Ashley Collier Biol3500: 2009

Control of scarring in adult wounds by neutralising antibody to transforming growth factor beta.

Shah M, Foreman DM, Ferguson MW (1992).

Lancet339:213-214.

In this paper, the researchers injected the margins of healing dermal wounds in adult male rats with antibodies that neutralize transforming growth factor-Beta (TGF-β), known as neutralizing antibodies, or NAs. As a result, the wounds healed without scar formation but were as strong as scarred tissue.

The purpose of this experiment was to test whether early manipulation of the concentration of certain cytokines, such as TGF-β, would be effective in controlling scarring. The researchers chose TGF-β because it affects all phases of the healing process, and it induces scarring in fetal wounds, which is significant because fetal wounds normally heal without scarring and with a lesser cytokine response.

The experiment involved reducing the concentration of TGF-β within the healing wound by using NAs to inhibit its activity. First, they obtained adult male rats and made four incisions on each. In each animal, one wound was not manipulated (control), another was injected with an irrelevant antibody (rabbit IgG) (sham control), the third was injected with TGF-β (positive control) and the final incision was injected with NAs to TGF-β. The wounds were analyzed using histology/immunocytochemistry, tensiometry, and biochemical analysis.

The tensiometry involved cutting strips from each wound at identical sites for all wounds and extending the strips until they ruptured. All strips ruptured at the site of the healing wound.

The biochemical analysis involved cutting strips, microdissecting the scar tissue or normal skin from underlying fat and muscle, drying and weighing the samples. The hydroxyproline content was measured to calculate the amount of collagen present (assuming collagen contains 13.6% hydroxyproline).

A week after wounding, NA-treated wounds contained much less collagen than the other wounds in the same animal. In the rats killed 14 days after wounding, the NA-treated wounds had much fewer macrophages and blood vessels than the other wounds. Despite this, there was no significant difference in tensile strength between NA-treated wounds and the control wounds. Fourteen days after wounding onwards all wounds had similar tensile strength and collagen content, but the NA-treated wounds had a normal orientation of their extracellular matrix and hence a permanent reduction in scarring. Additionally, the advantageous effects on scarring in the NA-treated wounds were not accompanied by a delay in wound healing or a reduction in wound strength.

The underlying mechanism is that the NAs inhibit the autocatalytic and autoinductive cascades of TGF-β amplification by neutralizing TGF-β such that it cannot interact with its own promoter to upregulate its own synthesis. Therefore, the early application of NAs to TGF-β leads to lower numbers of blood vessels, macrophages, and monocytes at the wound site.

With today’s hygiene and care, the speed of wound healing is not as essential as it was evolutionarily. Characteristics that expedited the healing process were probably selected for at the expense of scar quality. Cytokines can accelerate the healing process, and lowering cytokine concentration may inhibit wound healing. Today, however, the reduction of particular cytokines does not result in deleterious effects on the speed or strength of wound healing. Therefore, these findings suggest an important approach to control of scarring in normal wound healing.

Analysis of the Paper

I enjoyed this paper as it was very easy to read, and its clarity and concise presentation strengthened its readability. As a follow-up of this paper, I looked it up on Web of Science and discovered it had been cited 400 times since it was published in 1992. One of the most recent papers that cited it mentioned that a product has been developed based on this concept that is currently undergoing clinical trials (2). Avotermin is in clinical development for the improvement of scar appearance in the skin. To date, there are no data from studies of Avotermin applied to wounds that raise any safety concerns.

References

1. Shah, M., Foreman, D. M., & Ferguson, M. W. (1992). Control of scarring in adult wounds by neutralising antibody to transforming growth factor beta. Accessed 28 September 2009.
2. Laverty, H. G. et al. (2009). TGF-β3 and cancer: A review. Accessed 17 November 2009.

Origin and Function

When normal body tissue is damaged by more than just minor lesions, the body reacts to repair the damage by forming scars. When people think of scars, they usually think of scars on skin. These occur when the dermis is damaged (6). They can also form in response to infections such as chicken box or skin conditions such as acne (1). Scar tissue is formed as a natural part of healing to provide a protective barrier which, unfortunately, isn’t as functional as the original tissue (5). The scar can vary in appearance depending on how deep the wound was, where it is, how long it takes to heal, individual’s age, and the individual’s inherited tendency to scar (1). For example, young people often have more scarring than older because their bodies make stronger repairs (2). However, many scars fade over time and become less noticeable. To illustrate, striae (more commonly known as stretch marks) may be purple or red, and appear indented from normal skin at first, but over time they become lighter in colour and some almost disappear (3). There are also treatments available to minimize the appearance of scars, such as laser treatment, shown below.

Keloid scar on the chest before (A) and several months after (B) a second laser treatment. Courtesy of Tina S. Alster, MD. (7).

The Healing Process (14)

The healing process has four stages. The first two stages occur over 2-5 days. First is the hemostatic stage, in which blood vessels constrict, platelets aggregate, and clotting factors are relased. Next is the inflammation stage, which begins with the release of histamine from mast cells, resulting in dilated blood vessels. This allows antibodies and white blood cells to flow in. The proliferative stage takes place over the next 2 days to 3 weeks. During this stage, fibroblasts produce collagen fibers and repair the damaged blood vessels. The wound edges contract and epithelial cells start forming scar tissue over the moist and swollen wound surface. Finally, the remodeling stage begins, and continues over the next 3 weeks to 2 years of slow reorganization and strengthening of the collagen scar tissue, so that the would partially returns to its original appearance.

During the healing process, proper nutrition is vital (15). The body requires vitamin A for replacement and healing of epithelial tissues. B vitamins are used by enzymes, and vitamin C is essential for collagen production and maintenance, and blood vessel development.

Structure

Scar tissue is a type of fibrous connective tissue (5). It’s limited in function, including movement, circulation, and sensation, and it’s paler and denser because it has a limited blood supply (5). Scar tissue has a different texture & quality of the surrounding normal tissue (6). For example, scar tissue in the skin is inferior to healthy, normal skin because sweat glands are damaged or destroyed, hair does not grow back, and there is less resistance to ultraviolet radiation (5).

Scars are formed when fibroblasts become active in response to an injury (12). The fibroblasts multiply and make ground substance and fibers which compose the connective tissue. A scar is made up of the collagen fibers that are deposited by the fibroblasts. Collagen is very strong and tough.

Healing biopsy site on the skin a week after the incision. New epithelium has grown on the skin surface, and below this is granulation tissue with small capillaries and fibroblasts forming collagen. After a month, just a small collagenous scar will remain (13).

There are four main scar types: hypertrophic scars, keloids, striae, and atrophic scars (7).

Hypertrophic scars result from overproduction of collagen (6). They appear as a red lump, raised above the surrounding skin (6). They do not grow beyond the boundaries of the original wound, and they often improve in appearance after a few years (6).

Keloids are characterised by the presence of thick hyalinised collagen bundles and overproduction of extracellular matrix components (8). They can be more serious because they can become large, tumorous (although benign) growths (6). They’re completely harmless and non-contagious, but sometimes they become itchy or painful (6). Click here for a short video about keloids.A keloid scar composed of abnormally large collagen fibers and large fibroblasts (16).

Striae are formed when skin grows very rapidly such as in pregnancy (stretch marks), or when skin is put under tension during the healing process (usually near joints) (6). The microscopic structure of striae is not well known or agreed upon (9). Some describe the problem as related to the dermis, others to a unique type of scar tissue, and still others to the elastic fibers (9). Regardless, it is most often accepted that stretch marks represent an area in the dermis of the normal skin where the collagen has been stretched and ruptures then separates and the intervening gap is filled with scar tissue (9).

Atrophic facial scars are dermal depressions most commonly caused by collagen destruction during the course of an inflammatory skin condition such as acne (10). Atrophic scars usually respond well to laser resurfacing.

Atrophic scars before (left) and six months after (right) laser treatment (11).

Pathology (4)

When skin has been severely damaged, as a result of burn wounds or large, deep wounds, the inflammation stage of healing can last longer than usual. Sometimes, this causes abnormal scarring, which can result in keloids and hypertrophic scars. It has been clinically demonstrated that fibroproliferative disorders can develop as a result of prolonged inflammation and healing.

During normal wound healing, dermal fibroblasts migrate into the injury site. However, in extensive burn wounds, the fibroblasts may not be able to do this, and healing often leads to excessive extracellular matrix deposition in the dermis and development of hypertrophic scars.

An example of the abnormal scarring seen in postburn hypertrophic scars (erythematous, raised, pruritic, and inelastic tissue) is shown in the figure below. The physical characteristics of postburn hypertriphic scar tissue result from the deposition of a large amount of extracellular matrix (ECM) that is of altered composition and organization compared with normal dermis or mature scar. If left untreated, the collagen deposition may reorganize itself, adding functional impairment to the discomfort and cosmetic difficulties already suffered by burn patients.