Streptolysin O enhances keratinocyte migration and proliferation and promotes skin organ culture wound healing in vitro

Krzyzanowska

et al. 2010

Mol Med

Self Cite

134

Transforming growth factor β (TGFβ) is important in inflammation, angiogenesis, reepithelialization and connective tissue regeneration during wound healing. We analyzed components of TGFβ signaling pathway in biopsies from 10 patients with nonhealing venous ulcers (VUs). Using comparative genomics of transcriptional profiles of VUs and TGFβ-treated keratinocytes, we found deregulation of TGFβ target genes in VUs. Using quantitative polymerase chain reaction (qPCR) and immunohistochemical analysis, we found suppression of TGFβRI, TGFβRII and TGFβRIII, and complete absence of phosphorylated Smad2 (pSmad2) in VU epidermis. In contrast, pSmad2 was induced in the cells of the migrating epithelial tongue of acute wounds. TGFβ-inducible transcription factors (GADD45β, ATF3 and ZFP36L1) were suppressed in VUs. Likewise, genes suppressed by TGFβ (FABP5, CSTA and S100A8) were induced in nonhealing VUs. An inhibitor of Smad signaling, Smad7 was also downregulated in VUs. We conclude that TGFβ signaling is functionally blocked in VUs by downregulation of TGFβ receptors and attenuation of Smad signaling resulting in deregulation of TGFβ target genes and consequent hyperproliferation. These data suggest that application of exogenous TGFβ may not be a beneficial treatment for VUs.

Section: Discussionmentioning

confidence: 99%

“…Healthy skin samples were used to generate acute wounds as previously described (30)(31)(32)(33). Ex vivo human experimental wound models have been extensively used to study wound healing in human skin.…”

Section: Human Skin Ex Vivo Wound-healing Modelmentioning

confidence: 99%

Attenuation of the Transforming Growth Factor β-Signaling Pathway in Chronic Venous Ulcers

Krzyzanowska

et al. 2010

Mol Med

Self Cite

134

“…Interestingly, ligand-activated Ser 211 -phosphorylated-GR (GR-P) was localized in the nuclei of epidermal keratinocytes, suggesting endogenous hormone-induced activation of the receptor. To analyze the activity of the GC pathway in isolated epidermis without an external source of GCs, we incubated normal human skin on an air-liquid interface in basal, serum-free medium (40) in the absence of hydrocortisone for 24 h. Skin specimens were stained with GR-P antibody, and the signal was found in more than 30% of the nuclei of the cells in the epidermis of untreated human skin (Fig. 1A).…”

Section: Resultsmentioning

confidence: 99%

“…Cortisol Production Is Regulated during Acute Wound Healing-To further understand GC synthesis during wound healing, we used an established ex vivo human skin organ culture wound model (40). Acute wounds were maintained for 0, 4, 24, 48, and 96 h post wounding at the air-liquid interface, and the expression level of CYP11B1 was determined using realtime qPCR.…”

Section: Cyp11b1 An Enzyme Responsible For the Finalmentioning

confidence: 99%

Cortisol Synthesis in Epidermis Is Induced by IL-1 and Tissue Injury

Vukelic

Journal of Biological Chemistry

et al. 2011

Self Cite

176

196

Glucocorticoids (GCs) are known inhibitors of wound healing. In this study we report the novel finding that both keratinocytes in vitro and epidermis in vivo synthesize cortisol and how this synthesis regulates wound healing. We show that epidermis expresses enzymes essential for cortisol synthesis, including steroid 11 ␤-hydroxylase (CYP11B1), and an enzyme that controls negative feedback mechanism, 11␤-hydroxysteroid dehydrogenase 2 (11␤HSD2). We also found that cortisol synthesis in keratinocytes and skin can be stimulated by ACTH and inhibited by metyrapone (CYP11B1 enzyme inhibitor). Interestingly, IL-1␤, the first epidermal signal of tissue injury, induces the expression of CYP11B1 and increases cortisol production by keratinocytes. Additionally, we found induction of CYP11B1 increased production of cortisol and activation of GR pathway during wound healing ex vivo and in vivo using human and porcine wound models, respectively. Conversely, inhibition of cortisol synthesis during wound healing increases IL-1␤ production, suggesting that cortisol synthesis in epidermis may serve as a local negative feedback to proinflammatory cytokines. Local GCs synthesis, therefore, may provide control of the initial proinflammatory response, preventing excessive inflammation upon tissue injury. Inhibition of GC synthesis accelerated wound closure in vivo, providing the evidence that modulation of cortisol synthesis in epidermis may be an important regulatory mechanism during wound healing.Acute wound healing is a complex biological process mediated by a network of signaling pathways that coordinate multiple cellular processes, including cellular migration and proliferation, ultimately leading to barrier restoration (1). Wound healing is a tightly spatiotemporally regulated process, and changes in any component of this process can be detrimental, leading to further tissue damage or impairment of healing (2, 3).For example, timing of inflammatory response is essential; proinflammatory signals are important in the early stage of healing, but they can be detrimental if they persist (4, 5), underscoring a need for tight control of both stimulators and inhibitors during the wound healing process.Upon injury, keratinocytes are the first cells that respond (6, 7) by releasing pre-stored interleukin-1 (IL-1) (8). IL-1 has an autocrine and a paracrine function; that is, to activate keratinocytes and to alert the surrounding cells and tissues. The release of IL-1 by keratinocytes, with subsequent release of additional signaling molecules (9, 10), demarcates the proinflammatory phase of wound healing. In response to these signals, activated keratinocytes start migrating and proliferating (7,8,11). Successful repair after tissue injury requires resolution of inflammatory response, transitioning keratinocyte (HEK) from activated to differentiating phenotype. However, very little is known about endogenous epidermal signals that control keratinocyte activation cycle and inflammatory response.Epidermal keratinocytes have important immunol...

“…Wounds were created using 3-mm punch biopsies through the reticular dermis as previously described (29). They were maintained at the air-liquid interface using Dulbecco's modified Eagle's medium (Invitrogen) with antibiotics and antimycotics and stripped bovine serum.…”

Section: Methodsmentioning

confidence: 99%

Farnesyl Pyrophosphate Inhibits Epithelialization and Wound Healing through the Glucocorticoid Receptor

Vukelic

Journal of Biological Chemistry

et al. 2010

Self Cite

Farnesyl pyrophosphate (FPP), a key intermediate in the mevalonate pathway and protein farnesylation, can act as an agonist for several nuclear hormone receptors. Here we show a novel mechanism by which FPP inhibits wound healing acting as an agonist for glucocorticoid receptor (GR). Elevation of endogenous FPP by the squalene synthetase inhibitor zaragozic acid A (ZGA) or addition of FPP to the cell culture medium results in activation and nuclear translocation of the GR, a known wound healing inhibitor. We used functional studies to evaluate the effects of FPP on wound healing. Both FPP and ZGA inhibited keratinocyte migration and epithelialization in vitro and ex vivo. These effects were independent of farnesylation and indicate that modulation of FPP levels in skin may be beneficial for wound healing. FPP inhibition of keratinocyte migration and wound healing proceeds, in part, by repression of the keratin 6 gene. Furthermore, we show that the 3-hydroxy-3-methylglutaryl-CoA-reductase inhibitor mevastatin, which blocks FPP formation, not only promotes epithelialization in acute wounds but also reverses the effect of ZGA on activation of the GR and inhibition of epithelialization. We conclude that FPP inhibits wound healing by acting as a GR agonist. Of special interest is that FPP is naturally present in cells prior to glucocorticoid synthesis and that FPP levels can be further altered by the statins. Therefore, our findings may provide a better understanding of the pleiotropic effects of statins as well as molecular mechanisms by which they may accelerate wound healing.