PI Ries

Innovative therapeutic strategies in the treatment of sulfur mustard-evoked skin injuries: modulation of HIF-1α signaling and microRNA regulated pathways. 

Sulphur mustard (2,2´-dichlordiethylsulfide; SM) is an alkylating, highly toxic chemical agent that after exposure to humans causes severe inflammation and extensive blistering in skin and lung within several hours. The acute symptoms are followed by long term effects including impaired wound healing. SM has been used as a chemical warfare agent in various military conflicts during the twentieth century. Its relative ease of production and stockpiling together with its multiple incapacitating health effects make mustard gas a continuing threat by military conflicts and terrorist attacks. Identification of effective therapies for SM-induced injuries is the focus of research worldwide.

 

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SM upregulates miR-203 in keratinocytes under normoxia and hypoxia, and augments hypoxia-induced levels of miR-210. Antagomirs targeting miR-203 and miR-210 countermeasure SM-evoked cellular dysfunction (Deppe J. et al., Tox Letters, 2015).

 

The results of our research projects supported by the German Federal Ministry of Defense in 2005-2011 (M/SAB1/A001 and M/SABX/8A002) provided for the first time insight into molecular and cellular mechanisms that may contribute to the pathogenesis of SM injury. Our in vitro studies demonstrated that SM upregulates the release of proteinases from various skin cells. Remarkably, SM exposure of keratinocytes was shown to trigger the release of soluble factors from the cells which then induce an enhanced secretion of matrix metalloproteinase 9 (MMP-9) from fibroblasts. This may represent a SM-mediated pathomechanism that can lead to an increased degradation of basement membranes and thereby facilitate blister formation in skin (Ries C. et al., Toxicology, 2009). Moreover, our findings indicated that SM triggers premature differentiation in keratinocytes via p38 MAP kinase activity which is negatively influenced by ERK1/2. These events may contribute to the impaired regeneration capacity of skin after exposure to SM (Popp T. et al., Toxicology Letters, 2011). Together, our results suggest the usefulness of MMP-9 and p38 MAP kinase-inhibitors when applied in a timely differentiated manner for the treatment of SM-induced acute and chronic symptoms.

Calcium (Ca2+) is an important regulator of keratinocyte differentiation in the epidermis. Investigating the underlying molecular mechanisms we discovered that Wnt5a/β-catenin signaling is involved in this process. Binding of Ca2+ to the calcium-sensing receptor on the cell surface elevates free intracellular Ca2+ and upregulates the expression and secretion of Wnt5a in these cells. Wnt5a then acts as an autocrine stimulus by increasing β-catenin stability and signaling activity that promotes keratinocyte differentiation. Importantly, our results show that application of Wnt5a to the cells and stimulation of β-catenin activity accelerates differentiation of keratinocytes. A deeper knowledge of this regulatory network may improve the development of novel therapeutic approaches to enhance epidermal regeneration in degenerative skin diseases (Popp T. et al., J Invest Dermatol, 2014).

In a continuing project (M/SABX/BA003) funded 2011-2014 we investigated the importance of oxygen-deficiency (hypoxia), especially the role of HIF-1α and microRNAs in the pathophysiology of SM. Under normal physiological conditions, wound-associated hypoxia is a timely-limited situation that acts as an important stimulus for proper healing and regeneration in skin. Hypoxia controls the function and behaviour of keratinocytes and fibroblasts by influencing the expression of various regulatory molecules including cytokines and proteinases. In this context, HIF-1α plays a key role because it is significantly upregulated during hypoxic conditions in the skin and thereby stimulates various processes including cell proliferation, migration, autophagy, and angiogenesis that facilitate wound healing. The results of our studies provide evidence that SM attenuates hypoxia-induced HIF-1α accumulation and target gene expression in human primary keratinocytes and dermal fibroblasts, thereby impeding the migratory potential of these cells. Furthermore we demonstrate, that the addition of IOX2, a synthetic inhibitor of PHD-2 activity, fully restores HIF-1α stability and cellfuntionality in SM-intoxicated keratinocytes and fibroblasts (Deppe J. et al., Arch Toxicol, 2015).

microRNAs (miRNAs) are a group of small non-coding RNA molecules that play key roles in the regulation of numerous physiological and pathological processes. In wound healing, miRNAs are involved in the control of inflammation, angiogenesis, and apoptosis especially by influencing the functions of keratinocytes and fibroblasts. We hypothesize, that SM influences miRNA expression profiles in keratinocytes which may contribute to defects in functions of these cells essential in normal wound healing. Our studies demonstrate for the first time that SM induces miR-203 expression in primary keratinocytes independent of oxygen levels, and augments miR-210 in these cells under a hypoxic state. We show that upregulation of these miRNAs contribute to SM-induced deficiencies in cellular viability, proliferation, and differentiation of keratinocytes. Furthermore, our results suggest that specific inhibitors of miR-203 and miR-210 (anti-miRs) might be useful to counterbalance disturbances of these processes under pathological conditions. Our findings may have implications for the development of novel therapies to improve re-epithelialization and wound healing in skin lesions of patients after exposure to SM (Deppe J. et al., Tox Letters, 2015).

 

 

 

 

 
 
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Direktor:

Prof. Dr. med. Christian Weber

Institute for Cardiovascular Prevention (IPEK)

 

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