Association of NF-κB and its downstream pathway with acute radiation-induced myocardial fibrosis in rats

Abstract

Objective To examine the pathological changes in the myocardial tissues such as inflammatory response and fibrosis in a rat model of acute radiation-induced heart damage (RIHD), and to explore whether NF-κB and its downstream pathway are associated with acute radiation-induced myocardial fibrosis. Methods Fourteen male adult Sprague-Dawley rats were randomly divided into control group and radiation group. Local heart irradiation was delivered to the precordial region of rats to establish an RIHD model in a single fraction with a dose of 20 Gy generated by a 6 MV linear accelerator. At 14 days after irradiation, the histopathological changes in myocardial and interstitial tissues were examined by HE staining; the distribution of collagen fibers was observed by Masson staining, and collagen volume fraction (CVF) was used as a semi-quantitative evaluation for myocardial collagen deposition, which was defined as the percentage of collagen area occupied in total area, and was compared using the independent-samples t test. The protein and mRNA expression levels of the NF-κB members p50 and p65 and the downstream pathway members hypoxia-inducible factor 1α(HIF-1α), connective tissue growth factor (CTGF), and type I (COL-1) were quantitatively analyzed by Western blot and qPCR, respectively. Results At 14 days after local heart irradiation, the radiation group showed significant myocardial edema and derangement, rupture of some myocardial cells, mild nuclear pyknosis, darkened nuclear staining, a small number of irregular nuclei, and myocardial interstitial inflammatory cell infiltration accompanied by increased fibroblast, as compared with the control group. The Masson staining showed that the collagen fibers in radiation group were widely distributed at the interstitial tissue and increased significantly compared with those in the control group; normal myocardial cells were in disordered array and had a tendency to be replaced by collagen fibers. The semi-quantitative analysis showed that radiation induced a significant increase in CVF (22.05% vs. 3.76%, P=0.003). Western blot and qPCR revealed that the protein and mRNA expression of p50, p65, HIF-1α, CTGF, and COL-1 was significantly higher in the radiation group than in the control group (all P <0.05). Conclusions The pathological features of acute RIHD include significant myocardial edema and myocardial interstitial inflammatory cell infiltration accompanied by increased fibroblasts and collagen fibers. Radiation exposure can activate NF-κB and cause the upregulation of HIF-1α and CTGF at both protein and mRNA levels, which may play an important role in the progression of radiation-induced myocardial inflammation to fibrosis. Key words: Radiation induced heart disease; Histopathology; Fibrosis; Gene expression; Protein expression