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R Gniadecki

Publications and source records attributed to R Gniadecki.

At least 19 recordsLinked to original sources
PubMed2006

Minimizing adverse side-effects of oral bexarotene in cutaneous T-cell lymphoma: an expert opinion.

Bexarotene is an oral retinoid therapy that is effective for the treatment of early and advanced-stage cutaneous T-cell lymphoma (CTCL) in patients who have failed on other therapies. However, bexarotene treatment is associated with unavoidable side-effects, in particular hypertriglyceridaemia and hypothyroidism, which are manageable with adequate concomitant medications and are reversible on cessation of treatment. A pragmatic strategy for minimizing bexarotene-associated hypertriglyceridaemia and hypothyroidism is suggested, based on data from the studies with bexarotene in CTCL and on day-to-day experience with this agent in the clinical setting. The strategy anticipates that these common adverse events are likely to occur and recommends the early use of preventive therapy to lower triglycerides and elevate thyroid hormone levels in the blood, followed by subsequent monitoring, dose adjustment during bexarotene treatment, and titration of the daily bexarotene dose from 150 to 300 mg m(-2), which is optimal for most patients. When further information becomes available on how bexarotene interacts with lipid metabolism and thyroid function, the management approach suggested here may need to be changed.

Anticarcinogenic Agents↗
PubMed2006

Are desmoglein autoantibodies essential for the immunopathogenesis of pemphigus vulgaris, or just "witnesses of disease"?

Pemphigus vulgaris (PV) is fascinating to dermatologists, epithelial biologists and immunologists alike, as its pathogenesis has been clarified to a much greater extent than that of most other organ-specific autoimmune diseases, and as it has provided abundant novel insights into desmoglein biology and pathology along the way. Historically, the most influential PV pathogenesis concept is that of Stanley and Amagai. This concept holds that autoantibodies against desmogleins are both essential and sufficient for epidermal blister formation (acantholysis) by impeding the normal functioning of these major adhesion proteins. However, as with most good theories, this landmark concept has left a number of intriguing and important questions open (or at least has not managed to answer these to everyone's satisfaction). Moreover, selected dissenting voices in the literature have increasingly called attention to what may or may not be construed as inconsistencies in this dominant PV pathogenesis paradigm of the recent past. The present debate feature therefore bravely rises to the challenge of re-examining the entire currently available evidence, as rationally and as undogmatically as possible, by provocatively asking a carefully selected congregation of experts (who have never before jointly published on this controversial topic!) to discuss how essential anti-desmoglein autoantibodies really are in the immunopathogenesis of PV. Not surprisingly, some of our expert "witnesses" in this animated debate propose diametrically opposed answers to this question. While doing so, incisive additional questions are raised that relate to the central one posed, and our attention is called to facts that may deserve more careful consideration than they have received so far. Together with the intriguing (often still very speculative) complementary or alternative pathogenesis scenarios proposed in the following pages, this offers welcome "food for thought" as well as very specific suggestions for important future research directions--within and beyond the camp of PV aficionados. The editors trust that this attempt at a rational public debate of the full evidence that is currently at hand will constructively contribute to further dissecting the exciting--and clinically very relevant!--immunopathogenesis of PV in all its complexity.

Animals↗
PubMed2003

Intracellular calcium pool emptying induces DNA synthesis in HaCaT keratinocytes.

Calcium signaling provides a central control mechanism for growth, differentiation and apoptosis of epidermal keratinocytes. Moreover, calcium signaling is important for carcinogenesis in view of the observations suggesting that emptying of intracellular stores in keratinocytes [e.g. by a selective blocker of calcium pump in the endoplasmic reticulum (ER), thapsigargin] facilitates skin cancer development. In this work, we analyzed whether calcium content in the intracellular stores is linked to HaCaT keratinocyte growth and apoptosis control. Treatment with thapsigargin caused calcium release from the intracellular pool and permanent pool depletion (up to 24 h) could be achieved using a high dose (1 micro M) of this inhibitor. HaCaT cells cultured in these conditions exhibited an increased rate of DNA synthesis, assessed by the BrdU incorporation assay. Moreover, a weak stimulation of involucrin (terminal differentiation marker) was observed. Studies where intracellular free calcium (Cai2+) was chelated with BAPTA [1,2-bis(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid] revealed that abrogation of thapsigargin-induced Cai2+elevation did not counteract its effects on DNA synthesis, but blocked thapsigargin-induced involucrin expression. Apoptosis was readily achieved by extracellular calcium chelation using EGTA [ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid], but was not observed after thapsigargin or BAPTA alone or in combination. In conclusion, depletion of intracellular calcium stores causes stimulation of keratinocyte proliferation independently of the elevation of Cai2+.

Apoptosis↗
PubMed2003

Expression of T-cell activation marker CD134 (OX40) in lymphomatoid papulosis.

BACKGROUND: CD134/OX40 and CD30 are transmembrane proteins from the tumour necrosis factor receptor (TNFR) family present selectively on activated T cells. TNFR-related proteins are crucially involved in the regulation of proliferation and survival of normal and malignant lymphohaematopoietic cells. CD30 has been used for the immunophenotyping and subclassification of cutaneous lymphomas; virtually nothing is known, however, about the expression pattern of CD134 in lymphoid skin malignancies. OBJECTIVES: To determine CD134 expression in cutaneous lymphoma and benign inflammatory disorders. METHODS: Biopsy material was obtained from patients with lymphomatoid papulosis (LyP, n = 42), mycosis fungoides (n = 21), Jessner's infiltrates (n = 10) and non-specific dermatitis (n = 14). The expression of CD134 and CD30 was scored after immunohistochemical staining with appropriate monoclonal antibodies. The proportion of G2 + S phase cells was determined by laser scanning cytometry from nuclei obtained from paraffin-embedded biopsies. RESULTS: Few, single and scattered CD134+ cells (< 10%) were observed in the benign inflammatory infiltrations and in mycosis fungoides. A subset of 16 patients with LyP presented with clusters of CD30+ CD134+ cells. There was no correlation between the magnitude of CD134 expression and the histological type or the proportion of G2 + S cells in LyP. CD134 immunoreactivity was lower than expected in patients with LyP and another lymphoid malignancy (P < 0.001, Fisher's exact test). CONCLUSIONS: CD134 is strongly expressed in a proportion (38%) of patients with LyP, but not in mycosis fungoides or benign lymphocytic infiltrations. Loss of CD134 expression in LyP may be a marker of an increased risk of second lymphoid malignancy.

Adolescent↗
PubMed2002

Calcipotriol for erythema annulare centrifugum.

: Erythema annulare centrifugum (EAC) is an uncommon inflammatory skin disease of unknown aetiology. No therapy is currently available. We describe a 73-year-old woman with a 3-year history of EAC that was resistant to topical and systemic glucocorticoids, antifungals, and psoralen plus ultraviolet A treatment. After 3 months of treatment with topical calcipotriol the lesions cleared completely and did not recur during a 6-month follow-up period. Vitamin D analogues may be of value in the therapy of EAC.

Aged↗
PubMed2001

Changes in the ultrastructure of cytoskeleton and nuclear matrix during HaCaT keratinocyte differentiation.

The cellular scaffold that comprises nuclear matrix and cytoskeleton provides mechanical support for the cell and plays a crucial role in motility, cellular signaling, regulation of gene transcription and DNA replication. In this study we examined the structure of cytoskeleton and nuclear matrix in the keratinocyte cell line HaCaT using a recently developed technique, embedment-free electron microscopy. With this method the three-dimensional structure of cellular scaffold is visualized in the cells extracted from soluble proteins and the chromatin. In actively proliferating cells the cytoskeleton appeared to consist of a continuous meshwork of 10--15 nm filaments with a smaller amount of thin (5 nm) and ultrathin (1--2 nm) filaments. In contrast to what could be expected from earlier immunofluorescence and electron microscopy studies, the cytoskeleton in HaCaT keratinocytes did not consist of superposed autonomous networks of different filaments but was a highly integrated, continuous structure filling whole cytoplasmic territory. Moreover, cytoskeletons of adjacent cells were in a direct physical contact. Nuclear matrix consisted of globular ribonucleoprotein aggregates attached to the meshwork of 20--40 nm filaments. Nuclear envelope was firmly fastened to the cytoskeleton. In keratinocytes induced to differentiation by calcium switch both the cytoskeleton and nuclear matrix were drastically rearranged and comprised a monomorphic, dense and regular meshwork of 10--15 nm filaments. Our data underscore the fact that in HaCaT keratinocyte monolayer in vitro, and probably also in the epidermis in vivo, the nuclear matrices and the cytoskeletons of adjacent cells seemed to form a continuous, highly ordered structure which is rapidly rearranged during cell differentiation. This feature may be crucial for the understanding of how the signal initiated by, e.g. mechanical forces generated through the cell--cell and cell--matrix interaction is transmitted to the nucleus producing ultimately changes in the pattern of gene expression.

Calcium↗
PubMed2001

Differences in activation of G2/M checkpoint in keratinocytes after genotoxic stress induced by hydrogen peroxide and ultraviolet A radiation.

Long-wave ultraviolet radiation (UVA) may cause extensive DNA damage via reactive oxygen species (ROS). In this study we examined whether UVA- and H2O2-mediated DNA damage have equivalent effects on the induction of G2/M phase checkpoint and cell cycle progression in a transformed keratinocyte cell line HaCaT. By employing single cell gel electrophoresis (comet assay) we determined the equipotent doses of UVA and H2O2 with respect to the induction of alkali-labile sites (an indicator of oxidative DNA decay). However, in contrast to H2O2 which caused a pronounced G2/M cell cycle arrest 24 h after treatment, UVA irradiation did not affect cell cycle progression. Increasing UVA doses up to 150 kJ/m2 did not affect cell cycle and proliferation whereas increasing H2O2 concentrations caused a cell cycle block or cell death. Cytometric analysis revealed that G2/M cell cycle arrest took place beyond the cyclin B1 restriction point. We conclude that the DNA damage induced by UVA is easily repaired and does not perturb cell growth, whereas the H2O2-induced damage leads ultimately to cell cycle arrest or cell death.

Cell Division↗
PubMed2000

[Why skin cancer?].

Basal cell carcinoma, squamous cell carcinoma, and malignant melanoma are the most prevalent types of skin cancer. Recent developments in skin cancer research have made it possible to identify environmental factors involved in cutaneous carcinogenesis and to describe the molecular pathology of skin cancer. The alterations in the sonic hedgehog signalling pathway are responsible for the development of basal cell carcinoma, whereas deactivation of the p53 protein is an important step in the pathogenesis of squamous cell carcinoma. The pathogenesis of malignant melanoma remains to be elucidated. This article reviews the current understanding of the carcinogenic role of ultraviolet radiation and briefly summarises the importance of other possible aetiological factors, such as oxidative stress, oncoviruses, diet, or immune suppression.

Carcinogens↗
PubMed2000

Role of mitochondria in ultraviolet-induced oxidative stress.

The biological effects of ultraviolet radiation (UV), such as DNA damage, mutagenesis, cellular aging, and carcinogenesis, are in part mediated by reactive oxygen species (ROS). The major intracellular ROS intermediate is hydrogen peroxide, which is synthesized from superoxide anion ((*)O(2)(-)) and further metabolized into the highly reactive hydroxyl radical. In this study, we examined the involvement of mitochondria in the UV-induced H(2)O(2) accumulation in a keratinocyte cell line HaCaT. Respiratory chain blockers (cyanide-p-trifluoromethoxy-phenylhydrazone and oligomycin) and the complex II inhibitor (theonyltrifluoroacetone) prevented H(2)O(2) accumulation after UV. Antimycin A that inhibits electron flow from mitochondrial complex III to complex IV increased the UV-induced H(2)O(2) synthesis. The same effect was seen after incubation with rotenone, which blocks electron flow from NADH-reductase (complex I) to ubiquinone. UV irradiation did not affect mitochondrial transmembrane potential (DeltaPsi(m)). These data indicate that UV-induced ROS are produced at complex III via complex II (succinate-Q-reductase).

Cell Line↗
PubMed2000

Laser scanning cytometry for comet assay analysis.

BACKGROUND: The comet assay (single-cell gel electrophoresis) is a sensitive method for evaluating nuclear DNA damage. Previously used evaluation methods for the comet assay are time consuming and have an inherent risk of biased selection of comets due to manual selection and categorization of comet images. Laser scanning cytometry (LSC), the principle of which is equivalent to flow cytometry, enables quantification of fluorescence emitted from the cells on a microscope slide. In the present study, we explored whether LSC could be used to determine the degree of DNA damage demonstrated by the comet assay. METHODS: DNA damage was induced by ultraviolet A irradiation of keratinocytes and visualized by the comet assay. The evaluation included (a) LSC determination of DNA-specific fluorescence in 1,000 comet heads (undamaged DNA), (b) image acquisition of comets by rescanning of the microscope slide, and (c) digital image analysis and computation of tail moment and DNA content in the comet tails. RESULTS: Cells with damaged DNA were observed in a sub-G(1) area because the comet head loses DNA to the tail. We found a strong inverse correlation between tail moment and DNA content per nucleus. CONCLUSIONS: LSC enables an automated method for cell recognition and evaluation of the comets, thus providing quantitative information about nuclear DNA damage without subjective selection of analyzed comets.

Cells, Cultured↗
PubMed2000

Structural association of Bax with nuclear matrix and cytomatrix revealed by embedment-free immunogold electron microscopy.

Bax is a cellular protein functioning as a promoter of apoptosis. It is ultrastructurally associated with mitochondrial membranes, where it participates in permeability transition pore formation. By employing embedment-free electron microscopy (EFEM), we present evidence that Bax is also associated with the nuclear matrix and cytomatrix of cultured human tumour cells (COLO 205, PA-1, U-373 MG). Extracted cellular scaffolds were probed with anti-Bax antibody using the immunogold electron microscopy technique. Bax immunoreactivity was found on 10-15 nm intermediate filaments of karyo- and cytoskeleton, stretched between the nucleus, nuclear lamina and cell periphery. Bax immunoreactivity was preferentially localized to certain areas of filaments (spot-like). The target molecules for Bax binding in the cellular matrix and their physiological significance remain to be established.

Cytoplasm↗
PubMed2000

Hydrogen peroxide is responsible for UVA-induced DNA damage measured by alkaline comet assay in HaCaT keratinocytes.

We investigated the role of different reactive oxygen species (ROS) in ultraviolet A (UVA)-induced DNA damage in a human keratinocyte cell line, HaCaT. UVA irradiation increased the intracellular levels of hydrogen peroxide (H2O2), detected by a fluorescent probe carboxydichlorodihydrofluorescein, and caused oxidative DNA damage, single strand-breaks and alkali-labile sites, measured by alkaline single cell gel electrophoresis (comet assay). Superoxide anion (O2*-) was a likely substrate for H2O2 production since diethyldithiocarbamate (DDC), a superoxide dismutase blocker, decreased the level of intracellular H2O2. Hydrogen peroxide was shown to play a central role in DNA damage. Increasing the intracellular levels of H2O2 with aminotriazole (AT) (a catalase blocker) and buthionine sulfoximine (BSO) (an inhibitor of glutathione synthesis) potentiated the UVA-induced DNA damage. Exogenous H2O2 was also able to induce DNA damage. Since H2O2 alone is not able to damage DNA directly, we investigated the significance of the H2O2-derived hydroxyl radical (*OH). Addition of FeSO4, that stimulates *OH formation from H2O2 (Fenton reaction) resulted in a twofold increase of DNA-damage. Desferrioxamine, an iron chelator that blocks the Fenton reaction, prevented UVA-induced DNA damage. We also employed a panel of less specific antioxidants and enzyme modulators. Sodium selenite (Na-Se) present in glutathione peroxidase and thioredoxin reductase and addition of glutathione (GSH) prevented DNA-damage. Tocopherol potently prevented UVA-and H2O2-induced DNA damage and reduced intracellular H2O2 -levels. Ascorbic acid reduced H2O2 production, but only partly prevented DNA damage. Singlet oxygen (1O2) did not seem to play an important role in the UVA-induced DNA-damage since the specific 1O2 scavenger sodium azide (NaN3) and the less specific 1O2 scavenger beta-carotene did not markedly prevent either DNA-damage or H2O2 production. In conclusion the conversion of H2O2 to *OH appears to be the most important step in UVA-induced generation of strand breaks and alkali-labile sites and the bulk H2O2 appears to originate from O2*- generated by UVA irradiation.

Cell Line↗
PubMed2000

Structure of cytomatrix and nuclear matrix revealed by embedment-free electron microscopy.

Embedment-free electron microscopy (EFEM) is a new method which allows the visualisation of cytoskeleton in whole-mounted cells. In this study we employed EFEM to investigate the structure of cellular scaffolds in glioma C6 cell line. The cells were extracted with Triton X-100 that dissolves phospholipids in the membranes and removes most of cytoplasmic soluble proteins. The DNA and nuclear histones were removed with DNase I and high-salt buffer, respectively. The remaining cellular frameworks were temporary embedded in diethylene glycol distearate (DGD), sectioned and observed in transmission and scanning electron microscope after the removal of DGD. The predominant structure was the extensive meshwork of 10-20 nm filaments in the cytoplasm (cytomatrix) and 15-30 nm filaments in the nucleus (nuclear matrix). The 5 nm filaments, presumably corresponding to the actin filaments, were present in the cytomatrix, but not in the nuclear matrix. Moreover, the ultrathin (3 nm) filaments, connecting other cytoskeletal components were detected. Those are possibly identical with the previously described plectin filaments. For the first time we report the occurrence of ultrathin filaments in the nuclear matrix. Thus, in a addition to the well known cytoskeletal components (microtubules, intermediate filaments, actin microfilaments) EFEM showed a new type of filaments (the ultrathin filaments) in the cytomatrix and nuclear matrix. Further immunocytochemical studies are needed to determine the biochemical identity of the filaments observed in EFEM.

Animals↗
PubMed2000

Resistance of senescent keratinocytes to UV-induced apoptosis.

Irradiation with ultraviolet (UV) triggers programmed cell death (apoptosis) in keratinocytes. This process is believed to protect against skin carcinogenesis since the cells with damaged DNA are selectively removed, limiting the likelihood of the development of a malignant keratinocyte clone. The p53 protein is able to detect mutation-bearing DNA fragments and is thus indispensable for the UV-induced apoptosis in the epidermis. Since age is a risk factor for the development of skin tumors we investigated whether ultraviolet induces apoptosis and p53 activation in senescent keratinocytes. Cultured senescent keratinocytes were irradiated with broad-band ultraviolet, apoptosis was assessed using TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling) technique and the p53 activation pattern was determined with Western blotting and immunofluorescent staining with a panel of anti-p53 antibodies recognising different conformational forms of the protein (PAb 122, PAb 240, DO-7). In senescent keratinocytes arrested in the G1 phase of cell cycle, ultraviolet irradiation (100-2000 J/m2) caused accumulation and nuclear translocation of p53. However, in contrast to young cells where UV induces apoptotic cell death in G1, apoptosis was not detected in senescent cells. There were subtle differences in the p53 activation pattern between senescent keratinocytes and known patterns in young keratinocytes and other cell types. In senescent keratinocytes a constitutional nuclear expression of p53 (conformational form recognized by PAb 240) was present and the p53 induction in response to ultraviolet radiation was rapid. Suppression of apoptosis in senescent keratinocytes may be an important mechanism responsible for enhanced skin carcinogenesis in old age.

Antibodies, Monoclonal↗