Emily Lynam

The effect of amphiphilic siloxane oligomers on fibroblast and keratinocyte proliferation and apoptosis

The formation of hypertrophic scars (HSF) is a frequent medical outcome of wound repair and often requires further therapy with treatments such as silicone gel sheets (SGS) or apoptosis-inducing agents, including bleomycin. Although widely used, knowledge regarding SGS and their mode of action is limited. Preliminary research has shown that small amounts of amphiphilic silicone present in SGS have the ability to move into skin during treatment. We demonstrate herein that a commercially available analogue of these amphiphilic siloxane species, the rake copolymer GP226, decreases collagen synthesis on exposure to cultures of fibroblasts derived from HSF. By size exclusion chromatography, GP226 was found to be a mixture of siloxane species, containing five fractions of different molecular weight. By studies of collagen production, cell viability and proliferation, it was revealed that a low molecular weight fraction (fraction IV) was the most active, reducing the number of viable cells present after treatment and thereby reducing collagen production as a result. On exposure of fraction IV to human keratinocytes, viability and proliferation were also significantly affected. HSF undergoing apoptosis after application of fraction IV were also detected via real-time microscopy and by using the TUNEL assay. Taken together, these data suggests that these amphiphilic siloxanes could be potential non-invasive substitutes to apoptotic-inducing chemical agents that are currently used as scar treatments.

Severe hypoxia and malnutrition collectively contribute to scar fibroblast inhibition and cell apoptosis.

This study aims to investigate whether severe hypoxia and malnutrition in scar tissue play key roles to induce hypertrophic scar regression. And scar‐derived fibroblasts were treated with moderate/severe hypoxia and malnutrition to model condition of proliferative and regressive scar (5%O2+5%FCS and 0.5%O2 + 0.5%FCS), and normoxia with well nutrition as control (10%O2 + 10%FCS). Our results demonstrated that severe hypoxia and malnutrition resulted in significantly reduced cell viability and collagen production, as well as HIF‐1, VEGF, TGF‐β1, and Bcl‐2 protein expression when compared with control, and cell apoptosis occurred. Therefore, the severe hypoxia and malnutrition in scar tissue contribute to fibroblast inhibition and cell apoptosis, which is correlated with scar regression.

A pre-clinical functional assessment of an acellular scaffold intended for the treatment of hard-to-heal wounds.

The majority of the population experience successful wound‐healing outcomes; however, 1–3% of those aged over 65 years experience delayed wound healing and wound perpetuation. These hard‐to‐heal wounds contain degraded and dysfunctional extracellular matrix (ECM); yet, the integrity of this structure is critical in the processes of normal wound healing. Here, we evaluated a novel synthetic matrix protein for its ability to act as an acellular scaffold that could replace dysfunctional ECM. In this regard, the synthetic protein was subjected to adsorption and diffusion assays using collagen and human dermal tissues; evaluated for its ability to influence keratinocyte and fibroblast attachment, migration and proliferation and assessed for its ability to influence in vivo wound healing in a porcine model. Critically, these experiments demonstrate that the matrix protein adsorbed to collagen and human dermal tissue but did not diffuse through human dermal tissue within a 24‐hour observation period, and facilitated cell attachment, migration and proliferation. In a porcine wound‐healing model, significantly smaller wound areas were observed in the test group compared with the control group following the third treatment. These data provide evidence that the synthetic matrix protein has the ability to function as an acellular scaffold for wound‐healing purposes.

Functional and mechanistic investigation of Shikonin in scarring.

Scarring is a significant medical burden; financially to the health care system and physically and psychologically for patients. Importantly, there have been numerous case reports describing the occurrence of cancer in burn scars. Currently available therapies are not satisfactory due to their undesirable side-effects, complex delivery routes, requirements for long-term use and/or expense. Radix Arnebiae (Zi Cao), a perennial herb, has been clinically applied to treat burns and manage scars for thousands of years in Asia. Shikonin, an active component extracted from Radix Arnebiae, has been demonstrated to induce apoptosis in cancer cells. Apoptosis is an essential process during scar tissue remodelling. It was therefore hypothesized that Shikonin may induce apoptosis in scar-associated cells. This investigation presents the first detailed in vitro study examining the functional responses of scar-associated cells to Shikonin, and investigates the mechanisms underlying these responses. The data obtained suggests that Shikonin inhibits cell viability and proliferation and reduces detectable collagen in scar-derived fibroblasts. Further investigation revealed that Shikonin induces apoptosis in scar fibroblasts by differentially regulating the expression of caspase 3, Bcl-2, phospho-Erk1/2 and phospho-p38. In addition, Shikonin down-regulates the expression of collagen I, collagen III and alpha-smooth muscle actin genes hence attenuating collagen synthesis in scar-derived fibroblasts. In summary, it is demonstrated that Shikonin induces apoptosis and decreases collagen production in scar-associated fibroblasts and may therefore hold potential as a novel scar remediation therapy.