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NIK-driven IL-23 production by myeloid cells is a key factor in the development of autoimmune inflammation

Nishada S. Ramphal, Xinyuan Liu, Ilaria Palagi, Rebecca Jasser, Alma N. Mohebiany, Matthias Klein, Frederike Westermann, Sinduya Krishnarajah, Tommy Regen, Tobias Bopp, Burkhard Becher, Ari Waisman

J Exp Med (2026) 223 (4): e20242294.

https://doi.org/10.1084/jem.20242294

Published: 27 February 2026

View article titled, NIK-driven IL-23 production by myeloid cells is a key factor in the development of autoimmune inflammation

Includes: Supplementary data

Images

NIK in CX3CR1-expressing cells drives EAE pathology. (A) Disease course of NIKΔCX3CR1 and littermate controls. (B) Maximum EAE score. Data in A and B are the cumulative data of 3 individual experiments. (C) Representative flow cytometry plots with mean percentages ±SEM, and the number of microglia (CD11b+CD45int), infiltrating CD45high and CD11b+CD45high immune cells into the CNS during the peak of EAE (15 dpi). (D–F) Number of CD4+CD90+ T cells, (E) MOG-responding cells (CD40L+CD44+), (F) and IL-17A, IFNγ, or GM-CSF by these T cells after a 6-h MOG35–55 antigen recall assay in the CNS (brain and spinal cord). (G–I) Number of CD4+CD90+ T cells, (H) MOG-responding cells (CD40L+CD44+), (I) and IL-17A, IFNγ, or GM-CSF by these T cells after a 6-h MOG35–55 antigen recall assay in the dLN and spleen. Duplets and dead cells were excluded before gating on shown cell populations. (J and K) EAE disease development and (K) maximum EAE score in NIKΔCD11c mice and littermate controls. Data are shown as the mean ± SEM and analyzed using two-tailed unpaired Student’s t test or two-way ANOVA with Šídák’s multiple comparisons test. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Each dot represents one mouse. Data (C–K) are from at least two independent experiments. dpi = days after immunization. dLN = draining lymph nodes.

NIK in CX3CR1-expressing cells drives EAE pathology. (A) Disease course of...

in NIK-driven IL-23 production by myeloid cells is a key factor in the development of autoimmune inflammation > Journal of Experimental Medicine

Published: 27 February 2026

Figure 1. NIK in CX3CR1-expressing cells drives EAE pathology. (A) Disease course of NIK^ΔCX3CR1 and littermate controls. (B) Maximum EAE score. Data in A and B are the cumulative data of 3 individual experiments. (C) Representative flow More about this image found in NIK in CX3CR1-expressing cells drives EAE pathology. (A) Disease course of...

Images

Less cytokine-producing T cells in secondary lymphoid organs before EAE onset. (A) Scheme of the adoptive transfer EAE model: Ly5.1 wild-type mice were immunized with MOG35–55/CFA and PTx; isolated cells from dLNs and spleen were cultured for 4 days with MOG35–55, IL-23, and anti-IFNγ before being injected into NIKΔCX3CR1 and littermate controls. (B and C) Passive transfer EAE disease course and (C) maximum EAE score of NIKΔCX3CR1 and littermate controls after adoptive transfer of MOG-activated wild-type T cells. (D–F) Experimental scheme: NIKΔCX3CR1 and littermate control mice were immunized with MOG35–55/CFA and PTx, and cells were isolated from the spleen and dLN at 8 dpi and restimulated with MOG for 6 h. (E) Representative flow cytometry plots and total cell number of CD4+CD90+ T cells and (F) MOG-reactive (CD44+CD40L+) T cells. (G) Representative flow cytometry plots and percentages of IL-17A, GM-CSF, and IFNγ-producing T cells. (H and I) 2D2 T cells were transferred into NIKΔCX3CR1 and littermate controls 1 day before MOG35–55 immunization. Six dpi, cells were isolated from the spleen and dLN and restimulated with PMA, ionomycin, and brefeldin A. (I) Representative flow cytometry plots of the spleen and the total cell number of transferred CD90.1+CD4+ 2D2 cells. (J) Percentages of 2D2 T cells that produce IL-17A, GM-CSF, or IFNγ in the dLNs and spleen and cytokine-producing CD4+ host cells, pregated on live/single cells. Data in graphs are shown as the mean ± SEM and analyzed using two-tailed unpaired Student’s t test or two-way ANOVA with Šídák’s multiple comparisons test. *P < 0.05, **P < 0.01, ****P < 0.0001. Each dot represents one mouse. Data are from at least three independent experiments. dpi = days after immunization.

Less cytokine-producing T cells in secondary lymphoid organs before EAE ons...

in NIK-driven IL-23 production by myeloid cells is a key factor in the development of autoimmune inflammation > Journal of Experimental Medicine

Published: 27 February 2026

Figure 2. Less cytokine-producing T cells in secondary lymphoid organs before EAE onset. (A) Scheme of the adoptive transfer EAE model: Ly5.1 wild-type mice were immunized with MOG_35–55/CFA and PTx; isolated cells from dLNs and spleen were More about this image found in Less cytokine-producing T cells in secondary lymphoid organs before EAE ons...

Images

scRNA-seq reveals dysregulation of antigen-presenting genes 4 days after MOG immunization in the dLNs. (A) Density plots overlaid on the UMAP of control (n = 4) and NIKΔCX3CR1 (n = 4) displaying clustering of myeloid cell type, excluding B, T, NK, and mast cells, pDCs, neutrophils, and granulocytes. (B) Heatmap displaying the top 10 highly expressed markers of each cluster in A with three of the top markers per cluster written out. (C) Proportions of each cell type within the UMAP of the control (n = 4) and NIKΔCX3CR1 (n = 4). (D) Antigen presentation cell signature score projected in each cluster, calculated with UCell. (E) 536 neighborhoods assigned by the Milo package overlaid on our UMAP of myeloid cell clusters. (F) Abundant neighborhoods across the different myeloid clusters. Red = more abundant in control, blue = more abundant in NIKΔCX3CR1, FDR = 10%. (G) Volcano plot of DEGs between NIKΔCX3CR1 and control in the migDC subset. Left is downregulated in NIKΔCX3CR1, and right is upregulated in NIKΔCX3CR1. (H) KEGG pathway enrichment analysis was performed on DEGs (adjusted P < 0.05 and |log2 fold change| >0.1 and <0.1) from the migDC clusters. (I and J) Volcano plot of DEGs between NIKΔCX3CR1 and control in the Mac subset and (J) MoMac subset. Left is downregulated in NIKΔCX3CR1, and right is upregulated in NIKΔCX3CR1. (K) KEGG pathway enrichment analysis was performed on DEGs (adjusted P < 0.05 and |log2 fold change| >0.1 and <0.1) from the Mac clusters (Macs/MoMacs).The dot plot shows enriched pathways ranked by significance. Circle size = count of DEGs found within each pathway; color = adjusted P value. Data in C are shown as mean percentage and analyzed using two-way ANOVA with Šídák’s multiple comparisons test. *P < 0.05, ***P < 0.001.

scRNA-seq reveals dysregulation of antigen-presenting genes 4 days after MO...

in NIK-driven IL-23 production by myeloid cells is a key factor in the development of autoimmune inflammation > Journal of Experimental Medicine

Published: 27 February 2026

Figure 3. scRNA-seq reveals dysregulation of antigen-presenting genes 4 days after MOG immunization in the dLNs. (A) Density plots overlaid on the UMAP of control (n = 4) and NIK^ΔCX3CR1 (n = 4) displaying clustering of myeloid cell type, More about this image found in scRNA-seq reveals dysregulation of antigen-presenting genes 4 days after MO...

Images

Reduced costimulatory marker expression on myeloid cells and reduced antigen-presenting capabilities. (A and B) UMAP map displaying 50,000 randomly sampled cells from the dLN of NIKΔCX3CR1 (n = 5) and littermate controls (n = 5) analyzed by flow cytometry, focusing on CD11b+ monocyte, Mac, and DC subsets 5 dpi. (B) Heatmap with median marker expression values for each population shown in the UMAP. (C) Relative frequencies of Ly6Chigh monocytes (Mono), Ly6Cint Mono, Ly6Clo Mono, ResDC2, MigDC2, MoDCs, Macs, other, pregated on CD11b+ cells, and dead cells, duplicates, T and B cells, neutrophils, and eosinophils were excluded. (D) Median (arcsinh-transformed) expression of CD80, CD86, MHCII, and PD-L1 across the eight identified myeloid cell populations. (E and F) Representative flow cytometry histograms showing CTV dilution of OT-II CD4+ T cells cocultured with either sorted Macs (T + Macs) (E) or migratory dendritic cells (T + migDCs) from dLNs (F) from control or NIKΔCX3CR1 mice. Accompanying graphs show the Division Index, reflecting the average number of divisions per input cell. Cells were pulsed with the indicated concentrations of OVA323–339 peptide for 1 h prior to coculture. Data are shown as the mean ± SEM and analyzed using two-way ANOVA with Šídák’s multiple comparisons test. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. dpi = days after immunization. Each dot represents one mouse. Data are from at least two independent experiments. CTV, CellTrace Violet.

Reduced costimulatory marker expression on myeloid cells and reduced antige...

in NIK-driven IL-23 production by myeloid cells is a key factor in the development of autoimmune inflammation > Journal of Experimental Medicine

Published: 27 February 2026

Figure 4. Reduced costimulatory marker expression on myeloid cells and reduced antigen-presenting capabilities. (A and B) UMAP map displaying 50,000 randomly sampled cells from the dLN of NIK^ΔCX3CR1 (n = 5) and littermate controls (n = 5) More about this image found in Reduced costimulatory marker expression on myeloid cells and reduced antige...

Images

NIK deletion in myeloid cells results in reduced Il23a, which is partially responsible for the resistance against EAE. (A–F) CD11b+ cells were isolated from the dLNs of NIKΔCX3CR1 and littermate controls and were cultured with or without anti-(α)CD40 or LPS for 1 or 4 h. Relative mRNA levels (ΔΔCt) of (B) Il6, (C) Il1b, (D) Il12a/p35, (E) Il23a/p19, and (F) Il10, and mRNA levels of each sample were normalized against the housekeeping gene HPRT and the medium control of the corresponding cytokine. Dotted line = medium control. (G) NIKΔCX3CR1 and littermate controls were immunized with MOG35–55/CFA and PTx; isolated cells from dLNs and spleen were cultured for 4 days with MOG35–55, IL-23, and anti-IFNγ before being injected into Rag−/− recipient mice. (H) Flow cytometry gating for CD44/GM-CSF and CD44/IL-17A of the donor cells before being injected into recipient mice, pregated on single/live/CD90+/CD4+ cells. (I) Passive transfer EAE disease course of recipients receiving control cells (Ctrl → Rag−/−) or NIKΔCX3CR1 cells (NIKΔCX3CR1 → Rag−/−). (J) AUC. (K) Day of onset. (L) Maximum disease score. Data in graphs are shown as the mean ± SEM and analyzed using two-way ANOVA with Šídák’s multiple comparisons test or two-tailed unpaired Student’s t test. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Each dot represents one mouse. Data are from at least two independent experiments. dpi = days after immunization. AUC, area under the curve.

NIK deletion in myeloid cells results in reduced Il23a, which is partially ...

in NIK-driven IL-23 production by myeloid cells is a key factor in the development of autoimmune inflammation > Journal of Experimental Medicine

Published: 27 February 2026

Figure 5. NIK deletion in myeloid cells results in reduced Il23a, which is partially responsible for the resistance against EAE. (A–F) CD11b⁺ cells were isolated from the dLNs of NIK^ΔCX3CR1 and littermate controls and were cultured with or More about this image found in NIK deletion in myeloid cells results in reduced Il23a, which is partially ...

Journal Articles

Central neurons encode interleukin-1β signals and mediate stress-induced inflammation

Okito Hashimoto, Tyler D. Hepler, Aisling Tynan, Alejandro Torres, Jian Hua Li, Michael Brines, Kevin J. Tracey, Sangeeta S. Chavan

J Exp Med (2026) 223 (4): e20252000.

https://doi.org/10.1084/jem.20252000

Published: 26 February 2026

View article titled, Central neurons encode interleukin-1β signals and mediate stress-induced inflammation

Images

Distinct neuronal populations in the BNST recapitulate IL-1β responses. (A) STPT of the whole mouse brain of TRAP2-tdTomato mice. Top lane: overview of the STPT data acquisition and registration; middle lane: representative coronal sections; and bottom lane: areas showing tdTomato+ neuronal cell bodies. (B) tdTomato+ cell count in indicated brain region in IL-1β (5 μg/kg; n = 6) and PBS-injected groups (n = 6). PVT, paraventricular nucleus of the thalamus; PBNc, parabronchial nucleus complex; LC, locus coeruleus; AP, area postrema; NTS: nucleus of the tractus solitarius. Data are represented as individual mouse data points with mean ± SEM. This experiment was completed once. One-way ANOVA with Tukey’s multiple comparison test. (C) Representative images showing immunofluorescence staining of the labeled cells (tdTomato, red) co-stained with DAPI (blue) in the BNST. tdTomato+ cell count in the dorsal and ventral BNST regions after exposure to IL-1β. Data are represented as individual mouse data points pooled from two independent experiments. Unpaired t test. Scale bar, 100 μm. (D) Schematic of TRAPing IL-1β–responsive BNST neurons and their reactivation in TRAP2 mice. (E) Representative images of BNST showing Gq-DREADD-mCherry–expressing cells (red) co-stained with c-Fos (green) after CNO administration. Scale bar, 20 μm. (F) Core body temperature for 6 h after reactivation with saline (black) as a control or CNO (red). (G) Minimum change in core body temperature at 1 h after injection. Data are represented as individual mouse data points pooled from two independent experiments. Unpaired t test. (H) ΔHR for 60 min after reactivation of IL-1β–TRAPed neurons with saline (black) or CNO (red). Saline, n = 8 mice; CNO, n = 10 mice, mixed-effects analysis with Šidák correction. (I) AUC of ΔHR after reactivation. Data are represented as individual mouse data points from three independent experiments. Unpaired t test. (J and K) Serum levels of IL-6 and corticosterone at 2 h after saline or CNO administration. Data are represented as individual mouse data points pooled from three independent experiments. Unpaired t test. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001. Box and whisker plots show the minimum, maximum, median, and 25th and 75th percentiles.

Distinct neuronal populations in the BNST recapitulate IL-1β responses. (A)...

in Central neurons encode interleukin-1β signals and mediate stress-induced inflammation > Journal of Experimental Medicine

Published: 26 February 2026

Figure 1. Distinct neuronal populations in the BNST recapitulate IL-1β responses. (A) STPT of the whole mouse brain of TRAP2-tdTomato mice. Top lane: overview of the STPT data acquisition and registration; middle lane: representative coronal More about this image found in Distinct neuronal populations in the BNST recapitulate IL-1β responses. (A)...

Images

Characterization of BNST cells. (A) snRNA-seq cataloging neuronal clusters in the BNST. A uniform manifold approximation and projection (UMAP) plot of transcriptomic data identified 17 neuronal clusters. This experiment was completed once. (B) Dot plots of average expression of Slc17a6 (Vglut2; glutamatergic neuronal marker), Slc32a1 (Vgat: GABAergic neuronal marker), corticotropin-releasing crh, sst, pdyn, and penk-expressing populations. Dot size: percent of cells in cluster and color: average expression levels. (C) ΔHR for 60 min after activation of CRH+, SST+, PDYN+, or PENK+ BNST neurons with saline (black) as a control or CNO (red) (crh-cre; saline, n = 5 mice, CNO, n = 5 mice, sst-cre; saline, n = 5 mice, CNO, n = 5 mice, pdyn-cre; saline, n = 5 mice, CNO, n = 5 mice, penk-cre; saline, n = 3 mice, CNO, n = 3 mice, mixed-effects analysis with Šidák correction). (D) Serum IL-6 levels at 2 h after activation with saline or CNO of CRH+, SST+, PDYN+, or PENK+ BNST neurons. (E) Serum corticosterone levels at 2 h after activation with saline or CNO of CRH+, SST+, PDYN+, or PENK+ BNST neurons. Data are represented as individual mouse data points pooled from two independent experiments. Unpaired t test. *P < 0.05, **P < 0.01.

Characterization of BNST cells. (A) snRNA-seq cataloging neuronal clusters...

in Central neurons encode interleukin-1β signals and mediate stress-induced inflammation > Journal of Experimental Medicine

Published: 26 February 2026

Figure 2. Characterization of BNST cells. (A) snRNA-seq cataloging neuronal clusters in the BNST. A uniform manifold approximation and projection (UMAP) plot of transcriptomic data identified 17 neuronal clusters. This experiment was completed More about this image found in Characterization of BNST cells. (A) snRNA-seq cataloging neuronal clusters...

Images

Adrenergic signaling mediates CRH+ BNST neuron effects. (A) Schematic for chemogenetic activation of IL-1β–responsive BNST neurons in TRAP2 mice. (B) Serum IL-6 levels at 2 h after reactivation with AR blockers (propranolol, SR59230A; both administered at 5 mg/kg i.p.). Data are represented as individual mouse data points pooled from two independent experiments. One-way ANOVA, Tukey’s multiple comparison test. (C) ΔHR for 60 min after reactivation of IL-1β–responsive BNST neurons with DMSO (black) as a control, propranolol (red), or SR59230A (blue) (DMSO, n = 5 mice; propranolol, n = 5 mice; SR59230A, n = 5 mice, mixed-effects analysis with Šidák correction). (D) AUC of ΔHR after reactivation of IL1-responsive BNST neurons with AR blockers. Data are represented as individual mouse data points pooled from two independent experiments. One-way ANOVA, Tukey’s multiple comparison test. (E) Serum corticosterone levels at 2 h after reactivation of IL1-responsive BNST neurons with AR blockers. Data are represented as individual mouse data points pooled from two independent experiments. One-way ANOVA, Tukey’s multiple comparison test. (F) Schematic for pharmacogenetic activation of BNST neurons in crh-cre mice. (G) Serum IL-6 levels at 2 h after activation of CRH-positive BNST neurons with AR blockers in crh-cre mice. (H) ΔHR for 60 min after activation of CRH+ BNST neurons with DMSO (black) as a control, propranolol (red), or SR59230A (blue) (DMSO, n = 4 mice; propranolol, n = 5 mice; SR59230A, n = 5 mice, mixed-effects analysis with Šidák correction). (I) AUC of ΔHR after activation with AR blockers in crh-cre mice. Data are represented as individual mouse data points pooled from two independent experiments. One-way ANOVA, Tukey’s multiple comparison test. (J) Serum corticosterone levels at 2 h after activation of CRH-positive BNST neurons with AR blockers in crh-cre mice. Data are represented as individual mouse data points pooled from two independent experiments. One-way ANOVA, Tukey’s multiple comparison test. *P < 0.05; **P < 0.01.

Adrenergic signaling mediates CRH⁺ BNST neuron effects. (A) Sch...

in Central neurons encode interleukin-1β signals and mediate stress-induced inflammation > Journal of Experimental Medicine

Published: 26 February 2026

Figure 3. Adrenergic signaling mediates CRH⁺ BNST neuron effects. (A) Schematic for chemogenetic activation of IL-1β–responsive BNST neurons in TRAP2 mice. (B) Serum IL-6 levels at 2 h after reactivation with AR blockers (propranolol, More about this image found in Adrenergic signaling mediates CRH⁺ BNST neuron effects. (A) Sch...

Images

BNST–PVN-RVLM neural signaling mediates IL-1β–induced changes in heart rate and IL-6. (A) Schematic for anterograde tracing of axonal projections and terminals from PBS-TRAPed or IL-1β–TRAPed BNST neurons. The AAV-hSyn-FLEx-mGFP-2A-synaptophysin-mRuby was injected into the BNST of TRAP2 mice. (B) Representative image for GFP+ axons and mRuby+ terminals in the PVN of PBS-TRAPed or IL-1β–TRAPed mice. The rightmost panel shows a higher-magnification view of the PVN in IL-1β–TRAPed mice. Arrowheads show regions with co-localization of mRuby and EYFP. Scale bars, 100 μm. (C) Schematic for anterograde tracing of IL-1β–TRAPed BNST neurons connected with PVN neurons. The AAV-pEF1a-DIO-FLPo-WPRE-hGHpA was injected into the BNST, and the AAV-Ef1a-fDIO-EYFP was injected into the PVN of TRAP2 mice. (D) Representative image for EYFP expression in the PVN, RVLM, and NTS. Arrowheads show neurons, and arrows show axonal projections with expression of EYFP. Scale bar, 100 μm for the PVN and RVLM. Scale bar, 200 μm for the NTS. (E) c-Fos expression in the RVLM after reactivation with saline as a control or CNO of IL-1β–responsive BNST neurons. Scale bar, 100 μm. (F) Schematic for activating the BNST–PVN neural pathway. The AAV-pEF1a-DIO-FLPo-WPRE-hGHpA was injected into the BNST, and the AAV-hSyn-fDIO-hM3D(Gq)-mCherry-WPREpA was injected into the PVN of TRAP2 mice. (G) Representative image of PVN showing Gq-DREADD-mCherry–expressing cells (red). Scale bar, 100 μm. (H) Serum IL-6 levels at 2 h after reactivation with saline as a control or CNO of the BNST–PVN neuronal pathway. Data are represented as individual mouse data points pooled from two independent experiments. Unpaired t test. (I) ΔHR for 60 min after reactivation of the BNST–PVN neuronal pathway: saline (black) or CNO (red) (saline, n = 7 mice; CNO, n = 10 mice, mixed-effects analysis with Šidák correction). (J) AUC of ΔHR after reactivation. Data are represented as individual mouse data points pooled from two independent experiments. Unpaired t test. (K) Serum corticosterone levels at 2 h after reactivation of the BNST–PVN neuronal pathway. Data are represented as individual mouse data points pooled from two independent experiments. Unpaired t test. *P < 0.05; **P < 0.01.

BNST–PVN-RVLM neural signaling mediates IL-1β–induced changes in heart rate...

in Central neurons encode interleukin-1β signals and mediate stress-induced inflammation > Journal of Experimental Medicine

Published: 26 February 2026

Figure 4. BNST–PVN-RVLM neural signaling mediates IL-1β–induced changes in heart rate and IL-6. (A) Schematic for anterograde tracing of axonal projections and terminals from PBS-TRAPed or IL-1β–TRAPed BNST neurons. The More about this image found in BNST–PVN-RVLM neural signaling mediates IL-1β–induced changes in heart rate...

Images

PVN signaling is necessary for mediating the effects of IL-1β–responsive BNST neurons. (A) Schematic for chemogenetic reactivation of IL-1β–responsive BNST neurons with ablation of IL-1β–responsive PVN neurons in TRAP2 mice. (B) Representative images showing IL-1β–responsive PVN neurons (c-Fos expression in green) after CNO administration in control dtA (−) and IL-1β–responsive neuron-ablated dtA (+) mice. Scale bar, 100 μm. (C) Serum IL-6 levels at 2 h after reactivation in naive mice, the control group, and with ablation of IL-1β–responsive PVN neurons. Data are represented as individual mouse data points pooled from three independent experiments. One-way ANOVA, Tukey’s multiple comparison test. (D) ΔHR for 60 min after reactivation of IL-1β–responsive BNST neurons with and without ablation of IL-1β–responsive PVN neurons: control (black) or dtA (red) (control, n = 5 mice; dtA, n = 5 mice, mixed-effects analysis with Šidák correction). (E) AUC of ΔHR after reactivation. Data are represented as individual mouse data points pooled from two independent experiments. Unpaired t test. (F) Serum corticosterone levels at 2 h after reactivation in naive mice, the control group without ablation, and with ablation of IL-1β–responsive PVN neurons. Data are represented as individual mouse data points from three independent experiments. One-way ANOVA. (G) Schematic for chemogenetic activation of CRH+ BNST neurons with ablation of CRH+ PVN neurons in crh-cre mice. (H) Representative images showing the expression of c-Fos (green) in the PVN after CNO administration in control dtA (−) and CRH+ PVN neuron-ablated dtA (+) mice. Scale bar, 100 μm. (I) Serum IL-6 levels at 2 h after reactivation in crh-cre naive mice, control mice without ablation, and mice with ablation of CRH+ PVN neurons. Data are represented as individual mouse data points pooled from two independent experiments. One-way ANOVA, Tukey’s multiple comparison test. (J) ΔHR for 60 min after chemogenetic activation of CRH+ BNST neurons with ablation of CRH+ PVN neurons: control (black) or dtA (red) (control, n = 5 mice; dtA, n = 7 mice, mixed-effects analysis with Šidák correction). (K) AUC of ΔHR after reactivation in crh-cre mice. Data are represented as individual mouse data points pooled from two independent experiments. Unpaired t test. (L) Serum corticosterone levels at 2 h after reactivation in crh-cre naive mice, control group without ablation, and with ablation of IL-1β–responsive PVN neurons. Data are represented as individual mouse data points pooled from two independent experiments. One-way ANOVA. *P < 0.05 and **P < 0.01.

PVN signaling is necessary for mediating the effects of IL-1β–responsive BN...

in Central neurons encode interleukin-1β signals and mediate stress-induced inflammation > Journal of Experimental Medicine

Published: 26 February 2026

Figure 5. PVN signaling is necessary for mediating the effects of IL-1β–responsive BNST neurons. (A) Schematic for chemogenetic reactivation of IL-1β–responsive BNST neurons with ablation of IL-1β–responsive PVN neurons in TRAP2 mice. (B) More about this image found in PVN signaling is necessary for mediating the effects of IL-1β–responsive BN...

Images

IL-1β–responsive BNST neurons are necessary for stress-induced inflammatory responses. (A) Schematic of ablating IL-1β–responsive BNST neurons bilaterally in TRAP2 mice and then exposure to restraint stress. (B) Serum IL-6 levels in TRAP2 mice: naive group without restraint stress, after 4-h restraint stress in the control dtA (−) and IL-1β–responsive BNST neuron-ablated dtA (+) mice. Data are represented as individual mouse data points from two independent experiments. One-way ANOVA with Tukey’s multiple comparison test. (C) ΔHR under restraint stress in IL-1β–responsive neuron-ablated group (red) and control group (black) (control, n = 5 mice; dtA, n = 5 mice, mixed-effects analysis with Šidák correction). (D) AUC of ΔHR under restraint stress conditions in the IL-1β–responsive BNST neuron-ablated group and control group. Data are represented as individual mouse data points pooled from two independent experiments. Unpaired t test. (E) Serum corticosterone levels after 4-h restraint stress. Data are represented as individual mouse data points pooled from two independent experiments. One-way ANOVA with Tukey’s multiple comparison test. (F) Schematic of ablating CRH+ BNST neurons bilaterally in crh-cre mice and then exposure to retrain stress. (G) Serum IL-6 levels at 4 h after restraint stress in crh-cre mice: naive group without restraint stress, control dtA (−), and CRH+ BNST neuron-ablated dtA (+) mice. Data are represented as individual mouse data points pooled from two independent experiments. One-way ANOVA with Tukey’s multiple comparison test. (H) ΔHR under restraint stress in control (black) and CRH+ neuron-ablated group (red) (control, n = 6 mice; dtA, n = 5 mice, P = 0.078, mixed-effects analysis with Šidák correction). (I) AUC of ΔHR under restraint stress. Data are represented as individual mouse data points pooled from two independent experiments. Unpaired t test. (J) Serum corticosterone levels at 4-h postrestraint stress in control, CRH+ BNST neuron-ablated group, and control group without restraint stress. Data are represented as individual mouse data points pooled from two independent experiments. One-way ANOVA with Tukey’s multiple comparison test. *P < 0.05, **P < 0.01, and ***P < 0.001.

IL-1β–responsive BNST neurons are necessary for stress-induced inflammatory...

in Central neurons encode interleukin-1β signals and mediate stress-induced inflammation > Journal of Experimental Medicine

Published: 26 February 2026

Figure 6. IL-1β–responsive BNST neurons are necessary for stress-induced inflammatory responses. (A) Schematic of ablating IL-1β–responsive BNST neurons bilaterally in TRAP2 mice and then exposure to restraint stress. (B) Serum IL-6 levels in More about this image found in IL-1β–responsive BNST neurons are necessary for stress-induced inflammatory...

Journal Articles

Allergic airway reactions rewired by PI3Kδ mutation

Lawrence P. Kane

J Exp Med (2026) 223 (4): e20252509.

https://doi.org/10.1084/jem.20252509

Published: 20 February 2026

View article titled, Allergic airway reactions rewired by PI3Kδ mutation

Journal Articles

Immune checkpoint inhibitor–induced myocarditis is dependent on CD8 T cell–derived TNF and TNFR2 signaling

Kathrynne A. Warrick, Anne Katrine Z. Johansen, Mengchi Jiao, Megan E. Linnemann, Irene Saha, Suh-Chin J. Lin, Charles N. Vallez, Thomas Hagan, Jeffery D. Molkentin, Chandrashekhar Pasare

J Exp Med (2026) 223 (4): e20251717.

https://doi.org/10.1084/jem.20251717

Published: 20 February 2026

View article titled, Immune checkpoint inhibitor–induced myocarditis is dependent on CD8 T cell–derived TNF and TNFR2 signaling

Includes: Supplementary data

Journal Articles

A PI3Kδ-Foxo1-FasL signaling amplification loop rewires CD4⁺ T cell signaling and differentiation

J Exp Med (2026) 223 (4): e20252154.

https://doi.org/10.1084/jem.20252154

Published: 20 February 2026

View article titled, A PI3Kδ-Foxo1-FasL signaling amplification loop rewires CD4+ T cell signaling and differentiation

Includes: Supplementary data

Images

$Immune checkpoint inhibition drives myocarditis in a novel mouse model. (A) Schematic diagram illustrating the DTG tetracycline–repressor system, driven by a myosin heavy chain, α isoform (MHC-α) promoter, for inducible overexpression of OVA in the heart. (B) Representative western blot for OVA from hearts of WT, tetracycline-regulated transactivator (tTA), OVA TG, OVA DTG, and OVA DTG + doxycycline (DOX) mice. β-Tubulin included for loading control. (C) Experimental regimen of early DOX administration to inhibit OVA expression during embryonic and postnatal development in DTG animals (DOX was removed at 5 wk of age). Transfer of 105 purified naive OTI T cells (injected retro-orbitally) was followed by the administration of αCTLA-4 and αPD-1 mouse monoclonal antibodies (dual ICI, 200 μg IP each). (D) Representative flow cytometry contour plots of CD45.1 congenically marked TCR Vα2+ OTI cells in the heart at day 9 following the adoptive transfer of OTI cells for myocarditis induction (pre-gated on live CD45.2+CD90.2+CD8+ cells). (E) Quantification of OTI expansion indicated by cells per mg of cardiac tissue. (F) Quantification of cardiac-infiltrating immune cells measured via flow cytometric analysis of CD45.2+ cells (n = 3). (G) Representative histological images of hearts stained with H&E from DTG mice receiving adoptive transfer (no ICI) and DTG mice receiving αCTLA-4 and αPD-1 (ICI) following OTI transfer (+dual ICI). Scale bar, 200 μm. (H) Kaplan–Meier survival curve following transfer of 106 OTI cells, with or without dual ICI administration (n = 8–9). (I) Representative echocardiograms performed at day 7 after adoptive transfer. (J) Echocardiographic analysis of fractional shortening (%FS) (n = 8–9). “D-1” indicates baseline %FS prior to adoptive transfer. (K) Heart weight-to-body weight (HW/BW) ratio (n = 6–8). (L) Representative electrocardiograms recorded at day 7 after transfer. All data points in graphs represent biological replicates. Statistical analysis performed via one-way ANOVA (E and F) and Student’s t test (H, J, and K). P value determined using Mantel–Cox log-rank (H). All data shown are mean ± SEM (E, F, J, and K). Data are representative of two to three independent experiments. **P < 0.01, ***P < 0.001; ns, not significant. Source data are available for this figure: SourceData F1.$

Immune checkpoint inhibition drives myocarditis in a novel mouse model. (A)...

in Immune checkpoint inhibitor–induced myocarditis is dependent on CD8 T cell–derived TNF and TNFR2 signaling > Journal of Experimental Medicine

Published: 20 February 2026

Figure 1. Immune checkpoint inhibition drives myocarditis in a novel mouse model. (A) Schematic diagram illustrating the DTG tetracycline–repressor system, driven by a myosin heavy chain, α isoform (MHC-α) promoter, for inducible overexpression More about this image found in Immune checkpoint inhibition drives myocarditis in a novel mouse model. (A)...

Images

PD-1 blockade induces de novo priming and differentiation of antigen-specific CD8 T cells. (A and B) Representative flow cytometry contour plots and cell numbers showing the kinetics of OTI T cell expansion in the MedLN at day 4.5, 6, or 9 after adoptive transfer. (C and D) Representative flow cytometry contour plots and cell numbers showing the kinetics of OTI T cell expansion in the heart at day 4.5, 6, or 9 after adoptive transfer. Quantified in D. n = 3 for no ICI condition; n = 5 for ICI condition (B and D). Statistical analysis performed via Student’s t test for each time point. (E and F) Representative gating of CD44hi, granzyme B+ (GrzmB) endogenous CD8 T cells (Endo. CD8 T cell), and OTI T cells from MedLN and heart at day 6 or day 9 after adoptive transfer (E). Quantified in F. (G) Intracellular IFNγ staining of Phorbol 12-myristate 13-acetate (PMA) and ionomycin-stimulated CD8 T cells isolated from MedLN or heart 5 h after stimulation in the presence of Brefeldin A. FMO indicates fluorescence minus one control. n = 4 DTG mice receiving OTI T cell transfer and ICI (both endogenous CD8 T cells and OTI T cells from each animal). (H) Representative flow cytometry plots indicating OTI (TCR Vα2+CD45.1+) T cell expansion in MedLNs at day 9 after adoptive transfer. Mice received three doses (200 μg) of αCTLA-4 antibody, αPD-1 antibody, both (ICI), or vehicle control on days 1–3. (I) Quantification of H (n = 3–5). Statistical analysis performed via one-way ANOVA. All data shown are mean ± SEM. *P < 0.05, **P < 0.01, and ****P < 0.0001; ns, not significant.

PD-1 blockade induces de novo priming and differentiation ...

in Immune checkpoint inhibitor–induced myocarditis is dependent on CD8 T cell–derived TNF and TNFR2 signaling > Journal of Experimental Medicine

Published: 20 February 2026

Figure 2. PD-1 blockade induces de novo priming and differentiation of antigen-specific CD8 T cells. (A and B) Representative flow cytometry contour plots and cell numbers showing the kinetics of OTI T cell expansion in the MedLN at day 4.5, 6, More about this image found in PD-1 blockade induces de novo priming and differentiation ...

Images

Cardiac-specific CD8 T cells drive innate inflammation and myocardial damage. (A) Quantification of cardiac-infiltrating myeloid cells (CD45.2+CD11b+ and CD45.2+CD11c+) and total T cells (endogenous and OTI T cells, CD45.2+CD90.2+) determined by flow cytometry analysis of the total heart on day 9 after adoptive transfer. (B) Delineation of myeloid subsets including myeloid DCs (CD11b+CD11c+), lymphoid DCs (CD11b−CD11c+), neutrophils (Neut) (CD11b+CD11c−Ly6chiLy6Ghi), inflammatory monocytes (Inflamm. mono) (CD11b+CD11c−Ly6chiLy6Glo), and macrophages (Macs) (CD11b+F4/80+). Statistics included in Fig. S4 B. (C) Representative immunohistochemistry for myeloid cells (CD68-white) and cell outlines with wheat germ agglutinin (WGA) (green) with DAPI-stained nuclei (blue) from ICI and control hearts at day 9 after OTI T cell transfer. Scale bar, 100 μm. (D) qRT-PCR analysis of Il1b and Il6 mRNA expression relative to Gapdh expression from bulk cardiac tissue (n = 3–6). (D and E) qRT-PCR analysis of Nppa, Nppb, and Myh7 mRNA expression from bulk cardiac tissue (n = 3–6). Statistical analysis performed via one-way ANOVA (D and E). Depicted as mean ± SEM. Data are representative of two independent experiments (A–E). (F) Schematic of experimental outline for bulk RNA-seq analysis of FACS-purified immune cells (CD45.2+CD90.2−) isolated from hearts of ICI-treated or untreated mice on day 9 after adoptive transfer. (G) Heatmap of inflammatory genes in sorted cardiac-infiltrating myeloid cells from mice treated with ICI following OT-I–adoptive transfer, compared with isotype antibody–treated control mice. The heatmap displays row-wise Z-scores of DESeq2-normalized RNA-seq counts. Each column represents a biological replicate sample grouped by treatment, while genes are grouped by pathways. Data are representative of a single experiment with three biological replicates per condition (G). *P < 0.05, **P < 0.01, and ***P < 0.001; ns, not significant.

Cardiac-specific CD8 T cells drive innate inflammation and myocardial damag...

in Immune checkpoint inhibitor–induced myocarditis is dependent on CD8 T cell–derived TNF and TNFR2 signaling > Journal of Experimental Medicine

Published: 20 February 2026

Figure 3. Cardiac-specific CD8 T cells drive innate inflammation and myocardial damage. (A) Quantification of cardiac-infiltrating myeloid cells (CD45.2⁺CD11b⁺ and CD45.2⁺CD11c⁺) and total T cells (endogenous and OTI T cells, CD45.2⁺CD90.2⁺) More about this image found in Cardiac-specific CD8 T cells drive innate inflammation and myocardial damag...

Images

CD8 T cell–derived TNF is required for ICI-induced myocarditis pathogenesis and mortality. (A) qRT-PCR analysis of Tnf mRNA expression from bulk cardiac tissue (n = 3–6). (B) Infiltration of leukocytes (represented as cell number per mg of tissue) quantified by flow cytometry at day 9 after adoptive transfer of either 105 TNF-sufficient (WT OTI) or 105 TNF-deficient (Tnf−/− OTI) T cells. (C) qRT-PCR analysis of Il1b mRNA expression from bulk cardiac tissue. (D) qRT-PCR analysis of Il6 mRNA expression from bulk cardiac tissue. n = 4 per condition. Statistical analysis performed via one-way ANOVA. Data depicted as mean ± SEM. (E) Experimental regimen for therapeutic targeting of TNF. Mice received standard three doses of ICI for myocarditis injection with additional administration of TNF neutralizing antibody (αTNF, 500 μg) on day 3 and day 6 following the adoptive transfer of 106 OTI cells. (F) Survival of DTG mice receiving Tnf+/+ (WT) OTI cells, with and without αTNF treatment, or Tnf−/− OTI cells following myocarditis induction with ICI. n = 4, WT OTI +ICI. n = 5, Tnf−/− OTI +ICI. n = 5, WT OTI +ICI +αTNF. (G) Representative electrocardiograms of mice following the development of fulminant myocarditis on day 7 after adoptive transfer. (H) Heart rate (beats per minute) averaged over a 2-min interval recorded at day 7 after transfer (n = 4–6). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001; ns, not significant.

CD8 T cell – derived TNF is required for ICI-induced myocarditis pathogenes...

in Immune checkpoint inhibitor–induced myocarditis is dependent on CD8 T cell–derived TNF and TNFR2 signaling > Journal of Experimental Medicine

Published: 20 February 2026

Figure 4. CD8 T cell – derived TNF is required for ICI-induced myocarditis pathogenesis and mortality. (A) qRT-PCR analysis of Tnf mRNA expression from bulk cardiac tissue (n = 3–6). (B) Infiltration of leukocytes (represented as cell number More about this image found in CD8 T cell – derived TNF is required for ICI-induced myocarditis pathogenes...

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