Lipoteichoic Acid Induces HO-1 Expression via the TLR2/MyD88/c-Src/NADPH Oxidase Pathway and Nrf2 in Human Tracheal Smooth Muscle Cells

Abstract

Heme oxygenase‐1 (HO‐1) is a stress‐inducible rate‐limiting enzyme in heme degradation, which confers cytoprotection against oxidative injury and provides a vital function in maintaining tissue homeostasis. Increasing reports have indicated that lipoteichoic acid (LTA) exerts as LPS as an immune system‐stimulating agent and plays a role in the pathogenesis of severe inflammatory responses induced by Gram‐positive bacteria infection. Here, we report that LTA is an inducer of HO‐1 and examine the signaling pathways by which LTA regulates HO‐1 expression in human tracheal smooth muscle cells (HTSMCs). LTA induced HO‐1 protein, mRNA expression, and HO‐1 promoter activity. LTA‐stimulated HO‐1 expression was attenuated by transfection with dominant negative mutants of Toll‐like receptor‐2 (TLR‐2) and MyD88, the NADPH oxidase inhibitor (DPI), the ROS scavenger (NAC), and transfection with siRNAs of Src and NF‐E2‐related factor 2 (Nrf2). LTA‐stimulated c‐Src phosphorylation, translocation of p47phox and Nrf2, and ROS production were attenuated by transfection with dominant negative mutants of TLR‐2, MyD88, and c‐Src, and by pretreatment with DPI or NAC. Further studies revealed that LTA induced TLR‐2, MyD88, c‐Src, and p47phox complex formation. These results demonstrated that in HTSMCs, LTA induced ROS generation through the TLR‐2/MyD88/c‐Src/NADPH oxidase pathway, in turn initiates the activation of Nrf2, and ultimately induces HO‐1 expression.

Saturated fatty acids activate TLR-mediated proinflammatory signaling pathways

“…of NADPH oxidase ( 31,32 ). Together, these results suggest that the responsiveness of cells to TLR4 agonists depends in part on ROS tone of cells in culture.…”

Lipopolysaccharide induces ICAM-1 expression via a c-Src/NADPH oxidase/ROS-dependent NF-κB pathway in human pulmonary alveolar epithelial cells

“…Total RNA was isolated using TRIzol according to the protocol of the manufacturer. The cDNA obtained from 0.5 g total RNA was used as a template for PCR amplification, as previously described (11). The primers used were as follows: ICAM-1:…”

“…Previous study also showed that lipoteichoic acid (LTA) induced the formation of a TLR2/MyD88/ TRAF6/c-Src complex in human airway smooth muscle cells (11). Thus we further investigated the physical association of TLR4, MyD88, TRAF6, and c-Src in LPS-induced ICAM-1 expression by immunoprecipitation using anti-c-Src and anti-TRAF6 antibodies subjected to Western blot.…”

“…Activated NADPH oxidase is a multimeric protein complex consisting of at least three cytosolic subunits of p47 phox , p67 phox , and p40 phox . Phosphorylation of p47 phox leads to a conformational change allowing its interaction with p22 phox (11). It has been demonstrated that p47 phox organizes the translocation of other cytosolic factors, hence its designation as "organizer subunit" (11).…”

“…NADPH oxidase is an enzymatic source for the production of reactive oxygen species (ROS) under various pathological conditions (13). The Src family kinases have been shown to regulate NADPH oxidase activation and ROS production (11). Thus, in this study, we investigated the roles of c-Src, NADPH oxidase/ROS, EGFR, and PDGFR in LPS-induced inflammatory responses, including ICAM-1 induction and monocyte adhesion.…”