Rudolf J Wiesner | Julia Boix Tarin - C 17

Upon aging, all organs of the body suffer from a progressive impairment in structure, morphology and function due to intrinsic factors. This contributes to a wide variety of skin alterations and pathologies such as skin atrophy and wound healing problems. The skin, as the outermost organ, acts as an external barrier that is exposed not only to such intrinsic, but also to extrinsic factors. The most common cause for external aging is UV irradiation (photoaging), which worsens and accelerates the aging process. Ultimately, long-term photoaging effects contribute to the formation of malignant skin tumors.

The accumulation of mitochondrial DNA (mtDNA) deletions has been related to aging in many cell types of the body (Larsson, 2010). In skin, mtDNA deletions were mostly found in the dermis, but not in the epidermis, and especially in areas exposed to UV-irradiation (Berneburg et al., 2004; Krishnan et al., 2006). Epidermal turnover is fast due to continuously proliferating keratinocytes, in contrast to dermal fibroblasts that rarely divide. We hypothesize that the proliferation rate is key for variable accumulation of mtDNA deletions in different tissues. Understanding these processes in skin will help us to understand the contribution of mtDNA defects to aging in general and will allow us to test interventions in this easily accessible organ.

1. Study the consequences of mtDNA deletions in adult mouse skin caused by photoaging and/or by genetic induction. Photoaging is represented by UV irradiation and mtDNA deletions are induced using a mouse model expressing a mutated mitochondrial helicase (K320E-Twinkle), which causes an accelerated accumulation of mtDNA deletions in patients (Suomalainen, 1997).
2. Understand why mtDNA deletions accumulate differentially in epidermis and dermis during photoaging.
3. Decipher the molecular mechanisms behind the differential mtDNA deletion accumulation process depending on the skin compartment by using mouse models which allow induction of K320E-Twinkle in the epidermis and dermis as well as photoaging.
4. Investigate whether mtDNA deletions delay the wound healing process and, in that case, if they contribute to the non-healing “status” of chronic human wounds.

Our group has generated a mouse model that allows the expression of a mutated form of the mitochondrial helicase Twinkle (K320E point mutation) in the tissue of our choice.

We investigated the effect of K320E-Twinkle in slowly- or non-dividing cells such as dermal fibroblasts, skeletal muscle fibers and cardiomyocytes. Our results indicate that indeed the two latter models resemble human aging with accumulation of mtDNA deletions and consecutive late aging phenotypes like muscle fiber replacement (Kimoloi et al., submitted) and severe arrhythmia, respectively (Fig. 1. Baris et al., 2015). Dermal fibroblasts present mtDNA depletion when proliferating in vitro and mtDNA deletions, but no obvious aging phenotype in vivo (collaboration with AG Eming; Knuever, Boix et al., in preparation).

When expressing K320E-Twinkle in chondrocytes of the bone growth plate, mice developed a short stature, similar to many patients suffering from mitochondrial disease due to mtDNA mutations, and later showed cartilage degeneration typical for advanced age (Holzer et al., 2019).

In contrast, in the highly proliferative epidermis, the K320E-Twinkle^epi keratinocytes show drastic mtDNA depletion instead of an accumulation of mtDNA deletions in vivo. This, however, does not lead to a severe epidermal phenotype but, surprisingly, to the early death of the mice with low levels of glucose and high lactate levels in blood and a severe inflammatory phenotype (Fig. 2. Weiland et al., 2018).

Those results, together with the fact that mtDNA deletions were not found in human UV-exposed epidermis, suggest that keratinocytes are protected from the accumulation of these mtDNA defects. The high epidermal proliferation rate may be key to prevent their accumulation. However, the unsolved question is: What are the molecular mechanisms that allow keratinocytes to escape from the accumulation of mtDNA defects?

Due to the early death of K320E-Twinkle^epi mice we could not investigate the consequences of mtDNA deletions in the epidermis of adult mice and its contribution to skin aging and inflammation, which is now part of this new proposal.

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2. Berneburg M, Plettenberg H, Medve-König K, Pfahlberg A, Gers-Barlag H, Gefeller O, Krutmann J. Induction of the photoaging-associated mitochondrial common deletion in vivo in normal human skin. J Invest Dermatol. 2004 122:1277-83.
3. Holzer T, Probst K, Etich J, Auler M, Georgieva VS, Bluhm B, Frie C, Heilig J, Niehoff A, Nüchel J, Plomann M, Seeger JM, Kashkar H, Baris OR, Wiesner RJ, Brachvogel B. Respiratory chain inactivation links cartilage-mediated growth retardation to mitochondrial diseases. J Cell Biol. 2019 218:1853-1870.
4. Kloepper JE, Baris OR, Reuter K, Kobayashi K, Weiland D, Vidali S, Tobin DJ, Niemann C, Wiesner RJ, Paus R. Mitochondrial function in murine skin epithelium is crucial for hair follicle morphogenesis and epithelial-mesenchymal interactions. J Invest Dermatol. 2015 135:679-689.
5. Krishnan KJ, Birch-Machin MA. The incidence of both tandem duplications and the common deletion in mtDNA from three distinct categories of sun-exposed human skin and in prolonged culture of fibroblasts. J Invest Dermatol. 2006 126:408-15.
6. Larsson NG. Somatic mitochondrial DNA mutations in mammalian aging. Annu Rev Biochem 2010 79:683-706.
7. Suomalainen A. Mitochondrial DNA and disease. Ann Med. 1997 29:235-46
8. Weiland D, Brachvogel B, Hornig-Do HT, Neuhaus JFG, Holzer T, Tobin DJ, Niessen CM, Wiesner RJ, Baris OR. Imbalance of Mitochondrial Respiratory Chain Complexes in the Epidermis Induces Severe Skin Inflammation. J Invest Dermatol. 2018 138:132-140.