Journal: Frontiers in Neurology

Article Title: HMGB1-TLR4 signaling-mediated neuroinflammation contributes to the pathogenesis of infantile epileptic spasms syndrome in rats

doi: 10.3389/fneur.2026.1848952

Figure Lengend Snippet: Behavioral results. Experimental grouping: Non-model group (pups from non-prenatal stress pregnant rats) was divided into blank control (BC, no treatment) and negative control (NC, normal saline instead of drugs); Model group (pups from prenatal stress pregnant rats, IESS model induced by prenatal stress + NMDA intraperitoneal injection) was divided into 5 subgroups: Model I (IESS+ACTH, 100 IU/kg intraperitoneal injection), Model II (IESS+anti-HMGB1, 300 μg/kg intraperitoneal injection), Model III (IESS+ACTH+anti-HMGB1), Model IV (IESS+NS, equal volume normal saline), Model V (IESS, positive control, no treatment); Detection indicator: the seizure latency:time from intraperitoneal injection of NMDA to the onset of epileptic seizures; seizure severity score were recorded after intraperitoneal injection of NMDA on P12 and P13, respectively. Statistical significance description: (* P < 0.05 vs. BC group, # P < 0.05 vs. NC group, & P < 0.05 vs. IESS+NS group, $ P < 0.05 vs. IESS group) (one-way ANOVA).

Article Snippet: The sections were permeabilized with 0.5% Triton X-100 at room temperature for 15 min. After washing with PBS, the sections were blocked with 5% BSA at 37 °C for 30 min. Primary antibody Rabbit Anti-HMGB1 (Abclonal, A19529, 1/200) was added dropwise to the slides and incubated at 4°C overnight; the next day, after the slides were rewarmed to room temperature, they were washed with PBS 3 times for 3 min each, and HRP-labeled goat anti-rabbit IgG (H+L) (ZB-2,301, Zhongshan Jinqiao, 1/200) was added dropwise and incubated at room temperature for 50 min. CY3-TSA (G1223, Servicebio, 1/1,500) was added and incubated at room temperature for 10 min in the dark.

Techniques: Control, Negative Control, Saline, Injection, Positive Control

Journal: Frontiers in Neurology

Article Title: HMGB1-TLR4 signaling-mediated neuroinflammation contributes to the pathogenesis of infantile epileptic spasms syndrome in rats

doi: 10.3389/fneur.2026.1848952

Figure Lengend Snippet: qPCR detection results of brain tissues. Experimental grouping: Non-model group (pups from non-prenatal stress pregnant rats) was divided into blank control (BC, no treatment) and negative control (NC, normal saline instead of drugs); Model group (pups from prenatal stress pregnant rats, IESS model induced by prenatal stress + NMDA intraperitoneal injection) was divided into 5 subgroups: Model I (IESS+ACTH, 100 IU/kg intraperitoneal injection), Model II (IESS+anti-HMGB1, 300 μg/kg intraperitoneal injection), Model III (IESS+ACTH+anti-HMGB1), Model IV (IESS+NS, equal volume normal saline), Model V (IESS, positive control, no treatment); Detection indicator: the mRNA expressions of HMGB1 and TLR4. Detection method: GAPDH was used as the internal reference, and the relative expression level of genes was calculated according to the 2 −ΔΔ Ct method; Statistical significance description: (* P < 0.05 vs. BC group, # P < 0.05 vs. NC group, & P < 0.05 vs. IESS+NS group, $ P < 0.05 vs. IESS group, % P < 0.05 vs. IESS+ACTH group, @ P < 0.05 vs. IESS+anti-HMGB1 group) (one-way ANOVA).

Article Snippet: The sections were permeabilized with 0.5% Triton X-100 at room temperature for 15 min. After washing with PBS, the sections were blocked with 5% BSA at 37 °C for 30 min. Primary antibody Rabbit Anti-HMGB1 (Abclonal, A19529, 1/200) was added dropwise to the slides and incubated at 4°C overnight; the next day, after the slides were rewarmed to room temperature, they were washed with PBS 3 times for 3 min each, and HRP-labeled goat anti-rabbit IgG (H+L) (ZB-2,301, Zhongshan Jinqiao, 1/200) was added dropwise and incubated at room temperature for 50 min. CY3-TSA (G1223, Servicebio, 1/1,500) was added and incubated at room temperature for 10 min in the dark.

Techniques: Control, Negative Control, Saline, Injection, Positive Control, Expressing

Journal: Frontiers in Neurology

Article Title: HMGB1-TLR4 signaling-mediated neuroinflammation contributes to the pathogenesis of infantile epileptic spasms syndrome in rats

doi: 10.3389/fneur.2026.1848952

Figure Lengend Snippet: WB detection results of brain tissues. Experimental grouping: Non-model group (pups from non-prenatal stress pregnant rats) was divided into blank control (BC, no treatment) and negative control (NC, normal saline instead of drugs); Model group (pups from prenatal stress pregnant rats, IESS model induced by prenatal stress + NMDA intraperitoneal injection) was divided into 5 subgroups: Model I (IESS+ACTH, 100 IU/kg intraperitoneal injection), Model II (IESS+anti-HMGB1, 300 μg/kg intraperitoneal injection), Model III (IESS+ACTH+anti-HMGB1), Model IV (IESS+NS, equal volume normal saline), Model V (IESS,positive control, no treatment); Detection indicator: the protein relative expressions of HMGB1 and TLR4. Detection method: the protein relative expressions levels in brain tissues detected by Western Blot (WB); Statistical significance description: ( * P < 0.05 vs. BC group, # P < 0.05 vs. NC group, & P < 0.05 vs. IESS+NS group, $ P < 0.05 vs. IESS group, % P < 0.05 vs. IESS+ACTH group, @ P < 0.05 vs. IESS+anti-HMGB1 group) (one-way ANOVA).

Article Snippet: The sections were permeabilized with 0.5% Triton X-100 at room temperature for 15 min. After washing with PBS, the sections were blocked with 5% BSA at 37 °C for 30 min. Primary antibody Rabbit Anti-HMGB1 (Abclonal, A19529, 1/200) was added dropwise to the slides and incubated at 4°C overnight; the next day, after the slides were rewarmed to room temperature, they were washed with PBS 3 times for 3 min each, and HRP-labeled goat anti-rabbit IgG (H+L) (ZB-2,301, Zhongshan Jinqiao, 1/200) was added dropwise and incubated at room temperature for 50 min. CY3-TSA (G1223, Servicebio, 1/1,500) was added and incubated at room temperature for 10 min in the dark.

Techniques: Control, Negative Control, Saline, Injection, Positive Control, Western Blot

Journal: Frontiers in Neurology

Article Title: HMGB1-TLR4 signaling-mediated neuroinflammation contributes to the pathogenesis of infantile epileptic spasms syndrome in rats

doi: 10.3389/fneur.2026.1848952

Figure Lengend Snippet: Expression levels of cytokines in brain tissues. Experimental grouping: Non-model group (pups from non-prenatal stress pregnant rats) was divided into blank control (BC, no treatment) and negative control (NC, normal saline instead of drugs); Model group (pups from prenatal stress pregnant rats, IESS model induced by prenatal stress + NMDA intraperitoneal injection) was divided into 5 subgroups: Model I (IESS+ACTH, 100 IU/kg intraperitoneal injection), Model II (IESS+anti-HMGB1, 300 μg/kg intraperitoneal injection), Model III (IESS+ACTH+anti-HMGB1), Model IV (IESS+NS, equal volume normal saline), Model V (IESS, positive control, no treatment); Detection indicator: expression levels of cytokines in brain tissues. Detection method: expression levels of cytokines in brain tissues detected by Enzyme-Linked Immunosorbent Assay (ELISA); Statistical significance description: (* P < 0.05 vs. BC group, # P < 0.05 vs. NC group, & P < 0.05 vs. IESS+NS group, $ P < 0.05 vs. IESS group, % P < 0.05 vs. IESS+ACTH group, @ P < 0.05 vs. IESS+anti-HMGB1 group) (one-way ANOVA).

Article Snippet: The sections were permeabilized with 0.5% Triton X-100 at room temperature for 15 min. After washing with PBS, the sections were blocked with 5% BSA at 37 °C for 30 min. Primary antibody Rabbit Anti-HMGB1 (Abclonal, A19529, 1/200) was added dropwise to the slides and incubated at 4°C overnight; the next day, after the slides were rewarmed to room temperature, they were washed with PBS 3 times for 3 min each, and HRP-labeled goat anti-rabbit IgG (H+L) (ZB-2,301, Zhongshan Jinqiao, 1/200) was added dropwise and incubated at room temperature for 50 min. CY3-TSA (G1223, Servicebio, 1/1,500) was added and incubated at room temperature for 10 min in the dark.

Techniques: Expressing, Control, Negative Control, Saline, Injection, Positive Control, Enzyme-linked Immunosorbent Assay

Journal: Frontiers in Neurology

Article Title: HMGB1-TLR4 signaling-mediated neuroinflammation contributes to the pathogenesis of infantile epileptic spasms syndrome in rats

doi: 10.3389/fneur.2026.1848952

Figure Lengend Snippet: Immunohistochemical detection results. Experimental grouping: Non-model group (pups from non-prenatal stress pregnant rats) was divided into blank control (BC, no treatment) and negative control (NC, normal saline instead of drugs); Model group (pups from prenatal stress pregnant rats, IESS model induced by prenatal stress + NMDA intraperitoneal injection) was divided into 5 subgroups: Model I (IESS+ACTH, 100 IU/kg intraperitoneal injection), Model II (IESS+anti-HMGB1, 300 μg/kg intraperitoneal injection), Model III (IESS+ACTH+anti-HMGB1), Model IV (IESS+NS, equal volume normal saline), Model V (IESS, positive control, no treatment); Detection indicator: the expression of inducible nitric oxide synthase (iNOS) and arginase 1 (Arg1) in the brain tissues (hippocampus). Detection method: Immunohistochemical detection; Statistical significance description: (* P < 0.05 vs. BC group, # P < 0.05 vs. NC group, & P < 0.05 vs. IESS+NS group, $ P < 0.05 vs. IESS group, % P < 0.05 vs. IESS+ACTH group) (one-way ANOVA).

Article Snippet: The sections were permeabilized with 0.5% Triton X-100 at room temperature for 15 min. After washing with PBS, the sections were blocked with 5% BSA at 37 °C for 30 min. Primary antibody Rabbit Anti-HMGB1 (Abclonal, A19529, 1/200) was added dropwise to the slides and incubated at 4°C overnight; the next day, after the slides were rewarmed to room temperature, they were washed with PBS 3 times for 3 min each, and HRP-labeled goat anti-rabbit IgG (H+L) (ZB-2,301, Zhongshan Jinqiao, 1/200) was added dropwise and incubated at room temperature for 50 min. CY3-TSA (G1223, Servicebio, 1/1,500) was added and incubated at room temperature for 10 min in the dark.

Techniques: Immunohistochemical staining, Control, Negative Control, Saline, Injection, Positive Control, Expressing

Journal: Frontiers in Neurology

Article Title: HMGB1-TLR4 signaling-mediated neuroinflammation contributes to the pathogenesis of infantile epileptic spasms syndrome in rats

doi: 10.3389/fneur.2026.1848952

Figure Lengend Snippet: Double immunofluorescence staining results of NeuN and HMGB1. Experimental grouping: Non-model group (pups from non-prenatal stress pregnant rats) was divided into blank control (BC, no treatment) and negative control (NC, normal saline instead of drugs); Model group (pups from prenatal stress pregnant rats, IESS model induced by prenatal stress + NMDA intraperitoneal injection) was divided into 5 subgroups: Model I (IESS+ACTH, 100 IU/kg intraperitoneal injection), Model II (IESS+anti-HMGB1, 300 μg/kg intraperitoneal injection), Model III (IESS+ACTH+anti-HMGB1), Model IV (IESS+NS, equal volume normal saline), Model V (IESS, positive control, no treatment); Detection indicator: the colocalization expression of NeuN+HMGB1 in the hippocampal tissues. Detection method: Double immunofluorescence staining; Statistical significance description: (* P < 0.05 vs. BC group, # P < 0.05 vs. NC group, & P < 0.05 vs. IESS+NS group, $ P < 0.05 vs. IESS group) (one-way ANOVA).

Article Snippet: The sections were permeabilized with 0.5% Triton X-100 at room temperature for 15 min. After washing with PBS, the sections were blocked with 5% BSA at 37 °C for 30 min. Primary antibody Rabbit Anti-HMGB1 (Abclonal, A19529, 1/200) was added dropwise to the slides and incubated at 4°C overnight; the next day, after the slides were rewarmed to room temperature, they were washed with PBS 3 times for 3 min each, and HRP-labeled goat anti-rabbit IgG (H+L) (ZB-2,301, Zhongshan Jinqiao, 1/200) was added dropwise and incubated at room temperature for 50 min. CY3-TSA (G1223, Servicebio, 1/1,500) was added and incubated at room temperature for 10 min in the dark.

Techniques: Double Immunofluorescence Staining, Control, Negative Control, Saline, Injection, Positive Control, Expressing

Journal: Frontiers in Neurology

Article Title: HMGB1-TLR4 signaling-mediated neuroinflammation contributes to the pathogenesis of infantile epileptic spasms syndrome in rats

doi: 10.3389/fneur.2026.1848952

Figure Lengend Snippet: Double immunofluorescence staining results of GFAP and HMGB1. Experimental grouping: Non-model group (pups from non-prenatal stress pregnant rats) was divided into blank control (BC, no treatment) and negative control (NC, normal saline instead of drugs); Model group (pups from prenatal stress pregnant rats, IESS model induced by prenatal stress + NMDA intraperitoneal injection) was divided into 5 subgroups: Model I (IESS+ACTH, 100 IU/kg intraperitoneal injection), Model II (IESS+anti-HMGB1, 300 μg/kg intraperitoneal injection), Model III (IESS+ACTH+anti-HMGB1), Model IV (IESS+NS, equal volume normal saline), Model V (IESS, positive control, no treatment); Detection indicator: colocalization expression of GFAP+HMGB1. Detection method: Double immunofluorescence staining; Statistical significance description: (* P < 0.05 vs. BC group, # P < 0.05 vs. NC group, & P < 0.05 vs. IESS+NS group, $ P < 0.05 vs. IESS group) (one-way ANOVA).

Article Snippet: The sections were permeabilized with 0.5% Triton X-100 at room temperature for 15 min. After washing with PBS, the sections were blocked with 5% BSA at 37 °C for 30 min. Primary antibody Rabbit Anti-HMGB1 (Abclonal, A19529, 1/200) was added dropwise to the slides and incubated at 4°C overnight; the next day, after the slides were rewarmed to room temperature, they were washed with PBS 3 times for 3 min each, and HRP-labeled goat anti-rabbit IgG (H+L) (ZB-2,301, Zhongshan Jinqiao, 1/200) was added dropwise and incubated at room temperature for 50 min. CY3-TSA (G1223, Servicebio, 1/1,500) was added and incubated at room temperature for 10 min in the dark.

Techniques: Double Immunofluorescence Staining, Control, Negative Control, Saline, Injection, Positive Control, Expressing

Journal: Frontiers in Neurology

Article Title: HMGB1-TLR4 signaling-mediated neuroinflammation contributes to the pathogenesis of infantile epileptic spasms syndrome in rats

doi: 10.3389/fneur.2026.1848952

Figure Lengend Snippet: Double immunofluorescence staining results of ED1 and HMGB1. Experimental grouping: Non-model group (pups from non-prenatal stress pregnant rats) was divided into blank control (BC, no treatment) and negative control (NC, normal saline instead of drugs); Model group (pups from prenatal stress pregnant rats, IESS model induced by prenatal stress + NMDA intraperitoneal injection) was divided into 5 subgroups: Model I (IESS+ACTH, 100 IU/kg intraperitoneal injection), Model II (IESS+anti-HMGB1, 300 μg/kg intraperitoneal injection), Model III (IESS+ACTH+anti-HMGB1), Model IV (IESS+NS, equal volume normal saline), Model V (IESS, positive control, no treatment); Detection indicator: the colocalization expression of ED1+HMGB1. Detection method : Double immunofluorescence staining; Statistical significance description : (* P < 0.05 vs. BC group, # P < 0.05 vs. NC group, & P < 0.05 vs. IESS+NS group, $ P < 0.05 vs. IESS group) (one-way ANOVA).

Article Snippet: The sections were permeabilized with 0.5% Triton X-100 at room temperature for 15 min. After washing with PBS, the sections were blocked with 5% BSA at 37 °C for 30 min. Primary antibody Rabbit Anti-HMGB1 (Abclonal, A19529, 1/200) was added dropwise to the slides and incubated at 4°C overnight; the next day, after the slides were rewarmed to room temperature, they were washed with PBS 3 times for 3 min each, and HRP-labeled goat anti-rabbit IgG (H+L) (ZB-2,301, Zhongshan Jinqiao, 1/200) was added dropwise and incubated at room temperature for 50 min. CY3-TSA (G1223, Servicebio, 1/1,500) was added and incubated at room temperature for 10 min in the dark.

Techniques: Double Immunofluorescence Staining, Control, Negative Control, Saline, Injection, Positive Control, Expressing