Mechanical Force and Actin Dynamics during Cutaneous Squamous Cell Carcinoma (cSCC) Progression: Opportunities for Novel Treatment Modalities

Cutaneous squamous cell carcinoma (cSCC) accounts for 25% of cutaneous malignancies diagnosed in the Caucasian population. Surgical removal in combination with radio- and chemotherapy is an effective treatment; however, prognosis for patients suffering from aggressive cSCC is still relatively poor. Increasing prevalence coupled with high mortality and morbidity in aggressive metastatic forms of cSCC highlights the need for development of novel targeted therapeutics. Metastasis is a complex process requiring dramatic reorganization of the cell cytoskeleton. Recent studies have highlighted the importance of mechanical forces and actin dynamics in cancer cells’ intrinsic ability to invade adjacent tissues, intravasate into vasculature, and ultimately metastasize. Tight regulation of the biochemical and mechanical properties of the actin cytoskeleton drives cellular processes involved in cSCC progression including polarity establishment, morphogenesis, and motility. Here we will provide a short introduction to disease pathogenesis, give an overview of the role of key regulatory proteins governing the mechanical forces and actin dynamics critical to cSCC progression, and describe the contribution of actin remodeling and actomyosin signaling to cSCC progression. We will also discuss how targeting protein regulating mechanical force and actin dynamics may have clinical utility in development of novel treatment modalities for patients suffering from aggressive cSCC

The Role of Actin Remodelling Proteins in Wound Healing and Tissue Regeneration

The actin cytoskeleton is an essential network of filaments that is found in all cells and has an important role in regulating cellular activities. The dynamic regulation of cytoskeletal synthesis, remodelling and function is critical for many physiological processes and is integral for the successful repair of wounds. Wound healing relies on the fine balance between cellular proliferation, adhesion and migration, resulting in tightly controlled equilibrium between tissue regeneration and fibrosis. The actin cytoskeleton regulates all these processes and is therefore an important factor contributing to the re-establishment of the skin barrier function, restoration of the skin anatomical structure and wound repair; however, it also inevitably results in scar formation. Regulation of the actin cytoskeleton is tightly controlled by several large protein families, which are discussed in this chapter. Members of the FERM superfamily of proteins, the filamin and tropomyosin families of actin-associated proteins as well as the gelsolin family of actin remodelling proteins are all important regulators of the actin cytoskeleton, which can affect different stages of wound healing. Targeted therapies against different proteins involved in cytoskeletal regulation may lead to novel therapeutic interventions aimed at improving wound healing and reducing scar formation

Effect of Flightless protein on skin architecture, cellular responses and Epidermolysis Bullosa.

Wound healing is an area of largely unmet medical need with patients often relying on wound management practice rather than specific therapies. Recent research in our laboratory has identified a cytoskeletal protein Flightless (Flii) as a negative regulator of wound healing. This highly conserved protein is important in development and has a unique structure allowing it to act as a multifunctional protein. Flii expression increases in response to wounding, inhibiting cellular migration and proliferation while its deficiency improves wound healing. The aim of this study was to investigate the effect of differential Flii expression on skin architecture, cellular responses during wound healing, adhesionmediated cell signaling and skin blistering associated with the genetic skin disorder Epidermolysis Bullosa (EB). Chapter 3 of this thesis describes the effect of differential Flii expression on skin architecture and formation of hemidesmosomes which anchor the skin layers. Using primary fibroblasts and keratinocytes with varying Flii expression this study investigated the effect of Flii expression on cellular adhesion, spreading and migration on different extracellular matrix substrates. The results presented in Chapter 3 also describe the effect of Flii neutralising antibodies on primary keratinocyte proliferation illustrating improved proliferation in response to decreased Flii expression. In Chapter 4 an incisional wound healing model was used to investigate the effect of differential Flii expression on different components of hemidesmosomes. Flightless was shown to regulate hemidesmosome formation through its effects on integrin-mediated cellular adhesion and migration. Using immunoprecipitation studies, Flii association with structural and signaling proteins present at the dermal-epidermal junction was investigated. Flii was found to form a cytoskeletal complex with talin, vinculin and paxillin suggesting its role in downstream signaling. The association of Flii with paxillin was further investigated in Chapter 5 where the effect of Flii over-expression on fibroblast adhesion and formation of adhesion structures was examined. Flii over-expression inhibited paxillin activation and the turnover of adhesion structures through down regulation of signaling proteins involved in cell adhesion signaling pathways. Chapter 6 of this thesis summarises the effect of Flii in skin blistering by utilizing both human samples and two mouse models of Epidermolysis Bullosa. Flii expression is significantly increased in response to skin blistering and its effects on integrin mediated cellular adhesion, migration and type VII collagen expression make Flii a negative contributor to blister formation. Decreasing Flii expression genetically or using neutralizing antibodies reduces skin blistering, improves cellular adhesion and decreases TGF-β mediated collagen contraction. In summary, Flii adversely affects skin strength and blister formation. Using a multidi-mensional approach of both in vitro and in vivo methodologies, human tissue and animal models this thesis reveals several novel findings and contributes to better understanding the involvement of Flii in both maintaining skin homeostasis and regulating wound repair. Flii is a novel target for development of mechanistic based therapy for improved wound healing. Findings presented in this thesis may open doors to significant changes in clinical practice and contribute to better therapeutic design by which would healing of blisters in patients with Epidermolysis Bullosa might be improved.Thesis (Ph.D.) -- University of Adelaide, School of Paediatrics and Reproductive Health, 201

Skin Integrity, Antimicrobial Stewardship and Infection Control:A critical review of current best practice

‘Skin integrity’ refers to intact, unbroken, and healthy skin. Disruption of skin integrity can be caused by intrinsic and extrinsic factors including altered nutritional status, vascular disease, diabetes, and tissue injury, and this is often associated with development of localised clinical infection. Skin health and hygiene is important for preventing wounds and development of localised clinical infection or sepsis. Clinical wound infection is an increasing problem in healthcare, with the potential for increasing the burden of antimicrobial resistance (AMR), if antimicrobials are overused to treat wound infection. In this review we discuss skin integrity and wound infection prevention and outline the guiding principles of antimicrobial resistance and antimicrobial stewardship for infection control. Additionally, we provide a critical review of current best practice, highlighting the pathway to guide management of patients at risk of infection development, and discuss the latest research progress on antimicrobial resistance and skin integrity.</p

Nanotechnological Advances in Cutaneous Medicine

Review articleWound healing is an area of unmet clinical need. Current treatments include occlusive dressings, hydrogels, and antimicrobials to control infection. However with the growing number of antibiotic-resistant bacteria and the increase in population age and clinical obesity, it is becoming proportionally harder to treat wounds with the drugs that have worked in the past. There is an urgent requirement for efficient mechanism-based treatments and more efficacious drug delivery systems. The potential of using nanoparticles as a drug delivery system has been identified and investigated. Nanoparticles have the ability to protect and carry drugs to specific targets in the body, enabling slower degradation, enhancing drug penetration, improving treatment efficacy with lower systemic absorption, and reducing unwanted side effects. Here we discuss the advantages and limitations of nanotechnology for the treatment of wounds and other cutaneous disorders.Jessica E. Jackson, Zlatko Kopecki and Allison J. Cowi

Bacteria-Activated Dual pH- and Temperature-Responsive Hydrogel for Targeted Elimination of Infection and Improved Wound Healing

Antibacterial treatment that provides on-demand release of therapeutics that can kill a broad spectrum of pathogens while maintaining long-term efficacy and without developing resistance or causing side effects is urgently required in clinical practice. Here, we demonstrate the development of a multistimuli-responsive hydrogel, prepared by cross-linking N-isopropylacrylamide with acrylic acid and loaded with ultrasmall silver nanoparticles (AgNPs), offering the on-demand release of Ag+ ions triggered by changes in the wound microenvironment. We demonstrate that this dual-responsive hydrogel is highly sensitive to a typical wound pH and temperature change, evidenced by the restricted release of Ag+ ions at acidic pH (7.4) (>90% release). The pH-dependent release and antibacterial effect show minimal killing at pH 4 or 5.5 but dramatically activated at pH 7.4 and 10, eliminating >95% of the pathogens. The in vivo antibacterial efficacy and safety showed a high potency to clear Staphylococcus aureus wound infection while significantly accelerating the wound healing rate. This multifunctional hydrogel presents a promising bacteria-responsive delivery platform that serves as an on-demand carrier to not only reduce side effects but also significantly boost the antibacterial efficiency based on physiological needs. It offers great potential to improve the way wound infections are treated with direct clinical implications, providing a single platform for long-lasting application in wound management

Lifting the Silver Flakes: The Pathogenesis and Management of Chronic Plaque Psoriasis

Psoriasis is a common chronic inflammatory skin condition in which patients suffer from mild to chronic plaque skin plaques. The disease manifests through an excessive inflammatory response in the skin due to complex interactions between different genetic and environmental factors. Psoriasis can affect the physical, emotional, and psychosocial well-being of patients, and currently there is no cure with treatments focusing primarily on the use of anti-inflammatory agents to control disease symptoms. Traditional anti-inflammatory agents can cause immunosuppression and adverse systemic effects. Further understanding of the disease has led to current areas of research aiming at the development of selective molecular targets to suppress the pathogenic immune responses

Flightless I regulates hemidesmosome formation and integrin-mediated cellular adhesion and migration during wound repair

Flightless I (Flii), a highly conserved member of the gelsolin family of actin-remodelling proteins associates with actin structures and is involved in cellular motility and adhesion. Our previous studies have shown that Flii is an important negative regulator of wound repair. Here, we show that Flii affects hemidesmosome formation and integrin-mediated keratinocyte adhesion and migration. Impaired hemidesmosome formation and sparse arrangements of keratin cytoskeleton tonofilaments and actin cytoskeleton anchoring fibrils were observed in FliiTg/+ and FliiTg/Tg mice with their skin being significantly more fragile than Flii+/− and WT mice. Flii+/− primary keratinocytes showed increased adhesion on laminin and collagen I than WT and FliiTg/Tg primary keratinocytes. Decreased expression of CD151 and laminin-binding integrins α3, β1, α6 and β4 were observed in Flii overexpressing wounds, which could contribute to the impaired wound re-epithelialization observed in these mice. Flii interacts with proteins directly linked to the cytoplasmic domain of integrin receptors suggesting that it may be a mechanical link between ligand-bound integrin receptors and the actin cytoskeleton driving adhesion-signaling pathways. Therefore Flii may regulate wound repair through its effect on hemidesmosome formation and integrin-mediated cellular adhesion and migration.

Lysosomal secretion of Flightless I upon injury has the potential to alter inflammation

Intracellular Flightless I (Flii), a gelsolin family member, has been found to have roles modulating actin regulation, transcriptional regulation and inflammation. In vivo Flii can regulate wound healing responses. We have recently shown that a pool of Flii is secreted by fibroblasts and macrophages, cells typically found in wounds, and its secretion can be upregulated upon wounding. We show that secreted Flii can bind to the bacterial cell wall component lipopolysaccharide and has the potential to regulate inflammation. We now show that secreted Flii is present in both acute and chronic wound fluid