Altered brain morphology after focal radiation reveals impact of off-target effects: implications for white matter development and neurogenesis

Abstract

Background: Children with brain tumors treated with cranial radiation therapy (RT) often exhibit cognitive late effects, commonly associated with reduced white matter (WM) volume and decreased neurogenesis. The impact of radiation damage in particular regions or tissues on brain development as a whole has not been elucidated.

Methods: We delivered whole-brain or focal radiation (8 Gy single dose) to infant mice. Focal treatments targeted white matter (anterior commissure), neuronal (olfactory bulbs), or neurogenic (subventricular zone) regions. High-resolution ex vivo MRI was used to assess radiation-induced volume differences. Immunohistochemistry for myelin basic protein and doublecortin was performed to assess associated cellular changes within white matter and related to neurogenesis, respectively.

Results: Both whole-brain and focal RT in infancy resulted in volume deficits in young adulthood, with whole-brain RT resulting in the largest deficits. RT of the anterior commissure, surprisingly, showed no impact on its volume or on brain development as a whole. In contrast, RT of the olfactory bulbs resulted in off-target volume reduction in the anterior commissure and decreased subventricular zone neurogenesis. RT of the subventricular zone likewise produced volume deficits in both the olfactory bulbs and the anterior commissure. Similar off-target effects were found in the corpus callosum and parietal cortex.

Conclusions: Our results demonstrate that radiation damage locally can have important off-target consequences for brain development. These data suggest that WM may be less radiosensitive than volume change alone would indicate and have implications for region-sparing radiation treatments aimed at reducing cognitive late effects.

Fig. 1

Planned dose distributions and biological verification of targeting in whole-brain (WB-Irr) and focal irradiation cohorts (AC-, OB-, and LV-Irr). (A) Dose distributions are represented as a colormap overlaid on top of a grayscale image of the average P16 CT image. Cyan colors represent a dose of ~8 Gy. The first and second columns indicate sagittal and axial views, respectively, with the ROI from the registered atlas outlined in white. (B) Representative sagittal sections of P16 mouse brains collected 2 h post-irradiation and stained with γ-H2AX (magenta) show the spatial distribution achieved in vivo (n = 3 mice per cohort). DAPI counterstain is shown in blue to indicate nuclei.

Fig. 2

Voxel-wise comparison of significant volume differences of adult mice (P63) after whole-brain or focal irradiation at infancy (P16) (q < 0.05). The first column shows mid-sagittal views of the average adult mouse brain with a colormap overlay showing significant percent volume differences of irradiated mice compared with controls. The second and third columns show the indicated axial views. White and yellow arrows indicate direct and off-target effects of radiation damage, respectively.

Fig. 3

Structure-wise comparison of volume differences after whole-brain versus focal irradiation. Normalized brain structure volumes of the (A) AC, (B) OBs, (D) GCL of the OBs, (E) LOT, and (F) AON of the OBs are displayed for all irradiation cohorts. (C) The location of the innermost GCL, the LOT, and the outermost GL are highlighted in a coronal view of the OBs. Error bars represent 95% confidence intervals; *q < 0.05 and **q < 0.01 relative to controls.

Fig. 4

Volume differences in central and cortical brain structures after whole-brain and focal irradiations. (A) The location of the AC: pars posterior (AC:pP), parietal cortex (PC), and corpus callosum (CC) in relation to the OBs and cerebellum (Cb) are shown in a mouse brain schematic. (B–D) Brain structure volumes of the AC: pars posterior (B), parietal cortex (C), and corpus callosum (D) for all cohorts are shown. Error bars represent 95% confidence intervals; *q < 0.05 and **q < 0.01 relative to controls.

Fig. 5

MBP staining intensity in P63 brains irradiated at infancy with whole-brain or focal irradiation. Quantification of MBP staining intensity for all irradiation cohorts is shown for the CC (A) and the AC (B). (C) Representative coronal sections are provided showing the AC and CC (n = 3 mice per cohort). Error bars indicate standard error of the mean. **q < 0.01 after one-way ANOVA with Tukey’s post-hoc test.

Fig. 6

The developmental impact of whole-brain and focal irradiations on neurogenesis in the SVZ and DG. Representative coronal sections of the SVZ (A) and the DG (B) in P63 mouse brains irradiated at infancy and stained for DCX (green; left column) (n = 3 mice per cohort). DAPI counterstain is shown in blue (right column). Quantification of DCX+ cell number is shown for the SVZ (C) and DG (D). Error bars indicate standard error of the mean. *q < 0.05, **q < 0.01, ***q < 0.001 after one-way ANOVA corrected with Tukey’s multiple comparisons test.