Protective Effect of Peripherally and Centrally Applied Ghrelin in the Ischemia-Reperfusion Induced Injury of the Small Intestine

Abstract

Purpose: Radiation therapy is an important modality for cancer treatment as well as chemotherapy and surgery. It has been reported that radiation proctitis occurs in 5 20% of cases treated with pelvic radiation for cancers such as cervical cancer and prostatic cancer. The most common symptom of radiation procitis is rectal bleeding, and it often impairs QOL and sometimes requires blood transfusion. The pathological findings of radiation proctitis are characterized by abnormal angiogenesis and the bleeding from the abnormal vessels. However, the mechanisms by which radiation proctitis develops in humans are totally unknown. Therefore, in this study, the expression profiles of angiogenic factors were analyzed to clarify their role in the etiology of radiation proctitis. Patients and Methods: Eight patients with radiation proctitis (M/F, 6/2; average age, 67 ± 10.4 years) and 8 healthy volunteers (M/F, 5/3; average age, 65.3 ± 7.6 years) were enrolled. Diagnosis of radiation proctitis was made according to the criteria by Cavci et al. Rectal biopsy samples were endoscopically taken and frozen. The protein lysates of the tissues were incubated with biotin-labeled antibody. Then, they were reacted with the antibody array membrane, incubated with streptavidin-horseradish peroxidase (HRP), and visualized using a luminescent image analyzer. Total RNA was extracted from the tissues and reverse transcribed into cDNA. Then, Taqman real-time PCR was performed to evaluate the mRNA level of each factor. Immunohistochemistry was performed using the labeled streptavidin biotin (LSAB) method. Results: Antibody array analysis revealed 2.127.31-fold higher expression levels of angiogenin, FGF1, endoglin, MMP-8, uPA and maspin in radiation proctitis tissues compared with normal rectal mucosa. The mRNA levels of angiogenin, FGF1, maspin, MMP-8, endoglin, and uPA were significantly higher in radiation proctitis tissues than in normal rectal tissues (P < 0.05), indicating that these angiogenic factors were overexpressed by transcriptional activation. Immunohistochemical staining showed strong expression of angiogenin and maspin in rectal epithelia, MMP-8 and uPA in infiltrating lymphocytes, FGF1 in fibroblasts, and endoglin in endothelial cells. The major angiogenic factor, VEGF, was not overexpressed in radiation proctitis tissue compared with the normal rectal mucosa (0.91 1.32-fold). Conclusion: Our results suggest that in radiation proctitis, MMP-8 and uPA cooperatively degrade the extracellular matrix (ECM) and basement membrane to provide space for angiogenesis. Simultaneously, angiogenin and FGF1 promote endothelial cell proliferation, and endoglin induces vessel formation, culminating in angiogenesis. Inhibitors of angiogenic factors such as angiogenin and FGF1 may be effective for treating radiation proctitis.