Low energy fluence CO2 laser treatment of lymphangiectasia

Lymphangiectasia is an uncommonly reported complication of lymphatic insufficiency. These dilatations of lymphatic vessels may be symptomatic, necessitating treatment While CO₂ laser ablation has been used with success in the treatment of lymphangiectasia, it is infrequently reported and previous laser protocols have relied on high irradiances. The successful use of low fluence CO* laser in the treatment of multiple lymphangiectases on the lower limb of a middle-aged Caucasian woman with unilateral chronic lymphoedema is described.     

Surgical treatment of chronic hidradenitis suppurativa: CO2 laser stripping-secondary intention technique

The objective of our study was to evaluate a surgical method of management of patients with hidradentitis suppurativa (HS), using a CO₂ laser technique for stepwise horizontal vaporization and peroperative monitoring of the removal of diseased tissue. Twenty-four patients were selected for treatment, 21 women and three men, who had suffered from HS for a mean of 13 years (range 1–28 years) and experienced more than three recurrences of suppurating lesions during the year prior to entering the study. The mean follow-up time after CO₂ laser surgery was 27 months (range 15–47 months), with clinical follow-up once or twice a week during the wound-healing period, and then at intervals of 1–3 months to evaluate recurrences and assess the end result. The patients' healing time was approximately 4 weeks (range 3–5 weeks). During the follow-up period two patients had recurrences in the treated areas. Twenty-two patients had no recurrences in the treated areas, but in four cases de novo suppurating lesions appeared 5–10 cm beyond the initial sites of surgery. Ten patients had exacerbations of HS lesion(s) in a region other than the treated site. Eight patients did not have any recurrences. Post-surgery results were satisfactory both cosmetically and with regard to quality of life. The CO₂ laser stripping-secondary intention technique is a rapid, efficient, and economical method for the treatment of HS. It allows simple treatment of early lesions which would otherwise have been treated using less effective local conservative remedies.     

Identification of CD4+, 2H4+ (T8γ+) suppressor-inducer cells in normal human epidermis and superficial dermis

Immunohistological staining of frozen sections of normal human skin demonstrated the presence of significant numbers of mononuclear cells expressing novel epitopes associated with CD4-positive suppressor-inducer functions. The cells were located around superficial vessels and within the basal layers of the epidermis and hair follicles. The antigen identified by the various antibodies has been shown to be functionally important in the induction of various suppressor cells capable of abrogating B cell responses to pokeweed mitogen. The presence in the skin of cells with possible down-regulatory functions in the immune response may be significant with respect to surveillance against neoplasms and control of appropriate responses to infectious agents.     

Characterization and activity of phospholipase A2 in normal human epidermis and in lesion-free epidermis of patients with psoriasis or eczema

The kinetic properties of phospholipase A2 isolated from single large specimens of normal human epidermis and 'uninvolved' (lesion-free) psoriatic epidermis were determined. The enzymes from the two sources behaved identically with respect to changes in protein concentration, Ca2+ concentration and pH, but the enzymes responded differently to changes in substrate concentration. Furthermore, the specific activity of the enzyme derived from lesion-free psoriatic epidermis was higher than that from normal epidermis under all conditions used. Increased specific activity of the enzyme in the lesion-free epidermis was also found when biopsy specimens taken from thirty-five patients with psoriasis vulgaris at varying severity were compared with biopsies of normal epidermis from thirty-one control volunteers (P less than 0.001). Mixing experiments, in which homogenates of lesion-free psoriatic epidermis and control epidermis were combined, suggested that the relatively low activity of the enzyme in normal epidermis was due to the presence of an inhibitor. As the activity of the enzyme was not elevated in the lesion-free epidermis from twelve cases of eczema, which is also an inflammatory condition of the epidermis and superficial dermis, it is suggested that the raised phospholipase A2 activity demonstrated in the lesion-free epidermis of psoriasis may play an important role in the pathogenesis of the disease.     

The influence of cutaneous factors on the transcutaneous pO2 and pO2 at various body sites

The transcutaneous partial pressures of oxygen (tcpO2) and carbon dioxide (tcpCO2) were measured at eight different sites in 10 adult male subjects with an electrode at a temperature of 44 degrees C. The mean tcpO2 values (mmHg) were significantly lower on the face (forehead 26.6, cheek 29.6) and the palm (26.4) than at other sites (60.6-69.6). The tcpCO2 values (mmHg) were only slightly higher on the face. Removal of the stratum corneum produced an average increase of the tcpO2 on the palm of 37.6 mmHg and on the forehead of 19.6 mmHg. However, in 10 children with an age range of 3-9 years, the difference in the mean pO2 between the cheek and forearm was very small. There was no significant difference in the cutaneous blood flow at 44 degrees C between the cheek, palm and forearm.     

Effects of lithium carbonate (Li2CO3) on in-vitro-cultured normal human skin explants

The early morphological changes induced by lithium carbonate, a well-known psoriasis-provoking drug, were studied on cultured skin. Normal human skin from patients undergoing mastectomy was cultured in the presence of 3 mM, 6 mM and 10 mM of Li2CO3 for 4 days. The morphological changes were then evaluated by three observers in a blind manner and their reports were matched and collated. The cultured skin in the presence of Li2CO3 showed cell crowding of keratinocytes in the lower part of the epidermis, indicating epidermal hyperplasia. Another striking finding was intercellular oedema and vacuolar alteration with formation of small cavities in the upper dermis. There was no evidence of parakeratosis or any other histological characteristic of psoriasis, except hyperproliferation of the epidermis. Based on our knowledge of mechanisms of lithium action, we proposed two competitive explanations for its action on the epidermis: i) that lithium acts directly on dividing cells of the epidermis; and ii) that it acts indirectly by altering epidermal barrier function. Although we lack definite proof, we suggest that the observed morphological changes, in particular the non-specific stimulus to epidermal proliferation, are the primary events which initiate the process that will ultimately lead to the development of psoriasis in a predisposed patient.     

Fractional CO2 laser: a novel therapeutic device upon photobiomodulation of tissue remodeling and cytokine pathway of tissue repair

Minimally ablative fractional laser devices have gained acceptance as a preferred method for skin resurfacing. Notable improvements in facial rhytides, photodamage, acne scarring, and skin laxity have been reported. The aim of the present work was to compare how different CO₂ laser fluences, by modulating the secretory pathway of cytokines, are able to influence the wound-healing process, and how these fluences are associated with different clinical results. Eighteen patients, all with photodamaged skin, were treated using a fractional CO₂ laser (SmartXide DOT, Deka M.E.L.A., Florence, Italy) with varying laser fluences (2.07, 2.77, and 4.15 J/cm²). An immunocytochemical study was performed at defined end points in order to obtain information about specific cytokines of the microenvironment before and after treatment. The secretory pathway of cytokines changed depending on the re-epithelization and the different laser fluences. Different but significant improvements in wrinkles, skin texture, and hyperpigmentation were definitely obtained when using 2.07, 2.77, and 4.15 J/cm², indicating fractional CO₂ laser as a valuable tool in photorejuvenation with good clinical results, rapid downtime, and an excellent safety profile.     

Loss of β2 microglobulin from the cell surface of cutaneous malignant and premalignant lesions

Biopsies from twenty-three patients with malignant skin lesions and from twenty-two patients with premalignant or benign skin lesions were stained for beta2 microglobulin by use of the immunoperoxidase technique. The results showed a significant loss of demonstrable surface beta2 microglobulin from the surface of malignant cells compared to benign, and a partial loss in the premalignant cases. This difference could prove to be a useful tool in the histological diagnosis of malignant skin lesions, and in assessing whether or not the lesion has been completely excised.     

1α,25-dihydroxyvitamin D3 modulates Ia antigen expression induced by interferon-γ and prostaglandin E2 production in Pam 212 cells

Recent investigations suggest that 1α,25-dihydroxyvitamin D₃ [1α,25(OH)₂D₃] has an effect on the regulation of immune responses and that keratinocyte (KC) expression of major histocompatibility complex class II antigens may be involved in immune responses. We investigated the modulation by 1α,25(OH)₂D₃ of Ia antigen expression induced by interferon-γ (IFN-γ) and prostaglandin E₂ (PGE₂) production in Pam 212 cells. 1α,25(OH)₂D₃ at 10^?6, 10^?8 or 10^?10m significantly decreased the levels of IFN-γ-induced Ia antigen expression in Pam 212 cells. Pam 212 cells produced PGE₂ and 1α,25-(OH)₂D₃ enhanced Pam 212 cell PGE₂ production. However, indomethacin (1, 5 and 10 μg/ml) did not abrogate the inhibitory effect of 1α,25(OH)₂D₃ on IFN-γ induction of Ia antigen expression in Pam 212 cells, indicating that the products of the cyclo-oxygenase pathway do not mediate 1α,25(OH)₂D₃ inhibition of IFN-γ induction of Pam 212 cell Ia antigen expression. Our studies suggest that in Pam 212 cells the levels of Ia antigen expression induced by IFN-γ and PGE₂ production are negatively and positively regulated by 1α,25(OH)₂D₃, respectively, and that 1α,25(OH)₂D₃ may play a role in the regulation of immune responses.     

The effect of tacalcitol (1,24(OH)2D3) on cutaneous inflammation, epidermal proliferation and keratinization in psoriasis: a placebo-controlled, double-blind study

The aim of the present study was to discover to what extent 1.24(OH)₂D₃ ointment (tacalcitol; 4 μg/g) can modulate epidermal proliferation and keratinization, and several aspects of inflamation. Ten patients with psoriasis vulgaris were included in a placebo-controlled, double-blind study, using 1,24(OH)₂D₃ ointment (4μg/g). Before, and after 8 weeks of treatment, punch biopsies were taken from lesions treated with the active agent and placebo-treated lesions. An immunohistochemical study was carried out using monoclonal antibodies against the hyperproliferation-associated keratin 16, against cycling nuclei, filaggrin, involucrin, T lymphocytes, Langerhans cells, CD14 and polymorphonuclear leucocytes (PMN). The Wilcoxon test for matched pairs was used for statistical analysis of results. The biopsies from the lesions treated with the active agent showed a statistically significant change towards normalization of all aspects of inflammation studied, and of epidermal proliferation and keratinization, but there did not appear to be any effect on Langerhans cells. The only parameter which showed a significant alteration in the placebo-treated lesions was the number of cycling nuclei in the epidermis (P≤0.02). However, the biopsies from the plaques treated with the active agent showed a greater decrease of cycling cells (decrease: M_active=70, M_placebo=53) and a lower P-value (≤0.01). We therefore conclude that at the cell biological level 1.24(OH)₂D₃ ointment (4μg/g) has a substantial effect on several cell types, with regard to inflammation, epidermal proliferation and keratinization, with the exception of Langerhans cells.     

Modulation of phospholipase A2 activity in extracts of lesion-free psoriatic epidermis by alkaline phosphatase and a protein phosphatase inhibitor

Phospholipase A₂ activity is raised in non-lesional psoriatic epidermis compared with normal epidermis. It has been shown that the activity of this enzyme is controlled by an inhibitory protein the inhibitory effect of which is increased by dephosphorylation. Treatment of epidermal extracts with alkaline phosphatase reduced the phospholipase A₂ activity, both in normal and in lesion-free psoriatic epidermis. Inclusion of pyrophosphate, a protein phosphatase inhibitor, in the homogenizing medium caused the activity of phospholipase A₂ in epidermal extracts from normal and lesion-free epidermis to be raised to the same high level. These results are consistent with the hypothesis that the raised phospholipase A₂ activity in psoriatic epidermis is due to hyperphosphorylation of an endogenous inhibitor as a result of defective control of a phosphorylation/dephosphorylation mechanism. The relevance of these findings to other work is discussed.     

Inhibition of normal and psoriatic epidermal phospholipase A2 by picomolar concentrations of recombinant human lipocortin I

Normal human and psoriatic epidermal phospholipase A2 (PLA2) was inhibited by human recombinant lipocortin I when the substrate was present in a several million-fold molar excess. Inhibition was not total, even at relatively high concentrations of lipocortin I. It is therefore suggested that human epidermis contains two species of PLA2: one that is controlled by lipocortin I and one that is not.     

Effects of H1 antagonists on the cutaneous vascular response to histamine and bradykinin: a study using scanning laser Doppler imaging

Histamine plays an important part in the cutaneous weal and flare response which underlies many allergic skin conditions. It has a direct effect on the local vasculature to promote vasodilatation and increase microvascular permeability and may also initiate the more widely spread neurogenic flare. Quantification of these responses and studies of the mediator mechanisms underlying them have been limited by the lack of appropriate techniques to investigate them. To address this we have used two relatively new techniques, scanning laser Doppler imaging (LDI) and dermal microdialysis to measure changes in skin blood flow and the release of histamine within the weal and flare, following intradermal injection of histamine or bradykinin. These measurements have been made both in the absence and presence of the H₁ receptor blockers cetirizine and loratadine. Scanning LDI of the inflammatory response revealed marked differences in both the development and steady state responses to the intradermal injection of histamine (1–3 μmol/L) and bradykinin (1 μmol/L). The development of the flare and the weal response to both histamine and bradykinin was significantly reduced by cetirizine but not by loratadine. The histamine-induced flare area fell by 57 ± 4% (mean ± SEM, n = 10, P < 0.001) after cetirizine and the area of the weal fell by 73 ± 11% (P < 0.009). Bradykinin-induced inflammatory responses were similarly reduced by cetirizine, the weal by 60 ± 16% (P < 0.02) and the flare by 61 ± 4% (P < 0.005). Measurement of histamine concentration in skin using microdialysis, in six subjects, confirmed that histamine levels rose in the dialysate collected from the weal to 310 ± 16 nmol/L following injection of histamine. Histamine levels also rose following bradykinin injection in some subjects (mean 147 ± 46 nmol/L, range 18–336). Little increase in histamine concentration was seen in the dialysate from the flare following injection of either histamine or bradykinin. The histamine concentration in dialysate from unprovoked skin was 4.19 ± 0.75 nmol/L. These data reveal differences in the dermal responses to different mediators when assessed using scanning LDI. They confirm that histamine is released within the weal but not the flare response to the intradermal injection of both histamine and bradykinin and that its effects on the local vasculature to cause the oedematous weal and the axon reflex-mediated flare are significantly attenuated by the H₁ antagonist cetirizine and to a lesser extent by the H₁ antagonist loratadine.     

Immunohistochemical localization of the Ca2+ binding S100 proteins in normal human skin and melanocytic lesions

The purpose of this study was to evaluate the expression of the Ca²⁺-binding S100 proteins S100A1. S100A2, S100A3, S100A4, S100A6 and S100B in normal skin. These immunohistochemical staining patterns were compared with those in melanocytic lesions. Parafin-embedded tissue of normal skin adjacent to 26 naevi. 39 primary cutaneous melanomas and 14 cutaneous melanoma metastases was incubated with polyclonal antibodies against the recombinant human S100 proteins (S100A1, S100A2, S100A3, S100A4, S100A6, S100B) using the alkaline phosphatase anti-alkaline phosphatase method. The S100A2 antibody stained the basal layer of the epidermis an hair follicles of normal skin. Four of 39 primary cutaneous melanomas were positive for S100A2, whereas none of the methastases or naevi showed any immunoreactivity. The S100A3 antibody only stained the inner root sheath cuticle of some hair follicles but no melanocytes or melanocytic lesions. Staining of S100A4 was weak and thus omitted to further analysis. S100A6 faintly labelled keratinocytes. Langerhans' cells, melanocytes and sweat glands. S100A6 immunoreaction was found in two of seven junctional naevi, five of seven compund naevi, and all dermal and blue naevi. There was an intense cytoplasmatic reaction for S100A6 in all primary cutaneous melanomas and in nine of 14 (64%) metastases. S100B was positive in melanocytes and Langerhans' cells, all primaries as well as in the metastases. S100A1 protein was not detected on any of the tissue specimens examined. Whereas S100B and S100A6 antibodies are useful markers for malignant melanoma, expression of S100A4 antibody is too low to be used for immunohistochemical staining. S100A1 and S100A3 antibodies are not expressed in melanocytic lesions and S100A2 is only found in selected tumours. The investigated S100 proteins, including S100B and S100A6, are also expressed in selected elements of normal skin. These findings are important for correct interpretation of staining patterns, when S100 antibodies are used as markers for melanoma or other tumours.     

Divalent cations (Mg2+, Ca2+) differentially influence the β1 integrin-mediated migration of human fibroblasts and keratinocytes to different extracellular matrix proteins

Abstract Directed migration of keratinocytes and fibroblasts is a fundamental prerequisite in wound healing. Cation-dependent affinity changes of integrins are responsible for cell adhesion to and deadhesion from extracellular matrix proteins and have been implicated in driving cell migration. The specific requirements for divalent cations in the integrin-dependent migration of human dermal fibroblasts and human epidermal keratinocytes to various extracellular matrix proteins have been studied in vitro using blindwell Boyden chambers. The migration of the tested cells to collagen type I was mediated by the α₂β₂ integrins, to fibronectin by the combined action of the α₃β₂ and the α₅β₁ integrin, and the migration of fibroblasts to laminin dependent both on the α₂β₁ and the α₆β₁ integrins. No migration of keratinocytes to laminin was detected. Mg²⁺ alone induced cell migration with an optimum at 2 mM for fibroblasts and at 10 mM for keratinocytes. Ca²⁺ alone at 2 mM only marginally enhanced fibroblast and keratinocyte migration. At higher concentrations Ca²⁺ suppressed the stimulatory Mg²⁺ effect. 2 mM Ca²⁺ combined with 2 mM Mg²⁺ showed an additive stimulatory effect on the migration of fibroblasts to fibronectin. These data suggest that extracellular divalent cations differentially influence the integrin-mediated cell migration. A concentration gradient of Mg²⁺/Ca²⁺, as reported in tissue injury, thus may play a regulatory role in cell migration required for tissue remodelling.