|
|||||||||||||||||
Organ
of the
|
GD Society for Dermopharmacy |
||||||||||||||||
Author Article Th. Förster, C. Jassoy, D. Petersohn, K. Schlotmann und M. Waldmann-Laue Systematic evaluation of new active substances and cosmetics Paper on the occasion of the 4th annual meeting of the Gesellschaft für Dermopharmazie (Dermopharmacy Society) in Freiburg on 24 May 2000
Phased Test Hierarchy
We can certainly regard skin models, and especially the in vitro skin-equivalent model, as worthy of inclusion among these revolutionary achievements. A skin-equivalent model is artificial skin that has been cultured in the laboratory. The structure of the artificial skin is similar to that of human skin, making it eminently suitable for cosmetics research purposes. Using skin ageing as an example, I will show how skin models can be meaningfully integrated into a test hierarchy for active substances and finished products in the cosmetics sector. This test hierarchy
involves three stages. The first is substance screening, involving fast
tests on simple skin cell cultures. At this stage a large number of potential
active substances are eliminated on the grounds that they are inadequately
effective. In stage 2 of the test hierarchy, the new in vitro skin models
are used to study cosmetic effectiveness under realistic conditions of
use. In stage 3, dermatological trials are carried out with a panel of
users. environment-related skin-ageing Aged skin can be recognized immediately by the presence of wrinkles, flaccidity and actinic keratosis. A histological section reveals some of the causes of these symptoms of ageing (Fig. 1 a and b).
Fig. 1
a and b with fibroblast cultures Fibroblasts can be
taken from the dermis and cultivated as a simple cell system in a monolayer
culture. These simple fibroblast cultures are suitable for investigating
harmful influences on fibroblasts and as a simple test system for screening
various cosmetic active substances with regard to their effectiveness
in influencing metabolic processes.
Effect of various peptide extracts on fibroblast metabolism Various peptide extracts were tested on 2- and 4-day-old fibroblast cultures to determine whether they promote proliferation and protein synthesis (Fig. 2). An extremely promising
soybean peptide 2 was identified from a range of different wheat, milk
and soybean peptides. It caused 20% activation of the protein synthesis
(4).
These harmful enzymes cleave collagen, the main component of the skin's connective tissue and thus cause wrinkles to form prematurely (5,6). A reduction in MMP-1 synthesis after the skin has been exposed to sunlight is thus a primary objective in the development of anti-ageing-products. An ideal anti-ageing substance is one that, even at a low concentration, inhibits the expression of collagenase MMP-1. The production of mRNA is the first and thus the most important step in the MMP-1 synthesis. Active substances that have an effect on mRNA production thus also automatically have an effect on the amount of protein and the enzyme activity of MMP-1. The quantification of the MMP-1 synthesis in sun-irradiated dermal fibroblasts was carried out by determining the amount of MMP-1 mRNA synthesized in the Northern Blot (5). As expected, the irradiation
of fibroblasts with simulated sunlight (UVA dosage of 10 J/cm2) caused
strong induction of the synthesis of MMP-1 mRNA. Retinyl palmitate and
antioxidants such as propyl gallate reduce the sunlight-induced expression
of MMP-1 very effectively by around 80% or 50 to 75% and thus prevent
photoageing of the skin.
New testing opportunities
Simple fibroblast
cultures enable active substances to be screened quickly and cost-efficiently
for a number of basic effects. For various reasons, however, the results
obtained with this simple cell model are not directly applicable to the
real in vivo situation. Simple skin cell cultures do not have a skin barrier
(stratum corneum), so the active substances are made available in the
culture medium. The concentration relationships are therefore completely
different and are not applicable to real application situations. Moreover,
only water-soluble or water-solubilized substances can be tested, but
not complex cream formulations. Finally it is only possible to study one
type of skin cell in this simple model, so interactions between stratum
corneum, epidermis and dermis and the corresponding skin cell types cannot
be taken into consideration.
After it has been
cultured, the skin-equivalent model lives for at least another 4 weeks,
during which time it remains virtually unchanged. During this time ageing
experiments can be carried out, e.g., with UV irradiation or ozone, or
tests of topical skin-protection or skin-care treatments. The main advantage
is that, just like normal human skin, these skin models have a stratum
corneum. The active substances can therefore be applied under realistic
conditions in a cream or gel basis. The active substances penetrate through
the stratum corneum into the skin, where they take effect.
After
it has been cultured, the skin-equivalent model lives for at least another
4 weeks, during which time it remains virtually unchanged. During this
time ageing experiments can be carried out, e.g., with UV irradiation
or ozone, or tests of topical skin-protection or skin-care treatments.
The main advantage is that, just like normal human skin, these skin models
have a stratum corneum. The active substances can therefore be applied
under realistic conditions in a cream or gel basis. The active substances
penetrate through the stratum corneum into the skin, where they take effect.
In view of the encouraging biochemical results for collagen- and GAG stimulation by the cytokine cream, it can be asked whether this biological effectiveness also results in a reduction in wrinkles. Clearly this is of interest to consumers. In a controlled study, 30 volunteers used the cytokine cream for a period of 4 weeks. At the start and end of the study the wrinkle status of each volunteer's skin was determined by FOITS (Fast Optical In-vivo topography of the skin). The wrinkle depth (roughness depth Rz) decreased by 16% on average. The individual topographic plots clearly show the evening out of the deep wrinkles in the critical zone around the corner of the eye (Fig. 7). The smoothing of the
wrinkles after treatment with the cream can be seen clearly. As a result,
in the subsequent questionnaire the volunteers rated skin smoothness (1.9),
skin structure (2.2), skin suppleness (1.9) and skin tautness (2.4) very
highly.
References 1. C. R. Lovell, K. A. Smolenski, V. C. Duance, N. D. Light, S. Young, M. Dyson, Type I and III Collagen Content and Fibre Distribution in Normal Human Skin during Ageing, Brit. J. Dermatol. 117 (1987) 419-428 2. J. Varani, J. R. L. Warner, M. Gharaee-Kermani, S. H. Phan, S. Kang, J. Chung, Z. Wang, S. C. Datta, G. J. Fisher, J. J. Voorhees, Vitamin A antagonizes decreased cell growth and elevated collagen-degrading matrix metalloproteinases and stimulates colllagen accumulation in naturally aged human skin, J. Invest. Dermatol. 114 (2000) 480-486 3. S. Shuster, M. M. Black, E. McVitie, The influence of age and sex on skin thickness, skin collagen and density, Brit. J. Dermatology 93 (1975), 639-643 4. C. Augustin, V. Frei, E. Perrier, A. Huc, O. Damour, An in vitro selection of new cosmetic active compounds: From screening tests on monolayered fibroblast culture to efficiency study on 3-D dermal equivalent, J. Appl. Cosmetol. 15 (1997), 1-11 5. K. Scharffetter, M. Wlaschek, A. Hogg, K. Bolsen, A. Schothorst, G. Goerz, T. Krieg, G. Plewig, UVA irradiation induces collagenase in human dermal fibroblasts in vitro and in vivo, Arch. Dermatol. Res. 283 (1991), 506-511 6. G. J. Fisher, S. C. Datta, H. S. Talwar, Z. Wang, J. Varani, S. Kang, J. J. Voorhees, Nature 379 (1996) 335-339 7. C. Augustin, C. Collombel, O. Damour, Use of in vitro dermal equivalent and skin equivalent kits for evaluating cutaneous toxicity of cosmetic products, In Vitro Toxicology 10 (1997), 23-31 8. V. Andre-Frei, E. Perrier, C. Augustin, O. Damour, P. Bordat, K. Schumann, T. Förster, M. Waldmann-Laue, A comparison of biological activities of a new soya biopeptide studied in an in vitro skin equivalent model and human volunteers, Int. J. Cosmet. Sci. 21 (1999), 299-311 top |
|||||||||||||||||
March 2001 | Copyright © 2000 - 2014 Institute for Dermopharmacy GmbH. Contact: webmaster@gd-online.de |