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ATP HYDROLYSIS REDUCES NEUTROPHIL INFILTRATION AND NECROSIS IN PARTIAL-THICKNESS SCALD BURNS IN MICE
Presenter: Sara De La Rosa, BS
Co-Authors: Bayliss J; Peterson JR; Eboda ON; Lisiecki JL; Rinkinen JR; Xi C; Su GL; Cederna PS; Wang SC; Levi B
Univerity of Michigan

Introduction: The inflammatory response following a burn injury plays a key role in wound healing and survival. Extracellular ATP, present in thermally-injured tissue, modulates this response. We hypothesize that removing ATP-dependent signaling at the burn site would reduce neutrophil infiltration and tissue necrosis.

Methods: Male C57 mice were subjected to 30% total body surface area partial-thickness scald burn by immersion for 18 s in a water bath at 60°C or 20°C (non-burn controls). In the treatment arm, an ATP hydrolyzing enzyme (ATPH, 400 mU/ml) was applied directly to the site immediately after injury. Skin was harvested after 24 h and 5 d. Elastase staining was used to quantify neutrophil infiltrate and activation. Extent of necrosis was measured from H&E slides. Ki-67 stains were quantified to determine cell proliferation. IFN-≤in 24 h skin samples was measured through qRT-PCR. Results represent the mean ( SD). Statistical significance was determined by ANOVA and Tukey test.

Results: At 24h, neutrophil elastase staining showed increased neutrophils in the burn group (20.45.3) compared to the control (v 13.45.2, p<.001) and ATPH (v 8.42.2, p<.001) groups (Fig. 1). Based on H&E staining, necrosis was less extensive in the ATPH group when compared to the burn group at 24 h and 5 d (Fig. 2). Ki-67 staining at 5d revealed significantly higher proliferation signal in the ATPH group than in the control group (p<.05). The intensity of elastase staining was reduced in the ATPH group when compared to the burn group at 24 h. IFN-≤ expression at 24 h in the ATPH group was significantly lower than in the burn group (0.450.11 v 5.92.5, p<.05, Fig. 3).

Conclusion: Our results support previous findings that extracellular ATP plays a role in neutrophil activity. We demonstrate that ATP hydrolysis at the burn site allays the neutrophil response to thermal injury and reduces tissue necrosis. This decrease in inflammation and tissue necrosis is at least partially due to IFN-≤ signaling. ATP hydrolyzing enzymes could be used as topical inflammatory regulators to quell the injury caused by inflammation.


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