Maintenance of distinct melanocyte populations in the interfollicular epidermis

Hair follicles and sweat glands are recognized as reservoirs of melanocyte stem cells (MSCs). Unlike differentiated melanocytes, undifferentiated MSCs do not produce melanin. They serve as a source of differentiated melanocytes for the hair follicle and contribute to the interfollicular epidermis upon wounding, exposure to ultraviolet irradiation or in remission from vitiligo, where repigmentation often spreads outwards from the hair follicles. It is unknown whether these observations reflect the normal homoeostatic mechanism of melanocyte renewal or whether unperturbed interfollicular epidermis can maintain a melanocyte population that is independent of the skin's appendages. Here, we show that mouse tail skin lacking appendages does maintain a stable melanocyte number, including a low frequency of amelanotic melanocytes, into adult life. Furthermore, we show that actively cycling differentiated melanocytes are present in postnatal skin, indicating that amelanotic melanocytes are not uniquely relied on for melanocyte homoeostasis.     

Measurement of protein metabolism in epidermis and dermis

We found that, in the rabbit ear, the dermal protein contains 75.5% of cutaneous phenylalanine and 97.9% of cutaneous proline; the remaining 24.5% of phenylalanine and 2.1% of proline are in the epidermal protein. This finding led us to develop two novel models that use phenylalanine and proline tracers and the rabbit ear to quantify protein kinetics in the epidermis and dermis. The four-pool model calculates the absolute rates of protein kinetics and amino acid transport, and the two-pool model calculates the apparent rates of protein kinetics that are reflected in the blood. The results showed that both epidermis and dermis maintained their protein mass in the postabsorptive state. The rate of epidermal protein synthesis was 93.4 +/- 37.6 mg x 100 g(-1) x h(-1), which was 10-fold greater than that of the dermal protein (9.3 +/- 5.8 mg x 100 g(-1) x h(-1)). These synthetic rates were in agreement with those measured simultaneously by the tracer incorporation method. Comparison of the four-pool and two-pool models indicated that intracellular cycling of amino acids accounted for 75 and 90% of protein kinetics in the dermis and epidermis, respectively. We conclude that, in the skin, efficient reutilization of amino acids from proteolysis for synthesis enables the maintenance of protein mass in the postabsorptive state.     

Characteristics on staining of leukocytes in experimental animals]

The staining characteristics of the peripheral blood cells from mouse, rat, guinea pig, rabbit, dog, marmoset and monkey were studied. In marmoset, it is easy to distinguish neutrophils from eosinophils by using the phosphate-buffered solution of pH 5 or 6. It was found in the special staining methods that neutrophil granules showed intense peroxidase and Sudan black B reactions in marmoset in comparison with those in the other species of experimental animals. Neutrophil granules rabbit was, however, intensely stained with esterase and acid phosphatase.     

Pharmacokinetics of lincomycin in experimental animals]

When administered parentally, lincomycin satisfactorily penetrated into the organs and tissues of experimental animals. Pronounced tropism of the antibiotic in the bone tissue was observed which provided its recommendation for the treatment of osteomyelitis. The antibiotic was excreted with urine and bile.     

Role of melanoma chondroitin sulphate proteoglycan in patterning stem cells in human interfollicular epidermis

Human interfollicular epidermis is renewed by stem cells that are clustered in the basal layer in a patterned, non-random distribution. Stem cells can be distinguished from other keratinocytes by high expression of beta1 integrins and lack of expression of terminal differentiation markers; they divide infrequently in vivo but form actively growing colonies in culture. In a search for additional stem cell markers, we observed heterogeneous epidermal expression of melanoma chondroitin sulphate proteoglycan (MCSP). MCSP was expressed by those keratinocytes with the highest beta1 integrin levels. In interfollicular epidermis, expression was confined to non-cycling cells and, in culture, to self-renewing clones. However, fluorescence-activated cell sorting on the basis of MCSP and beta1 integrin expression gave no more enrichment for clonogenic keratinocytes than sorting for beta1 integrins alone. To interfere with endogenous MCSP, we retrovirally infected keratinocytes with a chimera of the CD8 extracellular domain and the MCSP cytoplasmic domain. CD8/MCSP did not affect keratinocyte proliferation or differentiation but the cohesiveness of keratinocytes in isolated clones or reconstituted epidermal sheets was greatly reduced. CD8/MCSP caused stem cell progeny to scatter without differentiating. CD8/MCSP did not alter keratinocyte motility but disturbed cadherin-mediated cell-cell adhesion and the cortical actin cytoskeleton, effects that could be mimicked by inhibiting Rho. We conclude that MCSP is a novel marker for epidermal stem cells that contributes to their patterned distribution by promoting stem cell clustering.     

Mechanobiological dysregulation of the epidermis and dermis in skin disorders and in degeneration

During growth and development, the skin expands to cover the growing skeleton and soft tissues by constantly responding to the intrinsic forces of underlying skeletal growth as well as to the extrinsic mechanical forces from body movements and external supports. Mechanical forces can be perceived by two types of skin receptors: (1) cellular mechanoreceptors/mechanosensors, such as the cytoskeleton, cell adhesion molecules and mechanosensitive (MS) ion channels, and (2) sensory nerve fibres that produce the somatic sensation of mechanical force. Skin disorders in which there is an abnormality of collagen [e.g. Ehlers–Danlos syndrome (EDS)] or elastic (e.g. cutis laxa) fibres or a malfunction of cutaneous nerve fibres (e.g. neurofibroma, leprosy and diabetes mellitus) are also characterized to some extent by deficiencies in mechanobiological processes. Recent studies have shown that mechanotransduction is crucial for skin development, especially hemidesmosome maturation, which implies that the pathogenesis of skin disorders such as bullous pemphigoid is related to skin mechanobiology. Similarly, autoimmune diseases, including scleroderma and mixed connective tissue disease, and pathological scarring in the form of keloids and hypertrophic scars would seem to be clearly associated with the mechanobiological dysfunction of the skin. Finally, skin ageing can also be considered as a degenerative process associated with mechanobiological dysfunction. Clinically, a therapeutic strategy involving mechanoreceptors or MS nociceptor inhibition or acceleration together with a reduction or augmentation in the relevant mechanical forces is likely to be successful. The development of novel approaches such as these will allow the treatment of a broad range of cutaneous diseases.     

Epidermal proliferation characteristics are similar in the pilary canal of mouse hair follicles and in interfollicular epidermis

Proliferation characteristics of basal cells in the pilary canal of resting hair follicles were investigated and compared with corresponding parameters in interfollicular epidermis of hairless mice. The mitotic rates had similar 24-h means at both locations. Distinct circadian rhythms which showed phasing and amplitudes similar to that in interfollicular epidermis, were demonstrated by the 3H-TdR labelling index, the mitotic rate and the mitotic index. Influx of cells to and efflux of cells from the S phase were measured in the early morning and in the evening by a 3H-TdR double labelling method. The influx values were similar at both times of both locations. The efflux values recorded in the morning were more than twice the values seen in the evening in both the pilary canal and in interfollicular epidermis. The epidermal motitic rate in the pilary canal was depressed by epidermal extracts, and increased after adhesive tape stripping in the same way as in interfollicular epidermis. The results indicate no heterogeneity in cell proliferation characteristics between the two locations, and suggest that similar mechanisms are responsible for maintainance of growth equilibrium at both sites.     

Keratin metabolism in the epidermis and hair of mice with experimental diabetes]

An animal trial was performed using mice with streptozotocine-induced diabetes, with investigation of velocity of prekeratin and keratin biosynthesis and degradation using 14C-glycine, and evaluation of the content of -SH and -S-S groups in epidermal prekeratin. It has been found out that velocity of epidermal prekeratin and keratin in diabetic animals is higher than that in healthy group. SS and SH groups ratio in prekeratin in diabetic animals is 10 times as high as that in the control group. In the hair of diabetic mice an increased keratin turnover was observed as compared with the norm. The data testify that experimental diabetes manifests itself in increased intensity of keratin metabolism in epidermis and hair. These results may be used as the criteria in elaboration of non-invasive methods for diabetes diagnosis.