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Biomedical subjects

T Ruzicka

Publications and source records attributed to T Ruzicka.

At least 19 recordsLinked to original sources
PubMed1999

Cloning and characterization of hurpin (protease inhibitor 13): A new skin-specific, UV-repressible serine proteinase inhibitor of the ovalbumin serpin family.

Epidermal keratinocytes are the primary target of the midrange ultraviolet part (UVB, 280-320 nm) of terrestrial sunlight. Analysis of the resulting UV response at the transcriptional level by differential display PCR identified a formerly unrecognized large group of repressed genes. Among those UV-repressible genes, a novel serine proteinase inhibitor (serpin) termed hurpin (HaCaT UV-repressible serpin) has been identified. The isolated full-length cDNAs harbour a 1176 bp open reading frame encoding a potential protein with 391 amino acid residues and a predicted molecular mass of approximately 44 kDa. The novel serpin has nearly 59 % amino acid identity with the squamous cell carcinoma antigen 1 (SCCA1) and squamous cell carcinoma antigen 2 (SCCA2). In addition, it displays all of the structural features unique to the ovalbumin family of serpins (ov-serpins). The amino acid sequence of the hinge region in the reactive site loop suggests that hurpin has the potential for protease inhibition. The putative reactive center P1-P1'residues were identified as Thr356-Ser357 by alignment with other ov-serpins. The physiological target protease is unknown and the in vitro translated hurpin does not form SDS-stable complexes with a variety of known serine proteases. Expression of hurpin is restricted to epidermal cells where two distinct transcripts of 3.0 and 3.4 kb are detectable. Furthermore, expression of hurpin appears to be related to the activation or proliferation state of keratinocytes, since hurpin transcripts are more abundant in immortalized keratinocytes (HaCaT) and in cultured normal human keratinocytes, compared to the expression in normal skin. Moreover, in psoriasis, a skin disease characterized by hyperproliferation of keratinocytes and responsive to therapeutic UV irradiation, overexpression of hurpin is noted in psoriatic skin lesions compared to non-lesional skin.

Amino Acid Sequence↗
PubMed1999

Singlet oxygen mediates the UVA-induced generation of the photoaging-associated mitochondrial common deletion.

Mutations of mitochondrial (mt) DNA accumulate during normal aging. The most frequent mutation is a 4,977-base pair deletion also called the common deletion, which is increased in photoaged skin. Oxidative stress may play a major role in the generation of large scale mtDNA deletions, but direct proof for this has been elusive. We therefore assessed whether the common deletion can be generated in vitro through UV irradiation and whether reactive oxygen species are involved in this process. Normal human fibroblasts were repetitively exposed to sublethal doses of UVA radiation and assayed for the common deletion employing a semiquantitative polymerase chain reaction technique. There was a time/dose-dependent generation of the common deletion, attributable to the generation of singlet oxygen, since the common deletion was diminished when irradiating in the presence of singlet oxygen quenchers, but increased when enhancing singlet oxygen half-life by deuterium oxide. The induction of the common deletion by UVA irradiation was mimicked by treatment of unirradiated cells with singlet oxygen produced by the thermodecomposition of an endoperoxide. These studies provide evidence for the involvement of reactive oxygen species in the generation of aging-associated mtDNA lesions in human cells and indicate a previously unrecognized role of singlet oxygen in photoaging of human skin.

Cell Survival↗
PubMed1999

Tacrolimus: the drug for the turn of the millennium?

BACKGROUND: Tacrolimus has been shown to be a powerful suppressor of the immune system. It was introduced into clinical use to prevent allograft rejection and is now routinely used in kidney, liver, and heart transplantation. Recently, 2 double-blind multicenter studies demonstrated the therapeutic efficacy of topical and systemic tacrolimus in the inflammatory skin diseases atopic dermatitis and psoriasis. DATA SOURCE: MEDLINE was searched for relevant publications and combined with our own clinical, in vitro, and in vivo studies. STUDY SELECTION: All studies dealing with tacrolimus and dermatology were reviewed. DATA EXTRACTION: Publications with clinically relevant data were included in this review. CONCLUSIONS: Topical tacrolimus is a safe and effective therapeutic agent that may open a new era in the treatment of inflammatory skin diseases, particularly for patients with atopic dermatitis. Before its full potential in dermatology can be assessed, more clinical experience in treating children and comparison with the criterion standard of anti-inflammatory therapy, glucocorticosteroids, are needed.

Dermatitis, Atopic↗
PubMed1999

Fatty acid uptake by cultured human keratinocytes grown in medium deficient in or supplemented with essential fatty acids.

Epidermal linoleic acid, i.e. essential fatty acid (EFA), is essential for cutaneous barrier function. Cultured human keratinocytes, routinely used for studies of lipid metabolism, are grown in a keratinocyte serum-free medium (KSFM), under conditions that reveal EFA-deficient cells. Here, fatty acid (FA) uptake was analysed in human adult keratinocytes grown either under EFA-deficient conditions [KSFM supplemented with 10% FCS (A) or 1% UltroserG (B)] or EFA-supplemented conditions [KSFM supplemented with a devised FA cocktail (C) or evening primrose oil (D)]. The FA composition of the total cellular lipid and major lipid fractions was analysed by gas chromatography. Cells grown with supplements A or B balanced their EFA-deficient state primarily with oleic acid. Cells grown with supplements C or D normalized to the epidermal FA composition in vivo with raised linoleic and lower oleic acid contents. When cells were grown longer than 48 h with supplements C or D decreased cell growth was observed. FA uptake was curvilinear with preference for linoleic over oleic acid under all culture conditions. The uptake of linoleic acid by cells cultured with supplement B was twice the uptake of those cultured with supplement A, while the uptake of oleic acid was similar under both culture conditions. Oleic acid uptake of cells cultured with supplement C or D was lower. These results show that the uptake of linoleic, but not that of oleic acid, is influenced by the extracellular FA composition, and that EFA-supplemented keratinocytes compared to EFA-deficient cells might serve as an in vitro model for the study of EFA metabolism.

Adult↗
PubMed1999

Ultraviolet A1 (340-400 nm) phototherapy for cutaneous T-cell lymphoma.

BACKGROUND: The results of a recent study suggested that ultraviolet A1 radiation (UVA1R; 340-400 nm) phototherapy for atopic dermatitis works through induction of apoptosis in skin-infiltrating helper T cells, indicating the possibility that other helper T cell-mediated skin diseases may respond to UVA1R as well. OBJECTIVE: The purpose of this open pilot study was to assess the therapeutic effectiveness of UVA1 phototherapy for cutaneous T-cell lymphoma (CTCL). METHODS: UVA1 phototherapy was used as monotherapy in patients (n = 3) with histologically proven CTCL (stages IA and IB). For daily whole body UVA1 irradiations, either a high-dose (n = 2; 130 J/cm2 UVA1 per exposure) or medium-dose (n = 1; 60 J/cm2 UVA1) regimen was used. Therapeutic effectiveness was assessed clinically and histologically. RESULTS: In each of the 3 patients, skin lesions began to resolve after only a few UVA1 radiation exposures. Complete clearance was observed between 16 and 20 exposures, regardless of whether the high- or medium-dose regimen had been employed. CONCLUSION: These studies suggest that patients with CTCL stages IA and IB can be treated effectively with UVA1 phototherapy.

Aged↗
PubMed1999

Optimum porphyrin accumulation in epithelial skin tumours and psoriatic lesions after topical application of delta-aminolaevulinic acid.

Photodynamic therapy with topically applied delta-aminolaevulinic acid is used to treat skin tumours by employing endogenously formed porphyrins as photosensitizers. This study examines the time course of porphyrin metabolite formation after topical application of delta-aminolaevulinic acid. Porphyrin biosynthesis in human skin tumours (basal cell carcinoma, squamous cell carcinoma), in psoriatic lesions, and in normal skin was investigated. Skin areas were treated with delta-aminolaevulinic acid, and levels of total porphyrins, porphyrin metabolites and proteins were measured in samples excised after 1, 2, 4, 6, 9, 12 and 24 h. There was an increase in porphyrin biosynthesis in all tissues with maximum porphyrin levels in tumours between 2 and 6 h and in psoriatic lesions 6 h after treatment. The pattern of porphyrins showed no significant difference between normal and neoplastic skin, protoporphyrin being the predominant metabolite. The results suggest that optimum irradiation time for superficial epithelial skin tumours may be as soon as 2 h after application of delta-aminolaevulinic acid, whereas for treatment of psoriatic lesions an application time of 6 h is more suitable.

Administration, Topical↗
PubMed1999

Multiple basal cell carcinomas associated with hairy cell leukaemia.

We report the case of a caucasian woman who, between the ages of 49 and 51 years, developed multiple (> 20) basal cell carcinomas (BCC). There was no family history of BCC. No abnormalities in the human homologue of the Drosophila segment polarity gene patched (PTCH), glutathione S-transferases T1 and M1, or cytochrome P450 1A1 were detected by polymerase chain reaction (PCR)-based molecular analysis. There was, however, actinic damage of the skin in sun-exposed areas. The patient was diagnosed as having hairy cell leukaemia (HCL) at the age of 51 years, based upon leucocyte morphology as assessed by light and electron microscopy, tartrate-resistant acid leucocyte phosphatase (TRAP) staining, fluorescence activated cell scanning of peripheral blood leucocytes and bone marrow histology. As the leukaemia slowly progressed over a 3-month period, the patient developed four further BCCs. Given that HCL is characterized by a profound defect in T-cell function, it is conceivable that T-cell immune dysregulation can contribute to the pathogenesis of BCC, possibly enhancing the aetiological effect of ultraviolet irradiation.

Basal Cell Carcinoma↗
PubMed1999

The photoprotective effect of ascorbic acid, acetylsalicylic acid, and indomethacin evaluated by the photo hen's egg test.

The aim of the present study was to evaluate the supposed photoprotective effects of ascorbic acid, acetylsalicylic acid, and indomethacin by the photo hen's egg test, a recently developed new model for phototoxicity. Therefore, in three independent experimental settings the blood vessel system of the embryo's yolk sac of 24 incubated hens' eggs (2 test groups) were exposed to 60 mJ/cm2 ultraviolet B (UVB) to induce severe phototoxic damage. Before UVB irradiation, one of these test groups was exposed additionally to one of the test substances and the other one to 0.9% sodium solution alone. To exclude plain toxic reactions, two additional test groups were exposed only to 0.9% sodium chloride solution or to one of the test substances alone. Over a test observation period of 24 h, the embryo lethality as well as the morphological changes of the yolk sac blood vessel system were observed. Ascorbic acid led to a significant and remarkable reduction of the UVB-induced damage. Acetylsalicylic acid also showed a significant but lower photoprotective capacity. In contrast, indomethacin showed no photoprotective effects in the photo hen's egg test.

Animals↗
PubMed1998

Human eosinophils produce biologically active IL-12: implications for control of T cell responses.

The present study assessed the capacity of eosinophils (EOS) to synthesize the cytokine IL-12. Blood-derived, highly purified human EOS from six atopic patients and two nonatopic individuals were treated in culture with IL-4, IL-5, granulocyte-macrophage CSF, IFN-gamma, TNF-alpha, IL-1alpha, RANTES, and complement 5a, respectively. The expression of both IL-12 protein and mRNAs for the p35 and p40 IL-12 subunits was strongly induced in all donors by the Th2-like cytokines IL-4 and granulocyte-macrophage CSF and was also moderately induced by TNF-alpha and IL-1alpha. IL-5 treatment resulted in IL-12 synthesis in four atopic donors and one nonatopic donor, whereas IFN-gamma induced IL-12 synthesis in only two atopic donors. In contrast, RANTES exclusively induced mRNA for the p40 subunit without detectable protein release, and complement 5a had no effect on IL-12 mRNA or protein expression. EOS-derived IL-12 was biologically active, because supernatants derived from IL-4-treated EOS superinduced the Con A-induced expression of IFN-gamma by a human Th1-like T cell line. This activity was neutralized by anti-IL-12 Abs. In conclusion, EOS secrete biologically active IL-12 after treatment with selected cytokines, which mainly represent the Th2-like type. Consequently, EOS may promote a switch from Th2-like to Th1-like immune responses in atopic and parasitic diseases.

Cell Line↗
PubMed1998

Topical FK506 suppresses cytokine and costimulatory molecule expression in epidermal and local draining lymph node cells during primary skin immune responses.

Recently, it has been shown that the immunosuppressive macrolide lactone, FK506, exerts good therapeutic efficacy in inflammatory skin diseases. The aim of this study was to analyze the influence of topical FK506 on molecular (IL-1alpha, IL-1beta, IL-2, IL-4, IL-12 p35, IL-12 p40, macrophage inflammatory protein-2 (MIP-2), granulocyte-macrophage CSF (GM-CSF), TNF-alpha, and IFN-gamma) and cellular (I-A+/CD80+, I-A+/CD54+, I-A+/CD69+, I-A+/B220+, and CD4+/CD25+) events in epidermal (EC) and local draining lymph node (LNC) cells during primary contact hypersensitivity responses. Cytokine mRNA levels for IL-1alpha, IL-1beta, GM-CSF, TNF-alpha, MIP-2, and IFN-gamma in EC and for IL-2, IL-4, IL-12 p35, IL-12 p40, and IFN-gamma in LNC were increased and resulted in significant LNC proliferation during oxazolone-induced contact hypersensitivity. Topical FK506 treatment dose-dependently suppressed oxazolone-induced LNC proliferation. This effect was correlated with decreased IL-1alpha, IL-1beta, GM-CSF, TNF-alpha, MIP-2, and IFN-gamma mRNA expression within the epidermis and decreased IL-12 p35 and p40 mRNA expression in LNC. Further analysis of the LNC cytokine pattern revealed that the production of both Thl (IFN-gamma and IL-2) and Th2 (IL-4) cytokines was dramatically impaired after topical FK506 treatment. Flow cytometric analysis showed that topical FK506 decreased the population of epidermis-infiltrating CD4+ T cells and suppressed the expression of CD54 and CD80 on I-A+ EC and LNC during hapten-induced contact hypersensitivity. Furthermore, topical FK506 profoundly impaired oxazolone-induced up-regulation of CD25 expression on CD4+ LNC and dramatically decreased hapten-induced expansion of I-A+/B220+ and I-A+/CD69+ LNC subsets. In conclusion, these results give new insights into the mechanisms of action of topical FK506 treatment.

Administration, Topical↗
PubMed1998

Differential modulation of pro- and anti-inflammatory cytokine receptors by N-(4-trifluoromethylphenyl)-2-cyano-3-hydroxy-crotonic acid amide (A77 1726), the physiologically active metabolite of the novel immunomodulator leflunomide.

N-(trifluoromethylphenyl)-2-cyano-3-hydroxy-crotonic acid amide (A77 1726), the physiologically active metabolite of leflunomide, has been described to exert antiproliferative effects in vitro and anti-inflammatory actions in several animal models. Currently, its use is being evaluated in clinical trials in psoriasis, which is characterized by epidermal hyperproliferation and infiltration of inflammatory cells. We studied the effects of A77 1726 on growth and gene expression in cultured epidermal cells by 5-bromo-2'-deoxy-uridine (BrdU) incorporation, reverse transcriptase-polymerase chain reaction (RT-PCR), Northern blot hybridizations and flow cytometry. A77 1726 inhibited epidermal proliferation at concentrations above 5 microM after 24 hr. However, the cells were still fully viable at a concentration of 100 microM. The drug caused a dose-dependent reduction in the mRNA level of the type A receptor for the proinflammatory cytokine interleukin-8 (IL-8-RA) and, in contrast, induced gene expression of the receptor for the anti-inflammatory cytokine IL-10 (IL-10R) at the mRNA and protein levels. In addition, the mRNA and protein levels of the p53 gene, which is a negative cell cycle regulator, were up-regulated by A77 1726. These data suggest that A77 1726 exerts its anti-inflammatory action via the modulation of epidermal gene expression.

Aniline Compounds↗
PubMed1998

Missense mutations in SMOH in sporadic basal cell carcinomas of the skin and primitive neuroectodermal tumors of the central nervous system.

About one-third of sporadic basal cell carcinomas (BCCs) of the skin and 10-15% of primitive neuroectodermal tumors (PNETs) of the central nervous system show mutations in the PTCH tumor suppressor gene. The PTCH gene product (Ptch) functions as a transmembrane receptor for the Sonic hedgehog protein (Shh) and interacts with another transmembrane protein called Smoh. To further elucidate the significance of alterations in the Shh signaling pathway, we investigated 31 sporadic BCCs and 15 PNETs for the mutation and/or expression of SMOH, PTCH, SHH, and GL11. In addition, we fine-mapped the SMOH gene locus by fluorescence in situ hybridization to chromosomal band 7q32. Mutational analysis identified four BCCs with somatic missense mutations in SMOH affecting codon 535 (TGG==>TTG: Trp==>Leu) in three tumors and codon 199 (CGG==>TGG: Arg==>Trp) in one tumor. A missense mutation at codon 533 (AGC==>AAC: Ser==>Asn) was found in one PNET. PTCH mutations were detected in eight BCCs and one PNET. Two BCCs demonstrated mutations in both SMOH and PTCH. The majority of tumors showed an increased expression of SMOH, PTCH, and GL11 transcripts as compared with that of normal skin and nonneoplastic brain tissue, respectively. In contrast, only one BCC and one PNET expressed SHH mRNA at levels detectable by reverse transcription-PCR, and no SHH gene mutations were found. In summary, our results indicate that both PTCH and SMOH represent important targets for genetic alterations in sporadic BCCs and PNETs.

Adolescent↗
PubMed1998

Atopic eczema between rationality and irrationality.

A topic eczema is one of the most common chronic inflammatory skin diseases. Its treatment is a matter of intensive controversy between rational academic medicine and irrational alternative complementary medicine more than any other skin disease. Rational therapy is based on a sound pathophysiological foundation. The complex pathophysiology of the disease includes interactions between genetic predisposition and exogenous provocation factors, and treatment necessitates an integrated holistic approach. Despite excellent therapeutic results with the methods of conventional medicine, which uses mostly drugs of natural origin such as glucocorticosteroids, patients with atopic eczema and their parents frequently seek complementary treatments that use unproven and unvalidated diagnostic and therapeutic procedures. Treatments based on simplistic theories and irrational ideologies cause unnecessary long-term suffering to patients and are harmful because they withhold effective treatment modalities.

Combined Modality Therapy↗
PubMed1998

Photodynamic therapy in dermatology.

Photodynamic therapy (PDT) uses exogenously administered or endogenously formed photosensitizers activated by light to induce cell death via formation of singlet oxygen and other free radicals. Photodynamic therapy is increasingly used for the treatment of skin cancers and other indications. The efficacy of PDT depends on the structure of the photosensitizer, the administration modality, the light source, and the treatment procedure. We reviewed the most recent clinical and experimental developments in PDT research related to dermatology. The substrate under most intense investigation in PDT research is delta-aminolevulinic acid (ALA). Photodynamic therapy with topically applied ALA has been shown to be highly efficient in the treatment of cutaneous neoplasms by using intralesionally formed porphyrins as photosensitizers. For solar keratoses, best response rates have been described. delta-Aminolevulinic-PDT is also efficient in the treatment of superficial basal cell and squamous cell carcinomas. In addition, the fluorescence of ALA-induced porphyrins under a Wood light is highly selective in neoplastic cutaneous tissue and offers a useful technique in detecting and delineating skin tumors with ill-defined borders.

Administration, Cutaneous↗