Researchers monitor eye microcirculation using multiwavelength laser Doppler holography in time-frequency optical tomography OCT

(Image Credit: AdobeStock/crevis)

For proper vison function, the eyes must be adequately supplied with blood, and abnormalities in the microcirculation may expose dysfunctions in other arteries, which are difficult to examine. For the first time, scientists from the International Centre from Translational Eye Research (ICTER), operating within the Institute of Physical Chemistry of the Polish Academy of Sciences, used multiwavelength laser Doppler holography to assess blood flow in various layers of the human retina in vivo, which may impact the diagnosis of circulatory disorders.

According to an ICTER news release, researchers explained that spatio-temporal optical coherence tomography (STOC-T) is one option for fast and aberration-free 3-dimensional retinal imaging in vivo. In previous research, ICTER researchers used a multimode optical fiber, ie, one that at its end emits several hundred non-repeating spatial patterns in the cross-section of the beam (so-called transverse modes) to obtain hundreds of OCT images which, when added together, reduce undesirable effects, including speckle noise.¹

The researchers pointed out in the news release the data set obtained during the STOC-T study can be processed in such a way as to reveal blood flow in the human retina. Typically, the visualization of blood vessels requires at least 2 volumes. Subtracting them from each other allows researchers to determine voxels whose intensity changed during the measurement. From there, images of blood vessels are generated.

However, the researchers pointed out this approach requires very fast repetition times, which are not available in STOC-T. To solve this problem, ICTER scientists have developed a new method, called multiwavelength laser Doppler holography (MLDH), which allows the generation flow images from one volume, which may revolutionize the way of monitoring not only the microcirculation of the eye but also the condition of the entire body.¹

The research was carried out by Dawid Borycki, Egidijus Auksorius, Piotr Węgrzyn, Kamil Liżewski, Sławomir Tomczewski, Karol Karnowski and Maciej Wojtkowski from ICTER, and the results were published in the journal Biocybernetics and Biomedical Engineering.²

What is microcirculation?

According to the ICTER news release, microcirculation is the part of the cardiovascular system located between the arterial and venous systems. Microcirculation includes capillaries with a diameter of less than 150 μm. Arterial and venous elements are connected by "bridges" called metarterioles, from which some of the capillaries branch off. They contain the precapillary sphincters, which regulate blood flow through the capillaries. The function of microcirculation delivers nutrients, exchange gases and metabolites, as well as regulate thermal and humoral processes.¹

Retinal arteries, with unique accessibility, offer easy assessment of early vascular changes in vivo. Changes in retinal microcirculation mean global changes in the circulatory system, and therefore potential cardiac disorders. Additionally, pathological changes detected during the assessment of retinal microcirculation are one of the first signs of organ damage, which may precede, for example, proteinuria.

The researchers explained the retina is vascularized by two vascular systems: the choroid, which primarily supplies cones and rods; and the central retinal artery, which mainly feeds the nervous tissue in the inner layers. The 2 systems differ in the amount of blood flow, which is much higher in the choroid than in the retinal vessels. Moreover, in the choroid, there are noticeably lower differences in blood oxygenation levels among arterial and venous vessels. When assessing retinal microcirculation, it is very important to precisely determine the measurement site.²

The fundus of the eye has been assessed by ophthalmologists since the invention of the first ophthalmoscope in 1851 by Helmholtz. Those early rests were not accurate, but they offered an opportunity to assess the damage to the retinal microcirculation in the course of various diseases. In 1939, a 4-stage classification of hypertensive angiopathy and the relationship between subsequent stages of retinal vessels and an increased risk of a cardiovascular event were presented.

The study of retinal vessels has undergone evolved over the years. It has been especially noticeable in the last 30 years. Today, physicians and researchers have a number of tools available to assess the diameter of the vessel, the thickness of its wall, or the speed of blood flow based on the assessment of flowing erythrocytes or leukocytes. Another one just appeared.

Laser Doppler flowmetry and its modifications

Laser Doppler flowmetry (LDF) emerged as one of the first non-invasive methods for assessing retinal microcirculation. According to the news release, by the early 1980s, it began to be more widely used in the study of flows in tissues and organs. This method uses a helium-neon laser with a wavelength of 632,8 nm.¹

The process reflects light from red blood cells moving in the vessels and from the solid, motionless surface of the skin. LDF results are presented as erythrocyte flow values ​​expressed in arbitrary perfusion units (PU), as it is not possible to calibrate the measurement to physiological units. The researchers pointed out this is not an ideal method because it assumes that the examined area should remain completely still, otherwise, artifacts will be created that affect the result.¹

Moreover, the researchers noted in the ICTER news release an extension of LDF is scanning laser Doppler flowmetry (SLDF), which allows not only the assessment of retinal microcirculation parameters but also the morphology of the arterioles themselves. In turn, bidirectional laser Doppler flowmetry (BLDV) involves a complete assessment of the flow velocity of erythrocytes in the retina.

The researchers pointed out the Doppler spectrum of the laser can be decomposed to obtain the velocity distribution of moving cells. Recently, a similar approach was used to visualize in vivo velocity-resolved images of human retinal blood flow. For this purpose, laser Doppler holography (LDH) was introduced and used, in which the shifted Doppler optical field, backscattered from the retina, is detected using a holographic or interferometric full-field optical system.²

A new technique for imaging eye microcirculation

The researchers also noted in the ICTER news release that both LDF and LDH use light with a fixed wavelength. For this reason, both techniques in their original implementation do not provide detailed information about blood flow encoded in the optical field, which changes over time due to movement. A very interesting approach is the combination of dual-beam Doppler with optical tomography (OCT), which enables imaging and assessment of retinal layers. This then provides for simultaneous assessment of blood velocity and blood flow in the retinal vessels.

ICTER researchers recently demonstrated that by spatially modulating the phase of incident light, the laser's spatial coherence can be reduced. Using a technique called spatio-temporal optical coherence tomography (STOC-T), it is possible to obtain many different OCT images, which, when averaged, allow for the removal of noise and distortions. This approach allows for in vivo imaging of the choroid with high spatial resolution.

It turns out that the same dataset can also be used to extract dynamic images of blood flow in the human retina. Individual two-dimensional STOC-T images, after appropriate digital correction, can be used to increase time resolution and obtain flow images.

A team led by Dawid Borycki, PhD, developed and tested a method using STOC-T tomography to improve the visualization of blood flow in the human retina in vivo using the so-called multiwavelength laser Doppler holography (MLDH). It combines laser flowmetry with holographic multiwavelength detection, allowing non-invasive visualization and quantification of blood flow in various layers of the retina.

Borycki explained that this is possible at high blood cell flow rates and with high resolution. This combined approach enables effective assessment of eye microcirculation and, ultimately, extrapolation of the obtained results to the entire circulatory system.

“Our method enables the acquisition of two-dimensional images of blood flow en face from a stack of interferometric images with different wavelengths recorded in ~8.5 ms,” Borycki said in the ICTER news release. “This time is comparable to the time needed in the case of conventional optical OCT (assuming a scanning frequency of 100 kHz) to register a pair of repeated cross-sectional scans, from which a one-dimensional image of blood flow can be obtained.”

Borycki, from ICTER, is one of the authors of the study, according to the news release.

Researchers also explained the implementation of MLDH does not require any modification of the standard STOC-T tomography protocol because this method uses blood flow information from the same data set. As a result, MLDH can be considered a valuable extension of STOC-T tomography, which gives a complete picture of what is happening in the retina.¹

“We have developed and applied a novel signal processing pipeline for spatio-temporal optical coherence tomography (STOC-T), aimed at enhancing blood flow visualization within the human retina in vivo through Multiwavelength Laser Doppler Holography (MLDH),” the researchers concluded in the study. “By numerically minimizing the effects of laser tuning, we then employ Doppler analysis to effectively map the hemodynamics of the inner retina and choroid, notably without the need for any contrast agents. Importantly, the implementation of MLDH does not necessitate modifications to the standard acquisition protocol used for STOC-T, as it extracts blood flow information from the same dataset. Consequently, MLDH serves as a valuable complement to STOC-T, adding a dedicated processing pipeline for blood flow visualization.”

References:

1. A new way to monitor eye microcirculation. Multiwavelength laser Doppler holography (MLDH) in time-frequency optical tomography OCT (STOC-T). EurekAlert! Accessed July 16, 2024. https://www.eurekalert.org/news-releases/1049531

2. Dawid Borycki, Egidijus Auksorius, Piotr Węgrzyn, Kamil Liżewski, Sławomir Tomczewski, Ieva Žičkienė, Karolis Adomavičius, Karol Karnowski, Maciej Wojtkowski, Multiwavelength laser doppler holography (MLDH) in spatiotemporal optical coherence tomography (STOC-T), Biocybernetics and Biomedical Engineering, Volume 44, Issue 1, 2024, Pages 264-275, ISSN 0208-5216, DOI: https://doi.org/10.1016/j.bbe.2024.03.002.