Objective

We compared the application of artificial dermis composite tissue flaps and traditional prefabricated flaps in a rat model of exposed bone and tendon injury.

Methods

Sprague Dawley rats were randomly divided into two groups (n = 40 per group). Group A rats received artificial dermis composite tissue flaps and group B rats received traditional prefabricated flaps. Flap appearance, range of motion, degree of swelling, tissue histologic results, and imaging findings were compared between groups at 7, 14, 21, and 28 days.

Results

There was no difference in flap appearance, range of motion, or degree of swelling between groups. However, blood perfusion of the artificial dermis composite tissue flap was better than that of the traditional prefabricated flap; the artificial dermis was also found to be thicker than the traditional prefabricated flap.

Conclusions

The artificial dermis composite tissue flap is an ideal method for repairing exposed bone and tendon, and it displays repair effects comparable with those of the traditional prefabricated flap and may be a better alternative.

The repair of large skin and soft-tissue injuries, especially those involving deep blood vessels, nerves, exposed tendons and bone, or abdominal wall defects due to war and natural disaster conditions, is a difficult problem to solve.

The traditional prefabricated flap has shortcomings, such as its thinness and lack of wear resistance. Therefore, we compared the use of artificial dermis and traditional prefabricated flaps in a rat model of exposed bone and tendon injury to assess their repair potential and characteristics.

Methods

Experimental Materials and Animal Models

Eighty healthy Sprague Dawley rats (male, aged 6–8 weeks, 150–200 g) were provided by the Experimental Animal Center of Second Military Medical University, Shanghai, China, for this experiment. The artificial dermis Pelnac (Gunze Ltd, Kyoto, Japan) was used. Rats were randomly divided into two groups (n = 40 per group), with four subgroups of ten rats each to be assessed at four different time points (7, 14, 21, and 28 days).

Operative Details

Under isoflurane anesthesia, the perimeter of each rat's right hind ankle was measured. A 1-cm incision was made, and the bone and tendon were exposed to simulate a complex wound. A 1 × 3-cm skin graft was excised from the abdomen, placed in wet gauze, and set aside.

In group A, the artificial dermis was fixed with sutures to the wound, followed by skin graft placement. After 4 weeks, the complete artificial prefabricated flap was taken off of the surrounding fascial flap, and a fascial pedicle flap was formed using the superficial inferior epigastric artery and vein. A model of exposed tendon and bone was created. The fascial pedicle flap was tunneled subcutaneously and sutured. Then, the abdominal wound was sutured directly (Fig. 1). In group B, artificial dermis was not used, but all of the other steps were the same as in group A. All of the wounds were dressed with sterile gauze, and an adhesive bandage was used to fix the wound in a functional position.

Figure 1.

Operation process of group A. A, Artificial dermis, which is made the same size as the wound, covers the wound of the abdominal wall. B, Skin grafts were implanted in the artificial dermis surface. C, Suture fixation of the skin. D, After 28 days, the artificial dermis composite tissue flap is cut to be a pedicled fascia flap with the superficial artery and vein. E, Making a bone and tendon–exposed wound of the right leg. F, The inner side after the operation. G, The lateral side after the operation. (continued on next page)

Figure 1.

Operation process of group A. A, Artificial dermis, which is made the same size as the wound, covers the wound of the abdominal wall. B, Skin grafts were implanted in the artificial dermis surface. C, Suture fixation of the skin. D, After 28 days, the artificial dermis composite tissue flap is cut to be a pedicled fascia flap with the superficial artery and vein. E, Making a bone and tendon–exposed wound of the right leg. F, The inner side after the operation. G, The lateral side after the operation. (continued on next page)

Figure 1.

continued

Figure 1.

continued

Outcome Measures

Flap Appearance.

Postoperative flap appearance (congestion, swelling, fluid infiltration, secretion, ulceration, necrosis, crusting, scar formation, and exposure of tendon, bone, or artificial dermis) was recorded at different time points (7, 14, 21, and 28 days) in each group.

Range of Motion.

Ankle range of motion (flexion minus extension angles) was measured at 28 days. The average of three measurements was used for the analysis.

Degree of Swelling.

Bilateral ankle joint circumference was measured at 7, 14, 21, and 28 days, and the average of three measurements at each time point was used.

Immunohistochemical Analysis.

Paraffin sections of the wounds were made, capturing the different time points studied (7, 14, 21, and 28 days). Anti-rabbit/mouse antibodies (REALTMEnVision+/HRP RABBIT/MOUSE; Dako Denmark A/S, Glostrup, Denmark) were used for CD31+ immunohistochemical staining. Slices were observed under high magnification with an optical microscope (ten slices, ×200) for the presence of vascular structures.

Masson Staining.

At postoperative day 28, 40 paraffin sections of the wounds were made (two slices per wound). After conventional Masson staining, the sections were viewed under high magnification with an optical microscope (×40) to determine flap thickness.

Imaging.

On postoperative day 28, rats were exsanguinated and infused with 10% barium sulfate (2 mL/min) via catheters placed into the inferior vena cava and abdominal aorta, respectively, until the death of the rat. Rats were immediately placed in a refrigerator at 4°C for 24 hours for radiographic imaging.

Statistical Analysis

The Student t test was used to compare mean values for range of motion, degree of swelling, and flap thickness. IBM SPSS Statistics for Windows, Version 18.0 (IBM Corp, Armonk, New York) was used for statistical analysis. A P < .05 was considered statistically significant.

Results

Flap Appearance

All of the flaps survived well in the postoperative period at all four time points, with a rich epidermal blood supply. There were no obvious signs of ulceration, infection, or artificial dermal exposure at each time point. Hair growth was seen after 21 days in all of the rats (Fig. 2).

Figure 2.

Flap from group A 7 days (A), 14 days (B), 21 days (C), and 28 days (D) after the operation. Flap from group B 7 days (E), 14 days (F), 21 days (G), and 28 days (H) after the operation. (continued on next page)

Figure 2.

Flap from group A 7 days (A), 14 days (B), 21 days (C), and 28 days (D) after the operation. Flap from group B 7 days (E), 14 days (F), 21 days (G), and 28 days (H) after the operation. (continued on next page)

Figure 2.

continued

Figure 2.

continued

Range of Motion

There was no significant difference in range of motion between right and left hind ankles in the same rats, or between groups, at postoperative day 28 (P > .05).

Degree of Swelling

Degree of swelling, as measured by ankle perimeter, was statistically significantly different between groups at different time points. In both groups, this difference was shown in the following comparisons: preoperative versus 7 days, 7 days versus 14 days, 14 days vs 21 days, and 21 days versus 28 days (P < .05). There was no significant difference in swelling comparing the preoperative and 28-day time points (P < .05). However, no significant difference was observed between groups comparing degree of swelling at the four time points.

Immunohistochemical Analysis

In group A, the artificial dermis was visible between the skin and fascia, with an obvious boundary and neovascularization on postoperative day 7. Over time, cell morphology and arrangement became less disordered, and neovascularization gradually increased. In group B at the same time point, fascial flaps also displayed increasing neovascularization (Fig. 3). CD31+ immunohistochemical-positive cumulative integrated optical density values were significantly different between groups at the 7- and 14-day time points (P < .05); this difference was no longer significant comparing flaps at 21 and 28 days (P > .05) (Figs. 4 and 5).

Figure 3.

Observation of CD31+ immunohistochemical staining of flap sections from groups A and B at different time points (×200). Red arrows indicate the vascular structures. Group A is shown at 7 days (A), 14 days (B), 21 days (C), and 28 days (D) after the operation. Group B is shown at 7 days (E), 14 days (F), 21 days (G), and 28 days (H) after the operation. (continued on next page)

Figure 3.

Observation of CD31+ immunohistochemical staining of flap sections from groups A and B at different time points (×200). Red arrows indicate the vascular structures. Group A is shown at 7 days (A), 14 days (B), 21 days (C), and 28 days (D) after the operation. Group B is shown at 7 days (E), 14 days (F), 21 days (G), and 28 days (H) after the operation. (continued on next page)

Figure 3.

continued

Figure 3.

continued

Figure 4.

CD31+ immunohistochemical staining integrated optical density (IOD) values of group A at different follow-up time points. *P < .05, 7 days versus 14 days. **P < .05, 14 days versus 21 days.

Figure 4.

CD31+ immunohistochemical staining integrated optical density (IOD) values of group A at different follow-up time points. *P < .05, 7 days versus 14 days. **P < .05, 14 days versus 21 days.

Figure 5.

The curve of CD31+ integrated optical density (IOD) values of groups A and B at different follow-up time points. Asterisk indicates P < .05.

Figure 5.

The curve of CD31+ integrated optical density (IOD) values of groups A and B at different follow-up time points. Asterisk indicates P < .05.

Masson Staining

Results of Masson staining for flap thickness at 28 days were significantly different between groups (mean ± SD: 1.94 ± 0.25 mm versus 1.19 ± 0.05 mm; P < .05), with a 63% increase in group A (Figs. 6 and 7).

Figure 6.

Observation of Masson staining flap sections of groups A (A) and B (B) 28 days after the operation (×40).

Figure 6.

Observation of Masson staining flap sections of groups A (A) and B (B) 28 days after the operation (×40).

Figure 7.

Flap thickness of groups A and B 28 days after the operation. *P < .05, group A versus group B.

Figure 7.

Flap thickness of groups A and B 28 days after the operation. *P < .05, group A versus group B.

Imaging

Radiographic assessment of vascular branches showed no significant differences between groups on postoperative day 28. Abdominal blood vessels and branches were visible bilaterally, without significant differences between limbs. However, it was noted on radiography that the right-sided flaps in group A were thicker than those in group B, with visible neovascularization in the artificial dermis (Fig. 8).

Figure 8.

Radiographs after perfusion of 10% barium sulfate in groups A and B 28 days after the operation. A, Right side of group A. B, Left side of group A. C, Right side of group B. D, Left side of group B.

Figure 8.

Radiographs after perfusion of 10% barium sulfate in groups A and B 28 days after the operation. A, Right side of group A. B, Left side of group A. C, Right side of group B. D, Left side of group B.

Discussion

Prefabricated flaps were first successful in animal experiments in the 1970s. With continued improvements in research models, these flaps have been increasingly applied in the clinical setting.10 There is great utility in constructing a new flap at a site where direct flap transfer is not possible.2  The original random flap can be constructed for use as a pivot flap to ensure a more adequate and stable blood supply. Despite the many advantages of prefabricated flaps, thickness is a limiting factor in its practical application.3  Prefabricated flaps cannot meet repair requirements such as greater range of motion and the ability to withstand friction due to lack of wear resistance and tendency to form contractures.4,5  Thus, increasing the thickness of prefabricated flaps has become a research direction. Other researchers have increased the flap thickness by adding the dermis, fascia, muscle, and other tissues or by injecting stem cells into the flap to promote the thickness.

Presently, artificial dermis, which is easy to obtain, is widely used in clinical practice due to its curative effect.6,7  Advantages of artificial dermis include increased tissue thickness as well as wear and contracture resistance.8,9  Herein, we combined the use of a prefabricated flap and artificial dermis in a rat model of open bone and tendon injury and compared the repair characteristics of artificial dermis composite tissue flaps with those of traditional prefabricated flaps. We found no obvious differences in flap appearance, range of motion, or degree of swelling between groups. However, blood perfusion was superior in the artificial dermis composite tissue flap compared with the traditional prefabricated flap. The reason for this may be the layer of artificial dermis present in the former type of flap, in which neovascularization and cell organization can occur, which together increase vascular density and flap thickness, which was confirmed by statistically significant comparisons.

Conclusions

In summary, the artificial dermis composite tissue flap is an ideal method for repairing exposed bone and tendon injuries, and it displays repair effects comparable with those of the traditional prefabricated flap and may be a better alternative due to the ease of acquisition of artificial dermis compared with harvested dermis. Its long-term repair effect requires further study.

Financial Disclosure: None reported.

Conflict of Interest: None reported.

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Author notes

*

Department of Plastic Surgery, Changhai Hospital, Second Military Medical University, Shanghai, China.