Lectin-like domain of thrombomodulin binds to its specific ligand Lewis Y antigen and neutralizes lipopolysaccharide-induced inflammatory response

Abstract

IntroductionSeptic shock syndrome resulting from excessive host immune responses induced by infectious organisms is a leading cause of death in hospitalized patients. [1][2][3] Pathophysiologic changes in sepsis involve the pathogen-induced uncontrolled release from immune cells, particularly monocytes and macrophages, of proinflammatory mediators. 4 Gram-negative bacterial infection is one of the major causes of systemic bacterial sepsis. 5 Lipopolysaccharide (LPS), a constituent of the Gram-negative outer membrane, is the leading cause of sepsis. LPS induces a rapid increase of proinflammatory mediators, leading to lethal systemic tissue damage and multiple organ failure, which mimics the inflammatory responses of septic syndrome. 6 In mammals, membrane-bound CD14 and toll-like receptor 4 (TLR4)-MD-2 participate in cellular recognition of LPS. 7 Binding of LPS to TLR4 triggers the activation of members of the mitogen-activated protein kinase (MAPK) pathway including p38, p42/p44 extracellular signal-regulated kinase (ERK1/2), and c-Jun N-terminal kinase (JNK). 8 In resting unstimulated cells, nuclear factor-B (NF-B), a heterodimeric complex composed of 50-and 65-kDa (p50/p65) protein subunits, 9 retains as an inactive complex bound to inhibitory B␣ (IB␣) in the cytoplasm. While the cells are under proinflammatory stimulation by LPS, phosphorylation and degradation of IB␣ permit NF-B nuclear translocation and promote the expression of inflammatory genes including inducible nitric oxide synthase (iNOS), tumor necrosis factor-␣ (TNF-␣), and others. 9Thrombomodulin (TM) is a 557 amino acid type I glycosylated transmembrane protein 10 with an NH 2 -terminal lectinlike region (domain 1; D1) followed by 6 epidermal growth factor (EGF)-like structures (domain 2; D2), an O-glycosylation site-rich domain (domain 3; D3), a transmembrane domain (domain 4; D4), and a cytoplasmic tail domain (domain 5; D5). TM domain 2 (TMD2) EGF-like structures are responsible for the anticoagulant activity of TM via the alteration of thrombin substrate specificity. TMD2-thrombin complex sequentially activates anticoagulant protein C inactivating procoagulant cofactors Va and VIIIa. 11 TM expression also occurs in keratinocytes, 12 polymorphonuclear neutrophils (PMNs), 13 monocytes, 14 and endothelial cells, 15 indicating additional functions of TM besides anticoagulation. 16 Indeed TM domains function as an adhesion molecule, 17 an angiogenic factor, 18 and an anti-inflammatory agent through protein C-dependent and -independent mechanisms. 16,19 Recently, anti-inflammatory activity of TM domain 1 (TMD1) was implied by observing that mice with a deleted TM lectinlike domain (TM LeD/LeD ) become more sensitive to LPS challenge through the suppressed expression of adhesion molecules via NFB and MAPK signaling pathways. 20 Moreover, mice with a mutation in the TM gene (TM pro/pro ) strongly reduce the capacity to generate activated protein C, an anti-inflammatory agent in treatment of sepsis. 21 Mice harboring the latter mutation display an u...

Thrombomodulin domain 1 ameliorates diabetic nephropathy in mice via anti-NF-κB/NLRP3 inflammasome-mediated inflammation, enhancement of NRF2 antioxidant activity and inhibition of apoptosis

“…These results show that, although intact THBD can activate protein C and activated protein C (APC) has anti-inflammatory and anticoagulation properties, the effect of THBDD1 in AAV-THBDD1-treated db/db mice was probably independent of APC. This finding is confirmed by our recent observation, in a mouse model of sepsis, that THBDD1 has APC-independent anti-inflammatory activity in vivo and inhibits neutrophil infiltration into the kidney [15]. Collectively, ours and several lines of research have suggested possible mechanisms for the direct anti-inflammatory effects of THBDD1, including: (1) interference with the binding of lipopolysaccharide to its receptor and inhibition of inflammatory mediator production by macrophages [15]; (2) suppression of adhesion molecule expression by neutrophils acting via the NF-κB and MAPK pathways [24]; (3) prevention of leucocyte activation by sequestration of HMGB1 protein [25]; and (4) interference with complement activation (C3a and C5a) via the classical and lectin pathways [26].…”

“…However, studies by our team [13] and others [14] have shown that THBD also has a protein C-and thrombinindependent physiological function. In agreement with this notion, we recently demonstrated that recombinant THBDD1 (rTHBDD1 ) can inhibit an inflammatory response by neutralising lipopolysaccharide through binding to Lewis Y antigen [15] and has anti-angiogenesis effects on cultured HUVECS [16]. We also demonstrated that cardiomyocytes are protected from apoptosis by THBD treatment as a result of its effect on the balance of B cell leukaemia/lymphoma 2 (BCL-2) and BCL-2-associated X protein (BAX) levels [17].…”

Recombinant Thrombomodulin Inhibits Lipopolysaccharide-Induced Inflammatory Response by Blocking the Functions of CD14

“…We also showed that rTMD1, but not the recombinant EGF-like domain plus serine/threonine-rich domain of TM (rTMD23), binds to LPS via the Lewis Y Ag (18). Because rTMD23 has six EGFlike domains, it acts as a novel angiogenic factor by facilitating endothelial cell proliferation and migration (22).…”

“…Because TM is a natural anticoagulant protein, recombinant human soluble TM protein (ART-123) effectively reduces disseminated intravascular coagulation (17). Furthermore, recombinant TM lectin-like domain (rTMD1) suppresses LPS-induced inflammation by binding directly to LPS and high-mobility group box 1 protein (18,19). Although the anti-inflammatory activity of activated protein C has been demonstrated (20), the pulmonary immune responses to respiratory pathogens and LPS in mice with strongly reduced protein C activation (TM pro/pro mice) are not different from those in wild-type mice (21), suggesting that TM can modulate the host inflammatory response through a protein C-independent mechanism.…”

The effects of nitric oxide on the immune system during Trypanosoma cruzi infection

“…More specifically, NO has been demonstrated to protect against infection from T. cruzi (Figs 1, 2) and other protozoa as Toxoplasma gondii, Leishmania major, Leishmania donovani, Plasmodium sp and Schistosoma mansoni (Adams et al 1990, Vespa et al 1994, Wynn et al 1994, James 1995, Stenger et al 1996, Murray & Nathan 1999, Brunet 2001. FurtherFurthermore, the killing activity of NO has also been shown to be effective in host defence against tumour cells (Huerta et al 2008) and alloantigens (Shi et al 2008).…”