Attenuation of Flightless I, an actin-remodelling protein, improves burn injury repair via modulation of transforming growth factor (TGF)-β1 and TGF-β3

Adams, Damian; Ruzehaji, Nadira; Strudwick, Xanthe L.; Greenwood, John; Campbell, Hugh D.; Arkell, Ruth M.; Cowin, Allison J.

doi:10.1111/j.1365-2133.2009.09296.x

Cited by 40 publications

(56 citation statements)

References 27 publications

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“…Diabetic wounds treated with FnAb showed accelerated wound closure with a 1.9-fold decrease in average wound size observed in day 3 and day 7 WT FnAb-treated diabetic wounds. Similar effects have been reported in previous studies where FnAb improved the rate of incisional [17], excisional [20] and burn wound repair [19]. Dose-dependent effects of FnAb on cellular proliferation have also previously been demonstrated [20] and the addition of FnAb to human chronic wound fluids can reduce their inhibitory effect on cell proliferation in vitro [18].…”

Section: Discussionsupporting

confidence: 68%

“…alcohol for 1 min each. Sections were brought to water, rinsed in ×1 PBS and placed into 250 ml target retrieval solution as described previously [19]. Primary antibodies (1:100) were applied and incubated overnight at 4°C.…”

mentioning

confidence: 99%

“…Detection was by species-specific secondary antibodies (1:200). AnalySIS software package (Munster, Germany) was used to measure fluorescence intensity per unit area as described previously [19]. Control immunostaining was carried out by omitting primary or secondary antibodies for verification of staining and non-specific binding.…”

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confidence: 99%

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Attenuation of flightless I improves wound healing and enhances angiogenesis in a murine model of type 1 diabetes

et al. 2013

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Aims/hypothesis Skin lesions and ulcerations are severe complications of diabetes that often result in leg amputations. In this study we investigated the function of the cytoskeletal protein flightless I (FLII) in diabetic wound healing. We hypothesised that overexpression of FLII would have a negative effect on diabetic wound closure and modulation of this protein using specific FLII-neutralising antibodies (FnAb) would enhance cellular proliferation, migration and angiogenesis within the diabetic wound.Methods Using a streptozotocin-induced model of diabetes we investigated the effect of altered FLII levels through Flii genetic knockdown, overexpression or treatment with FnAb on wound healing. Diabetic wounds were assessed using histology, immunohistochemistry and biochemical analysis. In vitro and in vivo assays of angiogenesis were used to assess the angiogenic response. Results FLII levels were elevated in the wounds of both diabetic mice and humans. Reduction in the level of FLII improved healing of murine diabetic wounds and promoted a robust pro-angiogenic response with significantly elevated von Willebrand factor (vWF) and vascular endothelial growth factor (VEGF)-positive endothelial cell infiltration. Diabetic mouse wounds treated intradermally with FnAb showed improved healing and a significantly increased rate of reepithelialisation. FnAb improved the angiogenic response through enhanced formation of capillary tubes and functional neovasculature. Reducing the level of FLII led to increased numbers of mature blood vessels, increased recruitment of smooth muscle actin-α-positive cells and improved tight junction formation. Conclusions/interpretation Reducing the level of FLII in a wound may be a potential therapeutic approach for the treatment of diabetic foot ulcers.

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Section: Discussionsupporting

confidence: 68%

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confidence: 99%

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Attenuation of flightless I improves wound healing and enhances angiogenesis in a murine model of type 1 diabetes

et al. 2013

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“…However, Flii is upregulated in tissue in response to injury in mice and we now show fibroblasts increase secretion of Flii in response to wounding (Adams et al, 2009;Cowin et al, 2007). The upregulation of Flii in response to injury seems counterproductive since increasing the levels of Flii leads to reduced cell migration, impaired healing and increased scar formation.…”

Section: Discussionmentioning

confidence: 93%

“…1A,B). We have previously shown that Flii is upregulated in mouse skin wound tissue in response to injury (Adams et al, 2009;Cowin et al, 2007). Since Flii levels increase in the wounds of mice, scratch wounds were created in a confluent layer of NIH3T3 fibroblasts to determine whether scratch wounding these cells could alter Flii secretion.…”

Section: Flii Secretion Is Both Constitutive and Regulatedmentioning

confidence: 99%

Flightless, secreted through a late endosome/lysosome pathway, binds LPS and dampens cytokine secretion

Lei

Franken

Ruzehaji

et al. 2012

Journal of Cell Science

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SummaryFlightless (Flii) is upregulated in response to wounding and has been shown to function in wound closure and scarring. In macrophages intracellular Flii negatively modulates Toll-Like Receptor (TLR) signalling and dampens cytokine production. We now show that Flii is constitutively secreted from macrophages and fibroblasts and is present in human plasma. Secretion from fibroblasts is upregulated in response to scratch wounding and lipopolysaccharide (LPS)-activated macrophages also temporally upregulate their secretion of Flii. Using siRNA, and wild-type and mutant proteins, we show that Flii is secreted by means of a late endosomal/lysosomal pathway that is regulated by Rab7 and Stx11. Flii contains 11 leucine-rich repeat domains in its N-terminus that have nearly 50% similarity to those in the extracellular pathogen binding portion of Toll-like receptor 4 (TLR4). We show secreted Flii can also bind LPS and has the ability to alter macrophage activation. LPS activation of macrophages in Flii-depleted conditioned medium leads to enhanced macrophage activation and increased TNF secretion compared with cells activated in the presence of Flii. These results show secreted Flii binds to LPS and in doing so alters macrophage activation and cytokine secretion, suggesting that like the intracellular pool of Flii, secreted Flii also has the ability to alter inflammation.

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Delivery of Flightless I Neutralizing Antibody from Porous Silicon Nanoparticles Improves Wound Healing in Diabetic Mice

Turner

McInnes

Melville

et al. 2016

Adv Healthcare Materials

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Flightless I (Flii) is elevated in human chronic wounds and is a negative regulator of wound repair. Decreasing its activity improves healing responses. Flii neutralizing antibodies (FnAbs) decrease Flii activity in vivo and hold significant promise as healing agents. However, to avoid the need for repeated application in a clinical setting and to protect the therapeutic antibody from the hostile environment of the wound, suitable delivery vehicles are required. In this study, the use of porous silicon nanoparticles (pSi NPs) is demonstrated for the controlled release of FnAb to diabetic wounds. We achieve FnAb loading regimens exceeding 250 µg antibody per mg of vehicle. FnAb-loaded pSi NPs increase keratinocyte proliferation and enhance migration in scratch wound assays. Release studies confirm the functionality of the FnAb in terms of Flii binding. Using a streptozotocin-induced model of diabetic wound healing, a significant improvement in healing is observed for mice treated with FnAb-loaded pSi NPs compared to controls, including FnAb alone. FnAb-loaded pSi NPs treated with proteases show intact and functional antibody for up to 7 d post-treatment, suggesting protection of the antibodies from proteolytic degradation in wound fluid. pSi NPs may therefore enable new therapeutic approaches for the treatment of diabetic ulcers.

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Attenuation of Flightless I, an actin-remodelling protein, improves burn injury repair via modulation of transforming growth factor (TGF)-β1 and TGF-β3

Abstract: Strategies aimed at reducing Flii expression, for example using neutralizing antibodies, may lead to improved burn outcomes.

Cited by 40 publications

References 27 publications

Attenuation of flightless I improves wound healing and enhances angiogenesis in a murine model of type 1 diabetes

Attenuation of flightless I improves wound healing and enhances angiogenesis in a murine model of type 1 diabetes

Flightless, secreted through a late endosome/lysosome pathway, binds LPS and dampens cytokine secretion

Delivery of Flightless I Neutralizing Antibody from Porous Silicon Nanoparticles Improves Wound Healing in Diabetic Mice

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