Selective inactivation of human neutrophil elastase by synthetic tannin

Tannins of natural or synthetic origin are well-known adjuvants in topical anti-inflammatory therapy of skin diseases. In this study, the influence of synthetic tannin on neutrophil accumulation, enzyme release, and on the proinflammatory activity of neutrophil-derived enzymes was investigated. The results show that synthetic tannin (Tamol) specifically inhibits the neutrophil serine protease human leukocyte elastase (HLE) in an irreversible manner with a half-maximal inhibitory concentration (IC50) of 0.3 microgram/ml. Exogenous protein partially abolished the tannin-dependent HLE inhibition (IC50 of Tamol at 1% protein-concentration:1.0 microgram/ml). Synthetic tannin did not influence the activities of other neutrophil enzymes like Cathepsin G, beta-glucuronidase, and myeloperoxidase. The specificity of Tamol for HLE was further substantiated by the lack of inhibition of other serine proteases. Additionally, Tamol had no effect on f-met-leu-phe-induced neutrophil chemotaxis and did not alter enzyme degranulation of neutrophils in response to f-met-leu-phe and opsonized zymosan. We conclude from our results that the anti-inflammatory properties of synthetic tannin may at least in part be due to inactivation of the proinflammatory protease HLE.     

Transient absence of C5a-specific neutrophil function in inflammatory disorders of the skin

Chemotactic migration, production of superoxide anion (O2-), and the release of beta-glucuronidase from azurophilic granules were determined in polymorphonuclear leukocytes (PMN) from 135 patients with infectious (e.g., pyoderma, acne conglobata, erysipelas) as well as noninfectious (psoriasis) skin diseases. Purified C5a and the formylated tripeptide FMLP were used as stimuli. In addition, longitudinal profiles of PMN activities were performed at daily intervals in several patients. There was a complete absence of PMN responses (chemotaxis, O2--production, and enzyme release) specifically induced by C5a in 25 patients suffering from various inflammatory diseases of the skin. In these patients PMN responsiveness for the tripeptide FMLP was either normal or increased. The C5a-dependent defect of PMN was transient and correlated with disease activity. When normal PMN were incubated with sera from C5a-defective patients, no inherent stimulatory or inhibitory activities compared to control sera were seen. Pretreatment of normal PMN in vitro with various concentrations of C5a failed to completely deactivate PMN without affecting FMLP dependent functions. These observations demonstrate the presence of a functional defect in circulating PMN during acute cutaneous inflammation. The in vitro experiments suggest transient blocking of C5a-dependent PMN functions by a cell-bound factor which seems not to be C5a or C5adesarg.     

Ultrastructural changes of the skin induced by human leukocyte elastase and cathepsin G.

Human neutrophil elastase and cathepsin G at a concentration of 10(-6) M were found to attack various substrates when normal skin biopsy specimens were incubated at 37 degrees C for 1 h with either of these enzymes. Elastase damaged primarily hemidesmosomes, leading to the epidermal cleavage from the dermis, whereas cathepsin G damaged the membrane structures. Both these neutral proteinases were highly specific to basal lamina of blood vessels. This indicates that neutrophil elastase and neutrophil cathepsin G may play different roles in various skin diseases related to enhanced activity and infiltration of neutrophils.     

Inhibitory effects of shale oils (Ichthyols) on the secretion of chemotactic leukotrienes from human leukocytes and on leukocyte migration

Ichthyols are sulfated shale oils with well-known anti-inflammatory effects in dermatologic diseases. Their possible mechanisms of action were studied by measuring chemotactic factor release from peripheral human leukocytes in vitro. Ichthyols caused no release of such factors by themselves but inhibited ionophore-induced release. After elution of the cell supernatants on reverse-phase high-pressure liquid chromatography, followed by analysis of the fractions in the chemotaxis assay and the radioimmunoassay, Ichthyols caused a reduction of lipids at marker positions for leukotriene B4 (LTB4) and 20-COOH-LTB4. The inhibition was also evident in the LTB4 radioimmunoassay, was dose- and time-dependent, and occurred in noncytotoxic concentrations of the agents. Ichthyols also inhibit the chemotactic response of neutrophils toward LTB4 and the unstimulated migration of cells. These inhibitory effects of Ichthyols on secretion of chemotactic arachidonate metabolites from leukocytes and on cell migration provide a plausible explanation for their anti-inflammatory activity.     

Recombinant human interleukin-8 is a potent activator of canine neutrophil aggregation, migration, and leukotriene B4 biosynthesis

Interleukin-8 (IL-8), formerly known as NAP-1, is formed by a variety of cells upon stimulation with IL-1 or tumor necrosis factor (TNF). The biologic activity of the cytokine involves activation of almost every neutrophil function studied so far in different species. In the present study, we compared the effects of recombinant human IL-8 (rIL-8) and the lipid mediators, leukotriene B4 (LTB4) and platelet-activating factor (PAF), on neutrophil functions in dogs. All three chemotactic factors induced neutrophil aggregation and chemotaxis, with rIL-8 being far more potent than LTB4 and PAF. The migration induced by rIL-8 was significantly greater than that observed towards LTB4 and PAF. In the aggregation assay, rIL-8 was shown for the first time to be a potent stimulant. The aggregation response was more persistent than that obtained with LTB4 and PAF and the potency of rIL-8 was greater. An intradermal dose-response study showed that rIL-8 is an extremely potent inducer of selective neutrophil infiltration in canine skin. The infiltration was more pronounced than following injection of LTB4 or PAF. It was proposed that the superior effect of rIL-8 was caused by a synergistic effect between injected rIL-8 and LTB4, which was shown to be produced in biologically active amounts by canine neutrophils stimulated with rIL-8. From a therapeutic point of view, the simultaneous presence of rIL-8 and LTB4 in inflammatory skin diseases highlights the need to develop drugs that inhibit the production and/or effect of both mediators.     

Analysis of the interaction of human C5a and C5a des Arg with human monocytes and neutrophils: flow cytometric and chemotaxis studies

C5a and C5a des Arg are potent complement-derived mediators that bind receptors on peripheral blood leukocytes and tissue-specific cellular elements to elicit and amplify inflammatory and immunomodulatory reactions. To study the interactions of C5a and C5a des Arg with these cells, fluorescein conjugates of these ligands were prepared by a new technique and their binding to monocytes, neutrophils, platelets, and endothelial cells was studied with flow cytometry. Fluoresceinated C5a produced neutrophil myeloperoxidase release and chemotaxis and also bound rabbit anti-C5a antibody much like native anaphylatoxin; likewise, fluoresceinated C5a des Arg demonstrated retention of biologic and antigenic activities. Both fluorescein-conjugated C5a and C5a des Arg bound to monocytes and neutrophils in a concentration-dependent, saturable, and homogeneous manner, but 10- to 15-fold higher concentrations of C5a des Arg were required to attain saturable binding of these leukocytes. Ligand binding was specifically inhibited by native purified human C5a in a concentration-dependent manner, while it was unaffected by C3a or N-formyl-methionyl-leucyl-phenylalanine-lysine. There was no evidence of a C5a receptor-negative subpopulation of monocytes or neutrophils. Moreover, comparative binding experiments with leukocytes from multiple normal volunteers showed that a greater percentage of monocytes than neutrophils bound C5a at less than saturable concentrations of ligand (P less than 0.05, 0.5 to 5.0 nM). A representative half-maximal binding of fluorescein-conjugated C5a (C5a des Arg) binding to monocytes and neutrophils was 1.2 nM (30 nM) and 2.6 nM (68 nM), respectively. In contrast, fluorescein-conjugated C5a did not specifically bind to human platelets or umbilical vein endothelial cells.     

Structural changes of human epidermis induced by human leukocyte-derived proteases

During the process of inflammation human neutrophils release potent serine proteases, such as human leukocyte elastase and cathepsin G. In psoriasis these enzymes are released within the epidermis. To investigate the destructive potential of neutrophil-derived serine proteases these were applied on viable human epidermis as well as full thickness human skin in vitro. Human leukocyte elastase and cathepsin G were found to dissociate keratinocytes from epidermal sheets in a time- and dose-dependent fashion. Significant keratinocyte dissociation was observed 4 h after application of 3 nM human leukocyte elastase. By electron microscopy of elastase- or cathepsin G-treated full thickness human skin, widening of the extracellular space followed by complete separation of keratinocytes without intradesmosomal cleavage was observed. In addition, cathepsin G induced membrane damage as well as destruction of intracellular organelles. Thus, neutrophil-derived serine proteases exert pronounced destructive potential in human epidermis in concentrations likely to appear in lesional psoriatic skin.     

Human leukocyte elastase induces keratinocyte proliferation by epidermal growth factor receptor activation

Epidermal hyperproliferation and neutrophil infiltration are major histopathological changes observed in psoriasis. Neutrophils contain human leukocyte elastase (HLE), which is released at sites of inflammation. HLE is present in psoriatic lesions and induces keratinocyte hyperproliferation in vitro and in vivo. To determine the molecular mechanisms linking a proteolytic effect of HLE and epidermal hyperproliferation, we examined the effects of HLE-induced signaling in human keratinocytes. Application of 100 nM HLE resulted in a transient calcium influx in FURA2-loaded human HaCaT keratinocytes observed by single-cell fluorescence imaging. The calcium signal was concentration dependent and was inhibited by addition of the HLE inhibitors elafin and secretory leukocyte protease inhibitor. The calcium signal was neither inhibited by pertussis toxin, cholera, or by pre-stimulation with trypsin. Incubation with the tyrosine kinase inhibitor genistein, a protein kinase C inhibitor, as well as incubation with neutralizing EGFR antibodies abolished the HLE-induced calcium influx. The supernatants of HLE-treated keratinocytes induced a calcium signal in separately cultured keratinocytes. This could be inhibited by the addition of anti-TGF-alpha antibodies. Application of HLE-induced keratinocyte proliferation, which could be inhibited by neutralizing of anti-EGFR and anti-TGF-alpha antibodies. Herein we demonstrate that HLE induces keratinocyte proliferation by proteolytic activation of an EGFR signaling cascade involving TGF-alpha.     

Factors secreted by untreated psoriatic monocytes enhance neutrophil functions

Monocytes stimulated with bacterial lipopolysaccharides (LPS) release mediators that induce increased responses of human granulocytes. Recently we showed that psoriatic monocytes can stimulate neutrophil chemotaxis, phagocytosis, and O2- production without addition of LPS and this effect is inhibited by cyclosporin A. We have now investigated the presence of cytokines in supernatants from cultures of psoriatic monocytes (resting monocytes). These cells were cultured for 24 h in endotoxin-free medium. Normal human neutrophils were then incubated for 1 h with the resulting supernatants (sMS, or conditioned media). The sMS from unstimulated psoriatic monocytes significantly enhanced neutrophil chemotaxis and superoxide anion production. The enhancing factors are protein in nature and require ongoing protein synthesis, demonstrated by the facts that the activity in conditioned medium is labile to heat denaturation at 100 degrees C for 10 min, is not produced by monocytes cultured in the presence of puromycin, and is proteinase sensitive. Additional evidence suggested that extremes of pH inhibit activity. None of the conditioned media treated in these ways activated neutrophils. The neutrophil function-enhancing factors derived from psoriatic monocytes are in part cytokines, including TNF and GM-CSF. The support for this conclusion is the higher level of TNF and GM-CSF in media conditioned by psoriatic monocytes than in media conditioned by normal human monocytes, the inhibition of TNF production and neutrophil stimulating activity by cyclosporin A, and the inhibition of neutrophil stimulating activity in conditioned media preincubated with anti-TNF and anti-GM-CSF antibodies. It is concluded that psoriatic monocytes spontaneously produce higher than normal levels of TNF alpha, GM-CSF, and, perhaps, other cytokines that might be responsible for the enhanced activity of psoriatic neutrophils.     

Inhibition by leukotriene B5 of leukotriene B4-induced activation of human keratinocytes and neutrophils

Leukotriene B5 (LTB5) that is generated enzymatically from eicosapentaenoic acid (EPA), was compared with arachidonic acid-derived LTB4 for its DNA synthetic effect on cultured human epidermal keratinocytes and for its chemokinetic effect on human blood neutrophils. Leukotriene B5 was much less potent than LTB4 in stimulating DNA synthesis and in inducing chemokinesis. Furthermore, the maximum response to LTB5 was only a mean of 38% that of LTB4 for mitogenesis and 70% that of LTB4 for chemokinesis. At an optimally active concentration of LTB4 (10(-10) M) the addition of LTB5 suppressed the enhancement by LTB4 of DNA synthesis in keratinocytes by a mean of 21%, 33%, and 54%, respectively, at 10(-9) M, 10(-8) M, and 10(-7) M LTB5. Leukotriene B5 inhibited to a lesser extent the maximum neutrophil chemokinetic response elicited by 10(-10) M LTB4 with mean inhibition of 10%, 20%, and 18%, respectively, by 10(-9) M, 10(-8) M, 10(-7) M LTB5; LTB5 was without effects on N-formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP)-elicited neutrophil chemokinesis and on thrombin-stimulated keratinocyte DNA synthesis. The dietary introduction of n-3 fatty acids, such as EPA, may reduce the epidermopoiesis and neutrophil migration evoked by LTB4 through decreases in generation of LTB4 and the capacity of LTB5 to inhibit the effects of LTB4.     

Distinct compartmentalization of immune cells and mediators characterizes bullous pemphigoid disease

Bullous pemphigoid (BP) is an autoimmune blistering disease characterized by recruitment of leucocytes into skin and release of damaging enzymes, resulting in epidermal detachment and blister formation. To better understand the role of leukotriene B4 (LTB4) and other inflammatory factors in BP pathophysiology, we conducted microscopic and immunohistochemical analyses of preserved skin biopsy sections and conducted flow cytometry and ELISA analyses of matched blood and blister fluid from BP patients. Neutrophils predominated in BP blister fluid, which also contained monocytes/macrophages and T cells, but few to no eosinophils and B cells. In contrast, BP skin histology showed a different pattern, with abundant neutrophils but eosinophils being the predominant immune cell type. LTB4 pathway and neutrophil activation markers were prevalent in BP skin lesions and strongly associated with perivascular neutrophils. Blister fluid neutrophils, monocytes/macrophages and eosinophils all exhibited increased surface expression of leukotriene A4 hydrolase and neutrophil elastase (P = .002 for both). Blister fluid was also enriched in interleukins (IL)-1α, IL-1β, IL-8, IL-10, IL-18, monocyte colony-stimulating factor (M-CSF) and vascular endothelial growth factor (VEGF). Our findings suggest differential leucocyte recruitment from blood into dermis and from dermis into blister, which correlates with disease activity, and presents potential new treatment opportunities for BP.     

Skin-infiltrating neutrophils following exposure to solar-simulated radiation could play an important role in photoageing of human skin

BACKGROUND: The pathophysiology of photoageing of the skin has been studied extensively. Matrix metalloproteinases (MMPs) originating from keratinocytes and fibroblasts are thought to play a primary role in this process. Although neutrophils are potent producers of a wide array of proteolytic substances and are present in sunburned skin, their contribution to the pathophysiology of photoageing has been described only in murine studies. OBJECTIVES: To determine the role of neutrophils in photoageing of human skin. METHODS: Healthy white-skinned volunteers were recruited and their sun-protected buttock skin was exposed to solar-simulated radiation (SSR) in dose-response and time-course studies. Punch biopsies were taken and the influx of neutrophils and the expression of neutrophil elastase and MMPs was studied using immunohistochemical techniques and in situ zymography. RESULTS: Neutrophil elastase and MMPs were detected only in skin irradiated with erythemogenic doses (> or = 1 minimal erythema doses) of SSR. Immunohistochemical double staining demonstrated neutrophils to be the major source of MMP-1, MMP-8 and MMP-9. In situ zymography showed elastase, collagenase and gelatinase enzyme activity in those cells. CONCLUSIONS: Our study suggests that neutrophils participate in the process of photoageing of human skin as they infiltrate the skin and release enzymatically active elastase (neutrophil elastase), collagenase (MMP-1) and gelatinase (MMP-9).     

Dapsone suppresses human neutrophil superoxide production and elastase release in a calcium-dependent manner

BACKGROUND: Dapsone (4,4'-diaminodiphenyl sulphone) is a powerful therapeutic tool in many skin diseases including neutrophilic dermatoses. The drug has an outstanding therapeutic efficacy against many skin diseases characterized by neutrophil-rich infiltrates; however, mechanisms of its action are poorly understood. OBJECTIVES: We investigated the effects of dapsone on respiratory and secretory functions of human neutrophils triggered by the chemotactic peptide N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP), the physiological agonist C5a, and phorbol myristate acetate (PMA). METHODS: Human neutrophils were isolated from venous blood obtained from healthy donors. We detected extracellular production of superoxide (O(2) (-)) by cytochrome C reduction assay, and intracellular production of O(2) (-) by flow cytometry. Neutrophil elastase release was measured by the cleavage of the specific elastase substrate N-methoxysuccinyl-Ala-Ala-Pro-Val-p-nitroanilide. Measurement of cytosolic free calcium concentration was performed using the calcium-reactive fluorescence probe, Fluo-3. RESULTS: Dapsone suppressed intra- and extracellular production of O(2) (-) and elastase release triggered by fMLP and C5a, but not by PMA. Both fMLP and C5a signalled the above pathways by inducing calcium influx, but PMA functions bypassed calcium influx. Dapsone was capable of antagonizing the induction of calcium influx. CONCLUSIONS: These findings suggest that one mechanism of the anti-inflammatory action of dapsone is inhibition of calcium-dependent functions of neutrophils including release of tissue-damaging oxidants and proteases in the affected skin.     

Effects of cepharanthin on neutrophil chemotaxis, phagocytosis, and reactive oxygen species generation.

The effects of cepharanthin on inflammatory parameters such as neutrophil chemotaxis, phagocytosis and reactive oxygen species (ROS) generation, were examined. Cepharanthin significantly decreased the levels of O2-, H2O2, and OH. generated by neutrophils. H2O2 and OH. generated in a cell-free, xanthine-xanthine oxidase system were also reduced in the presence of cepharanthin. However, the drug did not affect neutrophil chemotaxis or phagocytosis. The present study indicates that cepharanthin is an effective ROS scavenger, exerting its anti-inflammatory action by reducing the potent ROS species excessively generated in tissues and organs, especially at the sites of inflammation.     

Biologic properties of LTB4 and paf-acether in vivo in human skin

Using an improved skin chamber technique, the consequences of prolonged contact of leukotriene B4 (LTB4) and platelet-activating factor (paf-acether) with human dermis were evaluated quantitatively and kinetically in vivo. Leukocyte chemotaxis, histoenzymologic alterations, and modifications in vascular permeability were studied in two sets of experiments. In a first set of experiments, the dose-effect relationship of LTB4 and paf-acether on leukocyte migration was studied. LTB4 (3 X 10(-8) M to 9 X 10(-7) M) in Hanks' balanced salt solution (HBSS) elicited an intense dose-dependent and time-dependent neutrophil migration. Paf-acether, at the same concentration range, induced a significant increase in cell migration only at 9 X 10(-7) M and when diluted in HBSS containing 0.25% serum albumin (HBSS-BSA). Histoenzymologic analysis demonstrated that LTB4 in vivo induced degranulation of most of the neutrophils migrating through the dermis. Paf-acether caused mild degranulation of neutrophils and induced the appearance of degranulated basophils in dermal vessels. A second set of experiments was designed to study simultaneously the modifications in vascular permeability and cell migration induced by LTB4 and paf-acether, with or without prostaglandin E2 (both at a concentration of 3 X 10(-7) M in HBSS). Since spontaneous protein diffusion in HBSS progressively declined up to a plateau reached after 20 h (1.2 +/- 0.15 mg of proteins/cm2/2 h), these experiments were carried out after a 20-h equilibration period. Leukotriene B4 induced a late and slight increase in vascular permeability. Paf-acether did so intensely and transiently. Prostaglandin E2 significantly enhanced protein diffusion and neutrophil migration induced by LTB4 and, to a lesser extent, by paf-acether. Interestingly, despite the reintroduction into the skin chambers of freshly prepared solutions containing the mediators, leukocyte migration and protein diffusion progressively decreased during the experiments. This suggests the local production of anti-inflammatory factors that inhibit local mediators and thus regulate the inflammatory response.     

In vitro generation of chemotactic leukotrienes from unfractionated murine epidermal cells

Single cell suspensions of murine epidermal cells were studied for the generation of leukotrienes (LTs), using in vitro bioassays for chemotaxis, reverse-phase high-pressure liquid chromatography (HPLC), and radioimmunoassays (RIAs). A combination of arachidonic acid (AA) at 10(-3)-10(-4) M with the calcium ionophore A 23187 at 5 X 10(-6) M was the most potent stimulus, causing release of LTs within 10-30 min. Other stimuli, like the N-formyl-methionyl-leucyl-phenylalanine, at 10(-7) M and bradykinin at 10(-3) M, were less effective, and the tumor promotor phorbol-myristate-acetate (10(-5)-10(-8) M) caused no release at all. AA induced release at cytotoxic concentrations, but the other stimuli did not, and keratinocytes from different body regions were equally good sources of the LTs. In vivo or in vitro pretreatment of keratinocytes with UV radiation did not alter spontaneous or stimulated secretion of LTs, while pretreatment of cells with Ia, but not with Thy-1, monoclonal antibodies caused a moderate decrease of release. Analyses by HPLC indicated the release of 20-OH-LTB4 in addition to LTB4 in cell supernatants. Murine keratinocytes and epidermal dendritic cells serve therefore as a source of chemotactic leukotrienes after appropriate in vitro stimulation with agents that are known to play a role in cutaneous inflammation.     

Cathelicidin LL-37 induces the generation of reactive oxygen species and release of human alpha-defensins from neutrophils

BACKGROUND: Psoriasis is characterized by epidermal infiltration of neutrophils that destroy invading microorganisms via a potent antimicrobial arsenal of oxidants and antimicrobial agents. In contrast to atopic dermatitis, psoriasis exhibits low levels of skin infections due to the presence of antimicrobial agents, including cathelicidin LL-37. LL-37 kills a broad spectrum of microbes, and activates neutrophil chemotaxis. OBJECTIVE: To determine whether or not LL-37 could regulate additional neutrophil functions such as production of cytokines/chemokines, reactive oxygen species and release of neutrophil antimicrobial peptides. METHODS: Human peripheral blood neutrophils were used in this study. The production of interleukin (IL)-8 and release of alpha-defensins were analysed by enzyme-linked immunosorbent assay, and real-time polymerase chain reaction (PCR) was used to quantify alpha-defensin gene expression. Phosphorylation of mitogen-activated protein kinase (MAPK) was determined by Western blotting. The generation of reactive oxygen species was examined using flow cytometry, and intracellular Ca(2+) mobilization was measured using a calcium assay kit. RESULTS: LL-37 enhanced the production of IL-8 under the control of MAPK p38 and extracellular signal regulated kinase (ERK), as evidenced by the inhibitory effects of p38 and ERK1/2 inhibitors on LL-37-mediated IL-8 production. Furthermore, LL-37 induced phosphorylation of p38 and ERK. We also revealed that LL-37 stimulated the generation of reactive oxygen species dose- and time-dependently, most probably via NADPH oxidase activation and intracellular Ca(2+) mobilization. Finally, LL-37 induced both mRNA expression and protein release of alpha-defensins, known as human neutrophil peptide 1-3. CONCLUSION: Taken together, we suggest that in addition to its microbicidal properties, LL-37 may contribute to innate immunity by enhancing neutrophil host defence functions at inflammation and/or infection sites.     

Influence of eicosanoids on fibroblast chemotaxis and protein synthesis in vitro

Metabolism of fibroblasts plays a key role in wound healing, fibrosis, rheumatoid arthritis, and similar physiological and pathological processes. The regulatory influence of eicosanoids, an important class of inflammatory mediators, on fibroblast metabolism, in these processes is, to date, unclear. The aim of this study was to investigate the effect of some eicosanoids on chemotaxis and protein synthesis of fibroblasts in vitro. Of twelve eicosanoids tested, only 5(S)-HETE, LTB4, and 12(S)-HETE were active as chemo-attractants for fibroblasts. 5(S)-HETE was the most potent attractant. It exerted its maximal activity at 10(-10) mol/l. 12(S)-HETE and LTB4 caused similar dose dependent fibroblast chemotaxis with a maximum of activity at 10(-7) M and 5 x 10(-8) M, respectively. Hydroxylation of LTB4 on C20 or methylation of the carboxy group of 12(S)-HETE decreased reactivity of the parent compounds only slightly. Eicosanoid induced chemotaxis could be antagonized by 12(S)-HETE but not by the proteinaceous chemoattractants fibronectin, PDGF, or EGF. Receptors for peptide and eicosanoid mediated chemotaxis are thus different. Inhibition of collagen synthesis was observed in the presence of 5(S)-HETE and 12(S)-HETE while total protein synthesis was unaffected by 12(S)-HETE and augmented by 5(S)-HETE. These data suggest that certain eicosanoids specifically regulate fibroblast activities in wound healing and similar events of connective tissue reorganization.     

Lesional elastase activity in psoriasis, contact dermatitis, and atopic dermatitis.

Human leukocyte elastase (HLE) is a broad spectrum serine protease derived from neutrophils and macrophages. We developed an assay to determine HLE activity on the skin surface in patients with inflammatory skin diseases. HLE activity was absent in the skin of healthy controls. A massive increase of HLE activity was found in lesional skin of psoriasis (31 times), allergic contact dermatitis (55 times), and atopic dermatitis (35 times), but not in uninvolved skin of diseased patients. Therefore, this assay appears to represent a useful biochemical marker of epidermal inflammation. The presence of proteolytically active HLE in diseased epidermis, which is known to contain specific inhibitors of this enzyme, suggests a pathophysiologic role of this enzymatic activity in psoriasis, contact dermatitis, and atopic dermatitis.     

SUCCESSFUL TREATMENT OF A PATIENT WITH RECURRENT FURUNCULOSIS BY VITAMIN C: IMPROVEMENT OF CLINICAL COURSE AND OF IMPAIRED NEUTROPHIL FUNCTIONS

Background. Neutrophils play a critical role in host defense against a variety of microbial pathogens. There is much information to suggest a role for vitamin C in the physiology of neutrophils. Thus, the effects of vitamin C treatment were studied in a patient with a history of recurrent furunculosis who showed altered neutrophil functions. Methods. Superoxide generation was measured by cytochrome C reduction. Phagocytosis of opsonized zymosan by neutrophils and chemotaxis on agarose plates were determined. Results. Chemotaxis, phagocytosis, and superoxide generation of the patient's neutrophils were significantly lower than those of the matched control. Treatment with vitamin C (500 mg/day) for 30 days caused a dramatic clinical response and a significant improvement of all three neutrophil functions to values similar to those of the controls. Conclusions. We suggest that the patient described here had a temporary defect in neutrophil functions. The treatment with vitamin C probably prevented neutrophil oxidation, thus contributing to recovery of neutrophil function and arrest of furunculosis.