In vivo confocal microscopy of human skin: a new design for cosmetology and dermatology.

In-depth exploration of cellular structures in living human skin in situ is possible with the tandem scanning microscope (TSM). However, the rigid design of the microscope limited observations to the arms, hands, and fingers. A mobile version allowing the investigation of any parts of the body has been designed. The head containing the Nipkow disk and the optical path were the only part saved from the original TSM. This prototype can be used to observe, in real time, the different skin structures down to a depth of 200 microns and to measure the thickness of the different layers with micron precision level. The hydration of the stratum corneum (SC) could be assessed. For example, lengthy immersion of the hand in water led to an increase in SC thickness without affecting that of the living epidermis. Occlusive patch tests also showed that water and, even more so. propylene glycol, led to transient swelling of the SC. In dermatology, the example of psoriasis illustrated the value of the TSM for describing, measuring, and assessing pathologic skin changes. The availability of this noninvasive method for observing changes with time in a given skin site should prove useful for monitoring treatment efficacy. This tool opens up new insight for the investigation of cutaneous pathophysiology.

Three-dimensional microscopic biopsy of in vivo human skin: a new technique based on a flexible confocal microscope.

A new noninvasive microscopic technique of three-dimensional optical biopsy from in vivo human skin based on real-time confocal microscopy and computer reconstruction is demonstrated. A tandem scanning confocal microscope is a prototype of a mobile, flexible design for the in-depth microscopic exploration of the skin on the human body. The various skin layers were observed in real-time, at the subcellular level down to a depth of 200 microns with a vertical resolution of 2 microns. Rapid video recording of the Z-series through the ventral aspect of the forearm avoided shifts caused by subject movement and blood flow pulsations. Two video frames were averaged, and the average was digitized, providing a stack of 64 optical sections in 1-micron vertical steps. Three-dimensional reconstructions of in vivo human skin were obtained with sets of orthogonal slices, and slices at arbitrary planes through a volume containing the stack of slices. This method clearly shows the spatial relationships between the different cell layers. The use of orthogonal cutting planes is preferred because of its analogy with classical vertical sections of histopathology. Linear structures (surface lines) within the stratum corneum are described and their global orientations were determined by the use of Fourier transform analysis. En face optical sections constitute unusual views of this tissue, since typical pathohistological studies are based on sagittal (vertical) slices. The noninvasive optical microscopic technique provides a three-dimensional optical biopsy of in vivo human skin.

Mechanism of action of a lipophilic derivative of salicylic acid on normal skin.

BACKGROUND/AIMS: Keratolytics are agents used for a very long period of time to improve various skin disorders such as acne, hyperkeratoses, ichtyose etc. Very little is known about their mechanism of action on healthy skin. On man, the chronic application of a cosmetic cream containing a lipophilic derivative of Salicyclic acid (LSA) markedly improves the aspect and texture of the skin. Different methods were used to investigate the mechanisms of action of this new compound, compared to salicyclic acid. METHODS: Both non-invasive and histologic methods were used on the dorsal forearm of human volunteers treated with the products. Concerning the non-invasive methods, TEWL, silflo replica and confocal microscopy were used. On shave biopsies, various histometric parameters were measured by image analysis after different staining. The use of antibody MIB-1 reacting with the proliferating nuclear antigen Ki 67 allows one to measure the epidermis proliferation index. RESULTS: Compared to the excipient alone, presence of LSA 1% improves smoothness and firmness of the skin. The appearance in terms of clearness and healthy complexion is also improved. The thickening of all the living epidermis layers is obtained by both histometric measurement and confocal microscopy. This acanthosis is only recorded on the LSA-treated zones. The Ki 67 labelling study shows that LSA significantly increases the skin proliferation index. CONCLUSIONS: Salicylic acid, and more markedly its lipophilic derivative (LSA), appear to have a significative effect on the renewal of the living epidermis. This probably explains the cosmetic improvement of the skin obtained after a 1-month treatment with a cream containing this new molecule.

Effect of colchicine on atherosclerosis. III. Study of dermal elastic fibers by quantitative histochemistry, automated image analysis.

Computerized automatic-image analytical procedure was applied on dermal biopsies stained for elastin by a new procedure giving a completely white background and staining only the elastic fiber system. In arteriosclerotic hypertensive patients, a 3-4 months' treatment with 1 mg colchicine per day resulted in a significant (60-80%, p less than 0.01) increase of the dermal elastic fiber density both in the superficial papillary dermis and in the deep dermis. This result shows that the age-dependent increase of elastic fibers can be influenced by pharmacological means. The inhibition by colchicine of the synthesis and secretion of the fibroblast-derived metalloelastase-type protease could be a plausible explanation of this finding.

A selective histochemical method for the quantitative estimation of elastic fibers by computerized morphometric analysis. Effect of colchicin treatment.

A morphometric technique is reported that uses a new selective staining of the elastic system fibers in skin biopsy specimens to facilitate the quantitative evaluation of the volume fraction occupied by these elastic fibers in the tissue. The study of elastic fibers in the dermis of 30 patients, before and after six months of treatment with Colchicin, was carried out with a Quantimet 720 system. Preelastic (oxytalan and elaunin) fibers and mature elastic fibers were quantitated separately. Compared to the average volume fraction (surface occupied by the elastic fibers) before treatment with Colchicin (1.449 +/- 0.64%), the mean values after treatment were significantly increased (2.076 +/- 0.61%). The same results were found for the preelastic fibers: 0.807 +/- 0.51% before treatment and 1.025 +/- 0.54% after treatment. These results demonstrate the advantages of our monochromatic staining method for automatic quantitation of elastic fibers as well as the possibilities of the quantitative study of the elastic fibers in human dermis. This methodology should be applicable to other inherited or acquired diseases affecting skin elastic fibers as well as to other tissues containing elastic fibers.