The role of thrombomodulin lectin-like domain in inflammation

Thrombomodulin (TM) is a cell surface glycoprotein which is widely expressed in a variety of cell types. It is a cofactor for thrombin binding that mediates protein C activation and inhibits thrombin activity. In addition to its anticoagulant activity, recent evidence has revealed that TM, especially its lectin-like domain, has potent anti-inflammatory function through a variety of molecular mechanisms. The lectin-like domain of TM plays an important role in suppressing inflammation independent of the TM anticoagulant activity. This article makes an extensive review of the role of TM in inflammation. The molecular targets of TM lectin-like domain have also been elucidated. Recombinant TM protein, especially the TM lectin-like domain may play a promising role in the management of sepsis, glomerulonephritis and arthritis. These data demonstrated the potential therapeutic role of TM in the treatment of inflammatory diseases

Recombinant Zebrafish ␥-Glutamyl Hydrolase Exhibits Properties and Catalytic Activities Comparable with Those of Mammalian Enzyme

ABSTRACT: A cDNA encoding for zebrafish ␥-glutamyl hydrolase (␥GH) was cloned and inserted into a pET43.1a vector via SmaI and EcoRI sites and expressed in Rosetta (DE3) cells as a Nus-His-tag fusion enzyme (NH-z␥GH). After induction with isopropyl thiogalactoside, the enzyme was purified with a Ni-Sepharose column, and approximately 8 mg of pure enzyme was obtained per liter of culture. The primary sequence of the recombinant z␥GH was similar to mammalian ␥GH. Folate is an essential B vitamin and participates in the biosynthesis and metabolism of nucleic acids, proteins, several amino acids, methyl groups, many neurotransmitters, and some vitamins. Mammalian cells are unable to synthesize folates de novo and therefore depend on their food for the supply of folates. Naturally occurring folates are synthesized as poly-␥-glutamate forms (folylpolyglutamate) but are absorbed and transported most efficiently as folylmonoglutamates. The conversion of folylpolyglutamates in dietary food to folylmonoglutamates is catalyzed by carboxypeptidase II (EC 3.4.22.12) in mammals. In a recent study, ␥-glutamyl hydrolase (␥GH, EC 3.4.19.9), a lysosomal cysteine peptidase, was reported to be the enzyme responsible for hydrolyzing dietary folate in rat small intestine Consistent with this notion, the activity of ␥GH to hydrolyze the ␥-glutamyl peptide bonds of folylpolyglutamates has rendered this enzyme a potential target of antifolate chemotherapy and, at the same time, a primary component in regulating the intracellular levels of some antifolate drugs. Antifolate drugs, such as methotrexate, owe much of their effectiveness to being substrates for both folylpoly-␥-glutamate synthetase and ␥GH. Removal of ␥-linked glutamate residues decreases the retention and activity of these drugs. A polymorphism resulting in reduced catalytic activity of ␥GH was observed to be associated with greater accumulation of long-chain methotrexate polyglutamate forms The determination of individual folate derivatives in serum of patients receiving antifolate chemotherapy and in foods is an important current protocol. The first step in these determinations is converting folylpolyglutamates to folylmonoglutamates by ␥GH. Cur- The amino acid numbering used for z␥GH in this study is numbered starting from the first methionine in the full-length peptide with the signal peptide. Article, publication date, and citation information can be found a

Effects of Salvianolic Acid B on Protein Expression in Human Umbilical Vein Endothelial Cells

Salvianolic acid B (Sal B), a pure water-soluble compound extracted from Radix Salviae miltiorrhizae, has been reported to possess potential cardioprotective efficacy. To identify proteins or pathways by which Sal B might exert its protective activities on the cardiovascular system, two-dimensional gel electrophoresis-based comparative proteomics was performed, and proteins altered in their expression level after Sal B treatment were identified by MALDI-TOF MS/MS. Human umbilical vein endothelial cells (HUVECs) were incubated at Sal B concentrations that can be reached in human plasma by pharmacological intervention. Results indicated that caldesmon, an actin-stabilizing protein, was downregulated in Sal B-exposed HUVECs. Proteins that showed increased expression levels upon Sal B treatment were vimentin, T-complex protein 1, protein disulfide isomerase, tropomyosin alpha, heat shock protein beta-1, UBX domain-containing protein 1, alpha enolase, and peroxiredoxin-2. Additionally, Sal B leads to increased phosphorylation of nucleophosmin in a dose-dependent manner and promotes proliferation of HUVECs. We found that Sal B exhibited a coordinated regulation of enzymes and proteins involved in cytoskeletal reorganization, oxidative stress, and cell growth. Our investigation would provide understanding to the endothelium protection information of Sal B

Thrombomodulin Regulates Keratinocyte Differentiation and Promotes Wound Healing

The membrane glycoprotein thrombomodulin (TM) has been implicated in keratinocyte differentiation and wound healing, but its specific function remains undetermined. The epidermis-specific TM knockout mice were generated to investigate the function of TM in these biological processes. Primary cultured keratinocytes obtained from TMlox/lox; K5-Cre mice, in which TM expression was abrogated, underwent abnormal differentiation in response to calcium induction. Poor epidermal differentiation, as evidenced by downregulation of the terminal differentiation markers loricrin and filaggrin, was observed in TMlox/lox; K5-Cre mice. Silencing TM expression in human epithelial cells impaired calcium-induced extracellular signal–regulated kinase pathway activation and subsequent keratinocyte differentiation. Compared with wild-type mice, the cell spreading area and wound closure rate were lower in keratinocytes from TMlox/lox; K5-Cre mice. In addition, the lower density of neovascularization and smaller area of hyperproliferative epithelium contributed to slower wound healing in TMlox/lox; K5-Cre mice than in wild-type mice. Local administration of recombinant TM (rTM) accelerated healing rates in the TM-null skin. These data suggest that TM has a critical role in skin differentiation and wound healing. Furthermore, rTM may hold therapeutic potential for the treatment of nonhealing chronic wounds