and therapy such as nanotherapeutics, micro-implants, catheters and small medical tools also need to be precisely located and monitored while performing their function inside the human body. In this work, we show for the first time the real-time tracking of moving single micro-objects below centimeter thick tissue-mimicking phantoms, using multispectral optoacoustic tomography (MSOT). This technique combines the advantages of ultrasound imaging regarding depth and resolution with the molecular specificity of optical methods, thereby facilitating the discrimination between the spectral signatures of the micro-objects from those of intrinsic tissue molecules. The resulting MSOT signal is further improved in terms of contrast and specificity by coating the micro-objects’ surface with gold nanorods, possessing a unique absorption spectrum, which will allow their discrimination from surrounding biological tissues when translated to in vivo settings.

Medical imaging plays an important role in diagnosis and treatment of multiple diseases. It is a field which seeks for improved sensitivity and spatiotemporal resolution to allow the dynamic monitoring of diverse biological processes that occur at the micro- and nanoscale. Emerging technologies for targeted diagnosis and therapy such as nanotherapeutics, microimplants, catheters, and small medical tools also need to be precisely located and monitored while performing their function inside the human body. In this work, we show for the first time the real-time tracking of moving single micro-objects below centimeter thick phantom tissue and ex vivo chicken breast, using multispectral optoacoustic tomography (MSOT). This technique combines the advantages of ultrasound imaging regarding depth and resolution with the molecular specificity of optical methods, thereby facilitating the discrimination between the spectral signatures of the micro-objects from those of intrinsic tissue molecules. The resulting MSOT signal is further improved in terms of contrast and specificity by coating the micro-objects' surface with gold nanorods, possessing a unique absorption spectrum, which facilitate their discrimination from surrounding biological tissues when translated to future in vivo settings.

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.

Background & Aims Intestinal fibrosis is a long-term complication in inflammatory bowel diseases (IBD) that frequently results in functional damage, bowel obstruction, and surgery. Interleukin (IL) 36 is a group of cytokines in the IL1 family with inflammatory effects. We studied the expression of IL36 and its receptor, interleukin 1 receptor like 2 (IL1RL2 or IL36R) in the development of intestinal fibrosis in human tissues and mice. Methods We obtained intestinal tissues from 92 patients with Crohn's disease (CD), 48 patients with ulcerative colitis, and 26 patients without inflammatory bowel diseases (control individuals). Tissues were analyzed by histology to detect fibrosis and by immunohistochemistry to determine the distribution of fibroblasts and levels of IL36R ligands. Human and mouse fibroblasts were incubated with IL36 or control medium, and transcriptome-wide RNA sequences were analyzed. Mice were given neutralizing antibodies against IL36R, and we studied intestinal tissues from Il1rl2–/– mice; colitis and fibrosis were induced in mice by repetitive administration of DSS or TNBS. Bone marrow cells were transplanted from Il1rl2–/– to irradiated wild-type mice and intestinal tissues were analyzed. Antibodies against IL36R were applied to mice with established chronic colitis and fibrosis and intestinal tissues were studied. Results Mucosal and submucosal tissue from patients with CD or ulcerative colitis had higher levels of collagens, including type VI collagen, compared with tissue from control individuals. In tissues from patients with fibrostenotic CD, significantly higher levels of IL36A were noted, which correlated with high numbers of activated fibroblasts that expressed α-smooth muscle actin. IL36R activation of mouse and human fibroblasts resulted in expression of genes that regulate fibrosis and tissue remodeling, as well as expression of collagen type VI. Il1rl2–/– mice and mice given injections of an antibody against IL36R developed less severe colitis and fibrosis after administration of DSS or TNBS, but bone marrow cells from Il1rl2–/– mice did not prevent induction of colitis and fibrosis. Injection of antibodies against IL36R significantly reduced established fibrosis in mice with chronic intestinal inflammation. Conclusion We found higher levels of IL36A in fibrotic intestinal tissues from patients with IBD compared with control individuals. IL36 induced expression of genes that regulate fibrogenesis in fibroblasts. Inhibition or knockout of the IL36R gene in mice reduces chronic colitis and intestinal fibrosis. Agents designed to block IL36R signaling could be developed for prevention and treatment of intestinal fibrosis in patients with IBD.

The study aimed to evaluate the clinical feasibility of hybrid ultrasound/multispectral optoacoustic tomography (MSOT) for assessing microvascular dysfunction in systemic sclerosis (SSc). A handheld US/MSOT imaging system was applied for imaging patients diagnosed with SSc (n = 7) and healthy volunteers (n = 8). Semiquantitative MSOT values for deoxygenated (HbR), oxygenated (HbO(2)) and total haemoglobin (HbT) were analysed for subcutaneous finger tissue of both hands (8 fingers per subject, 120 fingers in total) and used to assess disease activity (progressive vs stable). Grouped data were compared by one-way nested analysis of variance, Tukey post-hoc test as well as student's t test were used for statistical analysis.Subcutaneous finger tissue of patients with SSc provided significantly lower MSOT values for HbO(2) (26.16 +/- 0.71 vs 38.2 +/- 1.54, P =.023) and HbT (55.92 +/- 1.62 vs 72.46 +/- 1.90, P =.018) compared to healthy volunteers. Patients with progressive SSc had significantly lower MSOT values compared to patients with stable disease and healthy volunteers.This pilot study shows the feasibility of MSOT imaging to resolve microvascular dysfunction in SSc as a marker of disease activity. By providing biological tissue properties not revealed by other imaging modalities, MSOT might help to grade SSc non-invasively and monitor early therapy response.

Graphical abstract, A common side effect of medication is gastrointestinal intolerance. Symptoms can include reduced appetite, diarrhea, constipation, GI inflammation, nausea and vomiting. Such effects often have a dramatic impact on compliance with a treatment regimen. Therefore, characterization of GI tolerance is an important step when establishing a novel therapeutic approach. In this study, Multispectral Optoacoustic Tomography (MSOT) is used to monitor gastrointestinal motility by in vivo whole body imaging in mice. MSOT combines high spatial and temporal resolution based on ultrasound detection with strong optical contrast in the near infrared. Animals were given Indocyanine Green (ICG) by oral gavage and imaged by MSOT to observe the fate of ICG in the gastrointestinal tract. Exponential decay of ICG signal was observed in the stomach in good correlation with ex vivo validation. We discuss how kinetic imaging in MSOT allows visualization of parameters unavailable to other imaging methods, both in 2D and 3D.

Reliably assessing tissue viability during surgery is of major importance in surgical procedures. The most basic requirement for viability is sufficient oxygen supply to the tissue. Therefore it is highly desirable to visualize in real-time the dynamic process of blood perfusion up to and within the microvasculature. A modality sensitive to structures in the range of few hundred micrometers and offering high contrast to the embedding tissue is then needed. To this end, a number of methods have been developed, but have had no significant impact on the clinical routine due to various deficiencies. In this paper we demonstrate the applicability of optoacoustic imaging, which combines ultrasonic resolution with strong optical contrast. A method for optoacoustic perfusion assessment, based on a local and repeatable injection of saline, was proposed and assessed ex-vivo on large pig bowels and in-vivo in mouse tails. The obtained dynamic perfusion images highlight the method's potential to enable immediate and quantitative assessment of tissue viability during surgery.

Multispectral optoacoustic tomography (MSOT) represents a new in vivo imaging technique with high resolution (~250 μm) and tissue penetration (>1 cm) using the photoacoustic effect. While ultrasound contains anatomical information for lesion detection, MSOT provides functional information based on intrinsic tissue chromophores. We aimed to evaluate the feasibility of combined ultrasound/MSOT imaging of breast cancer in patients compared to healthy volunteers. Imaging was performed using a handheld MSOT system for clinical use in healthy volunteers (n = 6) and representative patients with histologically confirmed invasive breast carcinoma (n = 5) and ductal carcinoma in situ (DCIS, n = 2). MSOT values for haemoglobin and oxygen saturation were assessed at 0.5, 1.0 and 1.5 cm depth and selected wavelengths between 700 and 850 nm. Reproducible signals were obtained in all wavelengths with consistent MSOT signals in superficial tissue in breasts of healthy individuals. In contrast, we found increased signals for haemoglobin in invasive carcinoma, suggesting a higher perfusion of the tumour and tumour environment. For DCIS, MSOT values showed only little variation compared to healthy tissue. This preliminary MSOT breast imaging study provided stable, reproducible data on tissue composition and physiological properties, potentially enabling differentiation of solid malignant and healthy tissue. • A handheld MSOT probe enables real-time molecular imaging of the breast. • MSOT of healthy controls provides a reproducible reference for pathology identification. • MSOT parameters allows for differentiation of invasive carcinoma and healthy tissue.

Item does not contain fulltext A crucial point for the management of pancreatic ductal adenocarcinoma (PDAC) is the decrease of R1 resections. Our aim was to evaluate the combination of multispectral optoacoustic tomography (MSOT) with fluorescence guided surgery (FGS) for diagnosis and perioperative detection of tumour nodules and resection margins in a xenotransplant mouse model of human pancreatic cancer. The peptide cRGD, conjugated with the near infrared fluorescent (NIRF) dye IRDye800CW and with a trans-cyclooctene (TCO) tag for future click chemistry (cRGD-800CW-TCO), was applied to PDAC bearing immunodeficient nude mice. 27 days after orthotopic transplantation of human AsPC-1 cells into the head of the pancreas, mice were injected with cRGD-800CW-TCO and imaged with fluorescence- and optoacoustic devices before and 2h, 6h and 24h after injection, before they were sacrificed and dissected with a guidance of FGS imaging system. Fluorescence imaging of cRGD-800CW-TCO allowed detection of the tumour area but without information about the depth, whereas MSOT allowed high resolution 3D identification of the tumour area, in particular of small tumour nodules. Highly sensitive delineation of tumour burden was achieved during FGS in all mice. Imaging of whole-mouse cryosections, histopathological analysis and NIRF microscopy confirmed the localization of cRGD-800CW-TCO within the tumour tissue. In principle, all imaging modalities applied here were able to detect PDAC in vivo. However, the combination of MSOT and FGS provided detailed spatial information of the signal, and achieved a complete overview of the distribution and localization of cRGD-800CW-TCO within the tumour before and during surgical intervention. This article is protected by copyright. All rights reserved.

The optic lobe of Drosophila houses about 60,000 neurons that are organized in parallel, retinotopically arranged columns. Based on the Golgi-staining method, Fischbach and Dittrich ([1989] Cell Tissue Res 258:441–475) determined that each column contains about 90 identified cells. Each of these cells is supposed to release one or two different neurotransmitters. However, for most cells the released neurotransmitter is not known. Here we characterize the vast majority of the neurons in the Drosophila optic lobe that release acetylcholine (Ach), the major excitatory neurotransmitter of the insect central nervous system. We employed a promoter specific for cholinergic neurons and restricted its activity to single or a few cells using the MARCM technique. This approach allowed us to establish an anatomical map of neurons with a cholinergic phenotype based on their branching pattern. We identified 43 different types of neurons with a cholinergic phenotype. Thirty-one of them match previously described members of nine different subgroups: Transmedullary (Tm), Transmedullary Y (TmY), Medulla intrinsic (Mi, Mt, and Pm), Bushy T (T), Translobula Plate (Tlp), and Lobula intrinsic (Lcn and Lt) neurons (Fischbach and Dittrich [1989]). Intriguingly, 12 newly identified cell types suggest that previous Golgi studies were not saturating and that the actual number of different neurons per column is higher than previously thought. This study and similar ones on other neurotransmitter systems will contribute towards a columnar wiring diagram and foster the functional dissection of the visual circuitry in Drosophila. J. Comp. Neurol. 519:162-176, 2011. © 2010 Wiley-Liss, Inc.

IMPORTANCE: Differential diagnosis of congenital vascular anomalies is challenging, and misdiagnosis is frequent. Vascular malformations are considered one of the most difficult vascular diseases to treat. A new imaging approach that visualizes anatomical features and quantitatively assesses molecular biomarkers noninvasively would aid diagnosis and monitoring of treatment response of vascular malformations. OBJECTIVE: To evaluate multispectral optoacoustic tomography (MSOT) for noninvasive assessment of molecular biomarkers for diagnosis and therapeutic monitoring of vascular malformations. DESIGN, SETTING, AND PARTICIPANTS: This pilot study examined 6 patients with arteriovenous malformation (AVM) and 6 patients with venous malformation (VM) diagnosed according to the classification system of the International Society for the Study of Vascular Anomalies. All patients underwent clinical hybrid MSOT/ultrasonographic (US) imaging before and after treatment at an interdisciplinary vascular malformations clinic by trained MSOT and US examiners. Examiners were blinded to the patient history and stage of disease. Data were collected from April 11 to 25, 2017, and analyzed from May 1 to October 31, 2017. INTERVENTIONS: Clinical hybrid MSOT/US imaging was performed before or within 1 week after endovascular embolization (for AVM) or percutaneous sclerotherapy (for VM). MAIN OUTCOMES AND MEASURES: Region-of-interest analysis of the lesion and contralateral healthy tissue revealed quantitative values for oxygenated (HbO(2)) and deoxygenated (HbR) hemoglobin by spectral unmixing of optoacoustic data acquired at multiple wavelengths. The HbO(2):HbR ratio was calculated for healthy tissue and for AVM and VM before and after treatment. RESULTS: Twelve patients (9 female and 3 male; mean [SD] age, 23 [18] years; age range, 6-59 years) with vascular malformations (6 with AVMs and 6 with VMs) were included. Significantly higher HbO(2):HbR ratios for AVMs (mean [SEM], 1.82 [0.08] vs 0.89 [0.03]; P

We present a hybrid preclinical imaging scanner that optimally supports image acquisition in both reflection-mode ultrasonography and optoacoustic (OA) tomography modes. The system comprises a quasi-full-ring tomographic geometry capable of the simultaneous dual-mode imaging through entire cross sections of mice with in-plane spatial resolution in the range of 150 and 350 μm in the respective OA and ultrasound (US) imaging modes with an imaging speed of up to 10 two-dimensional frames per second. Three-dimensional whole-body data is subsequently rendered by rapid scanning of the imaged plane. The system further incorporates rapid laser wavelength tuning for real-time acquisition of multispectral OA data, which enables studies of longitudinal dynamics as well as fast kinetics and biodistribution of contrast agents. In vivo imaging performance is demonstrated by label-free hybrid anatomical scans through living mice, as well as real-time visualization of optical contrast agent perfusion. By setting new standards for whole-body tomographic imaging performance in both the OA and pulse-echo US modes, the developed hybrid imaging approach is expected to benefit numerous applications where the availability of high-quality structural information provided by the tomographic reflection-mode US can ease interpretation of the functional and molecular imaging results attained by the OA modality.

Using optoacoustic excitation, a complete volumetric tomographic data sets from the imaged object can in principle be generated with a single interrogating laser pulse. Thus, optoacoustic imaging intrinsically has the potential for fast three-dimensional imaging. We have developed a system capable of acquiring volumetric optoacoustic data in real time and showcase in this work the undocumented capacity to generate high resolution three-dimensional optoacoustic images at a rate of 10 Hz, currently mainly limited by the pulse repetition rate of the excitation laser.

Cardiac imaging in small animals is a valuable tool in basic biological research and drug discovery for cardiovascular disease. Multispectral optoacoustic tomography (MSOT) represents an emerging imaging modality capable of visualizing specific tissue chromophores at high resolution and deep in tissues in vivo by separating their spectral signatures. Whereas single-wavelength images can be acquired by multielement ultrasound detection in real-time imaging, using multiple wavelengths at separate times can lead to image blurring due to motion during acquisition. Therefore, MSOT imaging of the heart results in degraded resolution because of the heartbeat. In this work, we applied a clustering algorithm, k-means, to automatically separate a sequence of single-pulse images at multiple excitation wavelengths into clusters corresponding to different stages of the cardiac cycle. We then performed spectral unmixing on each cluster to obtain images of tissue intrinsic chromophores at different cardiac stages, showing reduced sensitivity to motion compared to signal averaging without clustering. We found that myocardium images of improved resolution and contrast can be achieved using MSOT motion clustering correction. The correction method presented could be generally applied to other MSOT imaging applications prone to motion artifacts, for example, by respiration and heartbeat.

Melanoma accounts for less than 5% of skin cancer cases, yet it causes more than 75% of skin cancer death, and its incidence is growing faster than any other cancer in the world. Because melanoma metastasizes early into regional/sentinel lymph nodes (SLN), SLN excision (SLNE) is probably the most important diagnostic procedure for melanoma patients, as histology provides the most relevant prognostic factor for the survival of melanoma patients. However, 50% of excised lymph nodes show no evidence of metastasis; and, current histological protocols involve sampling only a small portion of the SLN, resulting in a relatively high false negative rate. Furthermore, SLNE exposes the patients to complications such as swelling, edema and future risk of infection. Therefore, a clear need exists to improve the sensitivity and specificity of SLN analysis. Multispectral optoacoustic tomography (MSOT) ulitizes the molecular specificity of optical imaging, while capitalizing on the high temporal and spatial resolution of ultrasound imaging. This method allows sensitive detection of optical markers such as melanin, which would identify potentially metastatic lymph nodes, and indocyanine green (ICG), which can potentially label SLN. In this study, 148 lymph nodes were excised from 65 melanoma patients (stage I - IV), and the lymph nodes were examined by MSOT ex vivo to guide the pathologist to examination of melanin-containing regions of the lymph node, thereby increasing the detection rate in histological analysis. Compared to standard histology, MSOT demonstrated a superior 100% sensitivity/47% specificity. Further, 21 melanoma patients were scanned with a handheld MSOT device in vivo. MSOT was able to detect sentinel lymph nodes using ICG specific contrast, with the added ability of non-invasive assessment of the melanin status prior to excision. In vivo MSOT measurements using a 2D and a 3D detector were compared with ultrasound, SPECT/CT, planar fluorescence imaging and ex vivo histology, with a promising concordance between in vivo MSOT measurements and in vivo and ex vivo gold standard assessments. MSOT represents a viability modality to improve histological analysis of excised SLN in melanoma patients, and it offers the ability to stage lymph nodes noninvasively, potentially reducing the necessity to excise lymph nodes in some melanoma patients. Citation Format: Stefan Morscher, Ingo Stoffels, Neal C. Burton, Jing Claussen, Thomas Sardella, Iris Helfrich, Uwe Hillen, Julia Leyh, Dirk Schadendorf, Matthias Gunzer, Joachim Klode. Sentinel lymph node detection and in vivo/ex vivo assessment of melanin distribution by means of multispectral optoacoustic tomography (MSOT) in patients with malignant melanoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5254. doi:10.1158/1538-7445.AM2015-5254

Invivo optical imaging modalities are mostly limited to cell cultures, superficial tissues, and intravital imaging since they lack either resolution or penetration depth.1 In contrast, optoacoustic (OA) imaging—combining features of optical and ultrasound imaging—has been used to visualize hemoglobin in depths of approximately 3 cm in patients with Crohn’s disease.2,3 Realizing an even higher resolution, raster-scanning OA mesoscopy (RSOM) provides intrinsic optical tissue contrast down to 10-20 μm resolution at still high penetration depths of several millimeters.