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K U Schallreuter

Publications and source records attributed to K U Schallreuter.

At least 19 recordsLinked to original sources
PubMed2006

Melanocortins in human melanocytes.

Human epidermal keratinocytes and melanocytes express proopiomelanocortins (POMC) and all of the enzymes for POMC processing, i.e. prohormone convertases PC-1 and PC-2 including the regulatory protein 7B2. In melanocytes POMC processing also occurs in the melanosome, a lysosome-derived organelle that specializes in the biosynthesis of melanin. Consequently, the autocrine synthesis and release of the key hormones ACTH, alpha and beta-MSH and beta-endorphin takes also place in melanocytes. All four hormones have been reported to promote the biosynthesis of eumelanin in melanocytes. ACTH and alpha-MSH bind to the melanocortin-1 receptor (MC-1-R) on the plasma membrane and activate the signalling pathway predominantly coupled to production of cAMP, and in some cell lines raising intracellular calcium levels. In the melanocyte this signalling is redundant due to the high expression of alpha1 and beta2-adrenoceptors. Downstream events increase melanocyte this signalling is redundant due to the high expression of a tyrosinase expression / activity to stimulate eumelanogenesis. Studies with rMC-1-R transfected COS cells showed that both ACTH and alpha-MSH bind to the receptor with similar or different affinity depending on the species (human vs mice). We have modelled the MC-1-R based on the X-ray crystal structure of a homologous 7 receptor rhodopsin. Docking studies with ACTH1-39, ACTH1-17 and ACTH11-17 and alpha-MSH1-13 revealed that all 3 ACTH peptides yield thermodynamically stable (key ACTH1-13 in-lock) complexes. Interestingly, alpha-MSH is predicted to only have a kinetic effect on the MC-1-R and beta-MSH has even a weaker affinity for the MC-1-R than alpha-MSH. Based on these results the relative importance of ACTH versus alpha-MSH in the human epidermis has been re-evaluated.

Adrenocorticotropic Hormone↗
PubMed2006

Glutamate receptors on human melanocytes regulate the expression of MiTF.

Glutamate is the major excitatory neurotransmitter in the central nervous system but has also important functions in the epidermis. It is involved in keratinocyte barrier function and in re-epithelialization processes after wounding. Recently, glutamate signalling has been suggested to be implicated in the development of melanoma. The present study examined the expression and functionality of metabotropic and ionotropic glutamate receptors on normal human melanocytes. We found that cultured melanocytes expressed the ionotropic glutamate receptors GluR2 and 4 [alpha-amino-3-hydroxy-5-methyl-4-isoxsazolepropionic acid (AMPA) receptors] and N-methyl-d-aspartate (NMDA) receptors 2A and 2C and possibly the metabotropic glutamate receptor 1. Melanocytes were also found to express specific glutamate transporters and decarboxylases, but appeared neither to produce nor to release l-glutamate. Stimulation with 10 or 100 microM AMPA or NMDA elevated intracellular calcium concentrations in melanocytes, and thus demonstrated the functionality of the glutamate receptors. Millimolar concentrations of l-glutamate did not induce melanocyte toxicity and had no stimulating effect on melanin production. However, blockage of AMPA and NMDA receptors with CFM-2, memantine or MK801 caused a rapid and reversible change in melanocyte morphology, which was associated with disorganisation of actin and tubulin microfilaments. After 24 h of treatment with the AMPA receptor inhibitor CFM-2, there was a sharp reduction in the expression of the crucial melanocyte differentiation and proliferation factor MiTF. The results of this study demonstrate a role for glutamate in melanocyte regulation that may have implications in melanocyte associated disorders.

Apoptosis↗
PubMed2005

In vivo and in vitro evidence for autocrine DCoH/HNF-1alpha transcription of albumin in the human epidermis.

The presence of albumin in the human epidermis has been reported more than a decade ago, but until now, it was assumed that this protein is synthesized in the liver and transported to the avascular skin. To our knowledge, transcription of albumin in the human epidermis was never considered. In this report, we present for the first time evidence for autocrine synthesis of albumin in the human epidermis in keratinocytes in situ and in vitro. Using double immunofluorescence labelling, we identified that albumin colocalized together with its transcription factor PCD/DCoH/HNF-1alpha in suprabasal keratinocytes in human full-thickness skin sections and in keratinocytes cultured in serum-free medium. Moreover, albumin and HNF-1alpha protein expression was confirmed by Western blotting in undifferentiated and differentiated keratinocytes as well as in human epidermal suction blister roof extracts. Reverse-transcriptase polymerase chain reaction analysis from human epidermal keratinocytes and epidermal suction blister roofs revealed the transcription of albumin. Using in vivo fluorescence excitation spectroscopy at the surface of human skin, we confirmed albumin as a major constituent yielding a lambda(max) at 295 nm, which was assigned to the single tryptophan 214 fluorophore in this protein. This in vivo result is in agreement with albumin concentrations of 10(-3) M, underlining the importance of this protein in epidermal homeostasis.

Albumins↗
PubMed2003

Molecular evidence that halo in Sutton's naevus is not vitiligo.

Both halo naevus and vitiligo are acquired leucodermas of unknown aetiology. To date a significant contribution of oxidative stress through accumulation of hydrogen peroxide (H2O2) has been documented in the pathomechanism of vitiligo but not in halo naevus. Both epidermal pterin-4a-carbinolamine dehydratase (PCD) and catalase are sensitive markers to follow H2O2 concentration-dependent deactivation of these proteins. In situ protein expression of PCD and catalase was examined in full-skin biopsies from skin phototype-matched controls (n=5), untreated and treated vitiligo patients (n=5) and patients with untreated halo naevus in association with vitiligo (n=3). Vitiligo was treated with pseudocatalase (PC-KUS) only. Catalase levels were determined in epidermal suction blister extracts using fast protein liquid chromatography (FPLC). In addition, epidermal H2O2 levels were followed in vivo by Fourier-transform Raman spectroscopy. The results of this study ruled out a contribution of H2O2 in the millimolar range in the depigmentation process of halo naevus as previously documented in vitiligo. Therefore, it can be concluded that both leucodermas exercise distinct concentration-dependent H2O2 signalling in their pathomechanisms.

Catalase↗
PubMed2001

Thioredoxin reductase - its role in epidermal redox status.

The human epidermis with an area of 1.8 m(2) is the outer most layer of the human body. Hence, this organ plays a pivotal role in the defence against reactive oxygen species (ROS) generated by UV or X-ray exposure, heat and other sources. Consequently, a plethora of defence mechanisms exist controlling the redox status in this compartment. The role of thioredoxin reductase (TR), thioredoxin (T) in antioxidant defence has gained widespread recognition. In the past it has been shown that thioredoxin protects against UVB-induced skin injury, as well as against peroxidative damage. Under normal conditions, TR reduces oxidised thioredoxin in the presence of NADPH. Reduced thioredoxin serves as an electron donor for thioredoxin peroxidase (TPx) which consequently reduces H(2)O(2) to H(2)O. In this context, it has been demonstrated that membrane associated TR correlates with different skin photo types I-VI (Fitzpatrick classification), where darker skin has significantly higher enzyme activity compared to very fair skin, underlining the importance of this system in ROS defence. Moreover, it was only recently demonstrated in vivo with non-invasive Fourier-Transform Raman spectroscopy that UVB generates H(2)O(2) in the epidermis in a dose-dependent manner. H(2)O(2) can oxidise the selenocysteine residue in the penultimate position of the carboxyl terminus of TR with a K(m) of 2.5 mM. This oxidation is followed by an upregulation of mRNA expression of the enzyme. Hence, it can be concluded that UVB generated H(2)O(2) induces TR. However, permanent H(2)O(2) levels induce the tumour suppressor p53 which in turn downregulates cytosolic TR. Therefore TR activities are under fine control by H(2)O(2). This conclusion is also supported by the observation that thioredoxin, the substrate for TR, migrates from the cytosol to the nucleus after UVB exposure. A new function for the TR/T/TPx system in epidermal cells has been discovered in the control of the important cofactor (6R)-L-erythro 5,6,7,8 tetrahydrobiopterin (6BH(4)) homeostasis. Full oxidation of 6BH(4) to 6 biopterin via H(2)O(2) can lead to a cytotoxic environment for epidermal melanocytes. This cascade of events is observed in the depigmentation disorder vitiligo, where millimolar levels of H(2)O(2) can accumulate in the epidermis of affected individuals, consequently leading to cellular vacuolation in this compartment.

Animals↗
PubMed2001

Epidermal H(2)O(2) accumulation alters tetrahydrobiopterin (6BH4) recycling in vitiligo: identification of a general mechanism in regulation of all 6BH4-dependent processes?

It has been shown in vivo that patients with the depigmentation disorder vitiligo accumulate hydrogen peroxide (H(2)O(2)) accompanied by low catalase levels and high concentrations of 6- and 7-biopterin in their epidermis. Earlier it was demonstrated that epidermal 4a-OH-tetrahydrobiopterin dehydratase, an important enzyme in the recycling process of 6(R)-L-erythro 5,6,7,8 tetrahydrobiopterin (6BH(4)), has extremely low activities in these patients concomitant with a build-up of the abiogenic 7-isomer (7BH(4)), leading to competitive inhibition of epidermal phenylalanine hydroxylase. A topical substitution for the impaired epidermal catalase with a pseudocatalase effectively removes epidermal H(2)O(2), yielding a recovery of epidermal 4a-OH-tetrahydrobiopterin dehydratase activities and physiologic 7BH(4) levels in association with successful repigmentation demonstrating recovery of the 6BH(4) recycling process. Examination of recombinant enzyme activities, together with 4a-OH-tetrahydrobiopterin dehydratase expression in the epidermis of untreated patients, identifies H(2)O(2)-induced inactivation of this enzyme. These results are in agreement with analysis of genomic DNA from these patients yielding only wild-type sequences for 4a-OH-tetrahydrobiopterin dehydratase and therefore ruling out the previously suspected involvement of this gene. Furthermore, our data show for the first time direct H(2)O(2) inactivation of the important 6BH(4) recycling process. Based on this observation, we suggest that H(2)O(2) derived from various sources could be a general mechanism in the regulation of all 6BH(4)-dependent processes.

Biopterins↗
PubMed2000

Melanocytes are not absent in lesional skin of long duration vitiligo.

This paper provides evidence that melanocytes are still present in the depigmented epidermis of patients with vitiligo even after stable disease of 25 years' duration. Melanocyte cultures were successfully established from depigmented epidermal suction blister tissue of all 12 randomly selected patients and these cells produced melanin. Even under in vitro conditions, vacuolation of melanocytes was demonstrated in five patients with active disease, which was reversible upon exogenous addition of bovine catalase to the culture medium. Full skin biopsies from 17 patients with vitiligo, obtained from depigmented and normally pigmented areas, confirmed the involvement of melanocytes, keratinocytes, and Langerhans cells in this disorder. In addition, the presence of clustered and single pre-melanosomes in basal and supra-basal keratinocytes of lesional and normal epidermis, as well as the retention of single melanocytes in lesional epidermis, was demonstrated by light and electron microscopy. Upon topical application of a narrow band UVB-activated pseudocatalase, vacuolation, granulation, and dilatation of the endoplasmic reticulum completely recovered, but the ectopic pre-melanosome shedding remained. Taken together, these observations indicate that melanocytes are never completely absent in the depigmented epidermis and that these melanocytes can recover their functionality in vivo and in vitro upon the removal of hydrogen peroxide. Furthermore, this study supports the concept that vitiligo involves the entire epidermal unit in both depigmented and 'normal' pigmented skin.

Adolescent↗
PubMed2000

In vivo delayed-type hypersensitivity in 109 patients with vitiligo.

BACKGROUND: Although the etiology of the depigmentation disorder vitiligo is still not completely understood, many investigators believe that an autoimmune reaction may play a major role. In this regard, T-lymphocyte-mediated immunity has been implicated frequently in the pathogenesis of the disease. Most studies have applied in vitro testing of cell-mediated immunity, however, rather than in vivo measurements. Therefore, our study was undertaken to define the cutaneous delayed-type hypersensitivity (DTH) in vivo reaction in association with the absence/presence of serum thyroid autoantibodies, which are a good representative marker for autoimmunity in patients with vitiligo. METHODS: DTH was evaluated in the normal pigmented skin of 109 vitiligo patients (29 men and 80 women) and in the depigmented skin of 27 of this group (5 men and 22 women) using the dermal application of seven common recall antigens together with a negative control. Individuals were considered to be hypoergic if the DTH sum score was </= 5 mm in women or </= 10 mm in men, or if they responded to only one or two antigens. RESULTS: The mean sum score was 10.2 +/- 8.4 with an average of 2.3 +/- 1.6 positive reactions in depigmented skin vs. a sum score of 12.4 +/- 9. 0 with an average number of 2.6 +/- 1.6 positive reactions in normal pigmented skin. There was no statistically significant difference between depigmented and normal pigmented skin using the paired t-test (P > 0.05). Further evaluation of these data showed no significant correlation between the presence of thyroid autoantibodies as well as selected clinical parameters and an aberration in cutaneous DTH. CONCLUSIONS: In contrast to earlier reports, our in vivo studies of cutaneous DTH reactions revealed no clinically significant aberrant cellular immunity in this patient group. These results indicate that the immune reaction in vitiligo may be only a secondary event in the pathogenesis of the disease.

Adolescent↗
PubMed2000

Pro-opiomelanocortin-related peptides, prohormone convertases 1 and 2 and the regulatory peptide 7B2 are present in melanosomes of human melanocytes.

Recently, it has been shown that alpha-melanocyte stimulating hormone can directly activate tyrosinase by removing the allosteric regulator 6(R)-L-erythro 5,6,7,8 tetrahydrobiopterin resulting in a stable alpha-melanocyte stimulating hormone/6(R)-L-erythro 5,6,7,8 tetrahydrobiopterin complex. As melanin production occurs in the melanosome, a specific organelle of the melanocyte, it seemed important to investigate whether these organelles themselves actually produce pro-opiomelanocortin-related peptides in their acidic environment. The presence of alpha-melanocyte stimulating hormone and adrenocorticotropin in the epidermis and melanocytes has been shown by several investigators. In order to follow possible pro-opiomelanocortin processing in the melanosome, human melanocytes were established in MCDB 153 medium and utilized for immunohistochemistry, immunogold electron microscopy, and western blotting. For this purpose antibodies against alpha-melanocyte stimulating hormone, adrenocorticotropin, prohormone convertases 1 and 2 (PC1 and PC2) and the PC2 regulatory protein 7B2 were used. Our results demonstrated the presence of the entire system for pro-opiomelanocortin processing in the melanosome. Considering the pH optima of these convertases, the results are in agreement with an autocrine intramelanosomal production of pro- opiomelanocortin-related peptides and an autocrine production and recycling of the cofactor 6(R)-L- erythro 5,6,7,8 tetrahydrobiopterin in melanocytes. Based on these novel observations, we would like to propose that the pigmentation process may not necessarily involve a melanocortin-1 receptor-mediated mechanism.

Blotting, Western↗
PubMed1999

Evidence for specific complex formation between alpha-melanocyte stimulating hormone and 6(R)-L-erythro-5,6,7,8-tetrahydrobiopterin using near infrared Fourier transform Raman spectroscopy.

The cofactor 6(R)-L-erythro-5,6,7,8-tetrahydrobiopterin (6BH(4)) and its 2 and 4 electron oxidation products 7,8-dihydro-L-biopterin and L-biopterin have been shown to form 1:1 complexes with the thirteen amino acid peptide alpha-melanocyte stimulating hormone (alpha-MSH). Hydrogen bonding to the pyrimidine ring of the cofactor has been established for glu(5) and his(6) of the hormone using Near Infrared Fourier Transform Raman spectroscopy. Binding of these pterins primarily involves the pyrimidine ring, although with the reduced pterins, 7,8-dihydro-L-biopterin and 6BH(4), there is evidence for pi orbital interaction with the pyrazine ring. It is proposed that this pi orbital interaction with the reduced biopterins and alpha-MSH could provide the basis for the observed stability of these pterins to oxidation by either molecular oxygen or photooxidation by UVB (290-320 nm) light. Our results suggest that the formation of the alpha-MSH/6BH(4) complex could play a major role in the control of all 6BH(4) dependent processes.

Biopterins↗
PubMed1999

alpha-MSH can control the essential cofactor 6-tetrahydrobiopterin in melanogenesis.

In the human epidermis both keratinocytes and melanocytes express POMC m-RNA. Immunohistochemical studies of both cell types demonstrate significantly higher levels of alpha-MSH in melanocytes than in keratinocytes. Both cell types also hold the full capacity for de novo synthesis/recycling of the essential cofactor (6R)-L-erythro-5,6,7,8-tetrahydrobiopterin (6BH4). 6BH4 is critical for the hydroxylation of the aromatic amino acids L-phenylalanine, L-tyrosine, and L-tryptophan, for nitric oxide production and in various immune modulatory processes. Recently it was shown that tyrosinase activity is regulated by 6BH4 through a specific allosteric inhibition. The tyrosinase/6BH4 inhibition can be activated by 1:1 complex formation between 6BH4 and alpha-MSH, but an excess of alpha-MSH over 6BH4 can inhibit tyrosinase due to complex formation by tyr2 in the alpha-MSH sequence. In both melanocytes and keratinocytes 6BH4 controls the L-tyrosine supply via phenylalanine hydroxylase (PAH). Recently we were able to show that the cellular uptake of L-phenylalanine and its intracellular turnover to L-tyrosine is crucial for melanogenesis. alpha-MSH can promote the production of L-tyrosine via PAH due to activation of the PAH tetramer to the more active dimer by removing 6BH4 from the regulatory binding domain on the enzyme. In conclusion, alpha-MSH can control (1) intracellular L-tyrosine formation from L-phenylalanine in both melanocytes and keratinocytes, and (2) tyrosinase activity, directly, in melanocytes.

Animals↗
PubMed1999

The importance of L-phenylalanine transport and its autocrine turnover to L-tyrosine for melanogenesis in human epidermal melanocytes.

A comparative study of (14)C-labeled l-phenylalanine and (3)H-labeled L-tyrosine uptake in cultures of human melanocytes (n = 8) and keratinocytes (n = 2) identified a significantly more rapid active transport for L-phenylalanine in melanocytes, whereas the slower uptake of L-tyrosine followed a concentration-dependent gradient, confirming facilitated diffusion rather than active transport. In addition, a significantly more efficient autocrine turnover of L-phenylalanine to L-tyrosine via intracellular phenylalanine hydroxylase was demonstrated in melanocytes. The incorporation of the newly synthesized radiolabeled L-tyrosine was directly followed into the end product melanin. The presence of L-phenylalanine in the culture medium produced 40% more melanin compared to an equivalent concentration of L-tyrosine alone. The transport of extracellular L-phenylalanine and its intracellular metabolism to L-tyrosine via intracellular phenylalanine hydroxylase are coupled to calcium uptake/efflux, whereas L-tyrosine uptake is calcium independent. Taken together, our results identified for the first time the importance of autocrine calcium-dependent active l-phenylalanine uptake/turnover in melanocytes as a major pathway for melanogenesis.

Autocrine Communication↗
PubMed1999

In vivo and in vitro evidence for hydrogen peroxide (H2O2) accumulation in the epidermis of patients with vitiligo and its successful removal by a UVB-activated pseudocatalase.

To date there is compelling in vitro and in vivo evidence for epidermal H2O2 accumulation in vitiligo. This paper reviews the literature and presents new data on oxidative stress in the epidermal compartment of this disorder. Elevated H2O2 levels can be demonstrated in vivo in patients compared with healthy controls by utilizing Fourier-Transform Raman spectroscopy. H2O2 accumulation is associated with low epidermal catalase levels. So far, four potential sources for epidermal H2O2 generation in vitiligo have been identified: (i) perturbed (6R)-L-erythro 5,6,7,8 tetrahydrobiopterin (6BH4) de novo synthesis/recycling/regulation; (ii) impaired catecholamine synthesis with increased monoamine oxidase A activities; (iii) low glutathione peroxidase activities; and (iv) "oxygen burst" via NADPH oxidase from a cellular infiltrate. H2O2 overproduction can cause inactivation of catalase as well as vacuolation in epidermal melanocytes and keratinocytes. Vacuolation was also observed in vitro in melanocytes established from lesional and nonlesional epidermis of patients (n = 10) but was reversible upon addition of catalase. H2O2 can directly oxidize 6BH4 to 6-biopterin, which is cytotoxic to melanocytes in vitro. Therefore, we substituted the impaired catalase with a "pseudocatalase". Pseudocatalase is a bis-manganese III-EDTA-(HCO3-)2 complex activated by UVB or natural sun. This complex has been used in a pilot study on 33 patients, showing remarkable repigmentation even in long lasting disease. Currently this approach is under worldwide clinical investigation in an open trial. In conclusion, there are several lines of evidence that the entire epidermis of patients with vitiligo is involved in the disease process and that correction of the epidermal redox status is mandatory for repigmentation.

Animals↗
PubMed1999

Serum selenium levels and blood glutathione peroxidase activities in vitiligo.

It has recently been shown that patients with vitiligo can accumulate epidermal hydrogen peroxide (H2O2) in association with low catalase levels. This study examined serum selenium levels and blood glutathione peroxidase activities in 61 patients and controls. The results showed high serum selenium levels in 56% of the patients. As at least one isoform of glutathione peroxidase requires selenium for its activity, enzyme activities were also evaluated. The overall results were not significantly different compared with controls, but further age-related analysis of the data indicated significantly lower activities in patients up to 46 years. As glutathione peroxidase can also efficiently degrade H2O2, the results of this study could indicate an additional impaired H2O2 metabolism in vitiligo.

Adolescent↗