Bacterial Challenge Stimulates Formation of Arachidonic Acid Metabolites by Human Keratinocytes and Neutrophils In Vitro

Abstract

Although the interactions of bacteria with keratinocytes induce the synthesis of various mediators, the capability of epithelial cells to form arachidonic acid mediators has not been studied, and therefore the first part of this study was initiated. The complex mixture of epithelium-derived mediators suggests that chemoattraction is not their only effect on neutrophils and that they may also affect neutrophil mediator synthesis. The effect of epithelium-derived mediators on neutrophil eicosanoide synthesis was evaluated in the second part of this study. We incubated human keratinocytes with human-pathogenic bacteria for 2 h and harvested the supernatants after 4, 6, 10, and 18 h of culture. Subsequently, the supernatants were coincubated for 5 min with human neutrophils with or without arachidonic acid. The formation of the arachidonic acid metabolites prostaglandin E 2 (PGE 2 ), leukotriene B 4 (LTB 4 ), 12-hydroxyeicosatetraenoic acid (12-HETE), and 15-HETE in keratinocytes and neutrophils was measured by reverse-phase high-pressure liquid chromatography. We demonstrated for the first time that keratinocytes produced significant amounts of LTB 4 and 12-HETE 4 to 6 h after bacterial challenge. Upon stimulation with epithelial supernatants, neutrophils produced significant amounts of PGE 2 , LTB 4 , 12-HETE, and 15-HETE throughout the observation period of 18 h, with a maximum synthesis by supernatants harvested 4 to 10 h after bacterial infection. The results of the study suggest that arachidonic acid mediator formation by epithelial cells following bacterial challenge may act as an early inflammatory signal for the initiation of the immune response. The epithelial supernatants were capable of inducing the formation of arachidonic acid mediators by neutrophils, which may have further regulatory effects on the immune response.Although the mechanisms controlling inflammation are poorly understood, the release of pro-and anti-inflammatory mediators plays a central role in the initiation, perpetuation, and limitation of this process. Bacterial infection is an important stimulus for the synthesis of inflammatory mediators in human tissues. Epithelial cells are the first cell line that interacts with bacteria and causes the increased expression and secretion of a number of cytokines and proinflammatory mediators (24). These mediators are characterized by their ability to chemoattract and activate polymorphonuclear leukocytes (PMNs), suggesting that an important function of epithelial cells is to initiate the mucosal influx of PMNs. The arachidonic acid metabolites leukotriene B 4 (LTB 4 ), 12-hydroxyeicosatetraenoic acid (12-HETE), and 15-HETE regulate PMN functions, including chemotaxis (6, 17, 33, 42), degranulation (13,38,41), and adherence (7), which may indicate the critical role of these metabolites for early events in inflammation.The synthesis of various inflammatory mediators by keratinocytes upon bacterial infection suggests that the induction of chemokinesis is not the only functional change that is induce...

The stage of native biofilm formation determines the gene expression of human β‐defensin‐2, psoriasin, ribonuclease 7 and inflammatory mediators: a novel approach for stimulation of keratinocytes with in situ formed biofilms

“…It has been reported that components of the arachidonic acid cascade, the leukotrienes, synthesized in epithelial cells, are potent inducers of inflammation after bacterial challenge (8). The results of the present study confirm this and show the induction of a key enzyme for the leukotriene B 4 synthesis – 5‐LO.…”

“…Various links have been revealed referring to interactions between the innate and the acquired immune systems, including antimicrobial peptides and regulatory enzymes of the arachidonic acid cascade (8, 26). Epithelial cells upon induction secrete both IL‐8 and leukotriene B 4 in response to A. actinomycetemcomitans (8). In contrast to IL‐8, the enzyme 5‐LO of the arachidonic acid cascade was upregulated by short‐time growth of biofilms.…”

The Prostanoid EP4 Receptor and Its Signaling Pathway

“…EP4 is involved in skin inflammation. It has been suggested that PGE 2 is upregulated within antigen-exposed skin (Ruzicka and Printz, 1982;Eberhard et al, 2002). Kabashima et al (2003) demonstrated that PGE 2 promoted skin immune responses by enhancing the migration and maturation of Langerhans cells through EP4 signaling.…”

Eicosanoids and Keratinocytes in Wound Healing

“…17 Multiple cells typically found in the wound bed are capable of eicosanoid synthesis. 3,8,18,19 Bacteria can induce synthesis of various eicosanoids, 20 although their contribution in acute and chronic wounds remains to be further studied. Free radical-induced conversion of AA into the isoprostanes, 21 which serve as prostaglandin analogs, augments the eicosanoid levels in the wound.…”