Gene and protein expression of proteinase-activated receptor-1, 2 in a murine model of acute graft vs host disease

Proteinase-activated receptors(PARs)are a novel subclass of seven transmembrane-spanning,G protein-coupled receptors.PAR-1 and PAR-2 are widely expressed in a variety of cells and are found to be involved in many physiological and pathological processes includ-ing inflammation and immune response.However,little is known about the function of PAR-1,2 in acute graft vs host disease(GVHD).In the present study,wefirst detected the expression of PAR-1,2 protein and mRNA in a murine model of acute GVHD using the methods of immunohis-tochemistry,Western blot and quantitative real-time polymerase chain reaction(PCR).Syngeneic hematopoie-tic stem cell transplantation(HSCT)mice served as controls.The relative gene expression level of PAR-1 was significantly increased in the skin,liver,small intestine of allogeneic HSCT mice(in skin:0.039�0.013 vs 0.008�0.002 of controls,P=0.009;in liver:0.165�0.006 vs 0.017�0.006 of controls,P=0.004;in small intestine:0.215�0.009 vs 0.016�0.002 of con-trols,P=0.003),but not in the stomach,lung and kidney of allogeneic HSCT mice(P>0.05).PAR-2 mRNA expression in the liver and small intestine of allogeneic HSCT mice(in liver:0.010�0.002 vs 0.003�0.001 of controls,P=0.008;in small intestine:0.006�0.001 vs 0.003�0.001 of controls,P=0.024)was increased significantly,but PAR-2 mRNA expression in the other organs(P>0.05)was not found to be significantly elevated.PAR-1,2 protein expression was in accordance with the mRNA expression,as shown by Western blot.Using immunohistochemistry the present study demon-strated that there was strong PAR-1,2 immunoreactivity in the epithelial cell and vascular endothelial cell of target organs of acute GVHD.Ourfindings of markedly increased expression of PAR-1,2 in target organs of acute GVHD suggest that PAR-1 and PAR-2 may play an important role in the pathogenesis of acute GVHD.

Significance of endothelial dysfunction in the pathogenesis of early and delayed radiation enteropathy

This review summarizes the current state of knowledge regarding the role of endothelial dysfunction in the pathogenesis of early and delayed intestinal radiation toxicity and discusses various endothelial-oriented interventions aimed at reducing the risk of radiation enteropathy. Studies published in the biomedical literature during the past four decades and cited in PubMed, as well as clinical and laboratory data from our own research program are reviewed. The risk of injury to normal tissues limits the cancer cure rates that can be achieved with radiation therapy. During treatment of abdominal and pelvic tumors, the intestine is frequently a major close-limiting factor. Microvascular injury is a prominent feature of both early （inflammatory）, as well as delayed （fibroproliferative） radiation injuries in the intestine and in many other normal tissues. Evidence from our and other laboratories suggests that endothelial dysfunction, notably a deficiency of endothelial thrombomodulin, plays a key role in the pathogenesis of these radiation responses. Deficient levels of thrombomodulin cause loss of vascular thromboresistance, excessive activation of cellular thrombin receptors by thrombin, and insufficient activation of protein C, a plasma protein with anticoagulant, anti-inflammatory, and cytoprotective properties. These changes are presumed to be critically involved in many aspects of early intestinal radiation toxicity and may sustain the fibroproliferative processes that lead to delayed intestinal dysfunction, fibrosis, and clinical complications. In conclusion, injury of vascular endothelium is important in the pathogenesis of the intestinal radiation response. Endothelial-oriented interventions are appealing strategies to prevent or treat normal tissue toxicity associated with radiation treatment of cancer.

Regulation of intestinal permeability: The role of proteases

The gastrointestinal barrier is-with approximately 400 m^2-the human body's largest surface separating the external environment from the internal milieu. This barrier serves a dual function: permitting the absorption of nutrients, water and electrolytes on the one hand, while limiting host contact with noxious luminal antigens on the other hand. To maintain this selective barrier, junction protein complexes seal the intercellular space between adjacent epithelial cells and regulate the paracellular transport. Increased intestinal permeability is associated with and suggested as a player in the pathophysiology of various gastrointestinal and extraintestinal diseases such as inflammatory bowel disease, celiac disease and type 1 diabetes. The gastrointestinal tract is exposed to high levels of endogenous and exogenous proteases, both in the lumen and in the mucosa. There is increasing evidence to suggest that a dysregulation of the protease/antiprotease balance in the gut contributes to epithelial damage and increased permeability. Excessive proteolysis leads to direct cleavage of intercellular junction proteins, or to opening of the junction proteins via activation of protease activated receptors. In addition, proteases regulate the activity and availability of cytokines and growth factors, which are also known modulators of intestinal permeability. This review aims at outlining the mechanisms by which proteases alter the intestinal permeability. More knowledge on the role of proteases in mucosal homeostasis and gastrointestinal barrier function will definitely contribute to the identification of new therapeutic targets for permeability-related diseases.

Visceral hypersensitivity in inflammatory bowel diseases and irritable bowel syndrome: The role of proteases

Proteases, enzymes catalyzing the hydrolysis of peptide bonds, are present at high concentrations in the gastrointestinal tract. Besides their well-known role in the digestive process, they also function as signaling molecules through the activation of protease-activated receptors(PARs). Based on their chemical mechanism for catalysis, proteases can be classified into several classes: serine, cysteine, aspartic, metallo- and threonine proteases represent the mammalian protease families. In particular, the class of serine proteases will play a significant role in this review. In the last decades, proteases have been suggested to play a key role in the pathogenesis of visceral hypersensitivity, which is a major factor contributing to abdominal pain in patients with inflammatory bowel diseases and/or irritable bowel syndrome. So far, only a few preclinical animal studies have investigated the effect of protease inhibitors specifically on visceral sensitivity while their effect on inflammation is described in more detail. In our accompanying review we describe their effect on gastrointestinal permeability. On account of their promising results in the field of visceral hypersensitivity, further research is warranted. The aim of this review is to give an overview on the concept of visceral hypersensitivity as well as on the physiological and pathophysiological functions of proteases herein.

Possible biological and translational significance of mast cells density in colorectal cancer

Mast cells (MCs), located ubiquitously near blood vessels, are descended from CD34+ hematopoietic stem cells. Initially, although their role has been well defined in hypersensitivity reactions, the discovery of their sharing in both innate and adaptive immunity has allowed to redefine their crucial interplay on the regulatory function between inflammatory and tumor cells through the release of mediators granule-associated (mainly tryptase and vascular endothelial growth factor). In particular, in several animal and human malignancies it has been well demonstrated that activated c-Kit receptor (c-KitR) and tryptase (an agonist of the proteinase-activated receptor-2) take pivotal part in tumor angiogenesis after the MCs activation, contributing to tumor cells invasion and metastasis. In this review, we focused on crucial MCs density (MCD) role in colorectal cancer (CRC) development and progression angiogenesis-mediated; then, we will analyze the principal studies that have focused on MCD as possible prognostic factor. Finally, we will consider a possible role of MCD as novel therapeutic target mainly by c-KitR tyrosine kinase inhibitors (imatinib, masitinib) and tryptase inhibitors (gabexate and nafamostat mesylate) with the aim to prevent CRC progression.