Biomedical subjects

M Detmar

Publications and source records attributed to M Detmar.

70 records · Page 4Linked to original sources

PubMed1990

Effects of recombinant tumor necrosis factor-alpha on cultured microvascular endothelial cells derived from human dermis.

We investigated the effects of recombinant human tumor necrosis factor-alpha (TNF) on cell proliferation and on expression of MHC class II antigens and intercellular adhesion molecule ICAM-1 in human dermal microvascular endothelial cells (HDMEC) derived from human foreskin. Second-passage HDMEC were treated with 0.1-10,000 U/ml TNF for up to 6 d, and cell growth was assessed by cell counts and a recently developed fluorogenic assay using 4-methylumbelliferyl heptanoate as a substrate. APAAP immunocytochemistry was performed using monoclonal antibodies against HLA-DR, HLA-DQ, and ICAM-1. TNF did not markedly inhibit the growth of HDMEC but induced expression of HLA-DR (1,000 U/ml and more) and of ICAM-1 (1 U/ml and more). Combination with interferon-gamma led to synergistic ICAM-1 induction. These results demonstrate a profound effect of TNF on the activation of dermal microvascular endothelial cells and suggest a major role of TNF in the mediation of leucocyte adhesion to endothelial cells of the skin microvasculature with possible implications for the initiation and maintenance of inflammatory skin processes.

Cell Adhesion Molecules↗

PubMed1990

Initial hyperproliferation and incomplete terminal differentiation of cultured human keratinocytes from lesional and uninvolved psoriatic skin.

Human epidermal keratinocytes (KCs) were isolated from lesional and from uninvolved skin of 8 patients with chronic plaque-like psoriasis and from the normal skin of 8 healthy volunteers. Primary KC cultures, grown on 3T3 cell feeder layers, were examined over a period of 4 weeks and their plating efficiency, colony growth area, DNA synthesis and ultrastructural cell differentiation were evaluated. Psoriatic KCs formed colonies one day earlier than non-psoriatic controls and proliferated faster during the first 2 weeks, as assessed by the mean colony growth area and 3H-thymidine incorporation. After 4 weeks, however, no significant differences were observed between the in vitro proliferation parameters of normal and psoriatic KCs. At the ultrastructural level, cultures of lesional psoriatic KCs consisted of more cell layers with adherent transitional cells and incomplete formation of cornified envelopes, even after 4 weeks, while KCs from uninvolved psoriatic skin were characterized by a transient delay of in vitro maturation. These results indicate that the characteristic hyperproliferation of psoriatic KCs may only be maintained over a short period of primary culture, whereas defective terminal differentiation of lesional psoriatic KCs was maintained throughout the culture period.

Biopsy↗

PubMed1990

[Preliminary findings on lectin binding in human follicular epithelium].

Glycoconjugates in the cell membranes of the outer and the inner root sheath of human anagen hair follicles were histochemically characterized by means of the following lectins: UEA-I, DBA, PNA, WGA, SBA, RCA, and Con A (avidin-biotin technique, counter-staining with methyl green). We observed intense labeling of the outer root sheath with UEA-I, DBA, Con A, and PNA, whereas the other markers were only weakly positive. VEA-I, DBA and SBA labeling was confiried to the upper two thirds of the outer root sheath. As to the inner root sheath, we found intense labeling with UEA-I, WGA, RCA, and SBA; all other lectins were only weakly represented. Our results suggest that the cell membranes of the outer and inner root sheaths of human hair follicles are characterized by the prevalence of different glycoconjugates; moreover, labeling with UEA-I, SBA, and DBA is obviously related to the process of differentiation within the follicle.

Cell Differentiation↗

PubMed1990

[Kyrle disease in juvenile diabetes mellitus and chronic renal failure].

We report on a 32-year-old female patient with chronic diabetes mellitus, type I, and chronic renal failure, who developed the typical clinical picture of hyperkeratosis follicularis et parafollicularis in cuteum penetrans (Kyrle's disease) within one year. Histological examination revealed a defective epidermal differentiation with hyper- and parakeratosis as well as premature keratinization as early as in the epidermal basal cell layer. Studies on lectin binding showed that the glycosylation process was impaired in both the epidermis and the basement membrane zone of the lesional skin. In addition, electron microscopic investigation revealed diabetic microangiopathy of the dermal vessels as well as marked ultrastructural alterations of the dermo-epidermal basal lamina. These findings confirm the association of diabetes mellitus with Kyrle's disease previously described; they make us suggest that Kyrle's disease might be characterized by a defective differentiation of the epidermis and the dermo-epidermal junction--due to some alteration of the underlying glycosylation processes--rather than by a local disorder of keratinization. Regarding the clinical manifestation of the disease, both diabetes mellitus and chronic renal failure may play a part as precipitating factors.

Adult↗

PubMed1989

Effects of azelaic acid on proliferation and ultrastructure of mouse keratinocytes in vitro.

The effects of azelaic acid (C9-dicarboxylic acid, AZA) on the proliferation and ultrastructure of neonatal NMRI mouse keratinocyte cultures were studied to clarify the mechanisms of AZA action on normal and diseased epidermis. The dose- and time-dependency of the drug effects on DNA synthesis was examined by 3H-thymidine incorporation into DNA and by autoradiography. Electron microscopy was used to detect the target cell organelles of the drug. Azelaic acid decreased DNA synthesis in a dose- and time-dependent manner with a 50% inhibitory concentration of 20 mM. The inhibition of DNA synthesis was already observed after 1 h of treatment, reached its maximum after 4 h, and was stable for 24 h. A complete reversibility of the inhibitory effects was observed within 2 h after discontinuation of the treatment, and, interestingly, a rebound effect occurred with a temporary increase of DNA synthesis. Furthermore, treatment with AZA reduced the RNA and protein synthesis of the cells. Electron microscopic evaluation of treated cultures showed early marked damage of the mitochondria, followed by dilation of the rough endoplasmic reticulum (RER). These alterations were completely reversible after discontinuation of the treatment. Our findings show that AZA exerts a dose- and time-dependent, reversible antiproliferative effect on keratinocytes, acting primarily on mitochondria and RER. The antiproliferative action of AZA could explain its beneficial effect in some skin disorders characterized by alteration of keratinocytic differentiation.

Animals↗

PubMed1989

A rapid fluorometric assay for the determination of keratinocyte proliferation in vitro.

In the present study we describe a simple technique for the determination of keratinocyte proliferation in vitro, based on the hydrolysis of a fluorogenic substrate by cell esterases. Normal and transformed human keratinocytes were grown in microtiter plates and were incubated with 4-methylumbelliferyl heptanoate after 3, 5, and 7 days. The fluorescence was quantified using an automatic fluorescence detection unit. The fluorescence showed a strong correlation with the cell number at various growth phases. In addition, the method reliably detected the growth inhibitory effect of recombinant interferon gamma on human keratinocytes. The fluorometric assay is a simple, fast and reliable method to assess cell number in keratinocyte cultures.

Cell Count↗

PubMed1989

Effects of azelaic acid on sebaceous gland, sebum excretion rate and keratinization pattern in human skin. An in vivo and in vitro study.

The effects of azelaic acid (AZA) on the epidermis of 47 individuals (12 with normal skin, 15 with seborrheic skin and 20 suffering from acne) and on in vitro cultured keratinocytes are reported. Topical application of a 20% AZA cream significantly improved the lesions of acne patients, but failed to induce clinically detectable changes in normal or seborrheic epidermis. Complementary investigations clearly showed that AZA treatment failed to induce specific changes in sebum composition, excretion rate, or in the size of sebaceous glands, but modified epidermal keratinization. Keratohyalin granules and tonofilament bundles were reduced in size and number, mitochondria were swollen and the rough endoplasmic reticulum of malpighian keratinocytes enlarged. The infundibular epidermis of acne individuals showed marked reduction of the horny layer thickness, widening of the horny cell cytoplasm, transitional corneal cells, normalization of filaggrin distribution, and the comedo contained few bacteria and spores. In vitro, AZA exerted marked time- and dose-dependent antiproliferative cytostatic effects on cultured keratinocytes, with a 50% inhibitory dose of 20 mM, decreased some keratinocyte proteins (highly soluble fractions S2, keratohyalin macroaggregate R2, and non-cross-linked fibrous protein S4) and a 95 kD and a 35 kD protein of the cytosolic fraction. Mitochondria were frequently damaged and the rough endoplasmic reticulum enlarged. Our results indicate that AZA is an antikeratinizing agent, displaying antiproliferative cytostatic effects on keratinocytes and modulating the early and terminal phases of epidermal differentiation.

Acne Vulgaris↗

PubMed1989

Isolation of human sebaceous glands and cultivation of sebaceous gland-derived cells as an in vitro model.

An experimental technique is presented as an in vitro model for the study of human sebaceous gland-derived cells. Intact sebaceous glands were isolated from full-thickness human skin after incubation in dispase (2.4 U/ml) and in deoxyribonuclease (0.02%) by using microsurgical instruments under microscopical observation of the epidermal underface. Subsequently, the ducts of the glands were removed, the isolated gland lobules were seeded on a 3T3-cell feeder layer in Dulbecco's modified Eagle's medium and Ham's F 12 medium (3:1) supplemented with fetal calf serum (10%), L-glutamine, antibiotics, epidermal growth factor (10 ng/ml), hydrocortisone (0.4 microgram/ml), and cholera toxin (10(-9) M), and were then cultivated in a CO2-incubator at 37 degrees C. After 2-3 wk cell outgrowths resulting from the periphery of the gland lobules were obtained and dispersed cells were passaged for three subcultures with or without 3T3-cell feeder layer. The cultured cells preserved in vitro morphologic characteristics and differentiation patterns comparable to those described for normal human sebocytes in vivo, with a high rate of viable cells. Their labeling pattern with MoAb showed close similarities to the pattern reported for sebocytes in vivo but differences to the pattern of keratinocytes in vivo and in vitro. In their cytoplasm oil red and nile red stained droplets were detected, and the observed density and distribution evidenced in vitro lipogenesis. The technique presented here may provide a promising model for further experimental studies on sebaceous gland cell development and function.

Antibodies, Monoclonal↗

PubMed1988

Blood flow and metabolism in heterotopic cerebellar grafts during hypoglycemia.

Hypoglycemia-induced disturbances of brain metabolism and neuronal injury exhibit a distinct predilection for forebrain structures, in particular the caudate-putamen, hippocampus and cerebral cortex, whereas the cerebellum is remarkably resistant. In an attempt to assess the biological basis of this differential regional vulnerability, we have used a neural transplantation technique to compare hemodynamic and metabolic changes in cerebellum during severe hypoglycemia with those in heterotopic cerebellar grafts. To this end, the cerebellar anlage of fetal rat brain (day 15 of gestation) was stereotactically transplanted into the vulnerable caudate-putamen. Following a differentiation period of 8 weeks the grafts had developed into an organotypic population of mature cells with laminar histoarchitecture. Host animals were then subjected to insulin-induced hypoglycemia. After 15 min of isoelectric EEG, blood flow was increased throughout the brain but residual glucose consumption was significantly higher in cerebellum (0.29 mumol/g per min) and cerebellar grafts (0.22 mumol/g per min) as a result of increased glucose extraction. Hypoglycemia caused a depletion of ATP in all brain structures except cerebellum where normal levels were maintained. Correlation of local ATP content and glucose utilization revealed a threshold-like decline of ATP at a glucose utilization rate of 0.27 mumol/g per min. ATP, in consequence, was normal in cerebellum but partially depleted in cerebellar grafts. It is concluded that the resistance of cerebellum to hypoglycemia is due to its capacity for higher glucose extraction at low blood glucose levels, and that this unique intrinsic property is preserved after heterotopic transplantation.

Adenosine Triphosphate↗

PubMed1987

Nucleolysis and diskography--an experimental study on interaction of chymopapain and contrast medium.

A test was carried out on 11 young, full sized mongrels to determine whether there is an interaction between chymopapain and the contrast agents iotrolan and iopamidol. A total of 75 intervertebral disks were punctured: nucleolysis alone was performed on 20, diskography with iotrolan and subsequent nucleolysis on 20, and diskography with iopamidol and subsequent nucleolysis on 10. Diskography alone was performed 10 times with iotrolan and 5 times with iopamidol. Aqua dest. was given intradiskally 5 times, and puncture was carried out 5 times without the administration of any substance. Following puncture, x-rays of the lumbar vertebral column were taken laterally: daily for the first 10 days, then weekly. Disk space narrowing typical of nucleolysis with chymopapain was found among the disks that were nucleolyzed only to the same extent as among those that had undergone diskography previously. There was no evidence of narrowing of the other disk spaces which had been punctured but not treated with chymopapain. On some of the dogs, CT and MRI examinations were carried out. The CTs showed a homogenous hypodensity in all of the disks, in which chymopapain had been injected. The MRI revealed a signal loss in all of the nucleolyzed disks. The results of short and long term follow up demonstrate that inhibition of chymopapain by iotrolan or iopamidol is not to be expected and therefore diskography prior to chemonucleolysis can be performed without danger of enzyme inactivation.

Animals↗

PubMed1987

Growth of human hair follicle keratinocytes in vitro. Ultrastructural features of a new model.

A simple experimental technique was developed to provide an in vitro model for the study of human follicular keratinocytes. Anagen-phase human hairs were plucked from the scalp of healthy individuals; the follicles were separated, plated on coverslips coated with collagen G, and cultivated in McCoy 5A Medium in a CO2-incubator at 37 degrees C. Light and electron microscopy after 1, 2, 3, and 6 weeks showed selective and progressive cell growth with keratinocyte differentiation, producing multilayered cultures of cells joined with fully developed desmosomes. Three distinct patterns of differentiation, leading to the formation of an incomplete horny layer, were seen. The particular arrangement of tonofilaments, the considerable amounts of cytoplasmic glycogen, and the absence of malpighian differentiation were ultrastructural indicators of the follicular origin of the cultured cell population, which most likely grew from the outer root sheath of the hair. This technique may provide a promising model on which to base further studies of hair biologic processes and hair growth.

Cell Differentiation↗

PubMed1987

Retinoids and keratinocyte differentiation in vitro.

Standard retinoids (e.g. etretinate, isotretinoin) are successful in the treatment of a variety of dermatological disorders. However, they are handicapped by an unfavorable ratio between activity and toxicity. In order to find more successful substances new promising compounds were recently synthesized. The aim of this study was to screen new monoaromates (etretin (Ro 10-1670) ), demethyletretin (Ro 12-7310), 13-cis-etretin (Ro 13-7652) and the polyaromates arotinoid (Ro 15-0778), arotinoid acid (Ro 13-7410) and arotinoid ethyl sulfone (Ro 15-1570) for their effect on keratinocyte differentiation in vitro and to compare these results with differentiation data of retinoids currently used clinically. It was found that the second generation of retinoids had a lower antikeratinizing potential than the third generation of retinoids. They markedly inhibited crosslinked keratins and envelope proteins. Arotinoid acid was found to be the most potent derivative. In contrast to this, differences with regard to the synthesis of keratohyalingranule macroaggregates were detected between the retinoid derivatives. Arotinoid acid and arotinoid sulfone stimulated this protein fraction, whereas arotinoid left this fraction uninfluenced. If the data obtained were considered the selection criterion for the future therapeutic application of the new arotinoids in psoriasis therapy, it should be more useful to employ arotinoid acid or arotinoid sulfone rather than arotinoid in clinical psoriasis studies.

Animals↗

PubMed1987

[Stability of cignolin (dithranol) in ointments containing tar with and without the addition of salicylic acid. Oxidation to danthron and dithranol dimer].

The stability of dithranol in white petrolatum was investigated over a period of 10 weeks in dithranol ointments containing liquor carbonis detergens (LCD) and/or coal tar, in the presence and absence of salicylic acid. Qualitative and semiquantitative analysis using high-pressure liquid chromatography (HPLC) showed that dithranol (0.1%, 0.5%, 1.0%) remained rather stable in white petrolatum for 10 weeks, regardless of the presence or absence of salicylic acid. The addition of 5% LCD or 5% crude coal tar resulted, however, in the rather rapid oxidation of dithranol into its products, danthron and dithranol dimer. The addition of 2% salicylic acid to these combined ointments did not stabilize the dithranol: rather, it reduced the amount of danthron but resulted in a considerable increase in the amount of dithranol dimer. The oxidation of dithranol into its products danthron and dithranol-dimer was almost complete in 0.1% ointments and ranged between 16.3% and 53.8% in 0.5% white petrolatum after 10 weeks. These findings may explain why the addition of tar minimizes dithranol-induced erythema, as reported in clinical studies. Such combinations may have an important influence on the antipsoriatic activity of dithranol and the appearance of brownish discoloration.

Anthralin↗

PubMed1986

Degeneration of granule cells following chronic phenytoin administration: an electron microscopic investigation of the mouse cerebellum.

Fifteen male mice (C57/Bl6J) were fed the liquid diet "Stardit" supplemented with vitamins together with phenytoin for 8 weeks; experimental animals and controls were pair-fed. After 8 weeks of treatment, the anesthetized animals were perfused with 3.5% glutaraldehyde. Tissue samples of the cerebral cortex (area 3), cerebellum (vermis), thalamus, hypothalamus, and liver were embedded in Araldite. All phenytoin-treated animals displayed a hepatomegaly. Semithin sections and ultrastructural investigations of the cerebellar vermis showed pyknoses of granule cells and an enlargement and swelling of parallel fibers in presynaptic areas in the molecular layer. The swollen axons showed an accumulation of tubular structures which represented proliferated smooth endoplasmic reticulum. Similar tubular structures were observed in hepatocytes of experimental animals. It is proposed that phenytoin caused an induction of the microsomal system of hepatocytes and granule cells which led to a proliferation of the smooth endoplasmic reticulum. The transport of these organelles to the axon terminals of parallel fibers via the axoplasmic flow is assumed to cause a swelling of the presynaptic area. A dying-back process may then lead to pyknosis of granule cells. Chronic phenytoin administration to mice is a new experimental model of neuroaxonal dystrophy.

Animals↗

PubMed1986

[Dicarboxylic acids inhibit the growth of keratinocytes in vitro].

The antiproliferative effect of three straight-chained saturated dicarboxylic acids was examined with neonatal mouse keratinocyte cultures. Adipic acid (C6), azelaic acid (C9), and sebacic acid (C10) were added to the cultures in concentrations ranging from 1 to 50 mmol/l. Proliferation was assayed by liquid-scintillation counting of 3H-thymidine incorporation into DNA and by autoradiography. Fifty percent inhibition of 3H-thymidine incorporation was observed with 50 mmol/l adipic acid, 20 mmol/l azelaic acid, and 10 mmol/l sebacic acid, respectively. The antiproliferative effect was completely reversible after cessation of treatment. Moreover, treated cultures then showed a rebound effect with increased DNA synthesis. These results show that dicarboxylic acids exert reversible antiproliferative effects on keratinocytes.

Adipates↗

PubMedDate not supplied

Vascular permeability factor/vascular endothelial growth factor: an important mediator of angiogenesis in malignancy and inflammation.

Vascular permeability factor (VPF), also known as vascular endothelial growth factor (VEGF), is a multifunctional cytokine that is overexpressed in many transplantable animal and autochtonous human cancers, in healing wounds, and in chronic inflammatory disorders such as psoriasis and rheumatoid arthritis. All of these entities are characterized by angiogenesis, altered extracellular matrix, and variable degrees of hypoxia. In addition, two VPF/VEGF receptors, flt-1 and kdr, are overexpressed by endothelial cells that line the microvessels that supply these tumors/inflammatory reactions. On the basis of these and other data, we have proposed a model of angiogenesis in which VPF/VEGF plays a central role: this model is applicable to tumors and also to the angiogenesis that occurs in non-neoplastic processes.

Animals↗