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Biomedical subjects

P E Van Erp

Publications and source records attributed to P E Van Erp.

15 recordsLinked to original sources
PubMed1998

Epidermal cell kinetics by combining in situ hybridization and immunohistochemistry.

Double labelling can serve as a useful tool for providing information about cell kinetics in normal and hyperproliferative tissues in general, and skin in particular. We have developed a double-labelling method that combines immunohistochemistry using the monoclonal antibody MIB1 and non-isotopic in situ hybridization using either a digoxigenin-labelled RNA probe specific for histone 3 mRNA sequences or a Fluorescein-labelled oligonucleotide probe specific for histone 2b, 3, 4 mRNA sequences. Double labelling was performed on normal, tape-stripped normal skin and psoriatic skin. The three proliferation markers were also examined by single labelling. The ratio of cells in the S-phase (Ns) and the growth fraction (Ncy) was determined. In normal skin, psoriatic skin and tape-stripped normal skin after 24 h and after 48 h, we calculated that 15%, 16%, 3% and 12% of growth fraction consisted of cells in the S-phase respectively. The S-phase lasts approximately 10 h, so the cell cycle time in normal and psoriatic skin is approximately 62.5 h. At present, the MIB1/H3 digoxigenin or MIB1/H2b-H3-H4 Fluorescein double-labelling technique cannot be used routinely. Therefore, in order to understand the cell kinetic processes better, experiments are recommended to optimize these methods. From a practical point of view and for reasons of specificity and sensitivity, we prefer the Fluorescein-labelled oligonucleotide probe method.

Cell Division↗
PubMed1998

Investigation on a novel and specific leukotriene B4 receptor antagonist in the treatment of stable plaque psoriasis.

The aim of the present study was to investigate the efficacy and clinical tolerability of the specific leukotriene B4 receptor antagonist VML295 in the treatment of stable plaque psoriasis. Immunohistochemical and flow cytometrical methods were used to assess the effects on inflammation and epidermal proliferation. VML295 in the treatment of chronic plaque psoriasis was shown to be safe and well tolerated. After treatment, there was a statistically significant difference between patients treated with VML295 and patients treated with placebo with respect to the leukotriene B4-induced CD11b up-regulation on the cell surface of polymorphonuclear leukocytes derived from peripheral blood. Ex vivo CD11b up-regulation in the VML295-treated group was completely inhibited after 7 days of treatment (P = 0.001). This effect persisted until the end of the treatment period (P = 0.004 on day 15 and P < 0.0001 after 4 weeks), whereas CD11b up-regulation in the placebo group remained unaffected. There was no statistically significant difference in the median psoriasis area and severity index between the treatment groups at the end of the treatment period. During treatment, no significant histological changes were observed in the markers for cutaneous inflammation and epidermal proliferation. Although not statistically significant, a tendency for the increased expression of some markers of cutaneous inflammation and epidermal proliferation was observed after 1 week of treatment with VML295, and a decreased expression of these markers was seen after 4 weeks of treatment with VML295. This observation could indicate anti-inflammatory effects of VML295 appearing between 2 and 4 weeks after the start of treatment.

Adult↗
PubMed1998

The effect of long-term treatment with tacalcitol on the psoriatic epidermis. A flow cytometric analysis.

During the last decade, novel analogues of 1alpha,25-dihydroxy vitamin D3 have been developed for the treatment of psoriasis. Recently, the efficacy of short-term treatment with the novel derivative tacalcitol (1alpha,24-dihydroxy vitamin D3) has been documented. However, data on the long-term effect of tacalcitol on psoriatic skin are sparse. In this study, we assessed the cell characteristics of the psoriatic epidermis after treatment with tacalcitol for up to 24 weeks. We investigated how long-term treatment with tacalcitol modulates the percentages of differentiated keratinocytes, inflammation cells and basal keratinocytes, and the percentage of cells in the SG2M phase in the basal cell population. From 11 patients who were treated with tacalcitol for up to 18 months, we obtained single-cell suspensions of a representative psoriatic lesion after 0, 8, 12, 18 and 24 weeks of treatment. A Psoriasis Area and Severity Index was performed at each visit as well. Cell suspensions were stained with markers for inflammation (Vim3B4), differentiation (RKSE60) and proliferation (TO-PRO-3 iodide) and analysed flow cytometrically. Clinically, patients improved significantly after 8 weeks of treatment. This clinical effect was preserved for the rest of the period of treatment with no further significant improvement. Proliferative activity also decreased significantly after 8 weeks of treatment. Proliferation did not show further significant decreases or habituation after 12, 18 and 24 weeks. For inflammation, no statistically reliable trends could be seen. Differentiation improved significantly after 8 weeks of treatment, but decreased again significantly after 12 weeks. In the period from 12 to 24 weeks, no further significant change was observed. We conclude that tacalcitol is an effective antipsoriatic drug. Prolonged treatment with tacalcitol will generally maintain improvement at the level reached after 8 weeks. Owing to the beneficial effect on both clinical state and proliferation, tacalcitol is likely to be an adequate maintenance therapy.

Anti-Inflammatory Agents↗
PubMed1996

Induction of normal and psoriatic phenotypes in submerged keratinocyte cultures.

Lesional psoriatic epidermis displays a number of phenotypic changes that are distinct from the differentiation program found in normal interfollicular epidermis. In psoriatic epidermis, keratinocytes are hyperproliferative and several differentiation-associated molecules are expressed that are absent in normal skin (e.g., cytokeratins (CK) 6, 16, and 17, and the epidermal proteinase inhibitor SKALP/ elafin). In addition, several molecules which are normally restricted to the stratum granulosum are strongly upregulated in the stratum spinosum (e.g., psoriasis-associated fatty acid binding protein (PA-FABP), psoriasin, involucrin, and transglutaminase). The aim of this study was to develop in vitro culture systems which (a) would allow to study the induction of normal and psoriatic differentiation pathways, and (b) would be amenable for screening of antipsoriatic drugs. Here we have investigated several models for induction of differentiation with respect to the expression of markers for the normal and psoriatic phenotype. Cell cycle parameters and expression levels of CK1, CK10, CK16, SKALP/elafin, transglutaminase, involucrin, psoriasin, and PA-FABP were assessed in these models using flow cytometry, immunocytochemistry, and Northern blot analysis. We observed that induction of differentiation with fetal calf serum resembled the psoriatic phenotype (sustained hyperproliferation; high levels of CK16, SKALP/elafin, transglutaminase, and involucrin; moderate psoriasin expression), whereas differentiation induced by growth factor depletion in a confluent culture resembled the normal differentiation phenotype (low proliferative rate; high expression levels of CK1 and CK10; moderate expression of involucrin and transglutaminase; low expression levels of SKALP/elafin and CK16; absence of psoriasin). We propose that these models can be used to study expression and pharmacological modulation of selected differentiation genes and the coordinated expression of sets of genes associated with epidermal differentiation programs.

Blotting, Northern↗
PubMed1996

The role of epidermal proliferation in the pathogenesis of psoriasis.

The micromorphological appearance of the psoriatic lesion is characterized by cutaneous inflammation, epidermal acanthosis and abnormal keratinization. The relevance of immunological factors has been reviewed in recent literature. Despite a longstanding interest of many centres in the pathogenesis of psoriasis, the position of epidermal proliferation, a key feature of the psoriatic plaque, remains to be established. The aim of the present review was to provide evidence from the literature and from our own work that enables us to answer the question to what extent epidermal proliferation might be a primary pathogenetic event or whether it represents a secondary phenomenon resulting from other abnormalities.

Animals↗
PubMed1996

Epidermal cell DNA content and intermediate filaments keratin 10 and vimentin after treatment of psoriasis with calcipotriol cream once daily, twice daily and in combination with clobetasone 17-butyrate cream or betamethasone 17-valerate cream: a comparative flow cytometric study.

Calcipotriol and corticosteroids, two therapy modalities frequently prescribed in the treatment of psoriasis, are often used in combination. The aim of the present study was to determine whether the cell biological response pattern of concurrent use of calcipotriol and corticosteroids is different from calcipotriol monotherapy. Forty patients with chronic plaque psoriasis were divided at random in four parallel groups and treated for 8 weeks with: (1) calcipotriol cream (50 micrograms/g once daily); (2) calcipotriol cream twice daily; (3) calcipotriol and clobetasone 17-butyrate (0.5 mg/g) creams; and (4) calcipotriol and betamethasone 17-valerate (1 mg/g) creams. Before and after treatment keratotome biopsies were taken and single cell suspensions prepared for flow cytometric analysis. Flow cytometric multiparameter quantification of markers for proliferation (TO-PRO-3), differentiation (antikeratin 10) and inflammation (antivimentin) was used to evaluate all four therapy modalities. A statistically significant decrease of the percentage of basal cells in S- and G2M-phase (proliferation) was obtained with all therapy modalities, except for calcipotriol monotherapy applied once daily. A significant reduction of the number of vimentin-positive cells (non-keratinocytes) was observed following combined treatment with calcipotriol and clobetasone butyrate. In contrast, monotherapy with calcipotriol had virtually no effect on the number of vimentin-positive cells. It can be concluded that: (i) calcipotriol monotherapy, applied once daily was less antiproliferative compared with twice daily applications of calcipotriol or the combined treatment with corticosteroids and that (ii) the combination of calcipotriol and corticosteroids proved to have a marked effect on the percentage of non-keratinocytes, in contrast to the modest effect of calcipotriol.

Administration, Topical↗
PubMed1994

TO-PRO-3 iodide: a novel HeNe laser-excitable DNA stain as an alternative for propidium iodide in multiparameter flow cytometry.

A new red emitting fluorophore, TO-PRO-3 iodide (TP3), which is best excited by an HeNe laser (633 nm), has been compared with propidium iodide (PI) for measuring relative DNA content. TP3, which has a peak absorbance at 642 nm and emission at 661 nm, has been tested on peripheral blood lymphocytes (PBL) and keratinocytes in a two-laser system. As an example, we present a three-color flow cytometric application utilizing TP3 in combination with fluorescein-isothiocyanate (FITC) and phycoerythrin (PE) conjugated to monoclonal antibodies in this paper. A subequilibrium concentration of 1 microM TP3, most preferably used in combination with RNase treatment, proved to be a powerful alternative for DNA amount determination. In human- and mouse-Balb/MK-keratinocyte populations with different S-phase fractions, PI and TP3 showed a good correlation. Finally, in the triple labelling experiment we clearly demonstrated that TP3 is readily applied to the analysis of binding of two antibodies and relative DNA content simultaneously.

Animals↗
PubMed1993

Topical treatment of psoriatic plaques with 1,25-dihydroxyvitamin D3: a cell biological study.

Calcitriol, 1 alpha,25 dihydroxycholecalciferol (1 alpha,25 (OH)2 D3) is a natural active vitamin D3 metabolite, which has been shown to have antipsoriatic efficacy. In vitro studies have demonstrated that calcitriol influences various aspects of inflammation, epidermal proliferation and keratinization. The aim of the present study was to determine to what extent calcitriol (3 micrograms/g in white petrolatum) affects these parameters in vivo. Using an immunohistochemical assessment of recruitment of cycling epidermal cells, filaggrin and involucrin expression, T-cell accumulation, polymorphonuclear neutrophil (PMN) accumulation, amount of endothelium and ICAM-1 expression, we demonstrated that: (i) modulation of all these parameters occurred during calcitriol treatment; (ii) there was early reduction of epidermal proliferation and PMN accumulation; (iii) the order of changes was comparable with the response to treatment with calcipotriol. In conclusion, at the cell biological level, calcitriol (3 micrograms/g in white petrolatum) has a substantial effect on various elements of the psoriatic lesion.

Calcitriol↗
PubMed1990

Markers for proliferation and keratinization in the margin of the active psoriatic lesion.

To study the development of the psoriatic lesion, biopsies were taken from the margin of spreading plaques and acute pinpoint papules. Consecutive sections across the margin were stained using different monoclonal antibodies to characterize epidermal growth (Ki-67) and abnormal keratinization (Ks8.12, RKSE60). All three immunohistochemical markers showed pronounced changes in the lesional skin with a clear transition to the uninvolved skin. The suprabasal Ks8.12 binding was the earliest change found in the epidermis, and its localization high in the suprabasal compartment indicates that metabolic dysregulation in this cell population was not a consequence of the recruitment process in the basal layer.

Adult↗
PubMed1989

A sequential double immunoenzymic staining procedure to obtain cell kinetic information in normal and hyperproliferative epidermis.

A sequential double immunoenzymic staining procedure was developed using the monoclonal antibody anti-BrdUrd and Ki67 in order to determine whether hyperproliferative skin disorders, such as psoriasis, are characterized by an increased growth fraction rather than a much shorter cell cycle time of all germinative cells. Ki67 binds to a proliferation-associated nuclear antigen in a variety of human cell types, and anti-BrdUrd can be used to identify DNA-synthesizing cells. Although in hyperproliferative epidermis the absolute numbers of BrdUrd-positive cells as well as Ki67-positive cells were grossly increased, the ratio of these values was not changed compared to the ratio found in the epidermis of the clinically uninvolved skin of psoriatic patients and in normal epidermis. This suggests an increased growth fraction in hyperproliferative epidermis. Our data show that immunohistochemical double-staining techniques can be a valuable tool in the study of cell cycle kinetics in epithelial tissues.

Antibodies, Monoclonal↗
PubMed1989

Epidermal hyperproliferation assessed by the monoclonal antibody Ks8.12 on frozen sections.

Using the monoclonal antibody Ks8.12 directed against keratins 13 and 16, we have studied psoriatic and trauma-induced hyperproliferation. In normal skin 40 and 64 h after sellotape stripping and in psoriatic lesions, a pronounced suprabasal staining by Ks8.12 was observed. In unstripped normal skin and in the clinically uninvolved skin of psoriatic patients some patchy staining limited to the basal layer was seen. The clear distinction between normal and hyperproliferative skin in keratin expression as detected by Ks8.12 indicates that this antibody may serve as a marker for epidermal proliferation in psoriasis.

Antibodies, Monoclonal↗
PubMed1989

Two binding sites for Ki67 related to quiescent and cycling cells in human epidermis.

The monoclonal antibody Ki67 (Ki67) binds to a nuclear antigen expressed by cycling cells of several human tissues and to the cytoplasm of the basal layer cells of squamous epithelia. We have used an immunohistochemical method to visualize the binding sites of Ki67 in normal and hyperproliferative epidermis. Cytoplasmic staining was present in the basal layer cells of normal epidermis, but was decreased in psoriatic and post-tapestripping epidermis. In sections of normal epidermis only a small minority of nuclei were positive, but sections of psoriatic epidermis and epidermis 40 and 48 h after tapestripping showed large numbers of positive nuclei in the basal and suprabasal layers. Since recent reports strongly suggest that the cell production rate is regulated by changes in the number of cycling cells, the hypothesis that Ki67 binds also in human epidermis to the nuclei of cycling cells is supported by the present findings.

Antibodies, Monoclonal↗
PubMed1981

Studies on the plasma membrane of normal and psoriatic keratinocytes. 3. Uptake of labelled sugars and their incorporation into glycoconjugates.

We report the uptake of four labelled sugars by keratinocytes isolated from normal epidermis, psoriatic 'uninvolved' skin and psoriatic lesions. Our findings include the following: (1) The rate of uptake of all sugars by the psoriatic lesion is increased. (2) This abnormally high uptake diminishes dramatically during 22 h incubation in vitro. (3) There is a striking abnormality in the metabolism of fucose by psoriatic keratinocytes; our data suggest an increased rate of incorporation of fucose into glycoconjugates.

Carbohydrate Metabolism↗
PubMedDate not supplied

CD 2394, a novel synthetic retinoid, initiates an embryonic type of differentiation in hyperproliferative skin.

In human skin, there are 2 types of epidermal differentiation: normal differentiation, characterized by keratin 10 expression, and alternative differentiation. Alternative differentiation may be regeneration-associated differentiation (keratin 6 and 16) or re-induction of embryonic differentiation (expression of keratin 13, 15 and 19). The purpose of this study was to investigate the effect of the novel synthetic retinoid CD 2394 on hyperproliferative human skin, with respect to embryonic differentiation in particular. The effects of CD 2394 were compared with untreated and vehicle-treated skin 48 h after tape-stripping. In a multiparameter flow cytometric assay, parameters of proliferation, normal differentiation, embryonic differentiation and inflammation were assessed. With respect to proliferation, treatment with CD 2394 resulted in a decreased number of cells in the G2M-phase. Normal differentiation was decreased in CD 2394 treated skin. Furthermore, most of the CD 2394 treated samples showed expression of keratin 13, which was not seen in the otherwise treated skin. A correlation between keratin 10 and keratin 13 expression could not be demonstrated. This study showed that CD 2394 is capable of inducing an embryonic pathway of differentiation, which is distinct from normal differentiation or regeneration-associated differentiation.

Acne Vulgaris↗