CSF1R inhibition during cranial radiotherapy reshapes glial dynamics via microglial loss, monocyte engraftment, and accelerated astrocyte reactivity

BackgroundCranial radiotherapy (cRT), a common treatment for central nervous system tumors, induces progressive cognitive problems in over half of patients. In mice, microglial depletion via CSF1R inhibition can mitigate this effect, but the underlying cellular mechanisms remain unclear. We hypothesized that CSF1R inhibition-induced microglial ablation and repopulation improves brain health by modulating microglial reactivity to radiotherapy, which attenuates glial responses to radiotherapy. MethodsNine-week-old male C57BL/6JRj mice received either a CSF1R-inhibitor supplemented diet (pexidartinib, PLX3397) or control diet, followed by fractionated CT-guided cRT (30 Gy) or sham treatment. The pexidartinib diet was discontinued 10 days post-radiotherapy. Animals were sacrificed at three intervals post-radiotherapy, allowing the assessment of temporal changes. Multiple brain regions were assessed by immunohistochemistry for markers of microglia, astrocytes, oligodendrocytes and proliferating cells. Microglial morphological changes were assessed using the semi-automated microglia morphology analysis pipeline mGlia. ResultsRadiotherapy alone reduced microglial numbers and induced a progressive reactive morphology; mild at 30 days and pronounced at 6 months post cRT. CSF1R inhibition before cRT markedly decreased microglial markers but increased general macrophage markers at 30 days and 6 months after cRT, consistent with monocyte-derived cell engraftment. Morphometric analysis revealed rapid and severe morphological change towards a reactive morphotype at 30 days that persisted until 6 months. Microglial depletion did not prevent loss of neurogenesis or oligodendrocyte progenitor cells (OPCs) and accelerated reactive astrogliosis, though partial OPC recovery in the hippocampus and thalamus was observed at 6 months. ConclusionCSF1R inhibition combined with cRT accelerates reactive gliosis and monocyte-derived macrophage engraftment without protecting vulnerable neural cell populations, though limited long-term OPC recovery occurred. Thus, with this set-up CSF1R inhibition-induced microglial ablation and repopulation does not improve overall brain health, but is beneficial for OPCs on the long term after cRT. Key pointsO_LIPexidartinib and cranial radiotherapy have synergistic effects on microglial ablation C_LIO_LIInfiltrating monocytes repopulate the irradiated brain once the pexidartinib diet is discontinued C_LIO_LIIn the absence of microglia, astrocytes show an accelerated reactivity to radiotherapy C_LIO_LILong-term after cranial radiotherapy oligodendrocyte progenitor cell repopulation is enhanced in the pexidartinib treated animals C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=145 SRC="FIGDIR/small/681366v1_ufig1.gif" ALT="Figure 1"> View larger version (59K): org.highwire.dtl.DTLVardef@1153d64org.highwire.dtl.DTLVardef@171a100org.highwire.dtl.DTLVardef@1013e1borg.highwire.dtl.DTLVardef@949893_HPS_FORMAT_FIGEXP M_FIG C_FIG