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Biomedical subjects

M Heenen

Publications and source records attributed to M Heenen.

At least 19 recordsLinked to original sources
PubMed1992

Measurement of S phase duration in human epidermis using cyclin immunostaining and 3H-thymidine pulse labelling.

Cyclin/PCNA (proliferating cell nuclear antigen) is a nuclear protein strongly associated with DNA replication sites. Under methanol fixation, cyclin immunostaining might serve as a specific marker for S-phase cells. Associated with single labelling with 3H-thymidine, this immunodetection represents an easy method for measuring S-phase duration. Its application to six samples of normal human epidermis allowed us to measure an S phase of 9.7 +/- 0.3 h, a result in fair agreement with a previous estimate of 10.2 h obtained from a classical double-labelling technique with two different concentrations of 3H-thymidine.

Culture Techniques↗
PubMed1992

In vitro infection of normal human keratinocytes by human papillomavirus type 1 followed by amplification of the viral genome in reconstructed epidermis.

Primary cultures of normal human keratinocytes were inoculated in vitro with human papillomavirus type 1 (HPV-1), the agent responsible for deep plantar warts. Upon transfer to dead de-epidermized dermis and growth at the air-liquid interface, keratinocytes reconstituted a pseudoepidermis. Under these highly differentiating conditions, HPV-1 DNA amplification was found to take place in the reconstructed epidermis, being detectable from 7 days after the transfer and persisting for at least 10 days thereafter. The extent of keratinocyte differentiation may be insufficient to allow a complete HPV infectious cycle.

Cell Differentiation↗
PubMed1992

Kinetics of the calcium induced stratification of human keratinocytes in vitro.

In a low concentration of calcium (0.1 mM), keratinocytes form a monolayer with about 30% of cells synthesizing involucrin. After addition of calcium to the culture medium to a concentration of 1.2 mM, the monolayer stratifies within 24 h, with a preferential migration of involucrin positive keratinocytes. In the present study, we tried to determine if keratinocytes control the decision to migrate at a distinct cell cycle point. A percentage labelled mitosis (PLM) curve was constructed for keratinocytes grown in low calcium medium and values for the length of the cell cycle (47 h), S phase duration (11 h) and G2+M period (6 h), were obtained. Monolayer cultures at 80% confluence were switched to high calcium concentration at various times (from 0 to 48 h), after pulse labelling with [3H]-thymidine. Based on the PLM data, the behaviour of cells known to be in S, G1 and G2 at the time of the migration stimulus were followed. No significant difference in the percentage of labelled suprabasal cells was found for any point of the cell cycle. For cells submitting to stratification, in S phase involucrin staining showed that about 60% of the [3H]-thymidine labelled cells were also involucrin negative. These results indicate that upward migration of keratinocytes in cultured epithelium can be triggered at all points in the cell cycle with equal probability and is not restricted to those cells that already contained involucrin.

Calcium↗
PubMed1992

Renewal and differentiation of keratinocytes cultured on dead de-epidermalized dermis.

Human keratinocytes grown at an air-liquid interface on dead de-epidermalized dermis exhibit a pattern of organization similar to that seen in vivo. Cell renewal is limited to the basal layer. The cell cycle time determined after 7 days of culture, using a percentage labelled mitoses (PLM) technique, was about 15 h. This result is comparable with published data for cultivated keratinocytes but is shorter than the parameter proposed for epidermis in vivo. Appearance of labelled cells in the granular layer was observed 4 days after pulse labelling. Despite this high cell renewal, a normal cell differentiation with expression of various keratinization markers was maintained.

Adult↗
PubMed1992

In vitro acantholysis induced by D-penicillamine, captopril, and piroxicam on dead de-epidermized dermis.

Drug-induced pemphigus has been recognized for 20 years, but the mechanisms leading to acantholysis are still unclear. It has recently been demonstrated that penicillamine, captopril, and thiopronin may produce acantholytic lesions, either by direct toxic or biochemical effect, in human skin explants. Our work confirms that penicillamine and captopril may induce acantholysis on the model of keratinocyte culture on dead, de-epidermized dermis. Moreover, it demonstrates that piroxicam, a new non-steroidal anti-inflammatory drug, of which one side effect is a pemphigus vulgaris-like eruption, is also able to produce in vitro acantholysis.

Acantholysis↗
PubMed1990

Spreading of psoriatic plaques: alteration of epidermal differentiation precedes capillary leakiness and anomalies in vascular morphology.

To approach the temporal relationship between alterations in keratinization and capillary leakiness in psoriasis, we studied the topography of these anomalies in spreading psoriatic lesions. Histological and immunohistochemical studies were performed on skin biopsies obtained from normal individuals and from psoriatic patients. In the latter case, biopsies were taken in uninvolved skin, in the center of lesions, and at the edge of evolving plaques (spanning uninvolved and involved skin). Alterations in epidermal differentiation were assessed by the distribution of filagrin, involucrin, and epidermal membrane-bound transglutaminase. Capillary leakiness was evaluated by the abundance of plasma proteins such as albumin, fibrinogen, and immunoglobulin G within the epidermis. Typical alterations of epidermal differentiation were already obvious at the edge of the lesions, in areas devoid of vessel abnormalities and leakiness, or significant cellular infiltration. These results strongly suggest that, during the formation of a psoriatic plaque, defects in keratinocyte differentiation precede the development of vascular anomalies.

Adult↗
PubMed1989

Reproduction of the characteristic morphologic changes of familial benign chronic pemphigus in cultures of lesional keratinocytes onto dead deepidermized dermis.

We report the in vitro reproduction of the classic histologic and ultrastructural features of familial benign chronic pemphigus (FBCP) by seeding suspensions of lesional keratinocytes onto healthy heterologous dead deepidermized dermis (DED). With the use of normal keratinocytes, control cultures showed a well-differentiated epidermis, with keratohyaline granules and lamellar bodies. To the best of our knowledge this is the first successful culture of FBCP with the use of dispersed lesional keratinocytes. The results suggest that in FBCP the epidermis is the site of the defect leading to acantholysis, without any dermal contribution.

Cell Division↗
PubMed1989

Decrease of ultraviolet-induced DNA injury in human skin by p-aminobenzoic acid esters.

The effect of a sunscreen containing a p-aminobenzoic acid (PABA) ester on ultraviolet-induced DNA injury was assessed on human epidermal cells by the measure of unscheduled DNA synthesis intensity. Our results show that DNA repair was reduced by approximately 50%. Therefore, sunscreens based on PABA esters might be expected to reduce photocarcinogenesis.

4-Aminobenzoic Acid↗
PubMed1988

DNA replication fork progression rate and temporal organization of S phase in normal epidermis and in basal cell carcinoma.

The double-pulse labeling technique for DNA fiber autoradiography was applied to epidermal cells from normal human skin and from human basal cell carcinoma (BCC). We aimed to measure the size and replication rate of the replication unit (RU) for both types of cell and to account, from these results, for our previous observation of a near doubling of S-phase duration in BCC, compared with normal skin. The mean RU size was 76 +/- 4 micron in BCC, not significantly different from the 68 +/- 6 micron value found in normal skin, so the mean of those two values (i.e., 72 micron), was used in further calculations. The rate of replication fork progression was 0.59 +/- 0.005 micron/min in the normal epidermis and 0.33 +/- 0.03 micron/min in BCC, corresponding to a replication time of the average RU equal to 61 min and 109 min, respectively. Thus, with an unchanged RU size in BCC, the observed 1.8-fold decrease in the rate of fork progression in the tumor can account entirely for our previous observation of a 1.8-fold increase in S-phase duration in this tumor, without requiring the assumption of any change in the temporal organization of DNA synthesis in the malignant cells. Considering S phase as an ordered process in which a major part, if not all, of the genome replicates at genetically determined times, we suggest that the clusters of replication units are, in turn, organized into temporally defined "sets". These sets are composed of all the clusters (whatever their chromosomal location) that are programmed to initiate replication during the same fraction of the S period. This hypothesis implies that DNA synthesis in a given set is triggered by some event coupled to progression of replication in the immediately preceding set. Based on a S-phase duration of 10.2 hours in normal skin and of 19.2 hours in BCC (our previous data), and assuming perfect synchrony and homogeneity of the clusters within each set and of each cluster's constitutive RUs, the minimum number of sequentially replicating sets, in both instances, can be estimated as roughly equal to 10.

Basal Cell Carcinoma↗
PubMed1987

Psoriasis: hyperproliferation cannot induce characteristic epidermal morphology.

A mathematical model of cell renewal in epidermis is proposed for describing how psoriatic lesions might develop, based on available cell kinetic data for normal and psoriatic epidermis. Our simulations clearly demonstrate that an increase in the turnover rate in the germinative cell population cannot alone induce the typical psoriatic tissue architecture (i.e. increased number of germinative cells). Two perturbations are needed to account for the morphology of clinically-stable psoriatic lesions. The first corresponds to a temporary disturbance of the steady state of the germinative layer, resulting in limited growth of this compartment; the second perturbation corresponds to a reduction in transit time in the differentiated compartment. Moreover, our simulation, based on a widely-accepted hypothesis of homeostatic control of tissue kinetics, demonstrates that the primary cause of typical psoriatic morphology is probably an alteration in epidermal maturation. In this view, depletion of differentiated cells at the surface is the stimulus for the increased cell production rate in the germinative population.

Cell Cycle↗
PubMed1979

Psoriasis and cell cycle: a computer simulation.

Computer simulation using a two-phase model of the cell cycle has demonstrated that a transitory shift in the normal balance between the rate of cell flow to the differentiated and that to the proliferative compartment is sufficient to cause a permanent increase in the germinative population, thus resembling the alteration found in psoriasis. This shows the coherence of the hypothesis that cell kinetic parameters need not necessarily be expected to differ in psoriasis as compared with normal epidermis. This theoretical demonstration raises the possibility that the psoriagenic agent need not necessarily be present as long as the symptoms of psoriasis persist. From a therepeutic point of view this has implications that are sifficiently evident to warrant communication of this work.

Cell Cycle↗
PubMed1979

[Cell cycle time of normal epidermis, psoriasis and some epidermal tumors (author's transl)].

Cell kinetic of human epidermis has been studied with different techniques of labeling by 3H-thymidine. A review of the literature for human epidermis, psoriasis and some epidermal tumors has been performed and discussed in the light of our own studies. If the cell cycle time seems to be slightly shorter in epidermal tumors in comparison to normal epidermis, this doesn't constitute the fundamental characteristic of the neoplastic lesions. Indeed the most specific alteration in the cell cycle parameters for precancerous and cancerous tissue is a lengthening of the S phase duration. This modification of the DNA synthesis time is perhaps of reflection of the depressed state of the chromatin of neoplastic tissues.

Cell Cycle↗