SummaryBordetella pertussis, the causative agent of whooping cough, adheres to human monocytes/macrophages by means of a bacterial surface-associated protein, filamentous hemagglutinin (FHA) and the leukocyte integrin, complement receptor 3 (CR3, ~xmfl2, CDllb/CD18). We show that an FHA Arg-Gly-Asp site induces enhanced R pertussis binding to monocytes, and that this enhancement is blocked by antibodies directed against CR3. Enhancement requires a monocyte signal transduction complex, composed of leukocyte response integrin (a~/33) and integrinassociated protein (CD47). This complex is known to upregulate CR3 binding activity. Thus, a bacterial pathogen enhances its own attachment to host cells by coopting a host cell signaling pathway.A tachment of the gram-negative bacterium Bordetella pertuss/s to host cells at or near the respiratory mucosal surface is a crucial feature of whooping cough pathogenesis in humans. Ciliated respiratory epithelial cells and leukocytes are the primary targets for adherence by this organism (1-3). This process leads to respiratory tract colonization, systemic intoxication, and altered host immune cell function. R pertussis attachment involves a bacterial surface-associated and secreted protein, filamentous hemagglutinin (FHA) 1, and host galactose-containing glycoconjugates (4-7). In addition, FHA recognizes a leukocyte B2-integrin, complement receptor type 3 (CR3, CDllb/CD18, ol~2) (8). The biologic significance of FHA-CR3 recognition and R pertuss/s binding to leukocytes in nature may reflect several possible outcomes, including competitive blockade of CR3 by secreted FHA, facilitated delivery of bacterial toxins to host leukocytes, and/or bacterial intracellular entry, survival, and persistence (9-13). A recent study suggests that cross-linking of the fibronectin 1 Abbreviations used in this paper: FHA, filamentous hemagglutinin; lAP, integrin-associated protein; LRI, leukocyte response integrin; MDM, monocyte-derived macrophages; PT, pertussis toxin; RGD, Arg-Gly-Asp.This work has been presented in part at the meeting of the American Federation for Clinical Research, Western Section, Carmel, CA, 5 February 1994, and submitted in abstract form to the Annual Meeting of the American Society for Microbiology, Las Vegas. receptor otsfll on human peripheral monocytes enhances CR3-mediated attachment ofR pertussis via FHA (14). Augmented fl2-integrin-binding activity can be elicited by a number of other receptor-ligand binding interactions, including CD14 recognition of the LPS-LPS binding protein complex (15) and a fl3-integrin-containing receptor signal transduction complex (description follows).FHA and an intrinsic host ligand for CR3, complement fragment iC3b, contain Arg-Gly-Asp (RGD) cell recognition sites. These tripeptide motifs often denote binding domains that are recognized by integrins (16--18). It was initially assumed that the FHA and iC3b RGD sites were directly recognized by CR3. This assumption was based on the following observations: (a) an FHA RGD site-directed muta...

BACKGROUND The authors investigated whether the presence of matrix metalloproteinase‐2 (MMP‐2) and its inducer, CD147, in cancerous esophageal lesions and surrounding tissue might help to predict patient prognosis. METHODS Tissue samples from 101 patients with esophageal squamous cell carcinoma were stained with anti‐CD147 and anti–MMP‐2 antibodies for immunohistochemical analysis. RESULTS CD147 was expressed in cancerous and dysplastic lesions, but not in normal tissue. In contrast, MMP‐2 was detected mainly in normal interstitial tissue adjacent to cancerous lesions, but it was detected also in cancerous lesions in some patients. Pathologic findings demonstrated that the intensity of MMP‐2 staining in normal tissue was associated positively with the depth of tumor infiltration and the stage of disease, whereas MMP‐2 staining in cancerous tissue was associated positively with vascular and lymphatic vessel invasion as well as with immature differentiation of cancer cells. Using a proportional hazard model, including information on CD147 staining patterns within cancerous lesions along with clinical cancer staging, improved the accuracy of predicting patient prognosis. CONCLUSIONS These results suggested that measurement of CD147 and MMP‐2 expression with simple immunohistochemical staining may enhance further the understanding of the pathophysiology of invading tumor cells and, when used in combination with cancer staging, may increase the ability of investigators to predict prognosis in patients with esophageal squamous cell carcinoma. Cancer 2004. © 2004 American Cancer Society.

Dermatophytes cause intractable superficial infections in humans. Arthroderma benhamiae, a zoophilic dermatophyte, triggers severe inflammatory responses in humans, while Trichophyton tonsurans, an anthropophilic dermatophyte, triggers minimal ones. Cytokines and other factors derived from keratinocytes play important roles in inflammatory and immune responses in the skin. The authors performed an in vitro investigation to determine the human keratinocyte cytokine profiles during dermatophyte infection. The human keratinocyte cell line PHK16-0b was infected with A. benhamiae or T. tonsurans for 24 h, and the cytokines secreted were analysed using a human cytokine antibody array. Marked differences were observed in the cytokine profiles of the cells infected with the two dermatophytes. A. benhamiae infection resulted in the secretion of a broad spectrum of cytokines, including proinflammatory cytokines, chemokines, and immunomodulatory cytokines. In contrast, T. tonsurans-infected keratinocytes secreted only limited cytokines, including eotaxin-2, interleukin (IL)-8 and IL-16. cDNA microarray analysis confirmed that A. benhamiae infection upregulated genes encoding IL-1β, IL-2, IL-4, IL-6, IL-10, IL-13, IL-15, IL-16, IL-17 and interferon (IFN)-γ, while T. tonsurans infection upregulated only a few genes, such as those encoding IL-1β and IL-16. RT-PCR demonstrated that infection by both dermatophytes enhanced IL-8 mRNA expression in keratinocytes. These results suggest that A. benhamiae-induced secretion of several cytokines from keratinocytes may be involved in a severe inflammatory response, and that the limited cytokine secretion from keratinocytes in response to T. tonsurans infection may result in a minimal inflammatory response in the skin. These cytokine profiles may aid in proving the clinical features of dermatophytosis.

Bordetella pertussis, the agent of whooping cough, is capable of invading human respiratory epithelial cells. In this study, we investigated the mechanisms by which B. pertussis invades the human lung epithelial cell line A549 and normal human bronchial epithelial (NHBE) cells. In vitro adhesion and invasion assays using both cell types with a virulent B. pertussis strain and its isogenic mutants revealed profound defects in a mutant deficient in filamentous hemagglutinin (FHA) expression. In addition, a mutant in which an FHA Arg-Gly-Asp (RGD) site had been changed to Arg-Ala-Asp had significantly diminished invasiveness, although its adhesiveness was comparable to that of the parental strain. Furthermore, a synthetic RGD-containing hexapeptide inhibited invasion of both cell types by the virulent strain. These results demonstrate that an RGD sequence of FHA is involved in B. pertussis invasion of epithelial cells in vitro. Monoclonal antibodies directed against human alpha5beta1 integrin, but not other integrins, blocked invasion, indicating that this integrin is involved in B. pertussis invasion. Taken together, these findings suggest that B. pertussis FHA may promote invasion of human respiratory epithelial cells through the interaction of its RGD sequence with host cell alpha5beta1 integrin.

The lipophilic yeast Malassezia is an exacerbating factor in atopic dermatitis (AD). Among organisms of the Malassezia species, Malassezia globosa and Malassezia restricta are particularly dominant on the skin of AD patients. However, the precise role of Malassezia yeasts in the pathophysiology of AD remains uncertain. Keratinocytes play a critical role in cutaneous inflammatory and immune responses by secreting cytokines. In this study, we attempted to determine the cytokine secretion profiles of human keratinocytes that were exposed to Malassezia yeasts. The human keratinocyte cell line PHK16-0b was cocultivated with M. globosa or M. restricta for 24 h, and the resulting cytokine secretion profile was analysed using a cytokine antibody array. The keratinocytes responded to the two Malassezia species with different Th2-type cytokine profiles, i.e. M. globosa induced IL-5, IL-10 and IL-13 secretion from the keratinocytes, whereas M. restricta induced IL-4 secretion. Similar results were obtained with primary normal human epidermal keratinocytes. cDNA microarray analysis confirmed that IL-5, IL-10, and IL-13 mRNAs were induced only by M. globosa, while IL-4 mRNA expression was induced only by M. restricta. These findings suggest that M. globosa and M. restricta play a synergistic role in triggering or exacerbating AD by stimulating the Th2 immune response.