Acquired Immunity Is Not Essential for Radiation-Induced Heart Dysfunction but Exerts a Complex Impact on Injury

Figure 1. Salt-Sensitive (SS) wild-type (WT) rats have increased concentrations of circulating interleukin 2 (IL-2) and IL-13 after localized cardiac radiation. Adult SS WT female rats were administered either 24 Gy of localized cardiac radiation in one fraction or sham radiation. (A) A statistically significant increased concentration of IL-2 was detected in plasma 10 weeks post-radiation therapy (RT) compared to sham-treated. (B) Statistically significant increases in IL-13 were detected in plasma at 10 weeks post-RT compared to plasma from age-matched sham-treated rats. Plasma was collected at the time of harvest at one or 10 weeks post-RT (n = 7–10/group, * p < 0.05).

Figure 2. The left ventricles in SS rats have increased T-cell infiltration after localized cardiac radiation compared to sham-treated rats. Adult SS WT female rats were administered either 24 Gy of localized cardiac radiation in one fraction or sham radiation. Representative images of cluster of differentiation 3 (CD3) staining in heart tissue of rats given either (A) sham or (B) 24 Gy RT. Representative images of CD8α staining in heart tissue of rats given either (C) sham or (D) 24 Gy RT. T-cell population staining was quantified as the average number of cells per high power field for CD3 (E) and CD8α (F) stained cell populations (values are means ± standard error of the mean (SEM); n = 3–5/group, * p < 0.01). All images are at 40× magnification.

Figure 3. SS interleukin-2 receptor gamma chain knockout (IL2RG−/−) rats exhibit no significant differences in pleural or pericardial effusions after 24 Gy localized heart RT treatment, compared with the SS WT immune-competent rats. Adult female rats were treated with 24 Gy of image-guided cardiac radiation therapy (RT). Pericardial effusions were given a score of 0–4 during echocardiogram at three months and five months post-RT for (A) females and (B) males, with the group average scores displayed. Pleural effusions were collected and quantified at harvest, and normalized to body weight with the (C) females having larger amounts of effusion compared to (D) males, but no statistical difference was seen between the SS IL2RG−/− and immune-competent SS WT rats. No control rats had pericardial or pleural effusions (not shown). Values are means ± SEM; n = 9–15 rats/group. No p-values comparing the WT versus IL2RG−/− rats were <0.05.

Figure 4. Immune-competent SS WT female rats exhibit cardiac hypertrophy after cardiac radiation, but no hypertrophy occurs in female SS IL2RG−/− rats or in male WT or IL2RG−/− rats. Heart-to-body-weight ratios were measured at harvest in adult SS WT immune-competent and SS IL2RG−/− (A) female and (B) male rats that were given either sham or 24 Gy RT. At five months, hypertrophy was evident in the female immune-competent SS WT rats that received RT versus sham treatment. At five months, cardiac hypertrophy did not occur in the SS IL2RG−/− female rats. Male rats did not display hypertrophy after 24 Gy (B). Values were normalized to the sham-treated animal group average and are the means ± SEM. n = 8–15 rats/radiation group and 4–6 rats/control group; * p < 0.05.

Figure 5. Immune-compromised SS IL2RG−/− male and female rats receiving localized high-dose cardiac RT have echocardiogram parameters revealing worse heart function compared to SS WT rats. SS WT and IL2RG−/− rats were given 24 Gy × 1 localized cardiac RT and echocardiograms were performed, which revealed that the left-ventricular internal diameter at end-diastole (LVIDd) (A: female; B: male) and at end-systole (LVIDs) (C: female; D: male) were significantly increased at three months post-RT in SS IL2RG−/− versus SS WT rats, indicating worse function. In male rats, the significantly elevated LVIDs persisted at five months. End-systolic volume (ESV) was not significantly different in female rats, (E) but ESV was significantly elevated at three months post-RT in male IL2RG−/− versus WT rats (F), which can be associated with worse cardiac function. In females, poorer ejection fraction (EF) (G) and fractional shortening (FS) (I) values were seen in the IL2RG−/− versus WT rats post-RT (not statistically significant). In males, EF (H) and FS (J) were significantly decreased in IL2RG−/− rats at three months post-RT, and FS remained decreased at five months, indicating poorer function. Stroke…

Figure 6. Irradiated SS WT male rat hearts display trends of increased T-cell infiltration compared to sham-treated rats, but have less T-cell infiltration than SS WT female rats. Adult SS WT male rats were administered either 24 Gy of localized cardiac radiation in one fraction or sham radiation. Representative images of CD3 staining in heart tissue of rats given either (A) sham or (B) 24 Gy RT. Representative images of CD8α staining in heart tissue of rats given either sham (C) or 24 Gy RT (D). T-cell population staining was quantified as the average number of cells per high-power field for CD3 (E, p = 0.057 for sham versus 24 Gy) and CD8α (F) stained cell populations (values are means ± SEM; n = 3–5/group, * p < 0.01). Values are means ± SEM; n = 3–5 rats/group, * p < 0.05. All images are at 40× magnification.

Figure 7. SS IL2RG−/− immune-compromised rats versus SS WT immune-competent rats receiving fractionated 9 Gy × 5 cardiac radiation have slightly less cardiac damage measured via echocardiogram. Female SS IL2RG−/− and SS WT rats received 9 Gy × 5 localized cardiac radiation, and echocardiograms were performed at three and five months post-RT revealing that (A) left-ventricular internal diameter at end-diastole (LVIDd) was not significantly different between the groups at three or five months after 9 Gy × 5. (B) LVID at end-systole (LVIDs) and (C) end-systolic volume (ESV) were significantly lower in the SS IL2RG−/− rats versus SS WT rats at three months, which can be associated with improved cardiac function. The (D) ejection fraction (EF) and (E) fractional shortening (FS) were increased in SS IL2RG−/− versus SS WT rats, indicating improved function. However, these changes did not persist at five months post-RT. (F) Stroke volume (SV) was significantly elevated at five months post-RT in the IL2RG−/− rats compared to WT, also indicating improved cardiac function. Values are means ± SEM; n = 11/WT and n = 6/IL2RG−/−; * p < 0.05, # p < 0.01. Representative images of CD3 staining in heart…

Figure 8. The consomic SS.BN3 rats exhibit some differences in cardiac function from the SS rat strain when comparing immune-compromised (IL2RG−/−) versus WT rats after single-dose cardiac RT. After receiving 24 Gy of localized cardiac RT, only the male SS.BN3 IL2RG−/− rats had echocardiogram parameters indicating poorer heart function, while both male and female SS IL2RG−/− rats had parameters indicating poorer cardiac function compared to WT SS rats (Figure 5). The left-ventricular internal diameter at end-diastole (LVIDd) (A: female; B: male) was not significantly different in SS.BN3 IL2RG−/− versus SS.BN3 WT rats, and LVIDs (C: female; D: male), had significantly decreased values in IL2RG−/− rats only in males at three months post-RT (D). End-systolic volume (ESV) was not significantly different in female (E) but was increased at three months post-RT in male (F) SS.BN3 IL2RG−/− versus SS.BN3 WT rats. In females, ejection fraction (EF) (G) and fractional shortening (FS) (I) were not significantly different, whereas, in males, EF (H) and FS (J) were significantly decreased at three months post-RT in the SS.BN3 IL2RG−/− rats, indicating poorer cardiac function. Stroke volume (SV) was not…