Epidermal Cell Kinetics Research Articles

Comparison of intradermal injections of ]bromodeoxyuridine and tritiated thymidine in the in vivo measurement of epidermal cell turnover time in goats

Study of keratinisation defects such as seborrhoea may require investigation of epidermal cell kinetics in clinical cases. For this to be practically useful a safe, reliable and quick method is essential. This study compared epidermal cell kinetics in the goat following the use of intradermal injection of tritiated thymidine ([ 3H]TdR) and bromodeoxyuridine ( BURY) to label S phase nuclei. Turnover time for goat epithelial cells was not significantly different for the two agents; 26·3 ± 6·0 days for thymidine and 21·8 ± 5·2 days for burd It was concluded that intradermal injection of BURD will be a suitable technique for investigating epidermal cell kinetics in vivo and could have applications in a clinical situation because it is quicker and safer than [ 3H]TdR.

Benzoαaαpyrene-induced skin damage and tumor promotion in the mouse

Epidermal cell kinetics and DNA adduct levels, and skin morphological changes were measured following weekly topical applications for 29 weeks of high (16, 32 and 64 micrograms) benzo[a]pyrene (B[a]P) doses to female ICR/Harlan mice, in order to investigate the relationship of these parameters to the timing, incidence and morphology of the elicited tumors. During the tumor latency period, [3H]thymidine labeling index, mitotic index, epidermal cell stacking, incidence of pyknotic and dark basal keratinocytes and labeled mitoses were periodically measured, as were nuclear area and DNA content. DNA adducts in skin epidermis were measured by an ELISA method over a period of 9 weeks of single weekly applications of 64, 32, 16 or 8 micrograms B[a]P. There was an initial linear increase in DNA adducts with dose in the epidermis but the increase was much less steep above 32 micrograms/week. This did not correlate with the sharp rise in tumor response above the 32 micrograms/week dose rate. Cell kinetic changes in response to the 64 micrograms/week dose reached a plateau in the first few weeks of the tumor latent period. There was little epidermal hyperplasia but an associated dose-dependent increase in [3H]thymidine labeling index, mitotic index and incidence of pyknotic and dark cells. This evidence indicated that B[a]P produced extensive cytotoxicity and cell death with regenerative proliferation under these conditions. Giant keratinocytes occurred in all dose groups. Analysis of a labeled mitosis curve indicated that B[a]P produced a G2/M block. There was a marked inflammatory response in the dermis at all B[a]P doses. Mice were observed weekly for tumor formation. Virtually all of the tumors were papillomas on initial appearance and required an average of 8 weeks to convert to carcinomas. The substantial cell killing and regenerative proliferation, and the correspondence between the dose-response patterns for epidermal damage and tumors, together with the initial appearance of tumors in the benign form, a characteristic of the action of promoting agents, provided evidence that the tissue damage associated with the high dose levels of B[a]P used in this study reflected tumor-promoting activity in this mouse epidermal tumorigenesis model. The implication of the results for mathematical models of tumor formation are discussed.

Mathematical model analysis of mouse epidermal cell kinetics measured by bivariate DNA/anti‐bromodeoxyuridine flow cytometry and continuous [3H]‐thymidine labelling

In a previous study the epidermal cell kinetics of hairless mice were investigated with bivariate DNA/anti-bromodeoxyuridine (BrdU) flow cytometry of isolated basal cells after BrdU pulse labelling. The results confirmed our previous observations of two kinetically distinct sub-populations in the G2 phase. However, the results also showed that almost all BrdU-positive cells had left S phase 6-12 h after pulse labelling, contradicting our previous assumption of a distinct, slowly cycling, major sub-population in S phase. The latter study was based on an experiment combining continuous tritiated thymidine [( 3H]TdR) labelling and cell sorting. The purpose of the present study was to use a mathematical model to analyse epidermal cell kinetics by simulating bivariate DNA/BrdU data in order to get more details about the kinetic organization and cell cycle parameter values. We also wanted to re-evaluate our assumption of slowly cycling cells in S phase. The mathematical model shows a good fit to the experimental BrdU data initiated either at 08.00 hours or 20.00 hours. Simultaneously, it was also possible to obtain a good fit to our previous continuous labelling data without including a sub-population of slowly cycling cells in S phase. This was achieved by improving the way in which the continuous [3H]TdR labelling was simulated. The presence of two distinct subpopulations in G2 phase was confirmed and a similar kinetic organization with rapidly and slowly cycling cells in G1 phase is suggested. The sizes of the slowly cycling fractions in G1 and G2 showed the same distinct circadian dependency. The model analysis indicates that a small fraction of BrdU labelled cells (3-5%) was arrested in G2 phase due to BrdU toxicity. This is insignificant compared with the total number of labelled cells and has a negligible effect on the average cell cycle data. However, it comprises 1/3 to 1/2 of the BrdU positive G2 cells after the pulse labelled cells have been distributed among the cell cycle compartments.

Increased stratum corneum turnover induced by subclinical irritant dermatitis

The chronic effects of the irritant sodium lauryl sulphate (SLS) on stratum corneum (SC) barrier function, determined by transepidermal water loss (TEWL) measurements and on epidermal cell kinetics, estimated by stratum corneum turnover time (SCTT) determination (dansyl chloride staining method), were investigated in 18 healthy female volunteers. SLS (7.5%) was applied without occlusion for 20 min once daily, over a period of 3 weeks (5 days a week) on dansyl chloride-stained skin and on untreated skin. SCTT of untreated skin (19.3 +/- 0.8 days; mean +/- SEM) was not changed by daily treatment with water (control) (19.3 +/- 2.0) but was significantly reduced by SLS (10.9 +/- 0.6; P less than or equal to 0.0001; compared to controls). However, TEWL was increased in SLS-treated sites 1.5-fold after 4 days of treatment (5.3 +/- 0.6 vs. 3.5 +/- 0.3; P less than 0.001). At the end of the second week, TEWL was increased 2.6-fold and after 3 weeks TEWL was 3.3 times higher than in controls 13.0 +/- 1.6 vs. 3.9, P less than or equal to 0.0001). The intensity of SLS-induced irritation as measured by TEWL was significantly correlated with baseline TEWL (r = 0.50; P less than or equal to 0.02) and significantly negatively correlated with SCTT of SLS treated sites (r = -50; P less than or equal to 0.02) but not with SCTT of untreated skin (r = 0.19).

Kinetics of epidermal cells in skin from human cadavers.

Kinetics of epidermal cells in skin from human cadavers.

Effect of hexadecane on protein kinase C of pig epidermis.

The application of hexadecane on animals skin induces hyperkeratinization and hyperplasia of the epidermis, however, the initial mechanisms of the epidermal cell proliferation and keratinization by hexadecane stimulation remains unknown. Protein kinase C is reported to be one of the critical enzymes involved in proliferation and differentiation of various cells and tissues. Therefore we investigated the effects of hexadecane on protein kinase C in pig epidermis. Protein kinase C activity of the pig skin increased 10 min after topical application of hexadecane to the back of the pig, normalized at 30 min, and subsequently kept falling for 24 h. In studying hexadecane dropped on floating sliced pig skin in Krebs buffer, similar results were obtained for the short term. Immediately after the hexadecane treatment, protein kinase C activity was not altered as compared with that of the untreated skin. Thus, the alteration of the protein kinase C activity after the hexadecane treatment is not due to the direct effect of hexadecane on the enzyme, but is due to other as yet unknown mechanisms of epidermal cell kinetics in response to hexadecane stimulation. We discuss the mechanisms of protein kinase C activity alteration upon treatment with hexadecane.

Evaluation of Epidermal Cell Kinetics Following Freezing or Wounding of Mouse Skin and Their Potential as Initiators of Carcinogenesis

It has been shown that abrasion, and consequent regenerative hyperplasia, acts as a promoting agent in mouse skin carcinogenesis. The present experiments were designed to evaluate the possibility that ulceration and its consequent regeneration might also act as initiators. Female Sencar mice were used, and ulceration was induced either by the application of a frozen rod or by incision of the skin of the back. The time course of the ulceration, regeneration, and repair of the mouse skin following ulceration by either method was evaluated utilizing morphologic and autoradiographic techniques. The labeling index of the epidermis, using [3H]-thymidine and autoradiography, reached a maximum level 7 days after ulceration and the epidermal hyperplasia was most pronounced at days 7-14. The potential initiating activity of freeze ulceration or incision was evaluated by performing these procedures on 7-week-old female Sencar mice followed by promotion with 12-O-tetradecanoylphorbol-13-acetate (TPA) applied twice a week to the ulcerated areas, 5.2 micrograms in each application. Extending the total experimental observation to 1 year indicated that freeze ulceration and incision did not initiate carcinogenesis in the mouse skin when promoted with TPA.

Effects of ultraviolet radiation and immunosuppressive therapy on mouse epidermal cell kinetics.

The effects of immunosuppressive drugs on epidermal cell mitotic activity and the proliferative response of epidermis following ultraviolet radiation (UVR) were tested. Hairless (Skh-hr 1) mice were treated with immunosuppressive drugs at equivalent clinical doses with or without concomitant UVR (290-400 nm). Epidermal parameters measured were mitotic index (Im), rate of entry into mitosis (Fm), flash labelling index (FLI), rate of entry into DNA synthesis (Fs) and DNA content (flow cytometric analysis). In non-irradiated skin, prednisolone therapy depressed both mitotic activity and DNA synthesis; azathioprine and cyclosporin A had no effect; cyclophosphamide therapy increased the Fm and FLI values. Following repeated doses of UVR, there were enhanced mitotic activity and DNA synthesis in epidermis. Prednisolone therapy moderately depressed both proliferative responses; cyclophosphamide enhanced mitotic activity; azathioprine and cyclosporin A had no effect on these responses. The significance of these findings in relation to potential for increased susceptibility of skin to UV-induced carcinogenesis is discussed.

Cell Kinetic Basis for Pathophysiology of Psoriasis

Studies on the cell proliferation kinetics of psoriatic epidermal cells are presented and the results compared to similar studies for normal epidermis. The short 36-h duration of the psoriatic cell cycle (Tc) is confirmed with the first double-peaked fraction of labeled mitoses (FLM) curve in human subjects. The growth fraction of psoriasis using two experimental techniques approximates 100% within 36 h, confirming the rapid Tc found by the FLM method. The cell kinetic basis for the pathophysiology of psoriasis consists of at least 3 proliferative abnormalities in comparison to normal epidermis. By far the largest alteration is the shortening of the Tc from 311 to 36 h. There is also a doubling of the proliferative cell population in psoriasis from 27,000 to 52,000 cells/mm and an increase in the growth fraction from 60% to 100%. As a consequence of these abnormalities the psoriatic epidermis produces 35,000 cells/day from a proliferative compartment of 52,000 cells/mm2 surface area. This is a 28-fold greater production of cells than the 1,246 cells/day produced in normal epidermis. The biochemical or control factors leading to these kinetic differences continue to remain elusive.

Epidermal Growth in the Bottlenose Dolphin, Tursiops truncatus

Epidermal growth in two mature female bottlenose dolphins, Tursiops truncatus, was investigated by following the movement of a cohort of tritiated thymidine-labeled epidermal cells for 59 days. The majority of the cells migrated in a cluster which was estimated to reach the skin surface in 73 days. We calculate that the outermost cell layer is sloughed 12 times per day. Turnover time and sloughing rate are estimated to be 1.7 times longer and 8.5 times faster than the respective values for epidermal cell kinetics in humans. This apparent inconsistency of slow transit time and rapid sloughing rate is reconciled by the convoluted structure of the stratum germinativum in the dolphin which results in a ratio of germinatival to superficial cells of 876:1. The stratum germinativum of dolphin epidermis appears to lack morphologically distinct, spatially segregated subpopulations of anchoring and stem cells. Dolphin epidermis has a large capacity for cell population, relatively long turnover time, and rapid sloughing rate. The adaptive advantages of these characteristics are discussed.

DNA flow cytometry of the epidermis of patients with multiple epidermal carcinomas.

We used DNA flow cytometry to determine whether the uninvolved skin of twenty-four patients with multiple epidermal cancers differed from normal skin with regard to its epidermal cell kinetics. We also studied the relationship between the cell kinetics and the type of cancer, previous exposure to ionizing or UV radiation, previous arsenic intake, skin type and coexisting psoriasis. Both exposed and unexposed skin was studied. In thirteen patients the S and G2M fractions of a monoclonal tumour were compared with the values in unexposed and exposed skin. The S and G2M fractions increased progressively from unexposed skin to exposed skin to tumour tissue. Patients who had developed both squamous and basal cell carcinoma differed from those who had developed only basal cell carcinoma by having a significantly higher G2M fraction in the exposed skin, and significantly higher S and G2M fractions in the unexposed skin. Patients exposed to ionizing radiation tended to show higher G2M fractions in exposed skin compared with the remaining individuals (P = 0.06). The presence of psoriasis, exposure to UVR or sunlight and previous arsenic ingestion did not appear to affect the results.

Acute Effects of Ultraviolet Light B on Morphology and Epidermal Cell Kinetics in Human Skin Transplanted to Nude Mice

Grafts of human skin on nude mice were irradiated with varying doses of ultraviolet light B and at various intervals were subjected to histological examination and determination of the epidermal 3H-thymidine labelling index. The studies showed that the human skin in the foreign milieu of the nude mouse retained its ability to respond to phototoxic damage. The findings suggest that the nude mouse/human skin model could be a valuable tool in human photodermatological research.

Age-related changes of epidermal cell kinetics in the hairless mouse.

Cell kinetic variables in normal untreated hairless mice were studied in order to observe possible age-related changes. Generally, groups of 4 male and 4 female mice were subjected to study at various ages from one to 115 weeks. The number of basal and suprabasal cells per microscopic field was observed, and after injection of tritiated thymidine the mean labelling index, the average specific activity and the mean grain count were scored. After injection of Colcemid, the average number of Colcemid-arrested mitoses was counted. With flow cytometry the fraction of cells in S and in G2 + M was also observed. In general, both the number of suprabasal cells and the proliferative variables were significantly lower in the very young mice. They increased to slightly above normal values at about 20-22 weeks of age, and then fluctuated a little with two additional possible peaks at 40-50 and around 80 weeks, respectively, and two troughs some weeks after the peaks. However, this rhythmicity was slight and not significant. Thus the only significant age-related pattern was that very young mice have a thin epidermis and low proliferative variables. These values increase up to the age of 20 weeks, and from then on there are no obvious and significant alterations, only slight rhythmic undulations almost within normal limits. The importance of cell kinetic changes with age for epidermal carcinogenesis is discussed in relation to these observations.

Organ culture of adult human skin; effect of culture temperature.

ABSTRACTIn the organ culture of adult human skin, long‐term preservation of cultured epidermis was obtained when the culture temperature was shifted from 37°C to 32°C. The epidermal cell kinetics were compared for both culture temperatures using the findings of mitotic rate, mitotic distribution and autoradiographic labelling index with tritiated‐thymidine (3H‐TdR). As a result, it appeared that a lowered culture temperature controlled the cell cycle in vitro to maintain long‐term, active epidermal proliferation.

Cell Proliferation Kinetics in the Human Hair Root

The kinetics of cell proliferation in the matrix cells of human scalp hair were determined in vivo. Autoradiographic analysis was employed on biopsy specimens taken at specific time intervals after intradermal injection of tritiated thymidine (3H-TdR) into the scalps of volunteer subjects. The durations of the S and G2 phases (Ts and Tg2) were 11.1 and 3.6 hr respectively, obtained from the composite percent labeled mitosis curve. The labeling index of the hari matrix cells was 28.6% +/- 2.2%. The durations of mitosis and of the total cell cycle (Tm and Tgc) were calculated to be 0.8 hr and 38.8 hr respectively. The growth fraction of hair root germinative cells was estimated as 75-100% on the basis of "continuous" 3H-TdR exposure over the duration of the cell cycle. It is noted that the kinetics of human hair root cells are quite similar to the kinetics of psoriatic epidermal cells, suggesting that both tissues are approaching the maximum proliferative rate of keratinogenic epithelial cells.

Localization of the defect in skin diseases analyzed in the human skin graft-nude mouse model.

Human skin can be grown away from its donor for prolonged periods as grafts on congenitally athymic "nude" mice. This system has been used to analyze the defect in several skin diseases, specifically to localize the site of the defect to the skin itself or to the epidermal or dermal components of the skin. In order to validate the use of the nude mouse human skin graft system in the analysis of skin defects, we have demonstrated that a systemic metabolic defect which involves the skin, namely essential fatty acid deficiency, can be differentiated from a defect residing primarily in the skin itself. Skin-marker systems have been developed for use with the nude mouse-human skin graft model to document the identity of human skin grafts and epidermal and dermal components of the grafts after prolonged periods of growth on the nude athymic mice. Y-body, a small fluorescent segment of the Y-chromosome seen in interphase cells, is used as a sex marker and serves to distinguish sex differences between the graft and the mouse recipient or between skin components of the graft. The ABH "blood-group" antigens are present on differentiated epidermal cell surfaces and identify the grafted epidermis according to the blood groups of the donor. In previous studies, lamellar ichthyosis was shown to be well maintained after prolonged periods of growth on nude athymic mice, indicating that the defect in this disease resides in the skin itself. Recombinant grafts composed of normal and lamellar ichthyosis epidermis and dermis further localize the defect to lamellar ichthyosis epidermis. Psoriasis is well maintained on the nude mouse-skin graft model. The epidermal hyperplasia and hyperproliferative epidermal cell kinetics of psoriasis are manifested in the grafts of active psoriasis maintained for prolonged periods on the nude mice, but the inflammatory component of psoriasis is absent. Recombinant graft studies utilizing normal and psoriatic epidermis and dermis demonstrate psoriasis only in recombinant grafts composed of both psoriatic epidermis and dermis. These studies indicate that psoriasis requires both psoriatic epidermis and dermis for its expression.

Some Gerontologic Considerations in the Practice of Dermatology

Gerontology and geriatrics are areas of increasing importance in medicine today. Although changes in the appearance and behavior of human skin have long been noted to accompany aging, physicians have only recently begun to consider the impact of such an age-associated change on the manifestations of dermatologic disease and on the dermatologic management of elderly patients. Gerontologic studies in such fields as immunology, epidermal cell kinetics, DNA damage and repair, and therapeutics are potentially of great relevance to the practicing dermatologist and deserve increased emphasis within our specialty.

Epidermal cell kinetics in the nude mouse

The epidermal cell kinetics in nude mice is investigated by determining the mitotic rate and the mitotic count, the H3Tdr labelling index, and the proportion of basal cells in the different cell cycle phases by flow cytometry. The mitotic duration was calculated. The parameter values of the epidermal cell kinetics of the nude mouse are largely similar to those of the hairless mouse.

Epidermal cell kinetics and the fraction of tritium-labelled mitoses

Epidermal cell kinetics and the fraction of tritium-labelled mitoses

A new look at epidermal cell kinetics in psoriasis and other dermatoses

A new look at epidermal cell kinetics in psoriasis and other dermatoses