Therapeutic role of mesenchymal stem cells in second-degree burn wound repair: Integration with split-thickness skin grafts

Abstract

Background

Mesenchymal stem cells (MSCs) have shown significant potential in a variety of clinical applications for repairing and regenerating damaged tissues. MSCs can stimulate cellular processes such as fibroblast proliferation, migration, and endothelial angiogenesis, as well as modulate the host's immune response to favor more optimal wound healing. Therefore, the application of mesenchymal stem cell (MSC) therapy to severe burn wounds holds promise as a potential avenue for improved outcomes, especially in cases where alternative therapies may be limited.

Case presentation

We present the results of a specific case excluded from our previously reported phase I Clinical Trial, examining the safety of allogeneic bone marrow-derived stem cell therapy for deep 2nd-degree burn wounds. This patient had been discontinued from the trial as a clinical decision was made that the patient would best be served by subsequent split-thickness skin grafting, an exclusion criterion for the trial.

Conclusion

We report the benefits of local allogenic MSC therapy followed by an early split-thickness skin graft. We observed a reduction in scar formation with improved sensation to touch using MSC therapy. No adverse events or evidence of rejection were seen. Our case report provides evidence supporting the notion that employing stem cell therapy preceding grafting represents a promising and synergistic therapeutic strategy for effectively treating burn injuries.

Lay summary

Hypertrophic scarring is a common issue in burn wounds, despite surgery or non-surgical treatments, and it often leads to tight, restrictive scars that can cause significant discomfort and complications for patients. This report explores the safety of using bone marrow-derived mesenchymal stem cells (BM-MSCs) as a potential treatment for burn wounds, applied either directly to the skin or beneath the surface.

To date, no adverse side effects have been reported with these methods. The report details the case of a patient who first received stem cells under the skin, followed by a topical application, and later had a skin graft to treat a large, deep second-degree burn. The stem cells used were from a young healthy adult donor. The patient's progress was monitored for six months.

The findings suggest that combining donor stem cells with skin grafting could be a helpful way to treat burn injuries, particularly in cases of deep second-degree burns. This approach appears to enhance wound healing, facilitate faster recovery, and reduce complications like infections, which could lead to shorter hospital stays.

Introduction

The incidence of severe burns among those seeking medical attention was 11 million people globally in 2004.^1,2 In the United States, estimated medical costs and work-loss costs associated with burn-related injury totaled 2.6 billion dollars in 2010, according to the Centers for Disease Control. ³ Thermal injury leads to loss of the regenerative capacity of the skin and significant scarring in some cases. ⁴ Hypertrophic scars can lead to the formation of scar contractures and disfigurement, which may be debilitating to patients. Scar contracture rates have not changed even after improvements in burn care despite using treatments designed to mitigate the effects of hypertrophic scarring. ⁵ Rapid wound closure has been found to help overcome obstacles associated with thermal injury, including decreased chances of scarring and risk of infection, which correlates with patient survival and improved aesthetics. ⁶

Bone marrow mesenchymal stem cells (BM-MSC) can stimulate cellular processes to favor more optimal wound healing. ⁷ MSCs may augment local reparatory responses in wound healing by engaging host cells, such as fibroblasts, keratinocytes, macrophages, and progenitor cells at the injury site.^8–13 Furthermore, through the secretion of soluble factors, MSCs may induce angiogenesis, a critical step in wound healing.^8,14–16 Animal models in previous studies have shown that the use of transplanted MSCs in severe third-degree burns demonstrated appreciable enhancement of epithelialization of the burn wound without significant adverse events. ¹⁷ Our lab investigated the ability of MSCs to accelerate chronic wound healing with promising results. Therefore, the application of MSC therapy to severe burn wounds may present an opportunity for improved outcomes where alternative therapies are limited.

We present the results of a unique case enrolled but later excluded from our previously reported Institutional Review Board (IRB) approved phase I clinical trial, to determine the safety of allogeneic bone marrow-derived stem cell therapy for deep second-degree burn wounds.

Materials and methods

Clinical trial design

This clinical trial was a dose-escalation study, with MSC concentrations ranging from 2.5 × 10³ to 20 × 10³ cells/cm², and the patient was enrolled in the third group with an MSC dose concentration of 1 × 10⁴ cells/cm². ¹⁸ Each patient could receive up to two cell administrations within eight weeks, spaced at least 10 days. The treatment protocol aligns with MSCs used in chronic wound studies.^19–21 Immunologic safety was assessed by measuring serum cytokines (INFγ, IL-10, TNFα) after stimulating the patient's cells with donor MSCs (eBioscience: 88-7346-88). The subject was seen weekly during treatment, with standard care maintained throughout. Follow-up visits were scheduled at 1–4 week intervals until wound healing, and then monthly for six months to monitor healing and scar progression.

Due to the severity of the burn in this specific case, grafting was necessary after enrollment into the clinical trial and the two administrations of MSCs. This was the only case in this phase 1 clinical trial where a clinical assessment was made that subsequent grafting would best benefit the patient's outcome.

Bone marrow mesenchymal stem cells

The allogeneic bone marrow mesenchymal stem cells to treat the patient were obtained following the same source and procedure as the one reported by Schulman et al. in 2022. ¹⁸ All MSCs were delivered by subcutaneous injection using a 3 to 5-cc syringe fitted with a 19G needle.

Wound measurement using E-Z graph

The E-Z Graph (https://e-zgraph.com, San Diego, CA) system tracks burn healing progress after treatment. It includes a transparent graph with an adhesive backing for easy attachment to a worksheet. Simply place the completed graph tracing on the worksheet to document the wound. This system visually represents wound measurements and provides a permanent record of patient files (E-Z GRAPH).

Case presentation

A healthy 21-year old African American male came to the Jackson Memorial Hospital Ryder Trauma Center after sustaining multiple second-degree burns due to the explosion of the oil reservoir of his motorbike. He developed burns on his right thigh, right arm, and right side of his face, totaling 6.25% body surface area. Upon arrival, it was determined his right lower extremity was suitable for treatment, as it was a deep second-degree burn and wound healing was likely to be prolonged with potential for complication. Subsequently, the patient was screened and informed consent was reviewed and signed before participation in the study.

Initially, the patient's wound measure was 682 cm² (Figure 1(a)). Wounds were traced using a transparent graph sheet and area (cm²) was calculated by digital analytical software Image J. ²² This patient was enrolled in the 3^rd dose concentration group (1 × 10⁴ total cells/cm²). The treatment area (Figure 1(b)-outlined in black) was initially 438 cm² in size. The lower region of the leg, with an area measuring 244 cm², was treated with standard care procedures only. In this clinical trial, several clinical features will determine whether MSCs are administered to wounds by injection or applied topically. The burn surgeon concluded that the patient required injection treatment due to the significant size and severity of the wound and had anatomical features that could not be effectively sealed with occlusive dressings. The MSC treatment was applied by subcutaneous injection to the upper portion of the leg (Figure 1(c)). The wound was then wrapped with silicone absorbent foam dressing for 24 h after treatment. Two weeks after subcutaneous application, the treatment region open area measured 314 cm². The patient received a second dose of cells (1 × 10⁴ total cells/cm²) applied topically under transparent film dressing for 24 h (Figure 2(a) and (b)). The lower leg region did not receive topical treatment.

Figure 1.

(a) Initial assessment after enrollment into the study of the burn of the right lower extremity. The open area is 682 cm². (b) Evaluation of wound on the day of first treatment application, with the black outline showing where the treatment was applied (treatment area = 438 cm²). (c) The subcutaneous administration of the treatment to the superior portion of the wound.

Figure 2.

(a) Assessment of the wound on the day of the second treatment application, with the black outline showing where the treatment was applied (treatment area = 314 cm²). (b) Showing how the treatment is applied topically to the wound in a dependent approach.

The burn surgeon assessed the wound bed four days after the second application of MSCs and observed that the wound base appeared appropriate for grafting (Figure 3(a)). In particular, both the burn care and dermatology team noted the MSC-treated area (Figure 3(c)) demonstrated an improved granulation base as compared to the non-MSC-treated area (Figure 3(b)). The patient's deep second-degree burn wound underwent tangential excision and autografting 18 days after first treatment (subcutaneous) and four days after second treatment (subcutaneous).

Figure 3.

(a) Assessment of wound bed before grafting. (b) Close-up of the non-treated area. (c) A close-up of the treated area shows a significantly improved granulation base. It exhibits a healthier dermis with more prominent regions of granulation tissue and epithelialization, creating a more suitable bed for grafting.

Grafting: The wound was debrided on hospital day 4, at the same time as the first injection of MSCs. This was done with a Goulian Weck Skin Graft Knife Set and the excision was carried down to healthy appearing dermis, as noted by punctate bleeding. The thigh and below the knee areas were excised down to the same depth. At the time of grafting on hospital day 18, the wound was re-excised lightly to ensure a uniform wound bed. After tumescence with normal saline, the donor graft was taken from the ipsilateral thigh on the lateral side using a dermatome set at 12/1000's of an inch. No tumescence of the recipient site was used and no tourniquets were used. Post-operative dressings included Mepilex Ag (Molnlycke Healthcare US LLC, Anderson, SC 29624) directly on the graft secured with Kerlix gauze (Cardinal Health, 7000 Cardinal Place, Dublin, OH 43017) and a compression bandage and splinting to keep the knee in extension until postoperative day 4 when all dressings were removed and the patient was allowed to range the knee with physical therapy.

Physical examination during the follow-up one month after the graft procedure revealed a well-healed split-thickness skin graft and closed burn wounds (Figure 4(a)). All wounds healed with a mixture of hyper and hypopigmentation. At a six-month follow-up after the graft, the patient described decreased sensation to touch in non-treated areas and augmented sensation in the treated areas. The patient attributed this to the thickened skin overlying the area not being treated with MSCs. There was an aesthetic difference that was appreciated visually between the areas where MSC treatments were applied (Figure 4(b)) with a clear transition zone. The non-treated area was hypertrophic with multiple ridges and thickening compared to the treated areas (Figure 4(c) and (d)). These are features consistent with a hypertrophic scar. The patient maintained a full range of motion at the knee at the final follow-up one year after the injury.

Figure 4.

(a) One-month follow-up visit after split-thickness skin graft. (b) An assessment of the skin graft conducted five months after the procedure revealed that the graft below the knee exhibited hypertrophy, while the area above the knee did not show this issue. This suggests that using mesenchymal stem cells (MSC) with skin grafts may reduce the risk of hypertrophic scarring. (c) Non-treated area showing multiple ridges and hypertrophy. (d) Treated area showing smooth contour without hypertrophy.

Discussion and conclusion

There is a significant need for a treatment for burn wounds that promote rapid wound healing, hair regeneration, re-pigmentation, less scarring, and attenuation of the inflammatory processes. The timing of treatment is crucial for acute injuries like burns, as delays can lead to adverse outcomes. Allogeneic mesenchymal stem cells (MSCs) present a promising option for such treatments. ⁷ Although grafts are excellent barriers against infection and mechanical trauma, the long-term improvement in appearance has been modest, and the timing of grafting after injury is difficult. ²³ In this report, we outline the treatment of a portion of the burn wound with MSCs, allowing for a comparison of the areas of the wound where stem cell therapy was administered and where it was not administered. Several features of the treated area demonstrate a difference in wound healing outcomes. This case highlights the potential of initial stem cell treatment for a burn wound to optimize wound bed characteristics that may have resulted in early grafting and improve the outcome of split-thickness skin grafting. The patient exhibited no immune reactivity or graft site rejection following treatment.

The wound's progress was evaluated at each follow-up visit after initial treatment using a standardized and validated burn scar scale, known as the Patient and Observer Scar Assessment Scale (POSAS). ²⁴ Scores were based on established criteria from both observers and patients as outlined in our previous paper. ¹⁸ We noted a more significant decline in observer scores compared to the patient scores over time (Figure 5).

Figure 5.

The wound was assessed at every follow-up visit using the patient and observer scar assessment scale (POSAS). Over time, the observer scores displayed a more significant downward trend compared to the patient-reported scores.

This patient's wound demonstrated thermal injury due to hot oil with components of high-velocity impact injury, leading to significant soft tissue damage when compared to other thermal injuries. This is due to the oil's high boiling point, high viscosity, and combustibility. ²⁵ Therefore, we hypothesize that this injury may indicate more severe dermal damage than previously acknowledged, potentially leading to a longer healing process and an increased risk of serious adverse outcomes. ²³ This treatment protocol facilitated limited fibrosis and contracture, without complications or rejection.

The purpose of this clinical trial is to evaluate the safety of topical and subcutaneous application of BM-MSCs to burn wounds. So far, we have not encountered any associated adverse events. The observational findings in this unique case in our trial indicate that local allogenic MSC therapy facilitated early split-thickness skin grafting. This approach could be considered a synergistic therapeutic approach for treating burn injuries. Early grafting of thermal injuries can reduce hospital stays and decrease infectious complications. Severe burn injuries need a multidisciplinary approach that integrates cellular therapy, surgical procedures, and wound management to optimize patient outcomes. ²⁶ This case illustrates the effectiveness of combining MSC therapy with split-thickness skin grafts for treating burn injuries. The findings indicate that the MSC treatment enhances the wound bed for grafting, leading to a better functional and cosmetic outcome compared to the untreated area.