Modulation of cellular annexin I in human leukocytes infiltrating DTH skin reactions.
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Based on our previous studies showing endogenous annexin I being depleted from migrated neutrophils (PMN) in vitro, we have tested whether the levels of this glucocorticoid-regulated protein in PMN and mononuclear cells (PBMC) were modified after adhesion to endothelial monolayers in vitro and extravasation into skin blisters in vivo. In vitro, annexin I levels were depleted more significantly (-70%) in post-adherent PMNs than in monocytes (-25%) and lymphocytes (-50%, only in the positive fraction). In vivo, a significant time-dependent increase (approximately threefold, P < 0.05) in cell-associated annexin I was measured in PBMCs recovered from the blisters, whereas no significant changes were detected in extravasated PMNs. This was associated with annexin I release in the blister fluids (approximately 35 ng/mL), whereas no detectable protein was found in matched-paired plasmas. In conclusion, we report for the first time an activation of the annexin I pathway during an ongoing experimental inflammatory response in humans, which is differently regulated between PMNs and PBMCs. (+info)
Spontaneous regression of a bulla with the development of adenocarcinoma of the lung.
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Spontaneous regression of a bulla in the lung is rare. We describe a case of spontaneous regression associated with the development of adenocarcinoma of the lung in a 59-year-old male smoker. The bulla had begun to regress spontaneously at least six months before lung cancer was detected on a chest radiograph. He underwent left upper lobe lobectomy with mediastinal node dissection. The tumor arose within the bulla, extending along the bulla wall. He has been alive for more than eight years with no evidence of recurrence. This case suggests that spontaneous regression of a bulla should be recognized as one of the early radiographic signs of the development of lung cancer in patients with bullous lung disease. (+info)
Pemphigus vulgaris and pemphigus foliaceus antibodies are pathogenic in plasminogen activator knockout mice.
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Previous studies have suggested that urokinase plasminogen activator is required for blister formation in pemphigus vulgaris and pemphigus foliaceus. Other studies, however, have shown that downregulation of plasminogen activator does not inhibit blisters induced by pemphigus immunoglobulin G. To eliminate the possibility that small amounts of urokinase plasminogen activator might be sufficient for blister formation, we passively transferred pemphigus immunoglobulin G to urokinase plasminogen activator knockout neonatal mice. Pemphigus foliaceus and pemphigus vulgaris immunoglobulin G caused gross blisters and acantholysis in the superficial and suprabasal epidermis, respectively, to the same degree in knockout and control mice, demonstrating that urokinase plasminogen activator is not absolutely required for antibody-induced blisters. Some studies have shown elevated tissue-type plasminogen activator in pemphigus lesions. Tissue-type plasminogen activator, however, is not necessary for blister formation, because pemphigus foliaceus and pemphigus vulgaris immunoglobulin G caused blisters to the same degree in tissue-type plasminogen activator knockout and control mice. To rule out that one plasminogen activator might compensate for the other in the knockout mice, we bred urokinase plasminogen activator, tissue-type plasminogen activator double knockouts. After passive transfer of pemphigus foliaceus and pemphigus vulgaris immunoglobulin G these mice blistered to the same degree as the single knockout and control mice, and histology indicated blisters at the expected level of the epidermis. These data definitively demonstrate that plasminogen activator is not necessary for pemphigus immunoglobulin G to induce acantholysis in the neonatal mouse model of pemphigus. (+info)
The functional diversity of epidermal keratins revealed by the partial rescue of the keratin 14 null phenotype by keratin 16.
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The type I epidermal keratins K14 and K16 are remarkably similar at the primary sequence level. While a structural function has been clearly defined for K14, we have proposed that a function of K16 may be to play a role in the process of keratinocyte activation that occurs after acute injury to stratified epithelia. To compare directly the functions of the two keratins we have targeted the expression of the human K16 cDNA to the progenitor basal layer of the epidermis of K14 null mice. Mice null for K14 blister extensively and die approximately 2 d after birth (Lloyd, C., Q.C. Yu, J. Cheng, K. Turksen, L. Degenstein, E. Hutton, and E. Fuchs. 1995. J. Cell Biol. 129:1329-1344). The skin of mice expressing K16 in the absence of K14 developed normally without evidence of blistering. However, as the mice aged they featured extensive alopecia, chronic epidermal ulcers in areas of frequent physical contact, and alterations in other stratified epithelia. Mice expressing a control K16-C14 cDNA also rescue the blistering phenotype of the K14 null mice with only a small percentage exhibiting minor alopecia. While K16 is capable of rescuing the blistering, phenotypic complementation in the resulting skin is incomplete due to the multiple age dependent anomalies. Despite their high sequence similarity, K16 and K14 are not functionally equivalent in the epidermis and other stratified epithelia and it is primarily the carboxy-terminal approximately 105 amino acids of K16 that define these differences. (+info)
Penetration of moxifloxacin into peripheral compartments in humans.
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To characterize the penetration of moxifloxacin (BAY 12-8039) into peripheral target sites, the present study aimed at measuring unbound moxifloxacin concentrations in the interstitial space fluid by means of microdialysis, an innovative clinical sampling technique. In addition, moxifloxacin concentrations were measured in cantharides-induced skin blisters, saliva, and capillary plasma and compared to total- and free-drug concentrations in venous plasma. For this purpose, 12 healthy volunteers received moxifloxacin in an open randomized crossover fashion either as a single oral dose of 400 mg or as a single intravenous infusion of 400 mg over 60 min. An almost-complete equilibration of the free unbound plasma fraction of moxifloxacin with the interstitial space fluid was observed, with mean area under the concentration-time curve (AUC)(interstitial fluid)/AUC(total-plasma) ratios ranging from 0.38 to 0.55 and mean AUC(interstitial fluid)/AUC(free-plasma) ratios ranging from 0.81 to 0.86. The skin blister concentration/plasma concentration ratio reached values above 1.5 after 24 h, indicating a preferential penetration of moxifloxacin into inflamed lesions. The moxifloxacin concentrations in saliva and capillary blood were similar to the corresponding levels in plasma. Our data show that moxifloxacin concentrations attained in the interstitial space fluid in humans and in skin blister fluid following single doses of 400 mg exceed the values for the MIC at which 90% of isolates are inhibited for most clinically relevant bacterial strains, notably including penicillin-resistant Streptococcus pneumoniae. These findings support the use of moxifloxacin for the treatment of soft tissue and respiratory tract infections in humans. (+info)
Ectosomes released by human neutrophils are specialized functional units.
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Here we show that human polymorphonuclear leukocytes (PMN) release ectosomes independently of complement attack during their activation both in vitro and at the site of inflammation in vivo. Patterns of biotinylated proteins on the surface of PMN and on PMN-derived ectosomes indicated a specific sorting of cell surface proteins into and out of ectosomes. Ectosomes expressed clusters of complement receptor 1 (CR1), which allowed them to bind efficiently to opsonized bacteria. Myeloperoxidase and human leukocyte elastase, both stored within the azurophilic granules of PMN, were found to colocalize on ectosomes with CR1. Furthermore, myeloperoxidase colocalized with human leukocyte elastase. In contrast, not present on CR1-expressing ectosomes were CD63, a selective marker for the azurophilic granules, and CD14, which is located within the same granules and the secretory vesicles as CR1. Of the other complement regulatory proteins expressed by PMN, only CD59 colocalized with CR1, while CD55 and CD46 were almost absent. Ectosomes released by activated PMN at the site of inflammation may function as a well organized element (ecto-organelle), designed to focus antimicrobial activity onto opsonized surfaces. (+info)
Targeted inactivation of the type VII collagen gene (Col7a1) in mice results in severe blistering phenotype: a model for recessive dystrophic epidermolysis bullosa.
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Dystrophic forms of epidermolysis bullosa (DEB) are associated with mutations in the type VII collagen gene (Col7a1) which encodes the major component of anchoring fibrils. To develop a DEB animal model, type VII collagen deficient mice were generated by targeted homologous recombination. The targeting vector replaced exons 46-69 of Col7a1 with the neomycin-resistance gene, in reverse transcriptional orientation, resulting in elimination of most of the collagenous domain 1. Col7a1 heterozygous (+/-) mice were phenotypically normal. Mating of Col7a1 +/- mice revealed that Col7a1 null (-/-) mice, which were born with extensive cutaneous blistering, died during the first two weeks of life probably due to complications arising from the blistering. Transmission electron microscopy revealed subepidermal blistering below the lamina densa and absence of anchoring fibrils. Immunohistochemical staining with anti-human type VII collagen antibody stained the dermal-epidermal junction in control mice, but did not stain the skin of Col7a1 null mice. Collectively, the DEB mice recapitulate the clinical, genetic, immunohistochemical and ultrastructural characteristics of recessive DEB in humans. These mice provide an animal model to study the pathomechanisms of DEB and serve as a system to test therapeutic approaches, including gene replacement, towards the cure of this devastating skin disease. (+info)
BALB/c mice produce blister-causing antibodies upon immunization with a recombinant human desmoglein 3.
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Pemphigus vulgaris (PV) is an Ab-mediated autoimmune blistering disease of mucotaneous surfaces. Over 95% of the patients with PV express DR4 or DRw6, and the disease is characterized by the presence of autoantibodies directed against desmoglein 3 (Dsg 3), a protein expressed on keratinocytes. An appropriate animal model is required to understand immunoregulation and to address the role of immunogenetic components in the production of pathogenic Abs that are characteristic of PV. Therefore, we turned to the development of a mouse model. Four strains of female mice (BALB/c, DBA/1, SJL/J, and HRS/J) were screened for their ability to produce pathogenic anti-Dsg 3 Abs. We demonstrated that only BALB/c mice immunized with a full-length Dsg 3 can produce pathogenic Abs capable of causing acantholysis of human foreskin in culture and blistering in neonatal mice. This observation suggested that either H-2d or the BALB background contains the immunogenetic makeup necessary for the production of pathogenic anti-Dsg 3 Abs. No correlation was noted between a given isotype and the pathogenic potential of autoantibodies from different strains of mice. Similarly, the pattern of reactivity of Abs with a panel of 46 synthetic peptides that span the entire Dsg 3 failed to reveal any association between binding specificity and the pathogenic potential, and suggested that pathogenic Abs might recognize conformational epitopes. Moreover, our studies showed that the epitopes recognized by pathogenic Abs are contained within the extracellular Dsg 3. (+info)