In situ Transverse Rectus Abdominis Myocutaneous Flap: A Rat Model of Myocutaneous Ischemia Reperfusion Injury

My name is Marie Claire Edmonds. I'm a clinical research fellow at the University of Edinburgh. The objective of this video presentation is to demonstrate our init two transverse rectus abdominis myocutaneous flap as a model of myocutaneous isch reperfusion injury in the rat.

This is our surgical setup. The anesthetic rig and red induction box can be seen behind the operating table. The operating table is set up such that the animal is laid supine upon a homeo themic blanket used in conjunction with an external heat source with core temperature maintained via feedback through a rectal probe.

The surgical instruments are set up to the right of the animal and the gauze suture material, silicon and MicroPen to the left of the animal, both on tear off surgical fields. These instruments are only laid out once deflation is complete and the animal is then prepped and draped for surgery to conform with whom of it guidelines. First, the animal is anesthetized with 4%ISO fluorine.

Anesthesia is maintained at one to 1.5%ISO fluorine for the duration of the procedure. The anterior abdominal wall is shaved using an electric razor commercial delling cream is applied. This is important to get accurate reasonings with the laser doppler imaging machine.

The tating cream is removed and the area washed with warm sterile saline. 2%chlorhexidine in 70%isopropyl alcohol is then applied to the skin. This is allowed dry.

We advise you conform with local guidelines as to skin preparation. The flap shown will be based on the left deep superior epigastric vessels and the zones numbered as described by SCH Neal in 1982. For the human T tram flap, it is possible to base the flap on either the left or the right deep, inferior, or superior vessels.

Though it should be noted that the superior vessels are the dominant vessels in the rat. In contrast to the corresponding human anatomy, demarcating the flat boundaries identify the linear alba using the ZA sternum and tail as reference points. Mark the midline measure 0.8 centimeters below the ZA sternum and mark this point, draw a line perpendicular to the midline from the point you previously marked below the zip A sternum taking the midline at the center of the flap.

Mark one and two centimeters to the right and left of the midline. Draw a vertical line to parallel to the midline from these points. Measure four centimeters below the original horizontal line and draw another parallel to it.

Following these instructions, a four by four centimeter flap will be delineated consisting of four zones of equal dimensions. The zones are numbered as perle etal description, such that zone one is the inte overline directive abdom muscle directly supplied by the vascular pedicle. Zone two is integument overlying the contralateral rectus muscle.

Zone three is lateral to zone one and zone four to zone two. Once complete, the rat is transferred aseptically and under anesthesia to the laser doppler imaging machine where another anesthetic rig is in place To maintain anesthesia, laser doppler imaging could provide a non-invasive means to assess blood flow in the flap. A monochromatic light source is emitted from the scanner head.

This instant light shown in blue in this animation is scattered by the moving erythrocytes within the tissue. The degree of shift is related to the velocity of the erythrocytes. The shifted light shown in green in this animation is measured by the photodetector within the scanner head and profusion calculated profusion.

Thus measured is given in arbitrary units known as profusion units and the data displayed in an image much like a weather map, wherein profusion is graded from high to low and each value assigned to color, each instrument is calibrated such that all readings taken with the same scanner using the same setting can be compared. Using the software supplied with the scanner, a region of interest can be superimposed onto the LDI image and an average profusion of that area calculated. This permits total flap or individual zones to be assessed and compared.

After the LDI scan, return the rats supine to the home thermic blanket on the operating table, re scrubb hands and on fresh dead eye gloves drape the animal instead out surgical D drapes. And ask your assistant to perform a footpad pinch test to ensure adequate anesthesia before proceeding. Make an incision down.

The animal's left lateral marked edge. Make similar incisions along the horizontal lines to the left of the midline. Ensure hemostasis is established using a handheld cordy device.

Locate the fat overline, the left inferior erector sheath using forceps and fine iris scissors to open up a plane beneath this fat. Take care not to damage any perforators coming up through the left anterior recor sheath. The plane opened up by such dissection is that directly superior to the abdominal wall fascia.

Continue dissecting in this plane and around the demarcated boundaries. Next, dissect medially. As far as the lateral margin of the left rectus muscle, there is an obvious color change at this point from pink to white as highlighted by this white line.

The surgical field should now look like this image. Repeat the procedure on the contralateral side, but this time it can send the medial dissection to the linear alba midline and cauterize the perforators coming through the right anterior rector sheath. As they are visualized, achieve hemostasis, irrigate and place moist gauze over the raised flap.

The fasci cutaneous portions of the rat tram flap are thin enough to permit the flap to take it full thickness skin graft. To prevent this and to ensure that this is a true model of IRI, a thin flexible silicone sheet is placed underneath the fascial cutaneous portions of the flap. This step has been adopted by other researchers undertaking tram flap models in rats cut the silicon sheeting into ovals.

These should be large enough to cover most of the area beneath the fasci cutaneous portions of the flap. However, caution must be taken to ensure that the skin edge can be closed without any tension and there is no pressure on the perforators coming through the center of the left anterior erectus sheath. The silicon sheets are then sutured in place with six oh Vicryl covered with moist gauze return to the inferior margin of the left anterior erectus muscle cauterized the most inferior perforator seen cut a small window in the anterior rectus sheath using micro scissors and pointed curved gray forceps.

Blunt dissect slowly down the medial border of the left rectus muscle until their muscles thins out, but before their posterior rectus sheath is breached, repeat on the lateral side, then blunt dissect medially until the belly of the muscle is atop the curved edge of the forceps and the tips are free at the medial margin. Feed six centimeters of five etholone into the jaws of the forceps and tie off the inferior rector's muscle. In doing so, we now have a myocutaneous flap supplied by one vascular pedicle the deep superior epigastric vessels covered with moist gauze.

Using simple interruptive sutures, peg out the flap to reduce moisture loss as shown. Extend the wound superiorly just left of the zip sternum. Suture this tissue to the upper left quadrant to improve field of view.

Cut a window in the superior left anterior rectus sheath. Extend the wound medially until an obvious change in muscle fiber trajectory is seen from tightly packed vertical fibers of the rectus to OBL loosely packed fibers. Insert the curved forceps carefully between these muscle planes and extend laterally by blunt dissection.

Carefully cut down using micro scissors onto the closed upper surface of these forceps. Cutting through the belly of the left rectus abdo muscle to reveal the underlying deep left superior epigastric vessels using micro instruments and high power on the operating microscope. Carefully separate the artery and vein and strip off any surrounding fat.

Apply a traumatic LAN clamps to the artery and vein and start the timer to count down the 30 minute ischemic period. Irrigate the well created around the clamps and covered with moist gauze. Get your assistant to administer the buprenorphine and warmth fluid starting in the animal's upper left quadrant.

Suture down the flap using continuous subcutaneous sutures using six oh Vicryl. Continue along the inferior margin of the wound. Continue up the flap margin on the animal's right and the superior margin.

Stopping and tying off when you get to the zip a sternum. When the 30 minute ischemic period is finished, remove the clamps and irrigate the well with warm saline. Suture the cut edges of the rector sheath back in place with six oh Vicryl.

Taking great care not to damage the underlying vascular pedicle. Do not apply too much tension as this will lead to kinking of the vessels. Then complete the subcutaneous suturing.

Taking care to bury all knots below the skin. Clean the wound area and allow it to dry. Redraw the zones of the flap.

Re-scan the animal to obtain a postoperative assessment of profusion percentage area whole flat necrosis is shown in this figure N equals 10 mean an SEM are shown. Absolute perfusion is shown in this figure at the four time points. Preoperative postoperative 24 and 48 hours n equals 10 mean an SEM shown the laser doppler images and corresponding photographs at the four time points for three animals are shown here.

The anterior abdominal wall of the rat has significantly less subcutaneous fat than the anterior abdominal wall of most women undergoing T tram flap surgery for breast reconstruction. The model outlined in this text is designed specifically as a model of ischemia, brief profusion injury in myocutaneous flaps so that the effects of preconditioning therapies can be evaluated using skin necrosis and profusion as outcome measures. The procedure detailed in this article does not specifically model problems such as fat necrosis, which are incurred in human tramp flaps when flaps with significant fat components are deliberately harvested to create projection for large breast reconstructions.

Direct in vivo. Observation of the microcirculation is not demonstrated in this protocol, but has been described in the chma muscle model and in an osteo myocutaneous flap. The T tram model is a myocutaneous flap model.

If researchers are particularly interested in osteo myocutaneous flap, this model is not appropriate, but an alternative model has been described in the literature. We find this reliable and reproducible model of ischemia de profusion injury in a myocutaneous flap.