S. Wright Caughman

Neuropeptide regulation of human dermal microvascular endothelial cell ICAM-1 expression and function

There is increasing evidence that sensory nerves may participate in cutaneous inflammatory responses by the release of neuropeptides such as substance P (SP). We examined the direct effect of SP on human dermal microvascular endothelial cell (HDMEC) intercellular adhesion molecule 1 (ICAM-1) expression and function. Our results indicated that, although cultured HDMEC expressed mRNA for neurokinin receptors 1, 2, and 3 (NK-1R, NK-2R, and NK-3R), SP initiated a rapid increase in HDMEC intracellular Ca2+ levels, primarily by the activation of NK-1R. Immunohistochemistry studies likewise demonstrated that HDMEC predominantly expressed NK-1R. The addition of SP to HDMEC resulted in a rapid increase in cellular ICAM-1 mRNA levels, followed by a fivefold increase in ICAM-1 cell surface expression. This functionally resulted in a threefold increase in51Cr-labeled binding of J-Y lymphoblastoid cells to HDMEC. In vivo studies demonstrated a marked increase in microvascular ICAM-1 immunostaining 24 and 48 h after application of capsaicin to the skin. These results indicate that neuropeptides such as SP are capable of directly activating HDMEC to express increased levels of functional ICAM-1 and further support the role of the cutaneous neurological system in modulating inflammatory processes in the skin.There is increasing evidence that sensory nerves may participate in cutaneous inflammatory responses by the release of neuropeptides such as substance P (SP). We examined the direct effect of SP on human dermal microvascular endothelial cell (HDMEC) intercellular adhesion molecule 1 (ICAM-1) expression and function. Our results indicated that, although cultured HDMEC expressed mRNA for neurokinin receptors 1, 2, and 3 (NK-1R, NK-2R, and NK-3R), SP initiated a rapid increase in HDMEC intracellular Ca2+ levels, primarily by the activation of NK-1R. Immunohistochemistry studies likewise demonstrated that HDMEC predominantly expressed NK-1R. The addition of SP to HDMEC resulted in a rapid increase in cellular ICAM-1 mRNA levels, followed by a fivefold increase in ICAM-1 cell surface expression. This functionally resulted in a threefold increase in 51Cr-labeled binding of J-Y lymphoblastoid cells to HDMEC. In vivo studies demonstrated a marked increase in microvascular ICAM-1 immunostaining 24 and 48 h after application of capsaicin to the skin. These results indicate that neuropeptides such as SP are capable of directly activating HDMEC to express increased levels of functional ICAM-1 and further support the role of the cutaneous neurological system in modulating inflammatory processes in the skin.

Furunculosis Due to Mycobacterium mageritense Associated with Footbaths at a Nail Salon

ABSTRACT We report two cases of lower-extremity furunculosis caused by Mycobacterium mageritense. Both patients were patrons of the same nail salon, where they received footbaths prior to pedicures. M. mageritense bacteria isolated from two whirlpool footbaths were determined to be closely related to the patient isolates by pulsed-field gel electrophoresis.

Interferon γ-dependent Induction of Human Intercellular Adhesion Molecule-1 Gene Expression Involves Activation of a Distinct STAT Protein Complex

In response to interferon γ (IFNγ), intercellular adhesion molecule-1 (ICAM-1) is expressed on human keratinocytes, a cell type that is critically involved in cutaneous inflammation. An ICAM-1 5′ regulatory region palindromic response element, pIγRE, has been shown to confer IFNγ-dependent transcription enhancement. By electrophoretic mobility shift assays (EMSA), pIγRE forms a distinct complex with proteins from IFNγ-treated human keratinocytes, termed γ response factor (GRF). Binding of GRF is tyrosine phosphorylation-dependent, and mutations of pIγRE that disrupt the palindromic sequence or alter its spatial relationship abrogate GRF binding. Supershift EMSAs using antibodies to characterized STAT proteins suggest that GRF contains a Stat1α-like protein; however, non-ICAM-1 IFNγ-responsive elements (REs) known to bind Stat1α homodimers fail to compete for GRF binding in EMSA, and pIγRE does not cross-compete with these REs that complex with homodimeric stat1α. The pIγRE·GRF complex also displays a distinctly different electrophoretic mobility compared to that of IFNγREs complexed to homodimeric Stat1α. These findings indicate that a distinct complex containing a Stat1α-like protein mediates IFNγ-induced ICAM-1 gene transcription and identifies a subset of IFNγ-responsive genes that appear to be regulated by this complex.

CULTURE AND CHARACTERIZATION OF SINUSOIDAL ENDOTHELIAL CELLS ISOLATED FROM HUMAN LIVER

SummaryAlthough most vascular models use large vessel endothelial cells from human umbilical veins, there is marked heterogeneity among endothelial cells from different vascular beds and organs. More accurate modeling of endothelial involvement in liver diseases, including metastasis, may result from the use of human hepatic sinusoidal endothelial cells. Liver resection specimens were sectioned, then treated with a 1.2 U/ml dispase solution. The tissue slurry was mechanically disaggregated and separated by centrifugation on a Percoll density gradient. Cells were then cultured in an endothelial-specific media with growth factors. These techniques resulted in a homogeneous monolayer consistent with endothelial cells by light microscopy. An endothelial origin was further confirmed by the expression of Factor VIII, binding of Ulex lectin, and uptake of acetylated low density lipoprotein. Electron microscopy showed transcellular fenestrations consistent with a sinusoidal origin. These human hepatic sinusoidal endothelial cells were then studied for expression of the adhesion molecules CD31/PECAM, CD34, E-selectin, ICAM-1, L-selectin, LFA-3, P-selectin, and VCAM-1 plus the binding of wheat germ agglutinin lectin. The patterns of adhesion molecule expression and lectin binding by these cells are characteristic of hepatic sinusoidal endothelia. In this paper, we have described a method for isolation and culture of human cells with the morphologic and phenotypic characteristics of hepatic sinusoidal endothelia.

Neutrophilic Dermatoses Associated with Hematologic Disorders

The neutrophilic dermatoses are a group of disorders characterized histologically by dermal neutrophilic infiltrates, having no infectious or other identifiable etiology. These entities include acute febrile neutrophilic dermatosis, pyoderma gangrenosum, erythema elevatum diutinum, subcorneal pustular dermatosis, and neutrophilic eccrine hidradenitis. The neutrophilic dermatoses are frequently considered to be part of a continuous spectrum because they share many features including cutaneous neutrophilic infiltrates, similar clinical eruptions, systemic involvement, association with systemic diseases, and responsiveness to immunosuppressive agents.1 When evaluating patients with the clinical lesions of a neutrophilic dermatosis, infection, leukemic cell infiltration, and allergic contact dermatitis must be excluded as causes.2 The pathogeneses for these disorders are still unclear, but they are recognized paraneoplastic phenomena.3,4 The neutrophilic dermatoses may be associated with a variety of systemic disorders including myeloproliferative disorders, monoclonal gammopathies (especially IgA), inflammatory bowel disease, and rheumatoid arthritis.4 This report discusses the neutrophilic dermatoses and their associations with hematologic disorders. Acute Febrile Neutrophilic Dermatosis Acute febrile neutrophilic dermatosis (AFND) was described by Dr. Robert Sweet in 1964 and soon came to be known as Sweet’s syndrome.5 It is a clinically and pathologically distinct entity of unknown etiology6 and is characterized by fever, neutrophilia, erythematous plaques on the limbs, face and neck, and dense dermal neutrophilic infiltration.5 Additional criteria include absence of infection and responsiveness to corticosteroids.5 Although most cases are thought to be idiopathic,7 AFND can be associated with a variety of disorders, including malignancies, infections, autoimmune disorders, pregnancy, drugs, trauma, subacute thyroiditis, Behcet’s syndrome, acute renal failure, and human im

Sweet's syndrome in a patient with idiopathic progressive bilateral sensorineural hearing loss.

Acute febrile neutrophilic dermatosis, or Sweets syndrome, was first described in 1964 and consists of a triad of erythematous cutaneous plaques infiltrated by neutrophils in association with fever and leukocytosis. Sweets syndrome has been reported to be associated with conditions ranging from upper respiratory tract infections to inflammatory bowel disease to rheumatoid arthritis. We report a patient with a 2-year history of Sweets syndrome in whom idiopathic progressive bilateral sensory neural hearing loss (IPBSNHL) developed. IPBSNHL is a suspected immunologically mediated hearing loss first described by McCabe in 1979. On the basis of this association, hearing evaluation should be considered in the initial and subsequent examinations of the patient with Sweets syndrome.

Anti-metastatic prostacyclins inhibit the adhesion of colon carcinoma to endothelial cells by blocking E-selectin expression

Prostacyclins have long been shown to have anti-metastatic activity. One hypothesis is their modulation of cell adhesion molecule (CAM) expression by target organ endothelial cells. We have postulated that prostacyclin, its analogs, and mechanistic mimics decrease colon carcinoma adhesion to cytokine-stimulated endothelial cells by blocking endothelial expression of the adhesion molecule E-selectin, but not the vascular cell adhesion molecule-1 (VCAM-1). Cultured human microvascular endothelial cells (HDMEC) were pre-incubated with prostacyclin (PGI2), dibutyrl-CAMP (dbcAMP), forskolin (FOR), and/or iso-methylbutylxanthine (IBMX) for 15 min, then co-incubated with the cytokine tumor necrosis factor (TNF) for 4 h. HDMEC surface expression of E-selectin and VCAM-1 was evaluated by flow cytometry and ELISA. Adherence of 51Cr-labeled colon carcinoma cells to HDMEC monolayers was then determined. In parallel assays, HDMECs were incubated with anti-E-selectin and anti-VCAM-1 monoclonal antibody (1:100) prior to the addition of tumor cells. Prostacyclins, its analogs, and mimics significantly reduced E-selectin expression by HDMEC, while the reduction of VCAM-1 expression was much less pronounced. Prostacyclins also significantly decreased colon carcinoma adherence to stimulated HDMECs. The inhibition of E-selectin expression, but not VCAM-1 expression, corresponded to the reduction of tumor cell adherence. Prostacyclins effects on tumor adhesion were nullified by pre-incubation with E-selectin antibody. The inhibition of colon carcinoma adherence to cytokine-stimulated endothelial cells treated with prostacyclin, its analogs, and mimics appears to result from blocking endothelial E-selectin, but not VCAM-1, expression. These data support the hypothesis that prostacyclins may exert their anti-metastatic effect, in part, by inhibiting CAM-mediated adherence of colon carcinoma to endothelial cells in metastatic target organs.

Freshly isolated Thy-1+ dendritic epidermal cells express T cell receptor γδ-CD3

Abstract Within murine epidermis exists a population of Thy-1 + DEC which express membrane Thy-1 antigen, but lack CD8 or CD4 antigen. We examined freshly obtained non-cultured Thy-1 + DEC both by immunofluorescence and by biochemical techniques to identify the protein products of the T cell receptor (TCR) and the associated CD3 complex on these cells. Virtually all of the Thy-1 + DEC are brightly positive in CD3 expression with immunofluorescence using the monoclonal antibody 145-2C11. By immunoprecipitation, using this same antibody and polyclonal anti-TCR-γ antibody, the only TCR heterodimer detected on the freshly isolated Thy-1 + DEC is the γδ heterodimer. These findings suggest that in the phenotype and TCR expression, Thy-1 + DEC are analogous to CD8 − , CD4 − early fetal thymocytes.

The role of adhesion molecules in cutaneous inflammation.

The integrity and function of all tissues depend on the ability of their resident cells as well as cells that migrate through them to interact with each other and with matrix proteins in the tissues. These interactions rely in part on the regulated expression of cell adhesion molecules. Cell adhesion molecules are cell surface proteins, glycoproteins, or carbohydrates that play important roles in a wide variety of physiologic and pathophysiologic processes. These include inflammation, wound healing, tumor metastases, cell growth, development and differentiation, immunological responses, and leukocyte trafficking, among others. Cell adhesion molecules interact with their ligands (counterreceptors) on other cells or on structural proteins and allow binding to occur. These molecules have been the focus of tremendous scientific interest and effort in the last decade. The biochemical, functional, and molecular characterization of numerous cell adhesion molecules has enhanced our understanding of important physiologic and pathophysiologic processes that involve cell-cell and cell-matrix binding. This review will focus on a series of cell adhesion molecules relevant to the normal functioning of cutaneous cells or to the evolution of the inflammatory response in skin diseases. Selected cell adhesion molecules will be discussed in regard to their categorization in specific families, their function, and their regulation.

The regulation of cell adhesion molecule gene expression.

Our current understanding of the regulation of expression of the genes encoding various leukocyte adhesion molecules, and in particular their expression by specific cellular components of the skin, reflects in many ways our understanding of gene regulation in general. The questions that may be addressed with respect to particular adhesion molecule genes are in essence individual examples of the fundamental questions of biology: how, when and why do cells do what they do? Whether a biologist is primarily concerned with growth, differentiation, development, homeostasis, oncogenesis, immune or inflammatory responses, or any other aspect of normal or abnormal cell behavior, that behavior can be viewed as entailing two global, closely interrelated, biologic events: a signaling mechanism to or within the cell that commands a response, and differential gene expression that constitutes the response. Throughout the life of any organism, these events, in one form or another, are continuously occurring and recurring in various tissues to varying degrees. Leukocyte cell adhesion molecules play vital roles in the trafficking and localization of leukocyte subsets, and their rapid and exquisitely choreographed expression dictates in part the nature and extent of an inflammatory or immunologic response at particular sites of injury or threat (1-10, 21). As detailed in the accompanying article by Lawley et al., specific adhesion molecules, particularly those expressed by microvascular endothelial cells of