Disruption of CSF-1 receptor–mediated metal ion homeostasis in the murine brain promotes neurodegenerative disease
Violeta Chițu, Julia Alvarenga, Wenna Chen, David Reynolds, Yang Liu, Daqian Sun, Anders Sandell, Virginjia Danylaité-Karrenbauer, Per Uvdal, Iran A.N da Silva, Christophe Sandt, Oxana Klementieva, Ulf Johansson, Kavitha Subramanian Vignesh, Zbigniew K. Wszolek, Dennis W. Dickson, Jennifer T. Aguilian, Simone Sidoli, Deyou Zheng, E. Richard Stanley
Violeta Chițu, Julia Alvarenga, Wenna Chen, David Reynolds, Yang Liu, Daqian Sun, Anders Sandell, Virginjia Danylaité-Karrenbauer, Per Uvdal, Iran A.N da Silva, Christophe Sandt, Oxana Klementieva, Ulf Johansson, Kavitha Subramanian Vignesh, Zbigniew K. Wszolek, Dennis W. Dickson, Jennifer T. Aguilian, Simone Sidoli, Deyou Zheng, E. Richard Stanley
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Research Article Cell biology Neuroscience

Disruption of CSF-1 receptor–mediated metal ion homeostasis in the murine brain promotes neurodegenerative disease

Abstract

Dominant-inactivating mutations in the colony stimulating factor-1 receptor (CSF1R) cause CSF-1R–related leukoencephalopathy (CRL), an adult-onset neurodegenerative disease that is modeled in the Csf1r+/– mouse. CRL is caused by microglial dysfunction. However, the primary microglial deficit is unknown. To address this question, we employed single-nucleus RNA sequencing of brains from young Csf1r+/– mice without pathological or behavioral alterations. Reduction of CSF-1R signaling caused metal ion accumulation in brain macrophages, with concomitant activation of cell death and stress response pathways in oligodendrocytes and neuronal subpopulations. Reduction of metallothionein 1 (Mt1) and 3 (Mt3) gene expression was a common feature in glial and neuronal cells of Csf1r+/– mice. Overexpression of Mt1 restored metal ion homeostasis, normalized ROS production in microglia, and prevented the development of behavioral deficits, while Mt3 deletion had disease-enhancing effects. These findings demonstrate CSF-1R regulation of metal ion homeostasis via metallothioneins in the brain.

Authors

Violeta Chițu, Julia Alvarenga, Wenna Chen, David Reynolds, Yang Liu, Daqian Sun, Anders Sandell, Virginjia Danylaité-Karrenbauer, Per Uvdal, Iran A.N da Silva, Christophe Sandt, Oxana Klementieva, Ulf Johansson, Kavitha Subramanian Vignesh, Zbigniew K. Wszolek, Dennis W. Dickson, Jennifer T. Aguilian, Simone Sidoli, Deyou Zheng, E. Richard Stanley

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Figure 2

Csf1r+/– heterozygosity alters mitochondrial function and causes transitional metal ion accumulation in brain macrophages.

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Csf1r+/– heterozygosity alters mitochondrial function and causes transi...
(A) UMAP plot showing the distribution of WT and Csf1r+/–microglia in individual subclusters. (B) Representation of WT and Csf1r+/– nuclei in each subcluster. (C) Volcano plot showing DEGs between WT and Csf1r+/– microglia, indicating altered expression of transcripts encoding proteins involved in immune function, energy production and metabolism, and ion transport and homeostasis. (D) Expression of selected genes that define homeostatic/protective and inflammatory microglial states. (E) Examples of dysregulated transcripts related to ion transport, including metallothioneins 2 and 3 (Mt2 and Mt3). (F) Ingenuity Pathway Analysis–based (IPA-based) prediction of biological processes affected by Csf1r heterozygosity in microglia. The column labels indicate involvement of MTs in the process. (G) IPA-based prediction of pathways affected by Csf1r heterozygosity in microglia. Orange, activated; blue, inhibited. (H and I) Distribution of Zn2+ (H) and Cu+ (I) in the brains of young (2-month-old) WT and Csf1r+/– mice. Filled gray curves, unstained control; green lined unfilled curve, WT; filled orange curves, Csf1r+/–. Each symbol on the chart represents 1 mouse. Means ± SEM; Student’s 1-tailed paired t test. Note: Because CD11b is expressed in both microglia and perivascular macrophages, throughout this report we refer to cells isolated by CD11b expression as brain macrophages. However, ~90% of mononuclear phagocytes in the brain are microglia (79). (J) Representative XRF images showing the distribution and sulfur content-normalized abundance of Fe, Ca, Zn, and Cu in BMM at steady state in vitro. The color scale indicates the strength of the signal. (K) Example of partial colocalization of Fe-enriched (XRF, upper panel) and lipid-rich (OPTIR, lower panel) areas within a cell. The dotted lines delineate the boundaries of the cell and nucleus. (L) Quantification of the normalized abundance of Fe, Ca, Zn, and Cu in BMM (WT, black; Csf1r+/–, red). Each Csf1r+/– value is normalized to the corresponding WT value (set to 1). Box height: 75% to 25%, whiskers: 1 SD. *P < 0.05, ***P < 0.0005 (Student’s t test). Macrophages were obtained from 5 mice/genotype.