Name: multispectral optoacoustic tomography for functional imaging in vascular research
Uploaded: 2022-06-08T14:10:00
Description: Watch this Scientific Journal Video about Multispectral Optoacoustic Tomography for Functional Imaging in Vascular Research at JoVE.com

This research is funded by ALIES and COFAC principal providers of the technology under study, and by Funda&#231;&#227;o para a Ci&#234;ncia e a Tecnologia (FCT) through the grant UIDB/04567/2020 to CBIOS.

The experimental protocol was previously approved by the Ethics Committee of the University&#39;s School of Health Sciences (EC.ECTS/P10.21). Procedures fully respected the principles of good clinical practice defined for human research13. A convenient sample of six healthy participants of both sexes (n = 3 per sex) with a mean age of 32.8 &#177; 11.9 years old was chosen from the university community. Selected participants were required to be normotensive, non-smokers, and free of any medication ...

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This protocol emphasizes the working steps regarded as practical requirements to operate this new optoacoustic imaging instrument, from the adequate positioning (participant, probe) needed for 3D cup probe stabilization to image acquisition, ROI selection, and image reconstruction and analysis.
The proposed experimental approach, using &#34;instantaneous&#34; acquisitions together with images obtained under dynamic conditions, illustrates the interest and utility of this instrument in accessin...

Cardiovascular diseases are recurrent top causes of death worldwide and represent a huge burden for any health system1,2. Technology has been a major contributor to the expansion of our understanding of fundamental cardiac and vascular pathophysiology, providing more precise diagnostic tools and the possibility of early disease detection and more effective management. Imaging techniques offer the possibility to measure not only cardiac and major vessel performance but also, on a much smaller scale, to calculate the capillary density, local perfusion and volume, and endothelial dysfunction, among other characteristics. These technologies have offered the first quantitative insights into vascular biology with direct clinical application. Changes in capillary density, local perfusion reduction, or occlusion likely correspond to an ischemic condition, which helps to explain the growing role of imaging, becoming an indispensable tool in cardiovascular research and practice3,4,5.
In recent years, functional imaging has consistently set the pace in technological innovation, with ultrasound (US) near-infrared spectroscopy (NIRS), positron emission tomography (PET), computed tomography (CT), and magnetic resonance imaging (MRI) as some well-known examples. However, multiple concerns limit their application, from cost and patient safety (as well as comfort) to image contrast and resolution6,7. Optoacoustic tomography (OT) has recently emerged as a new direction in optical-based vascular research. This technology, centered on the detection of ultrasonic waves generated by thermoelastic expansion of the tissue impacted with ultrashort laser pulses, has been known for some time6,8. This physical reaction of heat development and tissue expansion evokes an acoustic signal detected by an ultrasound transducer. The use of pulses of light from visible to near-infrared and the absence of an acoustic background signal benefit the resolution depth. The detected contrast results from the most important chromophores present (hemoglobin or melanin). Compared with other technologies, OT has the advantages of (1) needing no contrast (label-free imaging), (2) better contrast and resolution with fewer artifacts than ultrasonography, and (3) lower price, and faster acquisition and ease of operation6,9,10,11.
Multispectral optoacoustic tomography (MSOT) is among the most recent generation of OT instruments. Built with a tunable optical parametric oscillator (OPO) pumped by an Nd:YAG laser providing excitation pulses, a 3D image is acquired by time-resolved signals detected from high-frequency ultrasonic excitation pulses at wavelengths from 680 nm to 980 nm with a repetition rate of up to 50 Hz12. The optoacoustic imaging platform provides the quantification of different chromophores in-depth (as low as 15 mm). Variables such as HbO2, Hb, and melanin are easily accessible. Other variables of interest, such as maximal Oxygenated Hemoglobin (Max HbO2) and Deoxygenated Hemoglobin (Max Hb), are also available. Reconstruction algorithms from the manufacturer&#39;s software allow the calculation of other variables such as vascular density (&#181;Vu), inter-unit average distance (&#950;Ad), and capillary volume (mm3).
The present study explores the essential operating aspects of this new system to understand better its practicalities and potential applications in cardiovascular preclinical research.&#160;

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			<td>Cuff</td>
			<td>PIC</td>
			<td>107001</td>
			<td></td>
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			<td>Drapes</td>
			<td>Pajunk</td>
			<td>021151-1501</td>
			<td></td>
		</tr>
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			<td>Ethanol 70%</td>
			<td>Sigma Aldrich</td>
			<td>EX0281</td>
			<td></td>
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		<tr>
			<td>Gogless</td>
			<td>Univet</td>
			<td>559G.00.00.201</td>
			<td></td>
		</tr>
		<tr>
			<td>Kimwipes</td>
			<td>Amoos</td>
			<td>5601856202331.00</td>
			<td></td>
		</tr>
		<tr>
			<td>MSOT</td>
			<td>iThera</td>
			<td>MSOTAcuity</td>
			<td></td>
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		<tr>
			<td>Stabilizing arm</td>
			<td>ITEM</td>
			<td></td>
			<td>Self designed and assemble</td>
		</tr>
		<tr>
			<td>Ultrasound gel</td>
			<td>Parker Laboratories</td>
			<td>308</td>
			<td></td>
		</tr>
		<tr>
			<td>Waxing cream</td>
			<td>Veet</td>
			<td>kkdg08hagd</td>
			<td></td>
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multispectral optoacoustic tomography for functional imaging in vascular research

Microcirculatory impairment has been recognized in various disease processes, underlying this growing theme within vascular research. In recent years, the development of live imaging systems has set the (analytical) pace in both basic and clinical research, with the objective of creating new instruments capable of providing real-time, quantifiable endpoints with clinical interest and application. Near-infrared spectroscopy (NIRS), positron emission tomography (PET), computed tomography (CT), and magnetic resonance imaging (MRI) are available, among other techniques, but cost, image resolution, and reduced contrast are recognized as common challenges. Optoacoustic tomography (OT) offers a new perspective on vascular functional imaging, combining state-of-the-art optical absorption and spatial resolution capacities (from micrometer optical to millimeter acoustic resolution) with tissue depth. In this study, we tested the applicability of multispectral optoacoustic tomography (MSOT) for functional imaging. The system uses a tunable optical parametric oscillator (OPO) pumped by an Nd: YAG laser, providing excitation pulses sensed by a 3D probe at wavelengths from 680 nm to 980 nm. Images obtained from the human forearm were reconstructed through a specific algorithm (supplied within the manufacturer&#39;s software) based on the response of specific chromophores. Maximal Oxygenated Hemoglobin (Max HbO2) and Deoxygenated Hemoglobin (Max Hb), Total Hemoglobin (HbT), and mean Oxygen Saturation (mSO2) to vascular density (&#181;Vu), inter-unit average distances (&#950;Ad), and capillary blood volume (mm3) may be measured using this system. The applicability potential found with this OT system is relevant. Ongoing software developments will surely improve the utility of this imaging system.

Data provided by optoacoustic imaging can be analyzed in post-processed export images (Figure 2) and plotted data (Figure 3). The purpose here was to introduce the operation of optoacoustic functional imaging and to explore its application in more commonly known vascular research. For that, we compared images acquired during rest and after a 200 mmHg occlusion of a major supplying artery (Figure 2). These observations can be quantif...

Watch this Scientific Journal Video about Multispectral Optoacoustic Tomography for Functional Imaging in Vascular Research at JoVE.com

Multispectral Optoacoustic Tomography for Functional Imaging in Vascular Research

The present protocol describes the acquisition of multispectral optoacoustic images of in vivo human skin vasculature. These include the quantification of hemoglobin and melanin, regarded as chromophores of interest for functional analysis.

Microcirculatory impairment has been recognized in various disease processes, underlying this growing theme within vascular research. In recent years, the ...

This protocol using optoacoustic technology enables us to assess a patient's vascular status in real time with functional imaging. And we can explore 3D image scans collected in static or dynamic conditions. The potential interest of such an approach is huge and impacts many health-related fields.The system provides life in vivo visualization of skin vascular structure at different depths. That means that we can access both skin plexus, the superficial and the deeper, although that requires some repeated experience from the operator With detailed, truly functional imaging, early detection and characterization of microvascular impairment is possible and can be integrated as a diagnostic tool or follow-up protocol to evaluate the progression of disease, and/or effects of treatment. To load subject information, begin by turning on the optoacoustic imaging equipment.While the equipment is warming up, introduce the Participant Information. The main Welcome window of the software opens to the Scan Overview. Introduce data after clicking on Patient ID, and finish the application by pressing on Select.Make sure the laser is ready by checking the equipment screen. Following the warm-up time, the laser status bar on the equipment screen must change from laser standby. Select the optoacoustic acquisition preset, hemoglobin, oxyhemoglobin, and melanin, in the examination screen.Press the laser switch foot pedal and wait for the laser power self-check. After a few seconds, a window appears with the current laser status with a checkup report Release this window by pressing the available OK button. Acclimatize the participant to the laboratory environment, choosing a comfortable position to minimize unnecessary movement.Ensure that the area to be scanned is previously cleaned. Apply a thin layer of ultrasound gel to the 3D cup. Image stabilization is achieved by holding the 3D cup in the desired imaging position at the volunteer forearm.After placing the 3D cup on the area of interest, partially lock the stabilizing arm lock for image acquisition. Select the anatomical area for baseline image acquisition. For exploratory purposes, the ventral forearm is recommended.Apply minimal pressure to the imaging site so that higher pressure readouts can be properly acquired. When the image stability is maximized, capture a snapshot of the area by pressing the Snapshot button on the touchscreen. Acquire the baseline control scan.To observe the post-occlusive reactive hyperemia maneuver, it's necessary to record a baseline acquisition with the deflated pressure cuff placed in the desired brachial area of the volunteer's arm. After following the previous steps, press and hold the laser foot switch pedal for continuous video acquisition, and pay attention to the View button on the touchscreen. The stabilized image will appear.Press Record to begin live image recording. Dynamic measurements are required to observe the post-occlusive reactive hyperemia maneuver. Inflate the cuff with suprasystolic pressure, and proceed to acquire images of the vasculature under pressure.To acquire a video to assess the impact of the pressure release on the imaged vasculature, open the pressure relief valve while acquiring the video. Follow the live image on the screen. Stop the recording by pressing the Stop button.The optoacoustic imaging platform will stop recording and automatically render the video to Preview mode. In the XY-plane, the melanin signal can be also observed in the planes YZ and XZ, indicating the epidermis limit. The occlusion of the brachial artery provokes some stasis in the larger vessels, visualized by the OT probe.In consequence, we detected an increase in the overall signals of reduced and oxidized hemoglobin shown as an increase of blue and red colors at axes XY, YZ, and XZ.The melanin signal confirms proper spectral unmixing as it remains constant during post-occlusive reactive hyperemia image acquisition. Unlike the recorded oxyhemoglobin and hemoglobin signals which change with the cuff pressure changes over time. It is critical to handle the 3D probe correctly to obtain good quality images.Mastering the handling of the probe is crucial for correct data acquisition and analysis. Here, we demonstrate how to use the system under static and dynamic conditions. In this case, we used the post-occlusive reactive hyperemia maneuver, which is a classical challenger to explore local adaptive mechanisms.We can apply the same strategies with different positionings, measuring in other body sites, and apply other challenges, of course. Established methodologies, such as laser doppler telemetry or photo plethysmography, although very useful, are single point measurements providing limited indiscriminate information partially related with perfusion. The functional information obtained with this system is not comparable from the area of interest to the depth and variables provided.Measurements here, are made non-invasively in vivo and in real time. The potential for research and diagnostics with clinical application is huge.

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