ROCTA-based assessment of retinal vascular reactivity will allow researchers and clinicians to investigate retinal vascular function at the capillary level in a noninvasive and safe manner. Because OCTA is an FDA-approved imaging modality, our technique may be applicable in future clinical trials as an end-point measure of retinal capillary function. A subject's retinal versicular reactivity likely reflects their retinal vascular integrity before clinically visible signs of damage are apparent, and therefore provides an early assessment of retinal vascular pathology.
The gas delivery apparatus can be easily adapted to most types of ophthalmic imaging devices and used in a wide array of consenting adults, subject populations, and retinal vascular diseases. Demonstrating the procedure with Dr.Ashimatey will be Elizabeth Corona, a lab ophthalmic service specialist, and Sam Kushner-Lenhoff, a medical student in my laboratory. To assemble the apparatus, first use an adapter to connect the Douglas bag to the clean bore tube.
Then, connect the clean bore tube to the three-way valve at the three-way valve selective inlet port. This combination will be called the air control unit. Next, use an appropriate adapter to fit an in-house rubber tube into the mouth port of the nonrebreathing valve.
Insert the elbow joint into the other end of the tubing. Connect the elbow joint to the gas delivery tubing. This setup will be called the nonrebreathing unit.
Use in-house rubber tubing in the appropriate adapters to connect the air control unit at the outlet port of the three-way valve to the nonrebreathing unit at the inlet port of the two-way nonrebreathing valve. Now, wrap sealing tape around any loose connections in the gas nonrebreathing apparatus to ensure a hermetic fit. Complete the setup by connecting the open end of the gas delivery tubing to a sterile mouthpiece.
The entire gas delivery setup can be seen here. Before beginning the experiment, disinfect the OCTA head and chin rest with an alcohol swab. Place a pulse oximeter on the subject's finger and begin monitoring the oxygen saturation levels and pulse.
Adjust the height of the OCTA setup so that the subject can easily rest their chin on the chin rest without overextending or flexing their neck. Enloop the gas delivery tubing with the mouthpiece attachment through the head and chin rest, with the mouthpiece facing the subject. Insert the mouthpiece into the subject's mouth and encourage the subject to practice breathing through the nonrebreathing setup to achieve familiarity with the apparatus.
Instruct the subject to take deep breaths to facilitate gas exchange and place the nose clip onto the subject to ensure that they are breathing through the mouthpiece. Switch the three-way valve to the appropriate channel as necessary. A room air control base line test is being performed here, so the valve remains in configuration two.
And note the time as the start of a gas inhalation. Carefully time the start of the image acquisition, as images obtained before one minute of gas exposure may not have had sufficient time to reflect the new gas provocation condition. Have the subject place their chin on the side of the chin rest opposite the eye being imaged and move their head forward until the forehead is in firm contact with the head rest.
With the subject facing forward and fixating on the target in the center of their view, center the scan observed in the iris view of the live image and use the left and right arrows to move the chin rest until the iris comes into focus. After confirming that the foveal dip is centered in the OTC scan, acquire an image. The scanning will usually last several seconds.
View the OCTA image after the completion of the scan to confirm that it is of an adequate quality and select Save, or rescan the eye as appropriate. When no more scans of the eye with this gas mixture are needed, allow the subject to sit back from the machine and remove the nose clip and mouthpiece. While the subject takes a break, fill the Douglas bag with the desired air mixture and then set the three-way valve to configuration two.
Repeat the procedure as just demonstrated, but remember to turn the three-way valve at the appropriate time. When all of the images have been acquired, discard the disposable elements of the setup and use an alcohol swab to wipe down the head and chin rest. Wipe the subject chair, OCTA table, and OCTA handles with a disinfectant wipe to remove any errant saliva, and disconnect the setup into its base components.
To empty the Douglas bag, roll the bag from the distal end toward the inlet port connected to the three-way valve and store the bag in an appropriate location for future retrieval. Next, fill a container large enough to submerge the gas delivery tubing with an appropriately diluted and well-mixed detergent disinfectant and soak the two-way nonrebreathing valve, the elbow joint, in-house rubber tubing, and tubing adapters in the disinfectant bath. After at least 10 minutes, thoroughly rinse all of the materials with water and air dry them on a paper towel on a clean countertop.
Then, put all of the dried parts into storage until the next analysis. Horizontal B-scans are often observed on the OCTA device. When properly aligned and viewed as a 2-D in-face image, these B-scans will constitute an OCTA angiogram.
The OCTA angiogram is what is exported from the device and is used for image analysis. To quantify this data in a way that allows for comparisons between subjects and among different conditions, a metric such as the vessel skeleton density can be computed from a skeletonized OCTA image. A heat map of these skeletonized images can then be used to qualitatively demonstrate the change in vessel density.
It is important to explain the procedure thoroughly to the subject. Have them practice breathing through the testing apparatus so they can ask any questions if needed. Any retinal or ocular imaging procedure can be readily adapted to allow retinal vascular reactivity measurements like ours.
This method can also be applied to any disease process that may affect vascular function in the eye. Monitor the subject and their vital signs closely and stop the experiment if you have any concerns or observe any signs of distress.
