MFG-E8 accelerates wound healing in diabetes by regulating “NLRP3 inflammasome-neutrophil extracellular traps” axis

Abstract

Sustained activation of NLRP3 inflammasome and release of neutrophil extracellular traps (NETs) impair wound healing of diabetic foot ulcers (DFUs). Our previous study reported that milk fat globule epidermal growth factor VIII (MFG-E8) attenuates tissue damage in systemic lupus erythematosus. However, the functional effect of MFG-E8 on “NLRP3 inflammasome-NETs” inflammatory loop in wound healing of diabetes is not completely elucidated. In this study, neutrophils from DFU patients are susceptible to undergo NETosis, releasing more NETs. The circulating levels of NET components neutrophil elastase and proteinase 3 and inflammatory cytokines IL-1β and IL-18 were significantly elevated in DFU patients compared with healthy controls or diabetic patients, in spite of higher levels of MFG-E8 in DFU patients. In Mfge8−/− diabetic mice, skin wound displayed exaggerated inflammatory response, including leukocyte infiltration, excessive activation of NLRP3 inflammasome (release of higher IL-1β, IL-18, and TNF-α), largely lodged NETs, resulting in poor angiogenesis and wound closure. When stimulated with high-dose glucose or IL-18, MFG-E8-deficient neutrophils release more NETs than WT neutrophils. After administration of recombinant MFG-E8, IL-18-primed NETosis of WT or Mfge8−/− neutrophils was significantly inhibited. Furthermore, NET and mCRAMP (component of NETs, the murine equivalent of cathelicidin LL-37 in human)-mediated activation of NLRP3 inflammasome and production of IL-1β/IL-18 were significantly elevated in Mfge8−/− macrophages compared with WT macrophages, which were also significantly dampened by the administration of rmMFG-E8. Therefore, our study demonstrated that as inhibitor of the “NLRP3 inflammasome-NETs” inflammatory loop, exogenous rMFG-E8 improves angiogenesis and accelerates wound healing, highlighting possible therapeutic potential for DFUs.

Bletilla striata Polysaccharide Promotes Diabetic Wound Healing Through Inhibition of the NLRP3 Inflammasome

“…As macrophage-mediated inflammatory cascades contribute the etiology of DFUs ( Zhang et al, 2017 ; Liu et al, 2019 ; Huang et al, 2020 ), we cocultured BMDMs with CMECs exposed to HG for 48 h in the presence or absence of BSP. As shown in Supplementary Figure S4 , compared to HG exposure alone, the levels of insulin stimulated p-Akt and p-GSK3β in CMECs decreased following they cocultured with BMDMs, indicating that the co-incubation with BMDMs impairs insulin sensitivity in CMECs exposed to HG.…”

“…Accumulating evidence suggests that chronic inflammation is key to the development of DFU ( Dinh et al, 2012 ; Mirza et al, 2013 ; Bitto et al, 2014 ; Noor et al, 2015 ; Kasiewicz and Whitehead, 2016 ). Notably, NLRP3 inflammasome activation in response to HG in DM, leads to IL-1β-evoked inflammatory cascades that contribute to the delay in DFU healing ( Mirza et al, 2014 ; Zhang et al, 2017 ; Liu et al, 2019 ; Huang et al, 2020 ). Recent report suggests that the sustained-activation of the NLRP3 inflammasome is a common feature in macrophages in the wounds of diabetic humans and mice, and blocking IL-1β activity using neutralizing antibodies, caspase-1 inhibitors and Nlrp3 or caspase-1 gene silencing can effectively improve DFU healing in mice ( Mirza et al, 2014 ).…”

“…Accumulating evidence indicates that DFU results from persistent inflammation caused by defective tissue repair responses in DM. Of note, the nucleotide-binding and oligomerization (NACHT) domain, leucine-rich repeat (LRR), and pyrin domain (PYD)-containing protein 3 (NLRP3) inflammasome-mediated inflammatory cascade has been suggested to contribute to the deficiencies in angiogenesis and ulcer healing in DM patients ( Mirza et al, 2013 ; Zhang et al, 2017 ; Liu et al, 2019 ; Huang et al, 2020 ).…”

“…Compared to non-diabetic patients, the expression of NLRP3, caspase-1 and IL-1β are significantly higher in the wound tissues of DFU patients ( Zhang et al, 2017 ). It is now recognized that the IL-1β-mediated inflammatory response can reduce insulin sensitivity and damage vascular endothelial cells, leading to defective wound healing ( Bitto et al, 2014 ; Zhang et al, 2017 ; Liu et al, 2019 ; Huang et al, 2020 ). Conversely, blocking either the NLRP3 inflammasome or IL-1β-mediated inflammatory reactions can effectively promote DFU healing ( Mirza et al, 2013 ; Bitto et al, 2014 ; Mirza et al, 2014 ; Huang et al, 2020 ).…”

NETosis in Wound Healing: When Enough Is Enough

“…NETosis dysregulation can also be induced by chronic inflammation, a mechanism responsible for the delayed wound healing process, as demonstrated in diabetic foot ulcer (DFU) patients, where NOD-like receptor protein (NLRP)-3 inflammasome-NETs axis was upregulated compared with both controls and diabetic patients with no ulcer [86,87]. Therefore, NETs have been recognized as markers of wound healing impairment in diabetic foot ulcers patients [88].…”