Effect of Topical Insulin on Wound Neutrophil Function
Effect of Topical Insulin on Wound Neutrophil Function
It has been reported that insulin regulates systemic inflammatory responses, whereas the regulation of traumatic inflammation by topical insulin has not been studied. The present study showed that low-dose topical insulin application decreased wound neutrophil infiltration and advanced wound neutrophil attenuation, which is closely related to insulin-regulated, decreased MIP-2 expression. Additionally, insulin enhanced the cellular functions of neutrophils in the wound area. These results demonstrate that insulin regulates traumatic inflammatory responses in a self-constriction manner through the regulation of both cell quantity and cell function.
Neutrophils assemble at the wounded area at the initiation wound healing. A day after injury, wound neutrophil infiltration was observed in both insulin- and saline-treated wounds. Neutrophil attenuation was observed at 3 days after injury in the saline group, whereas topical insulin advanced this phenomenon to 2 days after injury. Advanced neutrophil attenuation alleviated the negative effects of neutrophils, mainly the neutrophil-mediated tissue damage caused by a blast of enzymes and neutrophil-produced reactive oxygen (RO), which might be related to accelerated, insulin-induced healing.
MIP-2, a member of the CXC chemokines family, strongly induced neutrophil chemotaxis. At 2 days after injury with topical insulin application, traumatic MIP-2 expression significantly decreased. Similarly with the change of MIP-2, wound neutrophils had notably decreased. This observation suggests topical insulin regulated the inflammatory response in the wounded area by restraining wound neutrophil infiltration through inhibition of chemokine MIP-2 expression. Vascular permeability also regulates inflammatory cell recruitment. Despite regulating MIP-2 expression, insulin stabilizes vessel endothelial barrier function. More stable microvessels that undergo insulin treatment might hamper neutrophil transmigration to the wound area, and thus limit the quantity of neutrophils in the wound area. Meanwhile, insulin itself acts as a chemoattractant, recruits more macrophage infiltration, and facilitates macrophage phagocytosis (unpublished data), which could also be involved in advanced neutrophil attenuation.
MPO is a peroxidase enzyme most abundantly present in the azurophilic granules of polymorphonuclear neutrophils (PMN). MPO is a marker of PMN activation because MPO enzyme activity significantly correlates with PMN function.23 Activated PMNs release a significant quantity of ROS to induce lipid peroxidation of multiple unsaturated fatty acids in cell membranes via the respiratory burst. MDA is one of the products of lipid peroxidation; hence, the level of MDA marks the level of lipid peroxidation by ROS and further indicates how severely cells are damaged. The present study did not show significant differences in MPO and MDA levels between the insulin treatment group and saline control group. However, at 2 days after injury, the number of neutrophils in the wounds treated with insulin was significantly lower than in the wounds that received only saline treatment. Combining the results mentioned above, one might draw the conclusion that neutrophils in wounds with topical insulin receive enhanced cellular functions. Similar MPO and MDA levels with a lesser number of inflammatory cells indicate that insulin can boost neutrophil function and might help clean exogenous pathogens. Increased neutrophil function and restrained wound neutrophil infiltration, as well as advanced resolution of neutrophils, suggest a delicate regulation of insulin on wound inflammatory response during the healing process.
Although insulin affects the inflammatory response, the overall effects on wound healing are still mediated by repairing cells (keratinocyte and endothelial cells, and the like) and are revealed only in later phases of healing, as evidenced by the significant difference in the wound closure rate at the later and not the early stage of healing. Moreover, the differences in healing times and wound closure rates between the 2 groups, although statistically significant, appear to be minor. Statistical significance between groups strongly suggests the positive effects of insulin on healing. However, larger wounds that take longer to heal might magnify the difference between groups.
Injured patients with diabetes suffer from more infectious wounds and prolonged healing times. It has been reported that diabetic wounds have elevated neutrophils and prolonged neutrophil infiltration. Conversely, neutrophils in wounds of patients with diabetes are dysfunctional in terms of phagocytosis, migration, and bactericidal actions. The present findings of insulin-induced stimulation of neutrophil functions may help advance the clinical treatment of chronic, nonhealing diabetic wounds.
Discussion
It has been reported that insulin regulates systemic inflammatory responses, whereas the regulation of traumatic inflammation by topical insulin has not been studied. The present study showed that low-dose topical insulin application decreased wound neutrophil infiltration and advanced wound neutrophil attenuation, which is closely related to insulin-regulated, decreased MIP-2 expression. Additionally, insulin enhanced the cellular functions of neutrophils in the wound area. These results demonstrate that insulin regulates traumatic inflammatory responses in a self-constriction manner through the regulation of both cell quantity and cell function.
Neutrophils assemble at the wounded area at the initiation wound healing. A day after injury, wound neutrophil infiltration was observed in both insulin- and saline-treated wounds. Neutrophil attenuation was observed at 3 days after injury in the saline group, whereas topical insulin advanced this phenomenon to 2 days after injury. Advanced neutrophil attenuation alleviated the negative effects of neutrophils, mainly the neutrophil-mediated tissue damage caused by a blast of enzymes and neutrophil-produced reactive oxygen (RO), which might be related to accelerated, insulin-induced healing.
MIP-2, a member of the CXC chemokines family, strongly induced neutrophil chemotaxis. At 2 days after injury with topical insulin application, traumatic MIP-2 expression significantly decreased. Similarly with the change of MIP-2, wound neutrophils had notably decreased. This observation suggests topical insulin regulated the inflammatory response in the wounded area by restraining wound neutrophil infiltration through inhibition of chemokine MIP-2 expression. Vascular permeability also regulates inflammatory cell recruitment. Despite regulating MIP-2 expression, insulin stabilizes vessel endothelial barrier function. More stable microvessels that undergo insulin treatment might hamper neutrophil transmigration to the wound area, and thus limit the quantity of neutrophils in the wound area. Meanwhile, insulin itself acts as a chemoattractant, recruits more macrophage infiltration, and facilitates macrophage phagocytosis (unpublished data), which could also be involved in advanced neutrophil attenuation.
MPO is a peroxidase enzyme most abundantly present in the azurophilic granules of polymorphonuclear neutrophils (PMN). MPO is a marker of PMN activation because MPO enzyme activity significantly correlates with PMN function.23 Activated PMNs release a significant quantity of ROS to induce lipid peroxidation of multiple unsaturated fatty acids in cell membranes via the respiratory burst. MDA is one of the products of lipid peroxidation; hence, the level of MDA marks the level of lipid peroxidation by ROS and further indicates how severely cells are damaged. The present study did not show significant differences in MPO and MDA levels between the insulin treatment group and saline control group. However, at 2 days after injury, the number of neutrophils in the wounds treated with insulin was significantly lower than in the wounds that received only saline treatment. Combining the results mentioned above, one might draw the conclusion that neutrophils in wounds with topical insulin receive enhanced cellular functions. Similar MPO and MDA levels with a lesser number of inflammatory cells indicate that insulin can boost neutrophil function and might help clean exogenous pathogens. Increased neutrophil function and restrained wound neutrophil infiltration, as well as advanced resolution of neutrophils, suggest a delicate regulation of insulin on wound inflammatory response during the healing process.
Although insulin affects the inflammatory response, the overall effects on wound healing are still mediated by repairing cells (keratinocyte and endothelial cells, and the like) and are revealed only in later phases of healing, as evidenced by the significant difference in the wound closure rate at the later and not the early stage of healing. Moreover, the differences in healing times and wound closure rates between the 2 groups, although statistically significant, appear to be minor. Statistical significance between groups strongly suggests the positive effects of insulin on healing. However, larger wounds that take longer to heal might magnify the difference between groups.
Injured patients with diabetes suffer from more infectious wounds and prolonged healing times. It has been reported that diabetic wounds have elevated neutrophils and prolonged neutrophil infiltration. Conversely, neutrophils in wounds of patients with diabetes are dysfunctional in terms of phagocytosis, migration, and bactericidal actions. The present findings of insulin-induced stimulation of neutrophil functions may help advance the clinical treatment of chronic, nonhealing diabetic wounds.