Experimental Free Flaps Research Articles

A New Method for Secure Cannulation of Blood Vessels in Experimental Free Flap Surgery

Dabernig, Jörg MD; Zetlitz, Elisabeth MRCS; Cotrufo, Stefano MD; Ampomah, Opoku MRCS; Russell, David MSc Author Information

Interstitial Fluid Pressure in Free Vascularized Skin Flaps of the Rat Epigastrium

The purpose of this study was to observe the magnitude and time course of changes in tissue edema in experimental free flaps. Thirty female Sprague Dawley rats underwent elevation and orthoptic transplantation of free epigastric skin flaps. Using a modified "wick in needle" technique, interstitial fluid pressure (IFP) was measured in the flaps at various times up to 2 weeks postoperatively. Increases in IFP to positive values were seen by 12 hours. IFP remained elevated for 3 to 5 days, after which they gradually returned to control values by 2 weeks. This is the first study to follow the time course and magnitude of changes in IFP and edema in free flaps. It is a useful animal model by which the hemodynamic effects of physical and pharmacological manipulation of flaps can be studied.

Free microvascular transfer of coccygeofemoral muscle in rabbits.

A new experimental free flap model, the cocygeofemoral muscle flap of the rabbit, is introduced in this study. The coccygeofemoral muscle is a superficially located triangular muscle originating from the sacrum and inserting into the lateral condyle of the tibia, and it abducts the thigh. Anatomical dissection and microangiographic studies revealed that it has a major and a minor pedicle, the former originating from the caudal gluteal artery, and the latter originating from the caudal femoral artery. Its major pedicle contains the 0.7-mm artery, 1-mm vein, and the single nerve originating from the inferior gluteal nerve. In flap studies in 10 animals, the distal minor pedicle was ligated, and muscle was dissected up to its major pedicle and denervated, and the free muscle was transferred to the contralateral inguinal region via end-to-end anastomosis to the recipient vessels (i.e., the femoral artery and vein). At postoperative day 7, the transferred muscles were harvested and histologically examined. Nine of 10 muscles were viable; the coccygeofemoral muscle free flap is believed to be an easy-to-dissect and bulky flap model, having a relatively thick pedicle. It may serve as a good model in free muscle flap studies.

Secondary ischaemia in experimental free flaps – treatment by long acting prostacyclin analogues

Secondary postoperative ischaemia due to venous occlusion is the most detrimental insult to free microvascular flaps. In an experimental rat free flap model the efficacy of long acting prostacyclin analogues iloprost (Ilomedin™) and cicaprost in venous occlusion induced postoperative ischaemia was studied.Free, microvascular groin flaps were transplanted to the neck and the draining veins were temporarily occluded on the first postoperative day for a total of 20 min. In the untreated control group, haemorrhagic flap necrosis occurred. Intravital microscopy after secondary ischaemia revealed flap areas without reperfusion. The functional vessel density was significantly reduced. Reperfused capillaries were tortuous and significantly dilated. After reperfusion the interstitial leakage of macromolecular dextran increased, indicating loss of microvascular endothelial integrity. Intra-arterial and intravenous applications of iloprost were able to diminish the ischaemic effects, giving a flap survival rate of 83%. Similar results were obtained by intravenous and enteral administration of cicaprost. Transcutaneous oxygen partial pressure measurements confirmed the viability of the surviving flaps. We conclude that both iloprost and cicaprost are effective in preventing venous occlusion induced failure of free microvascular groin flaps.

Free flap transplantation in mice

A new model is presented that adapts the standard experimental groin free flap model to the mouse. The femoral vessels upon which the microvascular anastomoses are based are very small (0.2-0.4 mm diameter in the artery), making this a technically challenging exercise. A 100% patency was achieved for flap replantation in ICR (outbred) mice. Success rates for flap transplantation in Balb/C (syngeneic) mice rose from 20 to 75% with modification of the anastomoses from end-to-end to end-to-side procedures. This model offers new avenues of investigation when combined with recently developed transgenic mouse models.

Uncommon and experimental microvascular free flaps in head and neck reconstruction.

Uncommon and experimental microvascular free flaps in head and neck reconstruction.

Effects of Tissue Expansion on Secondary Ischemic Tolerance in Experimental Free Flaps

The effects of tissue expansion on free flap tolerance and metabolic response to secondary ischemia were evaluated. A total of 178 male syngeneic Lewis rats were used: 28 in perfusion study and 75 donor and 75 recipient animals in flap survival study. Animals were organized in three experimental groups: control, sham operation, and expansion group. Sham group animals had the expander implanted but not insufflated. After 4 weeks of tissue expansion, 3 x 5-cm epigastric free flaps were transplanted to recipient animals. Twenty-four hours later, secondary ischemia was produced by 3-hour venous occlusion. Flap survival, perfusion, and enzyme activities were determined. Pre-expanded skin flaps had an increase in perfusion of approximately 700% as measured by fluorescein levels compared with control flaps (p < 0.001) and demonstrated a better success rate (76%) compared with those of the control (40%) (p < 0.05) and sham (28%) groups (p < 0.05). Glutathione peroxidase, glutathione reductase, and glucose 6-phosphate dehydrogenase of the antioxidant defense systems significantly increased in skin in both the sham and the expansion groups. In response to secondary ischemia, the control and sham groups exhibited a decrease in enzyme activities of the glutathione redox cycle, whereas the expansion group showed no significant changes from the elevated baseline activities. Tissue expansion improved flap tolerance to secondary ischemia by increasing flap circulation and probably by augmenting tissue metabolic response to oxidative stress.

Flushing the vessel ends: the effect on ischemic free flap survival.

During vascular anastomosis of experimental ischemic free flaps it is common to observe that stagnant blood in the vessel ends has clotted. A study was performed to investigate if flushing away the stagnant blood on division of the vessels would increase flap survival. Flushing did not influence flap survival. Clinically, clot is rarely seen, except in conditions of prolonged ischemia or damaged vessels. Current surgical practice does not involve flushing the vessel ends upon division of the vessels and this approach is affirmed by our results.

Effects of combined cold and hyperbaric oxygen storage on free flap survival

We have previously reported that hyberbaric oxygen (HBO) improved the survival rate of experimental free flaps. The purpose of this study was to evaluate the effects of combined hypothermia and HBO administered during storage on free flaps and on the xanthine oxidase system in rats. Epigastric skin flaps were stored cold for 48 and 72 hours either in room air or under HBO (2.9 atmospheres absolute, 100% oxygen) before free flap transfer. The success rates of free flaps were 80% (8/10) after 48 hours and 20% (2/10) after 72 hours of cold storage in room air. HBO produced no effect after 48 hours but significantly increased the success rate to 70% (7/10) after 72 hours of cold storage. Tissue hypoxanthine (plus xanthine) levels increased to 210% of normal after 48 hours of cold storage in room air and to 176% in HBO. Elevated hypoxanthine levels returned toward normal by 72 hours of cold storage in room air, while the increased levels remained under HBO. Xanthine oxidase activities significantly increased by 60 to 80% during 72 hours of room air storage. HBO treatment inhibited xanthine oxidase activity to 48% of normal by 72 hours of storage. Free flaps exhibited no significant alterations in GR and G6PDH activity after 48 hours of cold storage in room air or HBO. After 72 hours of cold storage, the room air control displayed a trend of decreasing GR activity and a significant 20% decrease in G6PDH activity, while HBO groups showed no significant alterations in both GR and G6PDH activity compared to normal. Protection of the antioxidative enzymes by hypothermia and inhibition of the xanthine oxidase activity by HBO appear to be one of the mechanisms of improved skin flap survival in free flaps.

The Additive Beneficial Effect of Uw Solution and Urokinase on Experimental Microvascular Free-Flap Survival

Pharmacologic manipulation of free flaps to enhance tolerance to ischemia has become a subject of great interest in the research literature. In an effort to improve survival, perfusion washout of experimental free flaps was performed following an episode of primary ischemia. The perfusates utilized were lactated Ringer's solution (LR), University of Wisconsin solution (UW), a high-molecular-weight medium used in organ preservation, and urokinase, a thrombolytic agent. Seventy-five rats were used in this study and divided into groups of 5 each. A 3 x 6-cm abdominal free flap based on the superficial inferior epigastric vessels was raised in each rat. The free flaps were subjected to either 12 or 18 hr of primary ischemia. Following the period of ischemia, perfusion washout was performed with either LR, UW solution, or urokinase at increasing concentrations alone or in combination with UW solution. Urokinase was first evaluated as a perfusate alone at increasing concentrations. In the 12-hr ischemia group, free-flap survival was shown to increase from 0 percent in the LR-perfused flaps to 20 percent, 60 percent, and 80 percent in flaps perfused with 12,500, 25,000, and 100,000 U of urokinase, respectively (p < 0.05). A similar increase in survival was demonstrated in the 18-hr ischemia group, where 0 percent, 20 percent, and 40 percent of flaps survived following perfusion with 12,500, 25,000, and 100,000 U of urokinase, respectively (p < 0.05). Urokinase was then perfused along with UW solution to evaluate the combined effect on flap survival.(ABSTRACT TRUNCATED AT 250 WORDS)

Uw solution as an experimental microvascular skin flap perfusate

UW solution has been found to be an effective organ perfusate for transplantation. Initial studies in experimental pedicle skin flaps have also demonstrated its unique effectiveness in prolonged ischemia. To understand better the limits of its preservation properties without the influence of endothelial clamp damage, we have undertaken to study the properties of UW solution in experimental microvascular free flaps. Control, lactated Ringer's, and UW solutions were utilized in pedicle and microvascular free flaps in Sprague-Dawley rats over varying periods of ischemia. UW solution demonstrated a clear superiority over all other solutions in both flap models. In addition there also was a significant prolongation of critical ischemia time in UW-treated free flaps compared to pedicle flaps.

Interposition connectors with heparin coating for vascular anastomosis of microsurgical tissue flaps

The intraluminal surface of interposition connectos which are introduced for the quick vascular connection of microsurgical free flaps was coated by heparin. This heparin coating proved to be stable at the polyurethane wall until the intraluminal surface of the interposition connectors with a length of 15 mm was coated by a layer of endothelial cells. 90% of the experimental venous connections didn't show a thrombus formation and were open for the blood flow 10 days postoperatively. The experimental free flaps which were connected by this way had survived 21 days postoperatively and showed a regular healing.

Microcirculatory Disturbances Following the Passage of Emboli in an Experimental Free-Flap Model

Following completion of arterial repair in an experimental free-flap model, platelet emboli have been observed passing through the microcirculation downstream. The purpose of this experimental study was to observe and quantitate changes in capillary perfusion occurring subsequent to these events. The isolated rat cremaster model was used. For 6 hours subsequent to surgical injury of the main artery in this model, the number of emboli and the number of perfused capillaries downstream were counted. In eight rats having an intentional arterial wall injury, emboli were consistently seen during the first hour of reflow. In the nine control animals having no arterial injury, no emboli were seen. The presence of emboli in the cremaster muscle, resulting from the arterial injury, was associated with a significant reduction in the number of perfused capillaries. We suggest that the observed decrease in capillary perfusion was associated with microemboli that produced an adverse effect for several hours after their initial presence in the circulation.

Microcirculatory Disturbances Following the Passage of Emboli in an Experimental Free-Flap Model

Microcirculatory Disturbances Following the Passage of Emboli in an Experimental Free-Flap Model

Experimental cutaneous free flap transfers in the horse

Equine limb wounds often heal slowly by epithelialization, and large scars are a frequent end result. In some ways, they resemble the wound associated with human tibial injuries. The literature indicates that previous investigators have failed to transfer free skin flaps successfully in the horse. In this paper, we review our experimental work with the deep circumflex iliac flap in the horse. Dissections of 20 cadavers confirmed the anatomical consistency of the flap. Four flaps survived well when elevated as island flaps, but five orthotopic and nine heterotopic free flap transfers all failed. The cause of failure is still unknown, but our experiments suggested that the horse must be highly susceptible to ischemic reperfusion injury.

Geometrical approach to the end‐to‐side anastomosis

We analyzed the procedure of performing an arterio- or venotomy in rats and changed it from parallel to the long axis to perpendicular to the long axis. Simultaneously, geometrical analysis of the shape of an arterio- or venotomy led to a new approach for the end-to-side anastomosis in experimental free flap transplantation. This new method was applied in 268 anastomoses. It necessitated only six sutures and resulted in 100% arterial early and late patency and in 99% venous early and late patency.

Recovery of sensation in free flaps.

A clinical study of touch, pain, warm and cold stimuli and two-point discrimination was performed in 27 free flaps four months to four years after the microsurgical procedure. There were 5 free skin flaps (2 with nerve suture), 15 musculocutaneous, 4 muscle-covered with split skin grafts and 3 osteomusculocutaneous flaps transplanted to various sites on the body. The results show full or nearly full recovery of touch and pain sensation in all free skin flaps. The musculocutaneous and osteomusculocutaneous free flaps developed good sensation if firmly grown onto the healthy recipient skin with normal sensation. Muscle flaps covered with split skin grafts and all flaps surrounded by scar tissue had a clinical absence of sensation. This study and our earlier findings of the regeneration of nerves in free skin grafts, in skin flaps and in experimental free flaps, lead us to suggest that the healthy denervated skin of the free flap provides a strong neurotrophic stimulus to the cut cutaneous nerves in the edges of the recipient skin. Cutaneous nerves freely regenerate in the loose subcutaneous tissue of the flap. We therefore conclude that all free flaps with skin islands have a potential for developing sufficient protective touch and pain sensation and even some superficial sensitivity.

Creating a Free Muscle Flap by Neovascularization: An Experimental Investigation

Experimental free muscle flaps were created by inducing transmural neovascularization. Free muscle flaps were developed in Sprague-Dawley rats through a three-stage microsurgical procedure. In the first stage, a muscle strip from the external oblique abdominus muscle was elevated and folded around the superficial epigastric vessels. At six days, the second stage was performed in which the muscle flap was detached from its origin and partially elevated, demonstrating that the muscle had developed an acquired terminal axial pedicle. At 12 days, the third stage was performed in which the muscle flap on its new pedicle was transplanted to a new site. Neovascular free flap survival was documented at intervals of three hours, one day, and six days post free transfer. Plastic casts of the muscle flap vascular tree were made to demonstrate neovascularization.

Direct anastomotic assessment for postoperative microvascular monitoring.

An experimental comparison of two techniques for direct monitoring of anastomoses postoperatively is presented. Arterial differential temperature measurements indicated arterial occlusion (n = 24) in island flaps by mean temperature decreases in the affected artery of 0.5 degrees = C, but temperature changes in cases of venous occlusion (n = 17) were small (mean, 0.2 degrees C), inconsistent, and not of predictive value. The technique did not distinguish success (n = 4) from failure (n = 2) in experimental free flaps, and occlusion of deep arteries produced little or no change in the arterial wall temperature. In contrast, an implanted ultrasound Doppler probe reliably indicated arterial and venous occlusion in island flaps (n = 10), because in both circumstances the mean arterial Doppler signal invariably decreased to 0. The two could be distinguished between by changes in Doppler sound and instantaneous signal. In experimental free flaps, success (n = 10) was clearly distinguished from failure (n = 3) by characteristic patterns of Doppler sound and signal. Arterial differential thermometry had serious limitations, but implanted ultrasound Doppler showed great potential as a postoperative monitor applicable to all microvascular procedures.

Enhancement of blood flow in experimental microvascular free flaps.

Prolonged ischemia may result in tissue death because of the no-reflow phenomenon. By raising a flap 24 hours prior to subjecting it to ischemia, the authors have been able to significantly lengthen the ischemia interval prior to the onset of tissue death. These effects are believed to be the result of alteration of the no-reflow phenomenon.