iba1  (Proteintech)

Journal: iScience

Article Title: Cerebral organoid exosomes reversed behavioral deficits by repressing NLRP3-mediated neuroinflammation in stress models

doi: 10.1016/j.isci.2026.115069

Figure Lengend Snippet: OExo alleviates LPS-induced microglia activation by suppressing NLRP3 expression (A) Representative images of IBA-1 immunostaining reveal the uptake of PKH26-labeled OExo by IBA-1-positive BV-2 microglia after 48 h of incubation. The white arrows highlight the presence of PKH26-labeled OExo surrounding the nuclei of microglia. (B) Representative images of immunostaining for CD206 (top) and iNOS (bottom) demonstrating BV-2 microglia treated with CON (PBS), OExo (50 μg/mL), LPS (100 ng/mL), and LPS combined with OExo (LPS+OExo) for 24 h. (C and D) Quantitative analysis of the fluorescence intensity of CD206 (C) and iNOS (D). n = 6. (E) q-PCR analysis of the pro-inflammatory cytokines IL-6, IL-1β, and TNF-α in BV-2 microglia treated with CON, OExo, LPS, and LPS+OExo, with glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as the internal reference gene. n = 9. (F) Representative WB images displaying the expression of CD206, Arg-1, and iNOS in BV-2 cells treated with CON, OExo, LPS, and LPS+OExo, with β-actin as the internal control. Full-length blots are presented in . (G) Statistical analysis of CD206, Arg-1, and iNOS expression levels from WB. n = 6. (H) Representative WB images illustrating the NLRP3, Casp-1, and IL-1β expression levels in BV-2 cells treated with CON, OExo, LPS, LPS+OExo, nigericin, and nigericin+LPS+OExo. nigericin, a NLRP3 activator, was utilized to inhibit the rescuing effects of OExo on LPS-induced NLRP3 activation. β-actin was used as the internal control. Full-length blots are presented in . (I) Statistical analysis of NLRP3, Casp-1, and IL-1β expression levels by WB. n = 4. (J) Representative WB images showing NLRP3 levels in BV-2 cells treated with CON, MExo, OExo, LPS, LPS+MExo and LPS+OExo. β-actin was used as the internal control. Full-length blots are presented in . (K) Statistical analysis of NLRP3 expression levels in (J). n = 4. All western blot quantifications are based on at least three biological replicates, with two to three technical replicates per experiment. Scale bars, 50 μm. DAPI was used for nuclear counterstaining in all images. Data are presented as mean ± SEM. Data are subjected to one-way ANOVA followed by the Tukey’s test. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001.

Article Snippet: IBA-1 , Protein tech; Wako , Cat# 10904-1-AP; Cat# 019-19741.

Techniques: Activation Assay, Expressing, Immunostaining, Labeling, Incubation, Fluorescence, Control, Western Blot

Journal: iScience

Article Title: Cerebral organoid exosomes reversed behavioral deficits by repressing NLRP3-mediated neuroinflammation in stress models

doi: 10.1016/j.isci.2026.115069

Figure Lengend Snippet: Intranasal administration of OExo attenuated microglia activation in the hippocampus in mouse stress models (A) Representative images of immunofluorescence staining for IBA-1 showing PKH26-labeled OExo co-localized with IBA-1-positive microglia in the hippocampus 24 h after intranasal administration of OExo. Nuclei were counterstained with DAPI. The region within the white dashed box is enlarged in the lower image. White arrows indicate PKH26-labeled exosomes surrounding DAPI-stained nuclei in IBA-1-positive microglia. (B) Low-magnification representative images of IBA-1 and DAPI staining showing the hippocampal subregions CA1, CA3, and DG. (C) Representative immunofluorescence images of IBA-1 in the hippocampal CA1, CA3, and DG subregions. Insets show magnified views of the regions within the dashed boxes. (D) Quantitative analysis of IBA-1-positive microglia in the CA1, CA3, and DG subregions. n = 6. (E) qPCR analysis of pro-inflammatory cytokines ( IL-6 , IL-1β , and TNF-α ) in the hippocampus. GAPDH was used as the internal reference gene. n = 9. (F) Representative western blot images of Arg-1 and iNOS protein expression in the hippocampus. β-actin was used as the loading control. Full-length blots are presented in . (G) Representative WB images of NF-κB, p-NF-κB, NLRP3, IL-1β, and Casp-1 protein expression in the hippocampus. β-actin was used as the loading control. Full-length blots are presented in . (H) Quantitative analysis of Arg-1 and iNOS protein expression from western blot data. n = 6. (I) Quantitative analysis of NF-κB, p-NF-κB, NLRP3, IL-1β, and Casp-1 protein expression from WB data. n = 6. (J) Representative double immunofluorescence staining of NLRP3 and IBA-1 in the hippocampus. The regions within the white dashed boxes are enlarged in the right image. White arrows indicate cells positive for both NLRP3 and IBA-1. (K) Quantitative analysis of cells positive for both NLRP3 and IBA-1 among IBA-1-positive microglia in the hippocampus. n = 6. Scale bars, 20 μm (upper) and 10 μm (bottom) in (A); 1,000 μm in (B); 100 μm in (C); and 20 μm in the magnified insets of (C) and 50 μm in (J). Data are presented as mean ± SEM. The interaction between CMS and OExo was evaluated using two-way ANOVA followed by Tukey’s test (details are shown in ). ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001.

Article Snippet: IBA-1 , Protein tech; Wako , Cat# 10904-1-AP; Cat# 019-19741.

Techniques: Activation Assay, Immunofluorescence, Staining, Labeling, Western Blot, Expressing, Control, Double Immunofluorescence Staining