Journal: Journal of neuroinflammation

Article Title: OTUD1 exacerbates sepsis-associated encephalopathy by promoting HK2 mitochondrial release to drive microglia pyroptosis.

doi: 10.1186/s12974-025-03480-w

Figure Lengend Snippet: Fig. 4 OTUD1 deficiency reduces pyroptosis of microglia in vivo.(A-B) The levels of Iba-1, NLRP3, Caspase-1 P20, and GSDMD-NT in the hippocampus were measured by western blot (n = 3/group). (C-F) TNF-α, IL-6, IL-1β and IL-18 were measured in the hippocampus by ELISA (n = 3/group). (G-H) Numbers of GSDMD-NT/Iba-1 positive cells in the hippocampus were measured by double immunofluorescence staining (n = 5/group). Scale bar, 20 μm. **p<0.01 vs. WT+Sham group, ##p<0.01vs. WT+CLP group.

Article Snippet: 8-week-old male C57BL/6J mice were used, including wild-type (WT) mice from the Hubei Province Center and OTUD1−/− mice from Cyagen Biosciences.

Techniques: In Vivo, Western Blot, Enzyme-linked Immunosorbent Assay, Double Immunofluorescence Staining

Journal: Journal of neuroinflammation

Article Title: OTUD1 exacerbates sepsis-associated encephalopathy by promoting HK2 mitochondrial release to drive microglia pyroptosis.

doi: 10.1186/s12974-025-03480-w

Figure Lengend Snippet: Fig. 6 OTUD1 promotes pyroptosis by facilitating the release of HK2 from mitochondria. (A) Potential networks of interacting proteins of OTUD1 revealed by PPI String software. (B-C) The expression of HK2 in primary microglia were measured by western blot (n = 3/group). (D-E) The colocalization of HK2 and VDAC1 was analyzed by immunofluorescence (n = 3/group). Scale bar, 8 μm. (F-G) ER Ca2+ of primary microglia located by ER-targeted plasmid CMV- ER-LAR-GECO1 were observed by flow cytometry (n = 3/group). (H) The content of cytosolic mtDNA was detected by qPCR (n = 3/group). **p < 0.01 vs. Control group; ##p < 0.01 vs. LPS/Nigericin group.

Article Snippet: 8-week-old male C57BL/6J mice were used, including wild-type (WT) mice from the Hubei Province Center and OTUD1−/− mice from Cyagen Biosciences.

Techniques: Software, Expressing, Western Blot, Immunofluorescence, Plasmid Preparation, Flow Cytometry, Control

Journal: Journal of neuroinflammation

Article Title: OTUD1 exacerbates sepsis-associated encephalopathy by promoting HK2 mitochondrial release to drive microglia pyroptosis.

doi: 10.1186/s12974-025-03480-w

Figure Lengend Snippet: Fig. 7 OTUD1 interacts with HK2. (A) Molecular docking predicts the possibility and binding sites of the interaction between OTUD1 and HK2. Scale bar, 5 μm. (B-D) 3D confocal imaging to detect the colocalization of OTUD1 and HK2 (n = 5/group). (E-F) Transfection of Flag-OTUD1 and Myc-HK2 plasmids into 293T cells, followed by exogenous Co-IP to detect the interaction between OTUD1 and HK2 (n = 3/group). (G) The interaction between OTUD1 and HK2 in BV2 cells was evaluated by endogenous Co-IP (n = 3/group).

Article Snippet: 8-week-old male C57BL/6J mice were used, including wild-type (WT) mice from the Hubei Province Center and OTUD1−/− mice from Cyagen Biosciences.

Techniques: Binding Assay, Imaging, Transfection, Co-Immunoprecipitation Assay

Journal: Journal of neuroinflammation

Article Title: OTUD1 exacerbates sepsis-associated encephalopathy by promoting HK2 mitochondrial release to drive microglia pyroptosis.

doi: 10.1186/s12974-025-03480-w

Figure Lengend Snippet: Fig. 8 OTUD1 regulates the activity of HK2 protein through K63-linked ubiquitination. (A-B) Domain mapping of OTUD1 and HK2. (C-D) Transfection of truncated forms of OTUD1 and HK2 in 293T cells, followed by Co-IP to detect the interaction domains between OTUD1 and HK2 (n = 3/group). (E) In vitro assay of HK2 deubiquitination by OTUD1 and OTUD1 C320S (n = 3/group). (F) Transfection of Flag-OTUD1, Myc-HK2, HA-Ub, HA-K48 and HA-K63 into293T cells, followed by western blot to detect the ubiquitination of HK2 (n = 3/group). (G) Transfection of Flag-OTUD1, Myc-HK2, HA-Ub, HA-K48R and HA-K63R into 293T cells, followed by western blot to detect the ubiquitination of HK2 (n = 3/group).

Article Snippet: 8-week-old male C57BL/6J mice were used, including wild-type (WT) mice from the Hubei Province Center and OTUD1−/− mice from Cyagen Biosciences.

Techniques: Activity Assay, Ubiquitin Proteomics, Transfection, Co-Immunoprecipitation Assay, In Vitro, Western Blot

Journal: Journal of neuroinflammation

Article Title: OTUD1 exacerbates sepsis-associated encephalopathy by promoting HK2 mitochondrial release to drive microglia pyroptosis.

doi: 10.1186/s12974-025-03480-w

Figure Lengend Snippet: Fig. 9 OTUD1 is closely associated with human sepsis and 28-day survival rate. (A) Expression levels of OTUD1 in sepsis patients compared to non-sepsis patients. (B) Relationship between OTUD1 expression levels and 28-day mortality in sepsis patients. (C) GSEA of high OTUD1 expression and low OTUD1 expression. (D-F) Pearson’s correlation analysis between OTUD1 and TNF-α, IL-1β, and IL-18 inflammatory factors.

Article Snippet: 8-week-old male C57BL/6J mice were used, including wild-type (WT) mice from the Hubei Province Center and OTUD1−/− mice from Cyagen Biosciences.

Techniques: Expressing

Journal: Journal of neuroinflammation

Article Title: OTUD1 exacerbates sepsis-associated encephalopathy by promoting HK2 mitochondrial release to drive microglia pyroptosis.

doi: 10.1186/s12974-025-03480-w

Figure Lengend Snippet: Fig. 10 Mechanistic schematic illustrating OTUD1-mediated deubiquitination of HK2 in driving microglia pyroptosis during SAE.

Article Snippet: 8-week-old male C57BL/6J mice were used, including wild-type (WT) mice from the Hubei Province Center and OTUD1−/− mice from Cyagen Biosciences.

Techniques: