An ongoing inflammatory response involving certain types of immune cells appears to be a driver in creating proud flesh in horse wounds, research in Belgium suggests.
Horses are prone to develop a wound-healing disorder on their limbs called exuberant granulation tissue, or proud flesh.
The exact mechanism for its formation remains unknown but the inflammatory response is considered an important contributor, researchers noted in the journal, Animals.
Proud flesh is composed of collagen, elastin, proteoglycans and hyaluronic acid. It is characterized by the ingrowth of new blood vessels and the presence of fibroblasts, keratinocytes and inflammatory cells.
In wounds to the lower limbs, proud flesh keeps proliferating beyond the margins of the wound bed, preventing the efficient growth of new skin as part of the healing process.
Comparative studies have already shown that differences in the immune response have a big impact on the formation of proud flesh. In contrast to horses, ponies seldom develop the problem.
Studies that compared the healing process of lower limb wounds of ponies with horses describe a short but fierce inflammatory response in ponies, whereas horse wounds are characterized by a lower but persistent inflammatory response.
Wound location seems to influence the development of proud flesh, which is typically seen on lower limbs. Again, this is probably related to a different immune response, with chronic inflammation a driver.
Other factors that can be associated with it are differences in collagen metabolism and angiogenesis (the formation of new blood vessels), improper bandaging, wound contamination and foreign bodies.
Ghent University researchers Charis Du Cheyne, Ann Martens and Ward De Spiegelaere set out in their study to investigate the inflammatory response in both horse wounds that generate proud flesh, and those that do not, over time.
In biopsies, they detected two types of immune cells — those involved in early inflammation (MAC387+ cells) and immune cells important in the later phases of inflammation (CD163+ cells).
After 19 days, there were higher numbers of immune cells in the wounds involving proud flesh compared with the control wounds. This suggests that wounds that involve proud flesh may not be able to proceed through further phases in the wound-healing process, or that the inflammation phase is prolonged, the trio reported.
There were, they said, significantly higher numbers of MAC387+ and CD163+ cells in the fibrotic regions of the proud flesh at the 19-day mark compared with the control wounds.
Normal wound healing involves clotting, an inflammatory response, a rebuilding phase and maturation, or remodeling. The persistently high amount of fibrosis-promoting CD163+ cells in proud flesh suggests that the wound healing processes in these wounds are arrested at the level of the proliferation (rebuilding) phase of healing.
Discussing their findings, the researchers said the inflammatory phases of wound healing can be subdivided into an onset phase and a resolution phase. In the onset phase, phagocytic neutrophils and M1 macrophages play a crucial role by clearing the wound bed of debris and pathogens.
After performing their duty, the neutrophils are ingested by the macrophages (a type of white blood cell). This action triggers the pro-inflammatory M1 macrophages to switch to anti-inflammatory M2 macrophages to resolve the inflammation and initiate the resolution phase of inflammation, which is crucial for the proliferation and remodeling of the healing tissue.
In the wounds that generated proud flesh, overall levels of MAC387+ cells remained high, indicative of a persistent inflammatory response.
The authors say it is challenging to identify different macrophage subsets due to a lack of unique markers.
“In the future, more research is needed on the different macrophage subtypes in horses. RNA studies on single-cell levels can help to identify the different cells and their subtypes that are important during the different phases of wound healing.”
Du Cheyne, C.; Martens, A.; De Spiegelaere, W. High Numbers of CD163-Positive Macrophages in the Fibrotic Region of Exuberant Granulation Tissue in Horses. Animals 2021, 11, 2728. https://doi.org/10.3390/ani11092728