Irsogladine maleate regulates neutrophil migration and E-cadherin expression in gingival epithelium stimulated by Aggregatibacter actinomycetemcomitans
Graphical abstract
Introduction
Periodontitis is an inflammatory condition caused by colonization of the gingival sulcus by periodontopathogenic bacteria. Aggregatibacter actinomycetemcomitans (A. actinomycetemcomitans) is a facultative gram-negative anaerobic coccobaccilus that has the capacity to ferment many sugars, including glucose and fructose, and is strongly implicated as a causative organism in periodontitis [1], [2], [3]. Several virulence factors from A. actinomycetemcomitans have been identified, including lipopolysaccaride, leukotoxin, cytolethal distending toxin, collagenase, and outer membrane protein [4], [5], [6], [7], [8]. In periodontitis, gingival epithelial cells actively contribute to inflammatory processes as they represent the first line of defense against microbial attack. Epithelial cells function as a mechanical barrier against invasion by pathogenic organisms and promote intercellular communication through cell–cell junction complexes, for example, tight junctions, adherens junctions, and gap junctions [9], [10], [11], [12]. In addition, epithelial cells produce inflammatory cytokines and anti-microbial peptides. Therefore, the interaction between epithelial cells and A. actinomycetemcomitans has been suggested to play a significant role in the initiation of periodontitis.
Irsogladine maleate (IM) is known to enhance gap junctional intercellular communication in cultured rabbit gastric epithelial and pancreatic cancer cells [13], [14], and is clinically used as an anti-gastric ulcer agent. Our previous studies have shown that IM counters A. actinomycetemcomitans-induced reduction of the gap junction intercellular communication and the expression of zonula occludens-1, which is a major tight junction structured protein, and obviates the A. actinomycetemcomitans-induced increase in interleukin (IL)-8 levels in the culture of human gingival epithelial cells (HGEC) [11], [12]. Furthermore, IM counters the IL-1β-induced suppression in gap junctional intercellular communication in HGEC [15]. Since IM seems to regulate inflammatory responses induced by the bacterial attack and cytokine stimulation of human epithelial cells, it may be a candidate preventive medicine for periodontal disease.
An accumulation of activated neutrophils in lesional areas is observed in all diseases, and is thought to be involved in the onset of inflammation. As inflammation progresses, the distraction of periodontal tissue occurs with deeper periodontal pockets. The development of periodontal disease seems to be related the progression of inflammatory cell infiltration into deeper periodontal tissues [16]. Previous reports showed that IL-8 is present in diseased human periodontal tissues [17], [18], [19], [20], and the levels of IL-8 in both periodontal tissue and GCF have been correlated with the disease severity [21]. In addition, the expressions of IL-8 in diseased tissue, especially in gingival epithelium, is correlated with the migration of PMNs [17], [22], [23], [24]. Taken together, as Graves et al. suggested, the development of periodontal diseases may be related to the progression of inflammatory cell infiltration into periodontal tissues [16]. Therefore, the blocking of neutrophil activity and regulation of CXC-chemokines represent candidate therapeutic strategies for inflammation.
The junctional epithelium is located at a strategically important interface at the base of the gingival sulcus. E-cadherin, a subclass of cadherin found in stratified squamous epithelium, plays a crucial role in maintaining the structural integrity and function of both adherens and desmosomal epithelial intercellular junctions [25]. In the junctional epithelium, E-cadherin, which is a key molecule involved in forming adherens junctions and desmosomes [25], is known to play an important role against bacterial invasion [26], [27], [28], although the reduction of E-cadherin was observed in inflamed gingival tissue [26], [29]. In addition, Porphyromonas gingivalis or A. actinomycetemcomitans decreased E-cadherin expression in cultured gingival epithelial cells [9], [10]. In gastric mucosal epithelium, the disruption of E-cadherin seems to cause epithelial permeability to decrease [30]. Thus, the breakdown of interconnecting epithelial cell adhesions was suggested to lead to the disruption of the epithelial cell barrier function. Recovery of the barrier function may cause prevent bacterial invasion.
Mitogen-activated protein (MAP) kinases play a central role in mediating intracellular signal transduction and regulating cell functions in HGEC [31], [32], [33]. Three distinct mammalian MAP kinases have been identified: extracellular signal-regulated kinase (ERK or p44/42 MAP kinase), c-Jun kinase or the stress-activated protein kinase (c-JNK or SAP kinase), and p38 MAP kinase. In this study, to clarify the effect and mechanism of IM, we focused on neutrophil migration, the barrier function, and involvement of MAP kinase in the gingival epithelium on stimulation with A. actinomycetemcomitans.
Section snippets
Reagents and antibodies
IM was supplied by Nippon Sinyaku (Kyoto, Japan). Humedia-KB2 medium was obtained from Kurabo (Osaka, JAPAN). Todd-Hewitt broth was obtained from BBLR (Cockeysville, MD). Yeast extract was from Difco Laboratories (Detroit, MI). SB203580 and PD98059 were purchased from Calbiochem (La Jolla, CA). Histopaque 1119 and Histopaque 1077 were purchased from Sigma (St. Louis, MO). ISOGEN was from Wako Pure Chemical Industries (Osaka, Japan). Goat anti-rat CINC-2α antibody, goat anti-mouse E-cadherin
Histological findings of rat gingival epithelium
We divided the preparations into 3 groups: non-applied control rats, A. actinomycetemcomitans-applied rats without IM, and IM-injected rats before A. actinomycetemcomitans application. HE staining indicates that A. actinomycetemcomitans application caused the dilatation of intercellular spaces, and the severe infiltration of PMNs into the gingival epithelium (Fig. 1B). On the other hand, in IM-injected rats before A. actinomycetemcomitans application, the gingival epithelium showed the minimal
Discussion
In the present study, we demonstrated that IM inhibited the A. actinomycetemcomitans-induced inflammatory response in gingival epithelium by suppressing neutrophil migration in vivo and in vitro. In addition, IM recovered the A. actinomycetemcomitans-induced reduction in E-cadherin, suggesting enhancement of the barrier function of gingival epithelium.
Chemokines are a strong inducer of neutrophil chemotaxis and classified into four sub-families depending on the number and spacing of the first
Acknowledgements
We thank the Nippon Shinyaku Co., Ltd. (Kyoto, JAPAN), for donating IM. This study was supported in part by a Grant-in-aid for the encouragement of Young scientists (B) (No. 20791613) from the Japan Society for the Promotion of Science, Japan.
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2018, Japanese Dental Science ReviewCitation Excerpt :Collectively, these findings indicate that irsogladine maleate enhances epithelial barrier function by regulating intercellular junctional complexes through structural proteins in the junctional complex. Irsogladine maleate has been shown to inhibit A. actinomycetemcomitans-induced inflammatory responses in the gingival epithelium by suppressing neutrophil migration in vivo and in vitro [54]. Consistent with this finding, irsogladine maleate reduced the A. actinomycetemcomitans- or P. gingivalis-induced increases in IL-8 and CXCL-1 levels in cultures of human gingival epithelial cells [54,56].
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Houttuynia cordata suppresses the Aggregatibacter actinomycetemcomitans-induced increase of inflammatory-related genes in cultured human gingival epithelial cells
2015, Journal of Dental SciencesCitation Excerpt :Furthermore, real-time-PCR indicated that H. cordata at 0.5 and 5 μg/mL inhibited the A. actinomycetemcomitans-induced increase in the expressions of inflammatory-related genes, IL-8, IL-6, ICAM-1, and MMP-3 in HGECs (Fig. 3). To explore the molecular mechanism of the suppressive effect of H. cordata, we assessed whether H. cordata regulated the phosphorylation of ERK because our previous study showed that an ERK inhibitor blocked A. actinomycetemcomitans-induced mRNA expression of IL-8, ICAM-1, and MMP-3 in HGECs.6,7 Interestingly, H. cordata abrogated the ERK phosphorylation induced by A. actinomycetemcomitans in HGECs (Fig. 4).
Irsogladine maleate regulates the inflammatory related genes in human gingival epithelial cells stimulated by Aggregatibacter actinomycetemcomitans
2013, International ImmunopharmacologyCitation Excerpt :These findings encouraged us to test this mucosal protective effect on periodontitis, revealing that IM counters the A. actinomycetemcomitans-induced reduction in gap-junctional intercellular communication in cultured human gingival epithelial cells [17,18]. Intriguingly, IM also suppressed A. actinomycetemcomitans-induced gingival inflammation by regulating neutrophil migration and E-cadherin expression in the gingival epithelium in vitro and in vivo [19,20]. Accordingly, since IM can regulate junctional function in the gingival epithelium, we concluded that it might prevent periodontitis.
Irsogladine maleate regulates barrier function and neutrophil accumulation in the gingival epithelium
2012, Journal of Oral BiosciencesCitation Excerpt :However, administration of irsogladine maleate, an inhibitor of p38 mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinase (ERK), help recover the A. actinomycetemcomitans-induced reduction in E-cadherin expression at the mRNA and protein levels, suggesting that p38 MAPK and ERK are involved in the reduction of E-cadherin expression [41]. In addition, the exposure of A. actinomycetemcomitans to HGECs induced p38 MAPK and ERK phosphorylation, which was inhibited by irsogladine maleate [41]. Furthermore, pretreatment with irsogladine maleate prevented the tumor necrosis factor (TNF)-α-induced reduction of transepithelial electrical resistance in HGECs [42].