Extended applications of the carbon dioxide laser for skin deepithelialization

The efficacy of the carbon dioxide laser for skin deepithelialization in humans was initially demonstrated using reduction mammaplasty as a clinical model. Twenty-two additional successful cases including use of this technique as an adjunct for other flap transpositions and overgrafting of skin grafts are described. These results document a role for the carbon dioxide laser in skin deepithelialization, where it provides an advantage when rigid skin immobilization is impossible or if there would be a risk to underlying structures if more conventional approaches were used.     

Morphogenetic interactions between rotated skin cuffs and underlying stump tissues in regenerating axolotl forelimbs

Rotation of skin cuffs 180° around the longitudinal axis of the underlying tissues in the axolotl forelimb results in a high percentage of multiple regenerates after amputation through the rotated skin. Similar results occur after rotation of only the anteroposterior (A-P) axis of the skin. Rotation of only the proximodistal (Pr-Ds) axis of the skin results in normal regenerates whereas dorsoventral (D-V) axial skin rotation results in single regenerates with some disturbances in symmetry. Rotation of anterior or posterior half cuffs of skin produces results similar to those obtained after A-P rotation of full skin cuffs, and rotation of dorsal or ventral skin halves duplicates the results obtained by rotating full skin cuffs about the D-V axis. Skin cuffs rotated for periods from 6 months to over 2 years before amputation are also capable of causing multiple regenerates to form. No significant difference in the percentage of multiple regenerates was seen after skin rotation and limb amputation through shoulder, upper arm, and forearm levels. X-Radiation (4000 r) of either the skin or underlying tissues before skin rotation resulted in single regenerates after amputation. If a strip of normal skin was turned perpendicularly to the long axis of the irradiated underlying stump tissues, the regenerative response was blocked. In some of the above experiments, regenerates with longitudinally duplicated upper arm and forearm segments appeared. It is postulated that normally both the skin and the underlying limb tissues can influence morphogenesis during regeneration and that they work in harmony. In contrast, rotated skin and the underlying tissues each exert a morphogenetic influence upon the regenerating limb, and the regenerate is not able to integrate these disharmonious influences. This is reflected in the highly abnormal morphology of the regenerates. The nature of the morphogenetic influence disrupted by skin rotation is not yet known.     

The use of flow cytometry and cell sorting in a human colon carcinoma model

We investigated the growth characteristics of a human colon carcinoma cell line, WiDr, grown in culture flasks and on chick embryonic skin (CES). WiDr cells labeled in vitro with bromodeoxyuridine (BrdU) and analyzed by combined propidium iodide/Hoechst 33258 fluorescence showed evidence of more BrdU incorporation in early S phase as compared to late S phase. When inoculated on the CES, WiDr cells multiplied and invaded the underlying skin. Morphologic examination showed that with extended culture WiDr cells on the CES undergo progressive structural organization with the development of acini and basal lamina, structures similar to those in in vivo tumors. WiDr cells were labeled with monoclonal antibody to carcinoembryonic antigen (CEA) and the brightest 2% of the population was sorted. When subsequently grown on the CES, the sorted cells formed significantly more acinar structures at 3 and 6 days of culture than an unsorted population grown for a comparable time.     

Axial pattern composite prefabrication of high-density porous polyethylene: experimental and clinical research

Currently, various alloplastic materials are being used for reconstruction of three-dimensional structures, and high-density porous polyethylene is so far the best and the most commonly used material. Various indications for high-density porous polyethylene have been defined for closure of craniofacial defects, correction of congenital anomalies, and aesthetic augmentations. A common property of various studies published so far is that after being fixed to the bone or underlying structures, high-density porous polyethylene has been covered primarily or by skin flaps. For reconstruction of complex three-dimensional structures such as the ear and nose, the success of current methods is limited by the thinness and pliability of the skin flap. In this study, the authors' aim was to investigate the graftability of high-density porous polyethylene after prefabrication with an axial pedicle and to explore possible clinical applications in light of the new data obtained. In the experimental study, three-dimensional implants (rectangular prism) carved from high-density porous polyethylene were prefabricated using bilateral superficial epigastric arteries and veins of 25 New Zealand rabbits. After a waiting period of 2 to 6 weeks in five groups, control samples were obtained and the prefabricated implants that had been left in place were directly grafted. The results showed that high-density porous polyethylene was vascularized 75 percent after 4 weeks and 90 percent after 5 weeks, and 95 percent of the grafts had survived after 8 weeks. In the clinical study, three nose defects, three ear defects, and one hard palate defect in seven patients ranging in age from 21 to 72 years were reconstructed using the same method. High-density porous polyethylene has been prefabricated and directly grafted for the very first time on a clinical basis. No serious complications have been observed, except for minimal graft loss in two patients. It is obvious that full-thickness skin grafts that are thinner than flaps will adapt better to the fine details of high-density porous polyethylene and will highly increase the detail obtained in the reconstruction of three-dimensional defects.     

Combined in vivo confocal Raman spectroscopy and confocal microscopy of human skin

In vivo confocal Raman spectroscopy is a noninvasive optical method to obtain detailed information about the molecular composition of the skin with high spatial resolution. In vivo confocal scanning laser microscopy is an imaging modality that provides optical sections of the skin without physically dissecting the tissue. A combination of both techniques in a single instrument is described. This combination allows the skin morphology to be visualized and (subsurface) structures in the skin to be targeted for Raman measurements. Novel results are presented that show detailed in vivo concentration profiles of water and of natural moisturizing factor for the stratum corneum that are directly related to the skin architecture by in vivo cross-sectional images of the skin. Targeting of skin structures is demonstrated by recording in vivo Raman spectra of sweat ducts and sebaceous glands in situ. In vivo measurements on dermal capillaries yielded high-quality Raman spectra of blood in a completely noninvasive manner. From the results of this exploratory study we conclude that the technique presented has great potential for fundamental skin research, pharmacology (percutaneous transport), clinical dermatology, and cosmetic research, as well as for noninvasive analysis of blood analytes, including glucose.     

Noninvasive diagnosis of melanoma with tensor decomposition-based feature extraction from clinical color image

We propose a method for feature extraction from clinical color images, with application in classification of skin lesions. Proposed feature extraction method is based on tensor decomposition of the clinical color image of skin lesion. Since color image is naturally represented as a three-way tensor, it is reasonable to use multi-way techniques to capture the underlying information contained in the image. Extracted features are elements of the core tensor in the corresponding multi-way decomposition, and represent spatial-spectral profile of the lesion. In contrast to common methods that exploit either texture or spectral diversity of the tumor only, the proposed approach simultaneously captures spatial and spectral characteristics. The procedure is tested on a problem of noninvasive diagnosis of melanoma from the clinical color images of skin lesions, with overall sensitivity 82.1% and specificity 86.9%. Our method compares favorably with the state of the art results reported in the literature and provides an interesting alternative to the existing approaches.     

Molecular mechanisms of action of different concentrations of ethanol in water on ordered structures of intercellular lipids and soft keratin in the stratum corneum

Ethanol (EtOH) is one of the bases in topically applied medicines that promote the skin permeation of drugs. Although the effects of EtOH have been attributed to structural modifications in the stratum corneum, the underlying mechanisms, especially the influence of different concentrations of EtOH, have not been examined extensively. Structural modifications in the stratum corneum of hairless mouse due to the application of EtOH/water mixture were herein investigated at the molecular level using synchrotron X-ray diffraction. The results revealed that all EtOH concentrations examined greatly modified the short lamellar structures containing the aqueous layer in intercellular lipids and the structure of keratin fibrils in corneocytes, which can take up hydrophilic compounds. However, the long lamellar and the hydrocarbon-chain packing structures were unaffected by EtOH. Changes to the short lamellar structures were not proportional to the concentration of EtOH. However, the keratin fibril structures changed gradually with increasing EtOH concentration. The X-ray diffraction experiments enabled the effects of different EtOH concentrations on the morphology of the stratum corneum to be assessed by using a number of experimental samples to avoid variations due to individual differences. The results indicated that alterations to the short lamellar structures appeared to be related to the skin permeability of drugs with the application of EtOH/water mixture, and monotonous structural changes in the keratin fibrils with an increase in EtOH concentration may contribute to this permeation as supplement. These results will be useful for the development of new drug formulations containing EtOH.     

Touch sense

Cutaneous mechanoreceptors are localized in the various layers of the skin where they detect a wide range of mechanical stimuli, including light brush, stretch, vibration and noxious pressure. This variety of stimuli is matched by a diverse array of specialized mechanoreceptors that respond to cutaneous deformation in a specific way and relay these stimuli to higher brain structures. Studies across mechanoreceptors and genetically tractable sensory nerve endings are beginning to uncover touch sensation mechanisms. Work in this field has provided researchers with a more thorough understanding of the circuit organization underlying the perception of touch. Novel ion channels have emerged as candidates for transduction molecules and properties of mechanically gated currents improved our understanding of the mechanisms of adaptation to tactile stimuli. This review highlights the progress made in characterizing functional properties of mechanoreceptors in hairy and glabrous skin and ion channels that detect mechanical inputs and shape mechanoreceptor adaptation.     

Touch sense: Functional organization and molecular determinants of mechanosensitive receptors

Cutaneous mechanoreceptors are localized in the various layers of the skin where they detect a wide range of mechanical stimuli, including light brush, stretch, vibration and noxious pressure. This variety of stimuli is matched by a diverse array of specialized mechanoreceptors that respond to cutaneous deformation in a specific way and relay these stimuli to higher brain structures. Studies across mechanoreceptors and genetically tractable sensory nerve endings are beginning to uncover touch sensation mechanisms. Work in this field has provided researchers with a more thorough understanding of the circuit organization underlying the perception of touch. Novel ion channels have emerged as candidates for transduction molecules and properties of mechanically gated currents improved our understanding of the mechanisms of adaptation to tactile stimuli. This review highlights the progress made in characterizing functional properties of mechanoreceptors in hairy and glabrous skin and ion channels that detect mechanical inputs and shape mechanoreceptor adaptation.     

Biophysics of thermal lesions

Experiments on rats showed that thermal irradiation caused a more marked elevation of subcutaneous temperature at the area of the skin separated from the underlying tissues by a felt plate than at the skin area separated from the surrounding and underlying tissues and immediately sutured to its place or the area of the uninjured skin. The authors believe that the data obtained indicated that the blood flow played no significant role in leading the heat from the skin in the action of the radial heat on it. Of much greater significance is the capacity of the underlying tissues to lead off and to accumulate heat.     

Monoclonal autoantibodies to different epithelial structures of the thymus gland obtained by the immunization with streptococcal group A antigens

By the indirect immunofluorescence method it was shown to which epithelial thymus structures monoclonal antibodies (mAT) reacting with the different epidermal structures are directed. These mAT related to the autoantibodies were obtained earlier, as a result of lymphoid cells polyclonal activation, by the immunization of BALB/c mice with streptococcal group A nonspecific protein antigens of the cell wall. It was shown that mAT A6/1, reacting with the basal layer of the skin epithelium are directed to the epithelium of the cortical and medullar thymus zones, which is regarded as the so called endocrinal epithelium. These mAT, by the study with immunoblotting method, react with the protein of mV SOkD, B5/1 mAT to the skin epithelium, on the thymus sections react with the single cells around the Hassel bodies.     

Simultaneous marker-assisted selection for multiple traits in autogamous crops

A method is presented for the selection of parents with the aim of obtaining improved genotypes in the progeny of a cross. The procedure is designed to select in several unrelated traits simultaneously and is based on the selection of molecular markers that are linked to QTLs. The method was compared with conventional phenotypic selection in simulation experiments for a number of genetic structures underlying the traits and several types of parental populations. Although the method in general provides good results, some of the underlying assumptions may be violated quite easily, thereby reducing the applicability of the procedure in practice. Received: 10 September 1999 / Accepted: 24 August 2000     

Operations on the middle ear and mastoid

From 1953 to 1957, inclusive, 373 patients were operated on for chronic mastoid disease at the Mayo Clinic. Mainly because of the extensive use of magnifying glasses and microscopes, through these years, closer attention has been given to the preservation or improvement of hearing by meticulous procedures on the structures of the middle ear and the use of skin grafts. It is stressed, however, that it is extremely important to eradicate the underlying disease. Illustrative cases to represent different types of procedures are presented. The author recommends closer attention to the condition of the round window in the future.     

OPERATIONS ON THE MIDDLE EAR AND MASTOID—Changing Aspects of Treatment in Chronic Active Disease

From 1953 to 1957, inclusive, 373 patients were operated on for chronic mastoid disease at the Mayo Clinic. Mainly because of the extensive use of magnifying glasses and microscopes, through these years, closer attention has been given to the preservation or improvement of hearing by meticulous procedures on the structures of the middle ear and the use of skin grafts. It is stressed, however, that it is extremely important to eradicate the underlying disease. Illustrative cases to represent different types of procedures are presented. The author recommends closer attention to the condition of the round window in the future.     

Expression of PTHrP and the PTH/PTHrP receptor in growing red deer antler

Antler growth is highly co-ordinated, so that trabecular bone and antler skin (velvet) develop together, at a rapid rate and in a manner reminiscent of their development in the fetus. Parathyroid hormone-related peptide (PTHrP) is expressed in both bone and skin, and is therefore a candidate to effect co-ordination between these tissues. The aim of this study was to localize the expression of PTHrP and its principal receptor, the parathyroid hormone/parathyroid hormone-related peptide receptor (PTH/PTHrPR), in antler ("spiker") of one-year-old red deer. Using immunohistochemistry and in situ hybridization, intense and overlapping expression of PTHrP and its receptor was seen in developing osseocartilaginous structures and in the underlying layers of velvet epidermis. PTHrP was located on both the cell surface and within the nuclei. Our results strongly suggest that PTHrP, acting via the PTH/PTHrPR and possibly other intracrine mechanisms, plays a central role in the co-ordinated regulation of cell division and differentiation of developing antler bone and skin.     

Numerical simulation of microneedles' insertion into skin

Microneedles have recently received much attention as a novel way for transdermal drug delivery. In this paper, a numerical simulation of the insertion process of the microneedle into human skin is reported using the finite element method. A multilayer skin model consisting of the stratum corneum, dermis and underlying hypodermis has been developed. The effective stress failure criterion has been coupled with the element deletion technique to predict the complete insertion process. The numerical results show a good agreement with the reported experimental data for the deformation and failure of the skin and the insertion force. The influences of the mechanical properties of the skin and the microneedle geometry (e.g. tip area, wall angle and wall thickness) on the insertion force are discussed. The numerical results are helpful for the optimum design of the microneedles for the transdermal drug delivery system.     

Numerical simulation of microneedles' insertion into skin

Microneedles have recently received much attention as a novel way for transdermal drug delivery. In this paper, a numerical simulation of the insertion process of the microneedle into human skin is reported using the finite element method. A multilayer skin model consisting of the stratum corneum, dermis and underlying hypodermis has been developed. The effective stress failure criterion has been coupled with the element deletion technique to predict the complete insertion process. The numerical results show a good agreement with the reported experimental data for the deformation and failure of the skin and the insertion force. The influences of the mechanical properties of the skin and the microneedle geometry (e.g. tip area, wall angle and wall thickness) on the insertion force are discussed. The numerical results are helpful for the optimum design of the microneedles for the transdermal drug delivery system.     

Comparative biochemical study of N-linked glycans from skin of a squid, Todarodes pacificus

For comparative biochemical interest, we analyzed the structures of N-glycans in a squid belonging to the Lophotrochozoa, one of the protostome clades. N-Glycans were prepared from squid skin by hydrazinolysis and re-N-acetylation followed by fluorescent tagging with 2-aminopyridine. The labeled N-glycans were purified, and their structures were determined by the two-dimensional HPLC mapping method combined with glycosidase digestions and mass spectrometry. We found that high mannose-type glycans, paucimannose-type glycans and complex-type glycans with a type-1 structure (Galbeta1-3GlcNAc) were dominant in squid skin. The complex-type glycans detected in the squid were similar to those in vertebrates, but have not yet been found in the Ecdysozoa, which is another protostome clade. However, paucimannose-type glycans are commonly found in the Ecdysozoa. Thus, the N-glycan structures of the squid belonging to the Lophotrochozoa have features common to those in vertebrates and the Ecdysozoa including insects and nematodes.     

Conditional ablation of integrin alpha-6 in mouse epidermis leads to skin fragility and inflammation

Hemidesmosomes (HDs) are essential anchorage junctions which mediate the firm attachment of epithelia to the underlying basement membranes, of which one main component is the integrin α6β4. These specific junctions are also able to trigger signalling pathways, via the recruitment and interactions of signalling molecules with HD components such as the cytoplasmic tail of the β4 integrin or the plakin plectin. HDs must also assemble and disassemble depending on the tissue context for example during tissue remodelling. Alterations of HD components or their loss result in skin blistering disorders known as epidermolysis bullosa. Since mice lacking integrin α6 die at birth with severe skin blistering, we have produced a mouse line in which epidermal deletion of integrin α6 can be controlled by tamoxifen injection. We observed that the deletion was mosaic, but that hairless skin such as ears, tails and paws were affected and showed chronic inflammation associated with hyperproliferation, and expression of laminin-111. Interestingly, two cytokines, amphiregulin and epiregulin, previously found increased in integrin α6 deficient cultured keratinocytes, were also increased here in the affected skin. In detached areas, we validate clearly that the absence of integrin α6 leads to a delocalisation of plectin, and the complete disappearance of HD structures.     

Type I Transglutaminase Accumulation in the Endoplasmic Reticulum May Be an Underlying Cause of Autosomal Recessive Congenital Ichthyosis

Type I transglutaminase (TG1) is an enzyme that is responsible for assembly of the keratinocyte cornified envelope. Although TG1 mutation is an underlying cause of autosomal recessive congenital ichthyosis, a debilitating skin disease, the pathogenic mechanism is not completely understood. In the present study we show that TG1 is an endoplasmic reticulum (ER) membrane-associated protein that is trafficked through the ER for ultimate delivery to the plasma membrane. Mutation severely attenuates this processing and a catalytically inactive point mutant, TG1-FLAG(C377A), accumulates in the endoplasmic reticulum and in aggresome-like structures where it is ubiquitinylated. This accumulation results from protein misfolding, as treatment with a chemical chaperone permits it to exit the endoplasmic reticulum and travel to the plasma membrane. ER accumulation is also observed for ichthyosis-associated TG1 mutants. Our findings suggest that misfolding of TG1 mutants leads to ubiquitinylation and accumulation in the ER and aggresomes, and that abnormal intracellular processing of TG1 mutants may be an underlying cause of ichthyosis.