Schutz vor oxidativem Stress in retinalen Geweben durch Tyrosinase und seiner Metaboliten in Kombination mit oraler Zinkaufnahme
Zusammenfassung der Projektergebnisse
The question whether tyrosinase and melanisation are involved in defence against stress has not been fully answered. Some findings indicate a protective role for tyrosinase in the adult RPE, since tyrosinase-transduced cells appeared morphologically reorganised and had improved phagocytic function compared to controls (Biesemeier et al, submitted). In addition, the transduction was not toxic to the ARPE 19 cells. The melanin precursor DHI has been found to have pro- antioxidative functions depending on its concentration both in vitro and in vivo experiments. While 10 µM DHI showed cytoprotective effects in cell culture experiments and photoreceptor function after intense light, 100 µM DHI induced cell death of RPE cells and photoreceptor cones (Heiduschka et al, 2007). Unfortunately, the influence of melanin and zinc in primary RPE cells could not be investigated, because lipofuscin accumulation in aged donor cells distorted the results (Biesemeier et al, 2008). However, in vivo experiments on pigmented and unpigmented rats showed that the antioxidative properties of both melanin and zinc are interrelated. Pigmented rats showed a decrease in their antioxidative properties and an increased lipofuscin accumulation after a zinc free diet. In contrast, albino animals showed no differences after a zinc free diet, but were altogether more susceptible to oxidative stress and had an overall increased lipofuscin accumulation when compared to pigmented individuals (Kokkinou et al, submitted). The hypothesis that melanogenesis can be induced in non-pigmented adult human RPE cells has been verified, since melanin synthesis was measured quantitatively with light and electron microscopy and qualitatively with HPLC analysis after transduction with the vector AdTyr. Furthermore, a pathway of melanogenesis is presented, which is different to the classical scheme described in the literature. This pathway lacks both typical premelanosomes and tyrosinase trafficking using Golgi-derived vesicles. Instead, a formation of tyrosinase at free ribosomes and trafficking of the soluble protein to melanosomal stages are indicated (Biesemeier et al, in print). Meanwhile, the given mechanism has also been observed in non-transduced RPE cells of adult cattle both in cell culture and in retinal explants. It is proposed that this pathway may occur in vivo, but has not been recognised yet, since it is different to the known scheme. Evaluating the influence of ROS phagocytosis on tyrosinase activity and melanogenesis, diverging results have been obtained for primary donor RPE cells and ARPE-19 cells, respectively. While primary cells of both human donors and bovine eyes were able to activate tyrosinase synthesis in response to phagocytosis (Julien et al, 2007; Schraermeyer et al, 2006), non-transduced ARPE-19 cells did not produce tyrosinase in response to feeding alone. Nevertheless, after induction of tyrosinase with the adenoviral vector AdTyr, they synthesised more tyrosinase in response to ROS feeding than merely transduced cells. Additionally, they were also able to synthesise more melanin (Biesemeier et al, submitted). The transfer of ingested material to melanosomes could not be confirmed for ARPE-19 cells. Nevertheless, in the present experiments, no lipofuscin was generated, demonstrating that the degradation machinery was not inhibited by the experimental design. Moreover, the absence of lipofuscin indicates a beneficial role for tyrosinase in ROS degradation. The observed findings may help to develop gene therapy of melanin-associated aged related diseases and of albinism disorders. A patent, dealing with tyrosinase, melanin and melanogenic precursors as medication against oxidative stress related disease has been obtained in the time course of the project. As a more obvious approach, tyrosinase may help to keep cultured RPE cells differentiated and to avoid loss of typical RPE functions as pigment building and phagocytosis.
Projektbezogene Publikationen (Auswahl)
- (2004) The pigmentation of human iris influences the uptake and storing of zinc. Pigment Cell Res 17: 515-518
Kokkinou D, Kasper HU, Bartz-Schmidt KU, Schraermeyer U
- (2005) Zinc uptake and storage: the role of fundus pigmentation. Graefes Arch Clin Exp Ophthalmol 243(10): 1050-1055
Kokkinou D, Kasper HU, Schwarz T, Bartz-Schmidt KU, Schraermeyer U
- (2006) Influence of zinc deficiency on pigmentation. (ESPCR, Barcelona, Spain, September 2006, Pigment Cell Res 19:5, 527, Abstract No. PP037
Schraermeyer U, Kokkinou D
- (2006) Pigmented and albino rats show different response to light as studied by electroretinography and visual evoked potentials. (ESPCR, Barcelona, Spain, September 2006) Pigment Cell Res 19:5, 531, Abstract No. PP048
Heiduschka P, Schraermeyer U
- (2006) Tyrosinase biosynthesis and trafficking in adult human retinal pigment epithelial cells. (ESPCR, Barcelona, Spain, September 2006) Pigment Cell Res. 19:5, 539, Abstract No. PP074
Julien S, Kociok N, Kreppel F, Brito V, Peters S, Henke-Fahle S, Blitgen-Heinecke P, Kokkinou D, Bartz-Schmidt KU, Kochanek S, Schraermeyer U
- (2006) Tyrosinase biosynthesis in adult mammalian retinal pigment epithelial cells. Exp Eye Res 83(2): 315-321
Schraermeyer U, Kopitz J, Peters S, Henke-Fahle S, Blitgen-Heinecke P, Kokkinou D, Schwarz T, Bartz-Schmidt KU
- Phagocytosis of rod outer segments induces melanogenesis in ARPE-19 cells. ISOCB Cambridge, UK, 09.09.2006
Schraermeyer U, Biesemeier A, Kreppel F, Kochanek S
- Protection of human retinal pigment epithelial cells from UV-A irradiation by zinc is melanin dependent. ESPCR, Barcelona, Spain, 25.09.2006, Pigment Cell Res 19:5, 522, Abstract No. PP021
Biesemeier A, Kokkinou D, Schraeremyer U
- (2007) Melanin precursor 5,6-dihydroxyindol: protective effects and cytotoxicity on retinal cells in vitro and in vivo. Toxicol Pathol 35(7):1030-8
Heiduschka P, Blitgen-Heinecke P, Tura A, Kokkinou D, Julien S, Hofmeister S, Bartz-Schmidt KU, Schraermeyer U
- (2007) Tyrosinase biosynthesis and trafficking in adult human retinal pigment epithelial cells. Graefes Arch Clin Exp Ophthalmol 245(10): 1495-1505
Julien S, Kociok N, Kreppel F, Kopitz J, Kochanek S, Biesemeier A, Blitgen-Heinecke P, Heiduschka P, Schraermeyer U
- (2008) Comparison of visual function in pigmented and albino rats by electroretinography and visual evoked potentials. Graefes Arch Clin Exp Ophthalmol 246(11):1559-73
Heiduschka P, Schraermeyer U
- (2008) The classical premelanosome, known from melanocyte pigmentation, is not essential for RPE melanogenesis (IPCC 2008 and IMCR 2008, Sapporo, Japan, May 2008) Pigment Cell Res. 21:2, 302, Abstract No. PP4-6
Biesemeier A, Kreppel F, Kochanek S, Schraermeyer U
- (2008) UV-A induced oxidative stress is more prominent in naturally pigmented aged human RPE cells compared to non-pigmented human RPE cells independent of zinc treatment. J Photochem Photobiol B 90(2):113-20
Biesemeier A., Kokkinou D., Julien S., Heiduschka P., Berneburg M., Bartz-Schmidt K.U., Schraermeyer U
- Das klassische Prämelanosom - Organell der Pigmentsynthese im Melanozyten - ist für die Melanogenese des RPE nicht essentiell notwendig. Symposium des Netzwerks Elektronenmikroskopie Tübingen (NET), Tuebingen, 26.05.2008
Biesemeier A, Schraermeyer U
- (2009) Combined analytical TEM for chemical analysis of ocular melanosomes (ESPCR, Münster, Germany, September 2009), Pigment Cell Res. 22:5, 686, Abstract No. PP20
Schraermeyer U, Biesemeier A, Eibl O
- (2009) Tyrosinase and phagocytosis of retinal outer segments together influence the morphology and melanogenesis in cultured human ARPE19 cells. (ESPCR, Münster, Germany, September 2009), Pigment Cell Res. 22:5, 686, Abstract No. PP23
Biesemeier A, Blitgen-Heinecke P, Schraermeyer U
- Lokalisierung von Tyrosinase mit Hilfe analytischer Elektronenmikroskopie. Symposium des Netzwerks Elektronenmikroskopie Tübingen (NET), Tuebingen, 09. 11. 09
Biesemeier A, Schraermeyer U, Eibl O
