International Journal of Radiation Oncology*Biology*Physics
Biology ContributionInhibition of Protease-activated Receptor 1 Ameliorates Intestinal Radiation Mucositis in a Preclinical Rat Model
Introduction
The intestine is an important dose-limiting organ during abdominal and pelvic radiotherapy. Despite advances in treatment delivery techniques, intestinal radiation injury (radiation enteropathy) remains an important obstacle to cancer cures and continues to adversely affect the quality of life of many cancer survivors. Clinically, intestinal radiation toxicity occurs as early (acute) or delayed (chronic) radiation enteropathy, with related but different underlying mechanisms. Pathologically, early radiation enteropathy (“radiation mucositis”) is characterized by epithelial barrier breakdown and mucosal inflammation, whereas delayed radiation enteropathy exhibits prominent vascular sclerosis and intestinal wall fibrosis.
Radiation enteropathy is the result of complex interplay among many dose- and time-dependent pathophysiologic processes, including inflammation, epithelial regeneration, tissue remodeling, and collagen deposition, as well as activation of the coagulation system and, notably, endothelial dysfunction. Endothelial cells are mechanistically involved in early and delayed radiation responses in many normal tissues, including the intestine 1, 2, 3. Exposure to ionizing radiation elicits profound changes in the endothelium, commonly referred to as “endothelial dysfunction.” Loss of thromboresistance due to down-regulation of thrombomodulin and expression of tissue factor, resulting in increased formation of thrombin, is a hallmark feature of radiation-induced endothelial dysfunction 1, 3, 4.
Thrombin is a multifunctional serine protease that plays a central role in blood clotting by converting fibrinogen to fibrin and activating platelets. In addition, thrombin also regulates cell proliferation, inflammation, and tissue remodeling through activation of protease-activated receptors (PARs) (5), a family of 4 G-protein coupled receptors: PAR1, PAR2, PAR3, and PAR4. Protease-activated receptor 1 is the most biologically relevant in inflammation and fibrosis and is prominently up-regulated in irradiated intestine, suggesting a role in radiation enteropathy (3).
Preclinical and clinical studies performed in our and other laboratories demonstrate that so-called “endothelial-oriented” interventions can ameliorate radiation toxicity in several different normal tissues. We have also shown that there is increased thrombin formation, fibrin deposition, and up-regulation of PAR1 in irradiated intestine 3, 4. Although direct inhibition of thrombin ameliorates both early and delayed intestinal radiation injury (4), it is not known to what extent this benefit is due to suppression of thrombin-mediated blood clotting (conversion of fibrinogen to fibrin and stimulation of platelet aggregation) and/or to suppression of PAR1-mediated cellular effects. Because PAR1 is absent in rat platelets (6), this question was addressed using a specific small-molecule PAR1 inhibitor, SCH602539, in a preclinical rat model of radiation enteropathy. The results suggest that, although PAR1 is indeed involved in the development of early radiation toxicity (radiation mucositis), non-PAR1-mediated thrombin effects seem to be important in the mechanisms of development of chronic intestinal radiation fibrosis.
Section snippets
Experimental design
A total of 120 male Sprague-Dawley rats, 43-49 days of age (Harlan, Indianapolis, IN) were housed in conventional cages with free access to tap drinking water and standard mouse chow. All experimental protocols were approved by the University of Arkansas for Medical Sciences Institutional Animal Care and Use Committee.
The rat surgical model for localized small-bowel irradiation was prepared as described previously (7). Briefly, rats underwent bilateral orchiectomy, and a loop of distal ileum
Effect of PAR1 antagonist on early and delayed radiation enteropathy in rats
Radiation-induced histopathologic changes in the vehicle (control) group were similar to those observed in other studies performed in our laboratory (8). Early alterations (2 weeks) consisted mainly of mucosal injury (as measured by mucosal surface area), reactive intestinal wall thickening (as measured by intestinal wall thickening), and inflammatory cell infiltration (as measured by number of myeloperoxidase-positive cells). The delayed changes (26 weeks) included loss of mucosal surface
Discussion
Our data strongly suggest that cellular PAR1 signaling is involved in early radiation enteropathy development (ie, radiation mucositis) but not in delayed injury (ie, fibrosis). These findings have substantial translational significance because (1) PAR1 inhibition has substantially fewer side effects than direct thrombin inhibition (12), and (2) PAR1 blockade inhibits tumor growth and metastasis (reviewed in reference 13), and thus blocking PAR1 to reduce normal tissue toxicity would not be
Conclusions
SCH602539, a selective, small-molecule, nonpeptide antagonist of PAR1, seems to attenuate early intestinal radiation mucositis. This protective effect is likely due to inhibition of the cellular, receptor-mediated thrombin effects rather than to inhibition of blood clotting. The underpinnings of the lack of efficacy of SCH602539 in ameliorating delayed radiation fibrosis need further investigation. Protease-activated receptor 1 blockade has antitumor effects and, compared with direct thrombin
Acknowledgements
This research is supported by the National Institutes of Health (Grant CA-71382), Merck & Company, Inc, and the Veterans Healthcare Administration.
References (20)
- et al.
Is the loss of endothelial thrombomodulin involved in the mechanism of chronicity in late radiation enteropathy?
Radiother Oncol
(1997) - et al.
Deficiency of microvascular thrombomodulin and upregulation of protease-activated receptor 1 in irradiated rat intestine: possible link between endothelial dysfunction and chronic radiation fibrosis
Am J Pathol
(2002) - et al.
Hirudin ameliorates intestinal radiation toxicity in the rat: support for thrombin inhibition as strategy to minimize side effects after radiation therapy and as countermeasure against radiation exposure
J Thromb Haemost
(2004) - et al.
Effects of accelerated fractionation on radiation injury of the small intestine: a new rat model
Int J Radiat Oncol Biol Phys
(1988) - et al.
Recombinant soluble transforming growth factor beta type II receptor ameliorates radiation enteropathy in mice
Gastroenterology
(2000) - et al.
Protease-activated receptor 1 antagonists prevent platelet aggregation and adhesion without affecting thrombin time
Eur J Pharmacol
(2010) - et al.
Endothelial apoptosis as the primary lesion initiating intestinal radiation damage in mice
Science
(2001) Thrombin signalling and protease-activated receptors
Nature
(2000)- et al.
Indications for the presence of an atypical protease-activated receptor on rat platelets
Ann Hematol
(2000) - et al.
Relationship between intestinal fibrosis and histopathologic and morphometric changes in consequential and late radiation enteropathy
Acta Oncol
(1996)
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Conflict of interest: Madhu Chintala is an employee of Merck & Company, Inc. Merck produces SCH602359.