Low Microrna-21 In Diabetic Wound Macrophages Impair Resolution Of Inflammation
AMITAVA Das, Mithun Sinha, Pradipta Banerjee, Nirupam Biswas, Savita Khanna, Chandan K. Sen, Sashwati Roy.
Indiana University School of Medicine, Indianapolis, IN, USA.
Background: Under conditions of diabetes, wound macrophages (wmϕ) suffer from compromised efferocytosis which impairs resolution of inflammation. Our previous work demonstrated that delivery of miR-21 to human monocyte-derived macrophages (MDM) enhances efferocytosis and helps resolve inflammation. Here we test the hypothesis that diabetic wound macrophages are deficient in miR-21 which causes persistent inflammation. Methods: wmϕ were isolated from clinically presented chronic wounds using a standardized process involving processing of negative-pressure wound therapy (NPWT) dressings. In mice, subcutaneously implanted PVA sponges were used to isolate wmϕ. Results: Both human and murine diabetic wmϕ showed significantly lower miR-21 levels compared to non-diabetics (p< 0.05; n=4). To determine the significance of lower miR-21 in wmϕ fate and resolution of inflammation, a myeloid cell-specific miR-21 knockdown mice (miR-21mϕΔ/Δ) was generated by crossbreeding mice carrying floxed miR-21 allele (miR-21fl/fl) with LysM-Cre mice. In wmϕ of miR-21mϕΔ/Δ, expression of pro-inflammatory markers were significantly higher, and those representing reparative phenotype were attenuated (p<0.05; n=4). Excisional wounds of miR-21mϕΔ/Δ mice showed increased abundance of neutrophils and elevated levels of inflammation related cytokines (p<0.05; n=4). Bioinformatics analyses predicted the neutrophil chemoattractant GRO-α to be a direct target of miR-21. Using 3’-UTR firefly luciferase reporter, this work validated GRO-α as a direct target of miR-21 (p<0.05; n = 4). Lower miR-21 in diabetic and miR-21mϕΔ/Δ wmϕ desilenced GRO-α (p<0.05; n=4) causing marked increase in neutrophil recruitment to the wound-site. Conclusion: This work recognizes a novel miR-21-GRO-α mechanism underlying diabetes related persistent inflammation.
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