In-Vitro Wound Contraction in the Horse
In-Vitro Wound Contraction in the Horse
Wound healing in the horse is often complicated by the formation of proud flesh in which exuberant granulation tissue is present. In these wounds, the granulation tissue hyperproliferates to an excessive level thereby preventing reepithelization from taking place. Indolent wounds are sometimes encountered; the wound produces little granulation tissue, it is inflamed, and it is often infected. These wounds have important welfare implications, and despite their prevalence little is known about their etiology. Wounds that occur on the body and trunk heal rapidly with obvious contraction, while wounds on the lower limb either fail to heal or heal slowly with little wound contraction. In-vitro fibroblast culture systems are routinely used to investigate wound contraction under a wide range of experimental conditions. These include the effects of radiation, inhibition of chronic inflammatory cell mediators, and the biocompatibility of wound management products. In this study, the contractile capacity of fibroblasts from lower-limb wounds and five other sites on the body of the horse were studied in vitro in order to learn more about differences in wound contraction. Fibroblasts were harvested, cultured, and resuspended in collagen gels. Contraction was assessed by measuring decrease in surface area of the gels. The limb fibroblasts demonstrated a greater rate of contraction than cells harvested from the other sites. Fibroblasts harvested from granulation tissue contracted the gels significantly faster than normal and L929 cells. Cell viability remained high throughout the experiments, and there was no significant difference in viability between the three cell types used at the end of the experimental period. These studies showed that during second-intention wound healing in the horse, the differences in wound contraction between wounds on the limbs and the body are caused by differences in the contractile capacity of fibroblasts, and extracellular matrix factors determine the extent of contraction during wound healing.
Horses commonly have complications in the healing of wounds on the lower limb regions. Upper body wounds, often extensive and deep, often heal well with few complications. The speed of healing in different regions in the body and the limb may be attributed to wound contraction. The work of Wilmink, et al., showed that ponies have a higher rate of contraction than horses. There have been similar variations seen in some human wounds, especially human leg ulcers (Figure 1), which fail to heal despite advanced therapeutic intervention. Equine chronic wounds that are exuberant or indolent have similarities to human leg ulcers. In the horse and man, the problems manifest themselves in the lower limb and are often age related.
(Enlarge Image)
These photographs illustrate similarities between nonhealing human and equine limb wounds: A) human venous leg ulcer; B) indolent equine wound.
(Enlarge Image)
These photographs illustrate similarities between nonhealing human and equine limb wounds: A) human venous leg ulcer; B) indolent equine wound.
Wound contraction is a major contributor to the healing process, and the rate of contraction of wounds has been calculated from experimental studies involving experimental wounds of a standard size and shape. It has been shown that wound contraction not only speeds up the healing process but enhances the tensile strength and cosmetic appearance of the healed wound. In chronic limb wounds, however, the opposite occurs, and the newly formed epithelium is fragile and lacks hair follicles.
Myofibroblasts differentiate from granulation tissue and develop ultrastructural and biochemical features of smooth muscle cells, including the presence of microfilament bundles and the expression of
-smooth muscle actin. Myofibroblasts play a role in wound contraction and are the main cell type implicated in the synthesis of extracellular components, including type I collagen and fibronectin. The contraction capacity of the cells and the local extracellular environment will determine the degree of contraction of the wound.
Contraction of fibroblast/collagen gels has been used as an in-vitro model for investigating the biological mechanisms of wound contraction and also the effects of various compounds aimed at stimulating (enhancing wound healing) or reducing (preventing scar formation) the rate of contraction. The advantage of using this model is that the fibroblasts are grown in a three-dimensional collagen gel culture where collagen is a component native to the wound environment.
The culture of fibroblasts in these gels more closely resembles growth in an in-vivo situation, and this is important when considering cellular interactions.
The aim of this study was to compare the contractile abilities of differentiated fibroblasts harvested from five different sites of the body and the limbs of horses. A similar comparison was made between tissue harvested from chronic granulating wounds, normal skin, and L929 cells. The contractile capacity of the cells was determined by a decrease in gel area.
Wound healing in the horse is often complicated by the formation of proud flesh in which exuberant granulation tissue is present. In these wounds, the granulation tissue hyperproliferates to an excessive level thereby preventing reepithelization from taking place. Indolent wounds are sometimes encountered; the wound produces little granulation tissue, it is inflamed, and it is often infected. These wounds have important welfare implications, and despite their prevalence little is known about their etiology. Wounds that occur on the body and trunk heal rapidly with obvious contraction, while wounds on the lower limb either fail to heal or heal slowly with little wound contraction. In-vitro fibroblast culture systems are routinely used to investigate wound contraction under a wide range of experimental conditions. These include the effects of radiation, inhibition of chronic inflammatory cell mediators, and the biocompatibility of wound management products. In this study, the contractile capacity of fibroblasts from lower-limb wounds and five other sites on the body of the horse were studied in vitro in order to learn more about differences in wound contraction. Fibroblasts were harvested, cultured, and resuspended in collagen gels. Contraction was assessed by measuring decrease in surface area of the gels. The limb fibroblasts demonstrated a greater rate of contraction than cells harvested from the other sites. Fibroblasts harvested from granulation tissue contracted the gels significantly faster than normal and L929 cells. Cell viability remained high throughout the experiments, and there was no significant difference in viability between the three cell types used at the end of the experimental period. These studies showed that during second-intention wound healing in the horse, the differences in wound contraction between wounds on the limbs and the body are caused by differences in the contractile capacity of fibroblasts, and extracellular matrix factors determine the extent of contraction during wound healing.
Horses commonly have complications in the healing of wounds on the lower limb regions. Upper body wounds, often extensive and deep, often heal well with few complications. The speed of healing in different regions in the body and the limb may be attributed to wound contraction. The work of Wilmink, et al., showed that ponies have a higher rate of contraction than horses. There have been similar variations seen in some human wounds, especially human leg ulcers (Figure 1), which fail to heal despite advanced therapeutic intervention. Equine chronic wounds that are exuberant or indolent have similarities to human leg ulcers. In the horse and man, the problems manifest themselves in the lower limb and are often age related.
(Enlarge Image)
These photographs illustrate similarities between nonhealing human and equine limb wounds: A) human venous leg ulcer; B) indolent equine wound.
(Enlarge Image)
These photographs illustrate similarities between nonhealing human and equine limb wounds: A) human venous leg ulcer; B) indolent equine wound.
Wound contraction is a major contributor to the healing process, and the rate of contraction of wounds has been calculated from experimental studies involving experimental wounds of a standard size and shape. It has been shown that wound contraction not only speeds up the healing process but enhances the tensile strength and cosmetic appearance of the healed wound. In chronic limb wounds, however, the opposite occurs, and the newly formed epithelium is fragile and lacks hair follicles.
Myofibroblasts differentiate from granulation tissue and develop ultrastructural and biochemical features of smooth muscle cells, including the presence of microfilament bundles and the expression of
-smooth muscle actin. Myofibroblasts play a role in wound contraction and are the main cell type implicated in the synthesis of extracellular components, including type I collagen and fibronectin. The contraction capacity of the cells and the local extracellular environment will determine the degree of contraction of the wound.
Contraction of fibroblast/collagen gels has been used as an in-vitro model for investigating the biological mechanisms of wound contraction and also the effects of various compounds aimed at stimulating (enhancing wound healing) or reducing (preventing scar formation) the rate of contraction. The advantage of using this model is that the fibroblasts are grown in a three-dimensional collagen gel culture where collagen is a component native to the wound environment.
The culture of fibroblasts in these gels more closely resembles growth in an in-vivo situation, and this is important when considering cellular interactions.
The aim of this study was to compare the contractile abilities of differentiated fibroblasts harvested from five different sites of the body and the limbs of horses. A similar comparison was made between tissue harvested from chronic granulating wounds, normal skin, and L929 cells. The contractile capacity of the cells was determined by a decrease in gel area.
