Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: A CSF1R mRNA (upper panel) and protein domain structure (lower panel). The boxes in the upper panel (mRNA) are exons, and the arrow indicates the CSF1R-T567M mutation. The arrow in the lower panel (protein) indicates the position of the T567M mutation in the CSF1R protein. S signal peptide, Ig immunoglobulin-like domain, TM transmembrane domain, JMD juxta-membrane domain, TKD tyrosine kinase domain, KID kinase insert domain. B Sanger sequencing electropherograms of control and CSF1R-MT cDNA derived from human blood. The red arrows indicate the point mutation site at T567M in the CSF1R-RD patient. C SH-SY5Y cells stably expressing CSF1R-WT (WT) and CSF1R-MT (MT) were collected for western blotting. D – G SH-SY5Y cells stably expressing CSF1R-WT and CSF1R-MT were treated with CSF1 for 0, 5, 15, and 40 min before western blot analysis of pY723-CSF1R, pY708-CSF1R, and pY546-CSF1R in ( D ). Statistical analysis for pY723 ( E ), pY708 ( F ), and pY546 ( G ) from ( D ), N = 3. H – L BV2 cells stably expressing CSF1R-WT and CSF1R-MT were then used for western blot analysis ( H ). Statistical analysis of CSF1R ( I ), pY546-CSF1R/CSF1R ( J ), IBA1 ( K ), and LC3-II/I ( L ) from ( H ), N = 3. Two-tailed Student’s t -test was used to compare the differences between the two groups for ( C ) and ( I – L ). Two-way ANOVA with Tukey’s post hoc test was used to compare different groups for ( E – G ). Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05 and ** p < 0.01.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: Mutagenesis, Membrane, Sequencing, Control, Derivative Assay, Stable Transfection, Expressing, Western Blot, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: CSF1R-T567M mutant (MT) and corrected (Ctrl) iPSCs derived from patients were used to differentiate into microglia (iMGL). A Representative images of P2RY12 + and TMEM119 + iMGL. Scale bar = 10 µm. B Comparison of area, branch number, and maximum branch length between Ctrl and MT iMGL with IBA1 + TMEM119 + staining. Cells from 50 fields of view were counted for each group. Scale bar = 5 µm. Statistical analysis of IBA1 + iMGL cell area ( C ), branches per cell ( D ), and maximum branch length ( E ) in ( B ). F Cytokine profiling assay after 24 h of stimulation with 100 ng/mL LPS, showing significant upregulation of candidate cytokines. RT-qPCR of CCL2 ( G ) and IL-18 ( H ) transcription in Ctrl and MT iMGL. N = 3. I Migration assay of Ctrl and MT iMGL after stimulation with ATP. White arrowheads indicate some of the migrated iMGL. Cells from 15 fields of view were counted for each group. Scale bar = 5 µm. J Effect of the CSF1R-MT on the phagocytic ability of green fluorescent latex beads (white arrowheads). The iMGL were stained with IBA1 (red signals). N = 8. Scale bar = 10 µm. K Effect of the CSF1R-MT on the phagocytic ability of the pHrodo-labeled myelin (red signals) in iMGL. Number of cells quantified: Ctrl = 76, MT = 61. Scale bar = 10 µm. Two-tailed Student’s t -test was used to compare the differences between the two groups. Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05 and *** p < 0.001.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: Mutagenesis, Derivative Assay, Comparison, Staining, Quantitative RT-PCR, Migration, Labeling, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: Three batches of D30 iMGL were used to conduct bulk RNA-sequencing. A Venn diagram showing co-expressed and independently expressed genes in Ctrl and MT iMGL. B Volcano plot of differentially expressed genes between Ctrl and MT iMGL. C Gene Ontology analysis of top-downregulated genes. Some biological processes, cellular components, and molecular functions are downregulated in the MT iMGL. D KEGG pathway analysis of downregulated genes. ECM-receptor interaction, axon guidance, and PI3K-Akt signaling pathway are downregulated in CSF1R-MT iMGL. E Gene Ontology analysis of upregulated genes. Immune receptor activity is upregulated in the MT iMGL. F qPCR analysis of NLRP1 and TUBB4A mRNA expression. Two-tailed Student’s t -test was used to compare the differences between the two groups, N = 3. Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05 and ** p < 0.01.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: RNA Sequencing, Activity Assay, Expressing, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: NPCs and neurons differentiated from the Ctrl and MT iPSCs were used to study the effect of CSF1R-MT on neuronal functions. A – C NPCs were stained with Ki67 and EdU. A White arrowheads indicate Ki67/EdU double-positive NPCs. Scale bar = 25 μm. Statistical analysis of Ki67-positive NPCs ( B ) and EdU-positive NPCs ( C ) in ( A ). Cells from 18 fields of view were counted for each group. D Western blot analysis of CSF1R and TUJ1 in Ctrl and MT iPSC-derived neurons. N = 3. E Representative images of Ctrl and MT iPSC-derived neurons (D19), stained by TUJ1. Sholl analysis of dendrite number per cell ( F ) and number of crossings ( G ) in ( E ). N = 7. Scale bar = 5 μm. Two-tailed Student’s t -test was used to compare the two groups. Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: Staining, Western Blot, Derivative Assay, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: A Representative images of each stage of cerebral organoid (CO) generation. B , C Immunofluorescence staining of MAP2 at day 60 in MT and Ctrl COs. N = 10. D – J D130 COs were used for whole-cell patch-clamp recordings. N = 11. The membrane capacitance ( D ), resting membrane potential (RMP) ( E ), and input resistance ( F ) were recorded. G Representative traces and quantification of voltage-gated currents in MT and Ctrl COs. H Analysis of neuronal firing ability in MT and Ctrl COs. I Representative traces and quantification of spontaneous excitatory postsynaptic current (sEPSC) in MT and Ctrl COs. J Representative traces and quantification of spontaneous inhibitory postsynaptic current (sIPSC) in CSF1R-MT and Ctrl COs. Two-tailed Student’s t -test was used to compare the differences between the two groups ( C , D – F , I , J ). Two-way ANOVA with Tukey’s post hoc test was used to compare the two groups ( G , H ). Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: Immunofluorescence, Staining, Patch Clamp, Membrane, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: A CSF1R mRNA (upper panel) and protein domain structure (lower panel). The boxes in the upper panel (mRNA) are exons, and the arrow indicates the CSF1R-T567M mutation. The arrow in the lower panel (protein) indicates the position of the T567M mutation in the CSF1R protein. S signal peptide, Ig immunoglobulin-like domain, TM transmembrane domain, JMD juxta-membrane domain, TKD tyrosine kinase domain, KID kinase insert domain. B Sanger sequencing electropherograms of control and CSF1R-MT cDNA derived from human blood. The red arrows indicate the point mutation site at T567M in the CSF1R-RD patient. C SH-SY5Y cells stably expressing CSF1R-WT (WT) and CSF1R-MT (MT) were collected for western blotting. D – G SH-SY5Y cells stably expressing CSF1R-WT and CSF1R-MT were treated with CSF1 for 0, 5, 15, and 40 min before western blot analysis of pY723-CSF1R, pY708-CSF1R, and pY546-CSF1R in ( D ). Statistical analysis for pY723 ( E ), pY708 ( F ), and pY546 ( G ) from ( D ), N = 3. H – L BV2 cells stably expressing CSF1R-WT and CSF1R-MT were then used for western blot analysis ( H ). Statistical analysis of CSF1R ( I ), pY546-CSF1R/CSF1R ( J ), IBA1 ( K ), and LC3-II/I ( L ) from ( H ), N = 3. Two-tailed Student’s t -test was used to compare the differences between the two groups for ( C ) and ( I – L ). Two-way ANOVA with Tukey’s post hoc test was used to compare different groups for ( E – G ). Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05 and ** p < 0.01.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: Mutagenesis, Membrane, Sequencing, Control, Derivative Assay, Stable Transfection, Expressing, Western Blot, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: CSF1R-T567M mutant (MT) and corrected (Ctrl) iPSCs derived from patients were used to differentiate into microglia (iMGL). A Representative images of P2RY12 + and TMEM119 + iMGL. Scale bar = 10 µm. B Comparison of area, branch number, and maximum branch length between Ctrl and MT iMGL with IBA1 + TMEM119 + staining. Cells from 50 fields of view were counted for each group. Scale bar = 5 µm. Statistical analysis of IBA1 + iMGL cell area ( C ), branches per cell ( D ), and maximum branch length ( E ) in ( B ). F Cytokine profiling assay after 24 h of stimulation with 100 ng/mL LPS, showing significant upregulation of candidate cytokines. RT-qPCR of CCL2 ( G ) and IL-18 ( H ) transcription in Ctrl and MT iMGL. N = 3. I Migration assay of Ctrl and MT iMGL after stimulation with ATP. White arrowheads indicate some of the migrated iMGL. Cells from 15 fields of view were counted for each group. Scale bar = 5 µm. J Effect of the CSF1R-MT on the phagocytic ability of green fluorescent latex beads (white arrowheads). The iMGL were stained with IBA1 (red signals). N = 8. Scale bar = 10 µm. K Effect of the CSF1R-MT on the phagocytic ability of the pHrodo-labeled myelin (red signals) in iMGL. Number of cells quantified: Ctrl = 76, MT = 61. Scale bar = 10 µm. Two-tailed Student’s t -test was used to compare the differences between the two groups. Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05 and *** p < 0.001.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: Mutagenesis, Derivative Assay, Comparison, Staining, Quantitative RT-PCR, Migration, Labeling, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: Three batches of D30 iMGL were used to conduct bulk RNA-sequencing. A Venn diagram showing co-expressed and independently expressed genes in Ctrl and MT iMGL. B Volcano plot of differentially expressed genes between Ctrl and MT iMGL. C Gene Ontology analysis of top-downregulated genes. Some biological processes, cellular components, and molecular functions are downregulated in the MT iMGL. D KEGG pathway analysis of downregulated genes. ECM-receptor interaction, axon guidance, and PI3K-Akt signaling pathway are downregulated in CSF1R-MT iMGL. E Gene Ontology analysis of upregulated genes. Immune receptor activity is upregulated in the MT iMGL. F qPCR analysis of NLRP1 and TUBB4A mRNA expression. Two-tailed Student’s t -test was used to compare the differences between the two groups, N = 3. Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05 and ** p < 0.01.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: RNA Sequencing, Activity Assay, Expressing, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: NPCs and neurons differentiated from the Ctrl and MT iPSCs were used to study the effect of CSF1R-MT on neuronal functions. A – C NPCs were stained with Ki67 and EdU. A White arrowheads indicate Ki67/EdU double-positive NPCs. Scale bar = 25 μm. Statistical analysis of Ki67-positive NPCs ( B ) and EdU-positive NPCs ( C ) in ( A ). Cells from 18 fields of view were counted for each group. D Western blot analysis of CSF1R and TUJ1 in Ctrl and MT iPSC-derived neurons. N = 3. E Representative images of Ctrl and MT iPSC-derived neurons (D19), stained by TUJ1. Sholl analysis of dendrite number per cell ( F ) and number of crossings ( G ) in ( E ). N = 7. Scale bar = 5 μm. Two-tailed Student’s t -test was used to compare the two groups. Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: Staining, Western Blot, Derivative Assay, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: A Representative images of each stage of cerebral organoid (CO) generation. B , C Immunofluorescence staining of MAP2 at day 60 in MT and Ctrl COs. N = 10. D – J D130 COs were used for whole-cell patch-clamp recordings. N = 11. The membrane capacitance ( D ), resting membrane potential (RMP) ( E ), and input resistance ( F ) were recorded. G Representative traces and quantification of voltage-gated currents in MT and Ctrl COs. H Analysis of neuronal firing ability in MT and Ctrl COs. I Representative traces and quantification of spontaneous excitatory postsynaptic current (sEPSC) in MT and Ctrl COs. J Representative traces and quantification of spontaneous inhibitory postsynaptic current (sIPSC) in CSF1R-MT and Ctrl COs. Two-tailed Student’s t -test was used to compare the differences between the two groups ( C , D – F , I , J ). Two-way ANOVA with Tukey’s post hoc test was used to compare the two groups ( G , H ). Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: Immunofluorescence, Staining, Patch Clamp, Membrane, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: A CSF1R mRNA (upper panel) and protein domain structure (lower panel). The boxes in the upper panel (mRNA) are exons, and the arrow indicates the CSF1R-T567M mutation. The arrow in the lower panel (protein) indicates the position of the T567M mutation in the CSF1R protein. S signal peptide, Ig immunoglobulin-like domain, TM transmembrane domain, JMD juxta-membrane domain, TKD tyrosine kinase domain, KID kinase insert domain. B Sanger sequencing electropherograms of control and CSF1R-MT cDNA derived from human blood. The red arrows indicate the point mutation site at T567M in the CSF1R-RD patient. C SH-SY5Y cells stably expressing CSF1R-WT (WT) and CSF1R-MT (MT) were collected for western blotting. D – G SH-SY5Y cells stably expressing CSF1R-WT and CSF1R-MT were treated with CSF1 for 0, 5, 15, and 40 min before western blot analysis of pY723-CSF1R, pY708-CSF1R, and pY546-CSF1R in ( D ). Statistical analysis for pY723 ( E ), pY708 ( F ), and pY546 ( G ) from ( D ), N = 3. H – L BV2 cells stably expressing CSF1R-WT and CSF1R-MT were then used for western blot analysis ( H ). Statistical analysis of CSF1R ( I ), pY546-CSF1R/CSF1R ( J ), IBA1 ( K ), and LC3-II/I ( L ) from ( H ), N = 3. Two-tailed Student’s t -test was used to compare the differences between the two groups for ( C ) and ( I – L ). Two-way ANOVA with Tukey’s post hoc test was used to compare different groups for ( E – G ). Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05 and ** p < 0.01.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: Mutagenesis, Membrane, Sequencing, Control, Derivative Assay, Stable Transfection, Expressing, Western Blot, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: CSF1R-T567M mutant (MT) and corrected (Ctrl) iPSCs derived from patients were used to differentiate into microglia (iMGL). A Representative images of P2RY12 + and TMEM119 + iMGL. Scale bar = 10 µm. B Comparison of area, branch number, and maximum branch length between Ctrl and MT iMGL with IBA1 + TMEM119 + staining. Cells from 50 fields of view were counted for each group. Scale bar = 5 µm. Statistical analysis of IBA1 + iMGL cell area ( C ), branches per cell ( D ), and maximum branch length ( E ) in ( B ). F Cytokine profiling assay after 24 h of stimulation with 100 ng/mL LPS, showing significant upregulation of candidate cytokines. RT-qPCR of CCL2 ( G ) and IL-18 ( H ) transcription in Ctrl and MT iMGL. N = 3. I Migration assay of Ctrl and MT iMGL after stimulation with ATP. White arrowheads indicate some of the migrated iMGL. Cells from 15 fields of view were counted for each group. Scale bar = 5 µm. J Effect of the CSF1R-MT on the phagocytic ability of green fluorescent latex beads (white arrowheads). The iMGL were stained with IBA1 (red signals). N = 8. Scale bar = 10 µm. K Effect of the CSF1R-MT on the phagocytic ability of the pHrodo-labeled myelin (red signals) in iMGL. Number of cells quantified: Ctrl = 76, MT = 61. Scale bar = 10 µm. Two-tailed Student’s t -test was used to compare the differences between the two groups. Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05 and *** p < 0.001.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: Mutagenesis, Derivative Assay, Comparison, Staining, Quantitative RT-PCR, Migration, Labeling, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: Three batches of D30 iMGL were used to conduct bulk RNA-sequencing. A Venn diagram showing co-expressed and independently expressed genes in Ctrl and MT iMGL. B Volcano plot of differentially expressed genes between Ctrl and MT iMGL. C Gene Ontology analysis of top-downregulated genes. Some biological processes, cellular components, and molecular functions are downregulated in the MT iMGL. D KEGG pathway analysis of downregulated genes. ECM-receptor interaction, axon guidance, and PI3K-Akt signaling pathway are downregulated in CSF1R-MT iMGL. E Gene Ontology analysis of upregulated genes. Immune receptor activity is upregulated in the MT iMGL. F qPCR analysis of NLRP1 and TUBB4A mRNA expression. Two-tailed Student’s t -test was used to compare the differences between the two groups, N = 3. Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05 and ** p < 0.01.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: RNA Sequencing, Activity Assay, Expressing, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: NPCs and neurons differentiated from the Ctrl and MT iPSCs were used to study the effect of CSF1R-MT on neuronal functions. A – C NPCs were stained with Ki67 and EdU. A White arrowheads indicate Ki67/EdU double-positive NPCs. Scale bar = 25 μm. Statistical analysis of Ki67-positive NPCs ( B ) and EdU-positive NPCs ( C ) in ( A ). Cells from 18 fields of view were counted for each group. D Western blot analysis of CSF1R and TUJ1 in Ctrl and MT iPSC-derived neurons. N = 3. E Representative images of Ctrl and MT iPSC-derived neurons (D19), stained by TUJ1. Sholl analysis of dendrite number per cell ( F ) and number of crossings ( G ) in ( E ). N = 7. Scale bar = 5 μm. Two-tailed Student’s t -test was used to compare the two groups. Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: Staining, Western Blot, Derivative Assay, Two Tailed Test

Journal: Cell Death Discovery

Article Title: CSF1R T567M mutation induces microglial dysfunction and synaptic impairment in patient iPSC-derived cerebral organoids of CSF1R-related disorder

doi: 10.1038/s41420-026-02995-2

Figure Lengend Snippet: A Representative images of each stage of cerebral organoid (CO) generation. B , C Immunofluorescence staining of MAP2 at day 60 in MT and Ctrl COs. N = 10. D – J D130 COs were used for whole-cell patch-clamp recordings. N = 11. The membrane capacitance ( D ), resting membrane potential (RMP) ( E ), and input resistance ( F ) were recorded. G Representative traces and quantification of voltage-gated currents in MT and Ctrl COs. H Analysis of neuronal firing ability in MT and Ctrl COs. I Representative traces and quantification of spontaneous excitatory postsynaptic current (sEPSC) in MT and Ctrl COs. J Representative traces and quantification of spontaneous inhibitory postsynaptic current (sIPSC) in CSF1R-MT and Ctrl COs. Two-tailed Student’s t -test was used to compare the differences between the two groups ( C , D – F , I , J ). Two-way ANOVA with Tukey’s post hoc test was used to compare the two groups ( G , H ). Data are presented as mean ± SD. The statistical significance levels were set at * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: The antibodies used in this study were CSF1R (Cell Signaling, #3152), phospho-CSF1R Tyr708 (Cell Signaling, #3080), phospho-CSF1R Tyr723 (Cell Signaling, #3151), phospho-CSF1R Tyr546 (Cell Signaling, #3083), β-actin (Santa Cruz, #AC-15), LC3 (Abcam, #ab243506), IBA1 (Fujifilm Wako, #019-19741), and TUJ1 (Millipore, #MAB1637).

Techniques: Immunofluorescence, Staining, Patch Clamp, Membrane, Two Tailed Test

Journal: bioRxiv

Article Title: Macrophages regulate meiotic initiation and germ cell clearance in the developing ovary

doi: 10.64898/2026.02.28.708733

Figure Lengend Snippet: (a) Representative sections of E14.5 ( n =4 independent gonads) and E18.5 ( n =4) ovaries stained for EdU, CD45, and F4/80. Arrowheads denote EdU⁺CD45⁺F4/80⁺ macrophages; arrows denote EdU⁺CD45⁺F4/80 − monocyte-like cells. (b) Representative sections of P14 ( n =5), P30 ( n =5), and P90 ( n =4) ovaries stained for EdU, MHCII, and CSF1R. Black arrowheads indicate EdU⁺MHCII + CSF1R − macrophages; black arrows indicate EdU⁺MHCII − CSF1R + macrophages; and white arrows indicate EdU⁺MHCII + CSF1R + macrophages. (c) Quantification of CD45⁺F4/80⁺ macrophages and CD45⁺F4/80⁻ monocyte-like cells per 0.1 mm² ovarian area at E14.5 and E18.5. (d) Percentage of EdU⁺ cells among macrophages and monocyte-like cells at E14.5 and E18.5. (e) Quantification of MHCII⁻CSF1R⁺, MHCII⁺CSF1R⁺, and MHCII⁺CSF1R⁻ macrophage subsets per 0.1 mm² ovarian area at P14, P30, and P90. (f) Percentage of EdU⁺ cells within each macrophage subset at P14, P30, and P90. Data are shown as mean +/− SD. * P <0.05, ** P <0.01, *** P <0.001 (two-tailed Student’s t -tests). Scale bars, 100 μm (overview) and 50 μm (higher magnification/insets).

Article Snippet: To transiently deplete yolk-sac-derived and fetal CSF1R + macrophages, pregnant C57BL/6J females were injected intraperitoneally with 3 mg anti-CSF1R monoclonal antibody (mAb; clone AFS98, Bio X Cell #BP0213) or rat IgG2a isotype control (Bio X Cell #BP0089).

Techniques: Staining, Two Tailed Test

Journal: bioRxiv

Article Title: Macrophages regulate meiotic initiation and germ cell clearance in the developing ovary

doi: 10.64898/2026.02.28.708733

Figure Lengend Snippet: (a) Scheme of the Csf1r -creER; Rosa -Tomato lineage-tracing strategy. (b) Representative E18.5 ovarian images ( n =5) showing Tomato-labeled CD45⁺F4/80⁺ macrophages and CD45⁺F4/80⁻ monocyte-like cells in each induction group. Arrowheads, Tomato⁺ macrophages; arrows, Tomato⁺ monocyte-like cells. (c) Percentage of Tomato⁺ cells among total F4/80⁺ macrophages at E18.5 following the indicated induction times. (d) Number of Tomato⁺CD45⁺ F4/80⁻ monocyte-like cells per 0.1 mm² ovarian area at E18.5. (e) Representative P30 and P90 ovaries ( n =4) stained for MHCII, CSF1R, and Tomato, illustrating the contribution of Csf1r + lineage cells to MHCII⁻CSF1R⁺, MHCII⁺CSF1R⁺, and MHCII⁺CSF1R⁻ macrophage subsets after induction at E8.5, E10.5 or E12.5. Black arrows denote Tomato⁺MHCII − CSF1R + macrophages; white arrows denote Tomato⁺MHCII + CSF1R + macrophages; and black arrowheads denote Tomato⁺MHCII + CSF1R − macrophages. (f and g) Percentages of Tomato⁺ cells within each macrophage subset at P30 (f) and P90 (g). Data are shown as mean +/− SD. * P <0.05, ** P <0.01, *** P <0.001 (two-tailed Student’s t -tests). Scale bars, 100 μm (overview) and 50 μm (higher magnification/insets).

Article Snippet: To transiently deplete yolk-sac-derived and fetal CSF1R + macrophages, pregnant C57BL/6J females were injected intraperitoneally with 3 mg anti-CSF1R monoclonal antibody (mAb; clone AFS98, Bio X Cell #BP0213) or rat IgG2a isotype control (Bio X Cell #BP0089).

Techniques: Labeling, Staining, Two Tailed Test

Journal: bioRxiv

Article Title: Macrophages regulate meiotic initiation and germ cell clearance in the developing ovary

doi: 10.64898/2026.02.28.708733

Figure Lengend Snippet: (a) Scheme of the Kit -creER; Rosa -Tomato lineage-tracing strategy. (b) Representative E18.5 ovarian images ( n =5) showing Tomato-labeled CD45⁺F4/80⁺ macrophages, CD45⁺F4/80⁻ monocyte-like cells, and PECAM1⁺ endothelial cells after induction at the indicated stages. Arrowheads denote Tomato⁺ macrophages; arrows denote Tomato⁺ monocyte-like cells; dashed outlines mark Tomato⁺PECAM1⁺ endothelial cells. (c) Percentage of Tomato⁺ cells among total F4/80⁺ macrophages at E18.5 in each induction group. (d) Number of Tomato⁺CD45⁺F4/80⁻ monocyte-like cells per 0.1 mm² ovarian area at E18.5. (e) Representative P30 ( n =4), P60 ( n =4), and P90 ( n =6) ovaries stained for MHCII, CSF1R, and Tomato, illustrating the distribution of Kit -lineage cells among MHCII⁻CSF1R⁺, MHCII⁺CSF1R⁺, and MHCII⁺CSF1R⁻ macrophage subsets following induction at E8.5, E10.5 or E12.5. Black arrows denote Tomato⁺MHCII − CSF1R + macrophages; white arrows denote Tomato⁺MHCII + CSF1R + macrophages; and black arrowheads denote Tomato⁺MHCII + CSF1R − macrophages. (f and g) Percentages of Tomato⁺ cells within each macrophage subset at P30 (f) and P90 (g) for E8.5 and E10.5 inductions. (h) Contribution of E12.5-labeled cells to MHCII⁻CSF1R⁺, MHCII⁺CSF1R⁺, and MHCII⁺CSF1R⁻ macrophages at P60. Data are shown as mean +/− SD. * P <0.05, ** P <0.01, *** P <0.001 (two-tailed Student’s t -tests). Scale bars, 100 μm (overview) and 50 μm (higher magnification/insets).

Article Snippet: To transiently deplete yolk-sac-derived and fetal CSF1R + macrophages, pregnant C57BL/6J females were injected intraperitoneally with 3 mg anti-CSF1R monoclonal antibody (mAb; clone AFS98, Bio X Cell #BP0213) or rat IgG2a isotype control (Bio X Cell #BP0089).

Techniques: Labeling, Staining, Two Tailed Test

Journal: bioRxiv

Article Title: Macrophages regulate meiotic initiation and germ cell clearance in the developing ovary

doi: 10.64898/2026.02.28.708733

Figure Lengend Snippet: (a) Scheme of embryonic Cx3cr1 -creER; Rosa -Tomato labeling. (b) Representative E14.5 ( n =3) and E18.5 ( n =4) ovarian sections stained for F4/80, CD45, and Tomato following E12.5 induction. Arrowheads indicate Tomato⁺CD45 + F4/80 + macrophages. (c) Representative P30 ( n =5) and P90 ( n =4) ovarian sections stained for MHCII, CSF1R, and Tomato, showing the persistence of embryonically labeled macrophages within MHCII⁻CSF1R⁺, MHCII⁺CSF1R⁺, and MHCII⁺CSF1R⁻ subsets. Black arrows denote Tomato⁺MHCII − CSF1R + macrophages and white arrows denote Tomato⁺MHCII + CSF1R + macrophages. (d) Scheme of postnatal Cx3cr1 -creER ; Rosa -Tomato labeling. Pups received TAM at P4 and P5 and ovaries were analyzed at P8 and P60. (e) Representative P8 ( n =4) and P60 ( n =4) ovarian sections from postnatally induced animals stained for MHCII, CSF1R, and Tomato. Black arrows denote Tomato⁺MHCII − CSF1R + macrophages, white arrows denote Tomato⁺MHCII + CSF1R + macrophages, and black arrowheads denote Tomato⁺MHCII + CSF1R − macrophages. (f) Quantification of percent Tomato⁺ cells among CD45⁺F4/80⁺ macrophages at E14.5 and E18.5 after E12.5 induction. (g) Percentages of Tomato⁺ cells within each macrophage subset at P30 and P90. (h and i) Percentages of Tomato⁺ cells within MHCII⁻CSF1R⁺ macrophages at P8 (h), and within MHCII⁻CSF1R⁺, MHCII⁺CSF1R⁺, and MHCII⁺CSF1R⁻ macrophages at P60 (i) after P4/P5 induction. Data are shown as mean +/− SD. * P <0.05, ** P <0.01, *** P <0.001 (two-tailed Student’s t -tests). Scale bars, 100 μm (overview) and 50 μm (higher magnification/insets).

Article Snippet: To transiently deplete yolk-sac-derived and fetal CSF1R + macrophages, pregnant C57BL/6J females were injected intraperitoneally with 3 mg anti-CSF1R monoclonal antibody (mAb; clone AFS98, Bio X Cell #BP0213) or rat IgG2a isotype control (Bio X Cell #BP0089).

Techniques: Labeling, Staining, Two Tailed Test

Journal: bioRxiv

Article Title: Macrophages regulate meiotic initiation and germ cell clearance in the developing ovary

doi: 10.64898/2026.02.28.708733

Figure Lengend Snippet: (a) Representative P30 ovarian sections from Ccr2 GFP/+ control ( n =6) and Ccr2 GFP/GFP KO ( n =6) mice stained for MHCII and CSF1R. Black arrows indicate GFP-negative MHCII − CSF1R + macrophages; white arrows indicate GFP-expressing MHCII + CSF1R + macrophages; and black arrowheads indicate GFP-expressing MHCII + CSF1R − macrophages. (b) Representative P60 ovarian sections from Ccr2 GFP/+ control ( n =4) and Ccr2 GFP/GFP KO ( n =4) mice stained for MHCII and CSF1R. (c and d) Percentages of GFP⁺ cells within MHCII⁻CSF1R⁺, MHCII⁺CSF1R⁺, and MHCII⁺CSF1R⁻ macrophage subsets in control and KO ovaries at P30 (c) and P60 (d). (e and f) Proportional distribution of GFP⁺ cells among MHCII⁻CSF1R⁻, MHCII⁻CSF1R⁺, MHCII⁺CSF1R⁺, and MHCII⁺CSF1R⁻ populations at P30 (e) and P60 (f). Data are shown as mean +/− SD. * P <0.05, ** P <0.01, *** P <0.001 (two-tailed Student’s t -tests). Scale bars, 100 μm (overview) and 50 μm (higher magnification/insets).

Article Snippet: To transiently deplete yolk-sac-derived and fetal CSF1R + macrophages, pregnant C57BL/6J females were injected intraperitoneally with 3 mg anti-CSF1R monoclonal antibody (mAb; clone AFS98, Bio X Cell #BP0213) or rat IgG2a isotype control (Bio X Cell #BP0089).

Techniques: Control, Staining, Expressing, Two Tailed Test

Journal: bioRxiv

Article Title: Macrophages regulate meiotic initiation and germ cell clearance in the developing ovary

doi: 10.64898/2026.02.28.708733

Figure Lengend Snippet: (a) Representative whole-mount images of E13.5 ovaries from C57BL/6J embryos exposed at E6.5 to either control rat IgG2a antibody or anti-CSF1R blocking antibody to deplete YS-derived macrophages. For F4/80, PECAM1, and FOXL2 staining, n =5 control and n =6 anti-CSF1R-treated independent gonads; for CD11b and CD45 staining, n =3 control and n =3 anti-CSF1R-treated independent gonads. Insets show higher-magnification views of boxed regions; dashed lines mark the gonad-mesonephros boundary. (b) qRT-PCR analysis of macrophage-associated genes ( Adgre1, Cx3cr1, Csf1r, Mrc1 ) and endothelial marker Cdh5 , showing fold change in gene expression in E13.5 anti-CSF1R-treated ovaries versus controls. (c) Representative E14.5 sections stained for SYCP3 and TRA98 showing increased meiotic germ cells in anti-CSF1R-treated ovaries ( n =4) compared with control ovaries ( n =4). (d) qRT-PCR analysis of germ cell and meiotic genes ( Kit, Pou5f1, Ddx4, Stra8, Sycp1, Sycp3, Syce1, Smc1b ), showing fold change in gene expression in E14.5 anti-CSF1R-treated ovaries versus controls. (e) Representative E18.5 sections stained for F4/80 and CD45, and for SYCP3 and FOXL2, showing recovery of F4/80⁺ macrophages and advanced meiotic progression in anti-CSF1R-treated ovaries compared with control ovaries. For F4/80 and CD45 staining, n =4 control and n =4 anti-CSF1R-treated independent gonads; for SYCP3 and FOXL2 staining, n =3 control and n =3 anti-CSF1R-treated independent gonads. Arrows indicate SYCP3⁺ dictyate-stage oocytes. (f) Percentage of SYCP3⁺ germ cells at the dictyate stage among total SYCP3⁺ cells at E18.5. Data are shown as mean +/− SD. * P <0.05, ** P <0.01, *** P <0.001 (two-tailed Student’s t -tests). Scale bars, 100 μm (overview) and 50 μm (higher magnification/insets).

Article Snippet: To transiently deplete yolk-sac-derived and fetal CSF1R + macrophages, pregnant C57BL/6J females were injected intraperitoneally with 3 mg anti-CSF1R monoclonal antibody (mAb; clone AFS98, Bio X Cell #BP0213) or rat IgG2a isotype control (Bio X Cell #BP0089).

Techniques: Control, Blocking Assay, Derivative Assay, Staining, Quantitative RT-PCR, Marker, Gene Expression, Two Tailed Test

Journal: bioRxiv

Article Title: Macrophages regulate meiotic initiation and germ cell clearance in the developing ovary

doi: 10.64898/2026.02.28.708733

Figure Lengend Snippet: (a) Representative E14.5 ovarian sections from control ( Csf1r -Cre; Csf1r flox/+ ) ( n =3) and Csf1r -KO ( Csf1r -Cre; Csf1r flox/flox ) ( n =3) embryos stained for F4/80 and CD45, showing loss of macrophages but persistence of CD45⁺F4/80⁻ monocyte-like cells in Csf1r -KO ovaries. Insets show higher-magnification views of boxed regions; dashed lines mark the gonad-mesonephros boundary. (b) Representative E14.5 ovarian sections stained for TRA98 and STRA8 showing unchanged total germ cell distribution but increased STRA8⁺ cells in Csf1r -KO ovaries compared with control ovaries. For TRA98 staining, n =4 control and n =4 Csf1r -KO independent gonads; for STRA8 staining, n =9 control and n =8 Csf1r -KO independent gonads. (c) Higher-magnification images of E14.5 control ( n =7) and Csf1r -KO ( n =6) ovaries stained for SYCP3 and DDX4 to illustrate meiotic staging (non-SYCP3, leptotene, zygotene). (d-f) Quantification of TRA98⁺ germ cell number per 0.1 mm² area (d), STRA8⁺ germ cell number per 0.1 mm² (e), and percentage STRA8⁺ cells among TRA98⁺ cells (f) at E14.5. (g) Percentage of meiotic substages (non-SYCP3, leptotene, zygotene) among DDX4⁺ germ cells at E14.5. (h) Representative E18.5 sections stained for EdU, F4/80, and CD45, showing absence of F4/80⁺ macrophages and reduced EdU incorporation in CD45⁺F4/80⁻ cells in Csf1r -KO ovaries ( n =6) compared with control ovaries ( n =4). (i) E18.5 control ( n =4) and Csf1r -KO ( n =6) ovarian sections stained for SYCP3 and DDX4 to illustrate meiotic substages (zygotene, pachytene, diplotene, dictyate). (j) Percentage of EdU⁺ cells among CD45⁺F4/80⁻ monocyte-like cells at E18.5 in control and Csf1r -KO ovaries. (k) Percentage of meiotic substages among DDX4⁺ germ cells at E18.5 in control and Csf1r -KO ovaries. Data are shown as mean +/− SD. * P <0.05, ** P <0.01, *** P <0.001 (two-tailed Student’s t -tests). Scale bars, 100 μm (overview) and 50 μm (higher magnification/insets), except in c and i, where scale bars are 50 μm (overview) and 10 μm (higher magnification/insets).

Article Snippet: To transiently deplete yolk-sac-derived and fetal CSF1R + macrophages, pregnant C57BL/6J females were injected intraperitoneally with 3 mg anti-CSF1R monoclonal antibody (mAb; clone AFS98, Bio X Cell #BP0213) or rat IgG2a isotype control (Bio X Cell #BP0089).

Techniques: Control, Staining, Two Tailed Test

Journal: bioRxiv

Article Title: Macrophages regulate meiotic initiation and germ cell clearance in the developing ovary

doi: 10.64898/2026.02.28.708733

Figure Lengend Snippet: (a) Representative images of E18.5, P3, and P10 ovaries from C57BL/6J embryos exposed at E6.5 and E14.5 to either control rat IgG2a antibody or anti-CSF1R blocking antibody to deplete fetal ovarian macrophages. For E18.5, n =4 control and n =4 anti-CSF1R-treated independent gonads; for P3, n =6 control and n =6 anti-CSF1R-treated independent gonads; for P10, n =4 control and n =4 anti-CSF1R-treated independent gonads. Insets show higher-magnification views of boxed regions; red and green dashed outlines demarcate the ovary. (b) Representative sections stained for DDX4, showing developmental germ cell attrition at E18.5, P3, and P10 in control and anti-CSF1R-treated ovaries. For each time point, n =4 control and n =4 anti-CSF1R-treated independent gonads. (c) Quantification of CD45⁺IBA1 + macrophages per 0.1 mm² ovarian area at E18.5, P3, and P10. (d) Quantification of CD45⁺IBA1 − monocyte-like cells per 0.1 mm² ovarian area at the indicated stages. (e) Quantification of DDX4⁺ germ cell number per 0.1 mm² ovarian area at E18.5, P3, and P10, showing reduced physiological germ cell loss after macrophage depletion. Data are shown as mean +/− SD. * P <0.05, ** P <0.01, *** P <0.001 (two-tailed Student’s t -tests). Scale bars, 100 μm (overview) and 50 μm (higher magnification/insets).

Article Snippet: To transiently deplete yolk-sac-derived and fetal CSF1R + macrophages, pregnant C57BL/6J females were injected intraperitoneally with 3 mg anti-CSF1R monoclonal antibody (mAb; clone AFS98, Bio X Cell #BP0213) or rat IgG2a isotype control (Bio X Cell #BP0089).

Techniques: Control, Blocking Assay, Staining, Two Tailed Test