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Method Article

An Experimental Human DIEP Flap Model to Investigate Preservation Strategies for Vascularized Composite Allografts and Free Flaps

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DOI:

10.3791/69392

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December 5th, 2025

 ,  ,  ,  ,  ,  ,  ,  ,  , 
1Department of Plastic and Reconstructive Surgery, Institut Universitaire Locomoteur et du Sport, Pasteur 2 Hospital, Université Côte d'Azur, 2Université Côte d'Azur, CNRS, LP2M, 3Anatomy Laboratory, University of Thessaly, 4Department of Nephrology, Dialysis and Kidney Transplantation, University Hospital of Nice

In This Article

Summary

This protocol describes the surgical technique for harvesting a human perforator flap model based on the deep inferior epigastric artery pedicle, intended for experimental research.

Abstract

Recently, preservation approaches such as cytoprotective agents injection, ex vivo machine perfusion, and supercooling have emerged as strategies to enhance long-term preservation of both standard and marginal organs by mitigating ischemic and hypoxic injury. Although encouraging, its application in the field of vascularized composite allotransplantation (VCA) remains largely confined to preclinical research. To date, most studies investigating VCA perfusion strategies have relied on animal models, particularly swine or rodent composites. While these models provide valuable mechanistic insights, their anatomical, immunological, and physiological differences limit reproducibility and translational relevance to human applications.

In this protocol, each surgical step required for the procurement of a human deep inferior epigastric perforator (DIEP) flap for preservation studies is described in detail. The perforator is transected above the fascia without any subfascial dissection, yielding a short yet sufficient pedicle for catheterization. This model takes advantage of discarded tissue from standard abdominoplasty procedures, posing no additional risk to the patient. Critical steps are outlined to ensure a functional flap is harvested without prolonging operative time or compromising patient safety. Functional imaging is subsequently performed to confirm flap viability prior to experimental use, and sequential biopsies may be performed to follow tissue integrity. This model is particularly suited for research involving muscle-sparing VCA procedures -- such as partial facial transplantation -- and may also have relevance for the study of autologous free flap preservation.

Introduction

Vascularized composite allotransplantation (VCA) represents a promising reconstructive solution for patients with complex tissue defect, particularly in the face and upper limbs1. However, VCA is highly susceptible to ischemia-reperfusion injury, which can compromise graft viability and long-term outcomes2. In this context, preservation approaches such as cytoprotective agents injection, ex vivo machine perfusion, and supercooling have emerged as strategies to extend preservation time, optimize graft quality, and enable viability assessment prior to transplantation3,4,5,6,7. While normothermic ex vivo perfusion of solid organs has been successfully translated from an experimental laboratory technique into clinical practice over the past decade8, with promising results, it remains largely confined to the research setting in the field of VCA9.

To date, most studies investigating VCA perfusion protocols have relied on animal models, particularly swine or rodent composites10. While these models offer valuable insights, they inherently limit reproducibility and translational relevance due to anatomical, immunological, and physiological differences11,12. Human tissue models provide more relevant insights, but fresh tissues from brain-dead donors are extremely difficult to access for research purposes, requiring complex logistics, strict regulatory approval, and raising ethical concerns13,14. An ideal alternative would be a non-cadaveric, ethically acceptable, human-based flap model that does not carry the complexity or risk of a full VCA procedure and is entirely safe for the patient. Importantly, it should also be easily accessible to facilitate reproducible, scalable research.

Herein, a human deep inferior epigastric perforator (DIEP) flap model for VCA preservation research is described, using discarded tissue from abdominoplasty procedures without any additional risk to the patient.

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Protocol

All patients received standard humane care in accordance with the Helsinki Declaration, and the protocol was approved by the institutional ethics committee (project No. IRB00014528_2025_32). The patient provided informed consent for the use of discarded tissue for research purposes.

1. Preoperative care

NOTE: These cutaneous perforator flaps, excluding fascia and muscle, are derived from surgical waste in patients, most often post-bariatric, and can only be harvested in cases of prior dermolipectomy with predictable umbilical transposition (i.e., a significant abdominal pannus). Planned liposuction during the procedure is an exclusion criterion. Concomitant diastasis repair is not a contraindication, as this procedure occurs after flap harvesting. The abdominal pannus generally provides at least partial coverage of the pubic area. Patients must have no history of thromboembolic events, and the body mass index (BMI) must be below 30 kg/m², as higher values are associated with obesity and an unfavorable risk-benefit ratio for elective non-vital procedures under general anesthesia. Previous abdominal surgery, except liposuction, is usually not a contraindication, provided that a perforator is identified by Doppler examination or directly visualized intraoperatively. This research procedure should also be avoided in true clinical unilateral DIEP free-flap reconstructions, which are already lengthy and complex operations where additional preoperative steps would be irrational and increase procedural risk.

  1. Perform the preoperative dermolipectomy marking with the patient in the standing position with a dermographic pen. Identify the midline from the pubic symphysis to the xiphoid process, and draw a transverse line approximately 7 cm above the vulvar commissure. Extend this line laterally about 7 cm on each side, gently curving it upward to join just below the anterior superior iliac spines.
  2. Identify the perforators around the umbilicus using a handheld acoustic Doppler probe (8-10 MHz) with the patient in the supine position.
    NOTE: Apply a small amount of sterile ultrasound gel to the skin to ensure proper acoustic coupling. Hold the probe at a 45° angle with light contact pressure to avoid dampening the signal. A strong, triphasic pulsatile sound indicates an arterial perforator. Mark each perforator on the skin with a sterile marker to establish the perforator map, as routinely done prior to DIEP flap harvest.
  3. Using a dermographic pen, outline a skin paddle in an elliptical shape of the desired dimensions, including the perforator along its medial border and extending on both sides of the umbilicus (Figure 1).
  4. Prepare and drape the surgical field from the xiphoid process to the upper third of the thighs, including the pubic region, using sterile surgical drapes as routinely performed in standard abdominal surgery.

Plastic surgery procedure planning and execution, preoperative diagram and surgical setup.
Figure 1: Preoperative evaluation before post-bariatric abdominal dermolipectomy with DIEP flap harvest. (A) Preoperative marking performed using an acoustic Doppler and a dermographic pen to identify and outline the paraumbilical perforators on each side of the midline. (B) The surgeon demonstrates the laxity of the abdominal skin and subcutaneous tissue to be resected, highlighting the adipocutaneous panniculus before incision along the horizontal marking line. Please click here to view a larger version of this figure.

2. Flap procurement

  1. Detach the umbilicus from the abdominal wall through a circumferential incision down to the hypodermis using a No. 15 scalpel blade.
  2. Place a long, loose, non-absorbable suture on either side of the umbilical margin.
  3. Dissect the umbilicus vertically using Mayo scissors, from the superficial to the deep plane, until it is completely freed while preserving its umbilical pedicle.
    NOTE: Assistant participation is critical at this step. Gillies retractors should be used to expose the umbilicus, while applying gentle longitudinal traction on the long non-absorbable suture to facilitate dissection. The fat surrounding the umbilical pedicle forms a firm cord that is easily palpable during umbilical isolation, allowing precise identification and preservation of the pedicle.
  4. Make a low transverse skin incision along the preoperative marking, approximately 7 cm above the vulvar commissure, gently curving toward the anterior superior iliac spines on each side, using a No. 15 or No. 21 scalpel blade.
  5. Raise the superior abdominoplasty flap in the subcutaneous plane, starting from the lower incision and progressing cranially toward the umbilicus. Use a fine-tip monopolar electrocautery set to approximately 50-70 °C and 80 W in coagulation mode to separate the anterior rectus fascia from the overlying subcutaneous tissue and skin while maintaining meticulous hemostasis throughout the dissection.
  6. As the dissection approaches the previously isolated umbilical pedicle, reduce the electrocautery temperature to approximately 20-30 °C, keeping the power set at 80 W in coagulation mode, and continue with fine dissection using Metzenbaum scissors to avoid pedicle injury.
  7. Identify and carefully isolate the two dominant paraumbilical perforators arising from the deep inferior epigastric system. Dissect each perforator circumferentially under direct vision using Stevens or small Metzenbaum scissors, preserving their vascular pedicles. Do not extend the dissection above the level of the anterior rectus aponeurosis (Figure 2).
    NOTE: Perforators are selected based on their size; those appearing larger and pulsatile under direct vision are preferred. Dissection around the umbilicus must be meticulous to avoid damaging these perforators.
  8. Divide the adipocutaneous panniculus longitudinally along the midline, from the center of the lower pubic incision up to the umbilicus, using a No. 15 blade followed by monopolar electrocautery set at approximately 50-70 °C.
  9. Continue the dissection cranially up to the xiphoid region and along the lateral costal margins with electrocautery set at approximately 50-70 °C, keeping the flap pedicles intact and not divided (Figure 3).
    NOTE: This step 9 can also be delayed for practical reasons if the pedicle makes the supraumbilical dissection difficult and may instead be performed after pedicle division at Step 12, depending on intraoperative conditions..
  10. Control minor bleeding with medium-intensity coagulation, using a standard monopolar electrocautery set to 50-60 °C and 80 W in coagulation mode to maintain precise hemostasis.
  11. Readjust the outline of the skin paddle with a dermographic pen, based on the initial markings, to ensure that the perforator is included along the medial edge of the flap on each side of the adipocutaneous panniculus that was divided longitudinally at step 8 (Figure 4).
    NOTE: To avoid bias due to differences in surface area exposed to the test solution, both flaps (if two are harvested) must be of identical size. The flap thickness is determined by the subcutaneous tissue of the flap and cannot be surgically reduced without risking compromise of flap integrity.
  12. Ligate the deep inferior epigastric artery perforator pedicle using resorbable 3-0 sutures or automatic clips and transect the perforator above the fascia, with no subfascial dissection. Record the time of the warm ischemia.
    NOTE: An approximate 10-min period of flap ischemia is observed before complete detachment and treatment with a cytoprotective agent injection.
  13. Advance the supraumbilical skin and subcutaneous fat downward, and secure the undermined abdominal flap to the inferior incision margin at the midline using a non-absorbable suture, leaving one end of the knot long.
    NOTE: Place the patient in a semi-flexed position during this step. Downward traction on the upper abdominal flap helps determine the amount of excess skin to be resected.
  14. Use the long end of the midline non-absorbable suture connecting the supraumbilical and pubic areas as a guide to draw, with a dermographic pen, the resection line on the excess adipocutaneous panniculus, extending laterally toward each anterior superior iliac spine, as typically performed during abdominoplasty. The DIEP flaps are located below this marked resection line (Supplementary Figure 1).
  15. Incise the flap along the preoperative markings using a No. 15 scalpel blade. Harvest the perforator flap following the pre-established design, using monopolar electrocautery set to approximately 50-60 °C and 80 W in coagulation mode (Figure 5).
    NOTE: This is one of the most delicate steps, as the perforator must be included within the flap. Assistance is recommended to provide gentle traction, while the operator maintains continuous visual control of the perforator throughout the dissection. Cold-knife dissection is not advised, as it may compromise flap integrity and lead to vascular leakage.
  16. Completely detach the perforator flap from the surrounding discarded tissue.
    NOTE: If two flaps are harvested and subjected to different treatments, they can be distinguished by placing a suture on one of them.
  17. Excise the redundant dermo-adipose tissue corresponding to the excess skin and fat removed during the abdominoplasty.
    NOTE: If the perforator flap is harvested after excision of the excess dermo-adipose tissue for practical reasons, use a sterile, single-use, battery-operated disposable cautery pen (large tip, approximately 1,200 °C) to complete the detachment. A return electrode cannot be used in this situation, as the tissue is fully separated from the patient. After harvesting the DIEP flaps, the surgical team hands the flap to the research team waiting in the operating room, then proceeds with the abdominoplasty by dissecting along the pre-marked resection lines to detach and remove the redundant adipocutaneous panniculus.

Surgical procedure demonstrating tissue dissection using retractors, medical operation context.
Figure 2: Intraoperative view of the abdominal dermolipectomy procedure with skeletonization of the deep inferior epigastric perforator pedicles. The image shows the operative field before supraumbilical dissection and longitudinal infraumbilical division. The two dominant bilateral paraumbilical deep inferior epigastric perforators are exposed and highlighted with surgical instruments. Please click here to view a larger version of this figure.

Surgical procedure, fascia layer exposure, abdominal surgery, medical operation, clinical technique.
Figure 3: Intraoperative view after supraumbilical dissection showing the bilateral deep inferior epigastric perforators. The perforators are not ligated, and their subfascial course and length can be better appreciated under gentle traction (white arrows). Please click here to view a larger version of this figure.

Surgical incision planning; pre-op markings on tissue, medical procedure comparison, diagram.
Figure 4: Intraoperative view showing the DIEP flaps outlined on the skin in an elliptical shape before ligation. (A) The skin paddle is adjusted and outlined in an elliptical shape using a dermographic pen (white circle) to ensure inclusion of the perforator along the medial edge of the flap on each side of the adipocutaneous panniculus. (B) Enlarged view showing the DIEP flap outline and the perforator marked with an "x" according to preoperative Doppler mapping (white arrow). Please click here to view a larger version of this figure.

Supplementary Figure 1: Downward traction on the upper abdominal flap helps determine the amount of redundant skin to be resected. The long end of the midline non-absorbable suture connecting the supraumbilical and pubic regions is used as a guide. Using a dermographic pen, draw the resection line along the course of the suture (white arrows) on the redundant adipocutaneous panniculus. Extend this line laterally toward each anterior superior iliac spine. The DIEP flaps are located below this marked resection line. Please click here to download this File.

Surgical planning and incision marking for DIEP flap procedure, ruler scale indicates dimensions.
Figure 5: Intraoperative view of the deep inferior epigastric artery perforator (DIEP) flaps during harvest, following pedicle ligation. (A) The DIEP flap, measuring approximately 10 cm × 6 cm, is outlined with a surgical marker after localization of the perforator. (B) Skin incision and flap harvest are performed along the pre-established markings. The initial incision is made with a No. 15 blade, followed by dissection using a monopolar electrocautery set at approximately 50-60 °C, and 80 W in coagulation mode, when the excess cutaneous-adipose tissue remains attached to the patient. When the tissue has been completely detached, a sterile, single-use, battery-operated disposable cautery pen (large tip, approximately 1,200 °C) is used to complete the separation. Please click here to view a larger version of this figure.

3. Preparation of the flap

  1. Dissect the vascular pedicle under magnification using microsurgical instruments, and identify both the artery and the vein (Figure 6). Gently open the arterial lumen with a microvascular dilator.
    NOTE: Use a dedicated sterile table equipped with microsurgical instruments to prepare the flap. Trim 2-3 mm from the arterial end to obtain a clean, regular edge suitable for catheterization.
  2. Weigh each flap at the time of harvest to allow later quantification of edema formation.
  3. Catheterize the artery using an 18-24 G cannula and secure it in the lumen with 5-0 silk suture ligation (Figure 7).
    NOTE: For this experiment, the flaps were catheterized using a 24-G cannula, as the study involved the evaluation of cytoprotective agents through a single injection. For ex vivo machine perfusion or extracorporeal membrane oxygenation (ECMO) applications, cannulation should be performed with a larger 18-20-G cannula. In case of catheter occlusion during ex vivo perfusion, flush gently with heparinized saline or replace the cannula to restore flow. Maintaining low perfusion pressure (<25 mmHg) helps prevent recurrence.
  4. Inject contrast agent intra-arterially through the catheter into the perforator artery, and evaluate the flap under fluoroscopy (Figure 8).
    NOTE: The contrast agent should render the small capillaries uniformly radiopaque throughout the flap, indicating an adequate perfusion territory supplied by the selected pedicle. At this stage, the presence of non-perfused areas may render the model unsuitable for further research use or require excision of the distal part of the flap. The harvested flaps can subsequently be used for experimental protocols, placed on a perfusion machine, ECMO device, or injected with a test solution as cytoprotective agents. Punch biopsies (ranging from 1 mm to 8 mm in diameter) can be performed at different time points without compromising flap integrity, allowing kinetic assessment of histological changes within the tissues.

Surgical graft process; A, tissue measurement; B, graft preparation; medical procedure diagram.
Figure 6: Post-harvest view of a research DIEP flap. (A) The harvested DIEP flap is shown with the vascular pedicle dissected and isolated using microsurgical forceps. The arterial end is trimmed by a few millimeters to obtain a clean cut, and the lumen is gently opened with a micro-dilator forceps. (B) Close-up view showing microsurgical forceps securing the dominant perforator artery, with a 24-G catheter inserted into the lumen for contrast agent injection. Please click here to view a larger version of this figure.

Surgical tissue samples with measurement scales and syringe for size assessment in medical procedure analysis.
Figure 7: Catheterization of the DIEP flap pedicle. (A) The catheter is secured in place with a 5-0 silk suture tied gently to avoid obstructing the lumen of the pedicle. (B) View of the catheter positioned without the syringe attached, showing the extent of catheter insertion into the vascular lumen. Please click here to view a larger version of this figure.

Coronary angiogram, heart vessel visualization, diagnostic imaging, medical research.
Figure 8: Post-harvest fluoroscopy of the DIEP flap demonstrating the vascular anatomy. (A,B) Contrast agent is injected through a 24-G catheter into the dominant perforator artery, depicting the branching pattern and the vascularized area, confirming flap viability on two different flaps. Please click here to view a larger version of this figure.

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Results

Eleven flaps were harvested from six female patients included in this study. Two senior plastic surgeons, experienced in flap surgery and abdominoplasty, performed the dissections. In one patient, the flap was harvested unilaterally due to a dissection issue. The harvested flaps had a mean weight of 198.6 ± 24.4 g (n = 11) and an average size of 10 cm × 6 cm, which corresponded to the sample requirements for the preservation study in which these flaps were used. The pedicle length, measured under gentle tension, averaged...

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Discussion

Human models help bridge findings from animal studies to clinical practice, improving the relevance and predictivity of preclinical research16,17,18. They must be carefully designed to ensure safety and ethical compliance, often limiting the scope and invasiveness of experiments17. Across the four harvested flaps, no additional dissection was necessary, and there was no significant prolongation of operati...

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Disclosures

The authors have nothing to disclose.

Materials

List of materials used in this article
NameCompanyCatalog NumberComments
1 mL and 5 mL Luer-Lok syringeBecton Dickinsonhttps://www.bd.com/en-us/products-and-solutions/products/product-page.309646
11.5” Medium and Small Premium Surgiclip II Auto Suture vessel clip applierCovidienN/A
22–24 G vessel cannula blunt tipMedtronic IncN/A
5-0 silk suture N/AAny compatible suture
A single-use, battery-powered disposable cautery pen Helpmedical, EuropeN/A1200 °C, large-tip
Adson ForcepsMPM106-2112A
Bipolar Coagulation ForcepsOlsen20-1320I
Disposable Scalpel #15 & #21SklarN/A
Eosin Microm MicrotechU/C0363
Faraboeuf retractorsMedicta InstrumentsN/A
Fine needle cauteryCormedicaN/A
OEC MiniView MaxGE HealthcareN/AFluoroscopy system
Forceps DilatorsWPI15910
Hematoxylin Microm MicrotechU/C0303
Iohexol 300 mg I/mLGE Healthcare Canada Inc.N/A
Micro scissorsWPI504492
Valleylab Monopolar DiathermyMedtronic Inc
Non-ionic iodinated contrast medium, Omnipaque 300 mg I/mLGE HealthcareN/A
Portable acoustic Doppler, 8 MHzParks Medical Electronicshttps://www.parksmed.com/
Saline solution 0.9%GenDepotS0600-101
Standard plastic surgery instrument setAesculapN/Aincluding scalpel handle No. 3, Metzenbaum and Mayo scissors, Adson and Gillies forceps, dissecting scissors, hemostats, and needle holders.
Strabismus scissorsSurtex102-4109
Surgical marking penCardinal health212PR
Sutures Ethilon 4.0 and 3.0Ethicon1667G
Tissue ForcepsMPM106-0511

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Vascularized Composite AllograftFree Flap PreservationHuman Perfusion ModelCytoprotective AgentsMachine PerfusionIschemic InjuryMicrovascular DissectionArterial CatheterizationHistological Analysis
An Experimental Human DIEP Flap Model to Investigate Preservation Strategies for Vascularized Composite Allografts and Free Flaps
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