rabbit il  (Bioss)

Journal: Hepatology Communications

Article Title: Arrb2 in hepatocytes promotes M2 macrophage polarization, ameliorates hepatic ischemia–reperfusion injury through upregulating metabolite 6-ketoLCA

doi: 10.1097/HC9.0000000000000916

Figure Lengend Snippet: 6-ketoLCA upregulated by Arrb2 in hepatocytes promotes M2 macrophage polarization, alleviating hepatic IRI. (A–E) Liver and serum samples from liver-specific Arrb2-CKO and WT mice with or without 6-ketoLCA treatment after I/R (n=6). (A) HE and TUNEL staining were analyzed microscopically. Scale bar: 200 μm; Suzuki score and necrosis area were examined. (B) Quantitative analysis of the bile acid metabolite 6-ketoLCA in liver tissue. (C) Liver function levels (ALT, AST, TBA, GGT) were analyzed between groups. (D) qRT-PCR analysis of mouse liver tissue mRNA expression for inflammatory factors (IL-6, TNF-α, IL-10, TGF-β). (E) iNOS, CD86, CD206, and CD163 expression in PMMs was assessed by WB analysis. (F) The CCK8 assay measured the viability of RAW264.7 cells exposed to varying concentrations and durations of 6-ketoLCA (n=3). (G, I) Co-culture of RAW264.7 and AML12 cells. (G) ELISA for culture medium supernatant inflammatory factors (IL-6, TNF-α, IL-10, TGF-β). (I) BCL2, BAX, and GAPDH expression in AML12 cells was assessed by WB analysis. (H, J) Flow cytometry detected the proportions of M1 (CD86, PE) and M2 (CD206, APC) macrophages treated with IL-4 and 6-ketoLCA. The ratio of M1/M2 macrophages was measured. All results are presented as mean ± SD, and statistical significance was assessed. * p <0.05, ** p <0.01, and *** p <0.001. Abbreviations: ALT, alanine transaminase; AST, aspartate transaminase; CKO, conditional knockout; GGT, gamma-glutamyl transferase; HE, hematoxylin and eosin; I/R, ischemia/reperfusion; IL-6, interleukin 6; IL-10, interleukin 10; IRI, ischemia–reperfusion injury; qRT-PCR, quantitative reverse transcription polymerase chain reaction; PMMs, primary mouse macrophages; TBA, total bile acids; TGF-β, transforming growth factor-β; TNF-α, tumor necrosis factor-α; TUNEL, terminal deoxynucleotidyl transferase–mediated dUTP nick-end labeling; WB, western blotting; WT, wild type.

Article Snippet: RAW264.7 were cultured in 10% FBS RPMI-1640 medium with 6-ketoLCA (100 ng/mL) and IL-4 (20 ng/mL) to induce polarization to M2-like macrophage. shRNA targeting TGR5 (GeneCopoeia, Rockville, MD, USA) was transfected into RAW264.7 cells utilizing Lipofectamine 3000 (Invitrogen, Carlsbad, CA, USA) in accordance with the manufacturer’s protocol to achieve TGR5 knockdown.

Techniques: TUNEL Assay, Staining, Quantitative RT-PCR, Expressing, CCK-8 Assay, Co-Culture Assay, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Knock-Out, Reverse Transcription, Polymerase Chain Reaction, End Labeling, Western Blot

Journal: Neurobiology of Stress

Article Title: Microglial SIRT6 confers protection against neuroinflammation-associated depression through NRF2-HO1 signaling

doi: 10.1016/j.ynstr.2026.100804

Figure Lengend Snippet: Hippocampal SIRT6 Expression Profile in a Lipopolysaccharide (LPS)-Induced Depression Model. (A) Western blot analysis of SIRT6 expression in the hippocampal brain tissues. D, day. n = 4 mice. (B) Immunostaining for SIRT6 in the hippocampal region three days post-LPS injection revealed an overall reduction in brain tissues, while its expression was concurrently increased specifically in microglia. This was quantified in two graphs showing total SIRT6 expression (left) and microglial SIRT6 expression (right), with arrowheads indicating SIRT6 localization in microglia. n = 4 mice. (C) Western blot analysis of SIRT6 expression in microglia sorted from the hippocampus by MCAS at 3 days post-LPS injection. n = 4 mice. Data are presented as mean ± SEM. Statistical analysis: One-way ANOVA with Tukey's post hoc test in A; Unpaired t -test for two-group comparisons in B-C.

Article Snippet: Membranes were blocked with 5% non-fat milk in TBST for 1 h and incubated overnight at 4 °C with primary antibodies: SIRT6 (12486, Cell Signaling Technology), NRF2 (ab62352, Abcam), HO-1 (ab68477, Abcam), NQO1 (ab80588, Abcam), KEAP1 (8047, Cell Signaling Technology), NLRP3 (AG-20B-0014, AdipoGen), cleaved caspase-1 (4199, Cell Signaling Technology), cleaved IL-1β (83186, Cell Signaling Technology), and β-actin (A1978, Sigma-Aldrich).

Techniques: Expressing, Western Blot, Immunostaining, Injection

Journal: Neurobiology of Stress

Article Title: Microglial SIRT6 confers protection against neuroinflammation-associated depression through NRF2-HO1 signaling

doi: 10.1016/j.ynstr.2026.100804

Figure Lengend Snippet: SIRT6 deficiency potentiates microglial activation in a LPS-induced depression model. (A) Immunostaining of GFAP and IBA1 in the hippocampus area in Sirt6 MCKO and Sirt6 fl/fl mice at day 5 post-LPS injection. n = 4 mice. (B-G) Quantification of (B) IBA1 + and (C) GFAP + cell counts, along with microglial morphology parameters: (D) number of branches, (E) average branch length, (F) total branch length, and (G) soma area. n = 4 mice. (H) qPCR analysis of TNF-α,IL-6, and IL-1β mRNA levels in in the microglial sorted from the hippocampus brain tissue. n = 4 mice. Data are mean ± SEM. Statistical significance between two groups was determined by an unpaired two-tailed Student's t-test.

Article Snippet: Membranes were blocked with 5% non-fat milk in TBST for 1 h and incubated overnight at 4 °C with primary antibodies: SIRT6 (12486, Cell Signaling Technology), NRF2 (ab62352, Abcam), HO-1 (ab68477, Abcam), NQO1 (ab80588, Abcam), KEAP1 (8047, Cell Signaling Technology), NLRP3 (AG-20B-0014, AdipoGen), cleaved caspase-1 (4199, Cell Signaling Technology), cleaved IL-1β (83186, Cell Signaling Technology), and β-actin (A1978, Sigma-Aldrich).

Techniques: Activation Assay, Immunostaining, Injection, Two Tailed Test

Journal: Neurobiology of Stress

Article Title: Microglial SIRT6 confers protection against neuroinflammation-associated depression through NRF2-HO1 signaling

doi: 10.1016/j.ynstr.2026.100804

Figure Lengend Snippet: Deletion of microglial Sirt6 inhibited the NRF2-HO1 signaling and worsened the peroxidation damage. (A) Gene Ontology (GO) analysis was performed on RNA-Seq data from microglia sorted from Sirt6 MCKO and Sirt6 fl/fl control mice. (B) TAC, MDA, SOD, and GSH/GSSG levels at 5 days after LPS injection. n = 4 mice. (C) Gene Set Enrichment Analysis (GSEA) of RNA-Seq data profiled from microglia sorted from Sirt6 MCKO and Sirt6 fl/fl control mice. (D-E) Analysis of NRF2-HO1 and associated signaling proteins in sorted microglia. Protein levels of NRF2, KEAP1, HO-1, NQO1, NLRP3, Cleaved Caspase-3, and Cleaved IL-1β were assessed by Western blot (D) and quantified (E). n = 4 mice. Data are mean ± SEM. Statistical significance between two groups was determined by an unpaired two-tailed Student's t-test.

Article Snippet: Membranes were blocked with 5% non-fat milk in TBST for 1 h and incubated overnight at 4 °C with primary antibodies: SIRT6 (12486, Cell Signaling Technology), NRF2 (ab62352, Abcam), HO-1 (ab68477, Abcam), NQO1 (ab80588, Abcam), KEAP1 (8047, Cell Signaling Technology), NLRP3 (AG-20B-0014, AdipoGen), cleaved caspase-1 (4199, Cell Signaling Technology), cleaved IL-1β (83186, Cell Signaling Technology), and β-actin (A1978, Sigma-Aldrich).

Techniques: RNA Sequencing, Control, Injection, Western Blot, Two Tailed Test

Journal: Neurobiology of Stress

Article Title: Microglial SIRT6 confers protection against neuroinflammation-associated depression through NRF2-HO1 signaling

doi: 10.1016/j.ynstr.2026.100804

Figure Lengend Snippet: Knockout of the Nrf2 in microglial aggravated the depression behavior and facilitated the microglial activation. (A) Experimental timeline depicting the sequential administration of tamoxifen and LPS, followed by behavioral assessment. (B) Western blot analysis confirming the successful knockout of Nrf2 in microglial cells of Nrf2 MCKO mice. n = 3 mice. (C-D) Quantification of the duration of immobility in the (C) TST) and (D) FST. n = 8 mice. (E) Levels of TAC, MDA, SOD, and the GSH/GSSG ratio were measured in Nrf2 MCKO and Nrf2 fl/fl mice 3 days after LPS challenge. (F) Immunostaining of GFAP and IBA1 in the hippocampus area of Nrf2 MCKO and Nrf2 fl/fl mice 3 days after LPS injection. (G-L) Quantification of (G) IBA1 + and (H) GFAP + cell counts, along with microglial morphology parameters: (I) number of branches, (J) average branch length, (K) total branch length, and (L) soma area. n = 4 mice. (M) qPCR analysis of TNF-α, IL-6, and IL-1β mRNA levels in the microglial sorted from the hippocampus brain tissue. n = 4 mice. (N) Western blot analysis was conducted to determine the protein levels of SIRT6 and NRF2 downstream signaling components in primary microglia purified from the mouse hippocampus. n = 4 mice. Data are mean ± SEM. Statistical significance between two groups was determined by an unpaired two-tailed Student's t-test.

Article Snippet: Membranes were blocked with 5% non-fat milk in TBST for 1 h and incubated overnight at 4 °C with primary antibodies: SIRT6 (12486, Cell Signaling Technology), NRF2 (ab62352, Abcam), HO-1 (ab68477, Abcam), NQO1 (ab80588, Abcam), KEAP1 (8047, Cell Signaling Technology), NLRP3 (AG-20B-0014, AdipoGen), cleaved caspase-1 (4199, Cell Signaling Technology), cleaved IL-1β (83186, Cell Signaling Technology), and β-actin (A1978, Sigma-Aldrich).

Techniques: Knock-Out, Activation Assay, Western Blot, Immunostaining, Injection, Purification, Two Tailed Test

Journal: Neurobiology of Stress

Article Title: Microglial SIRT6 confers protection against neuroinflammation-associated depression through NRF2-HO1 signaling

doi: 10.1016/j.ynstr.2026.100804

Figure Lengend Snippet: Sirt6 regulated the microglial activation and the depression behavior through Nrf2-HO1 signaling in vivo (A) Experimental timeline of tamoxifen administration, AAV9-Cx3cr1-Nrf2 injection, LPS challenge, and behavioral tests in Sirt6 MCKO mice. (B) Western blot analysis of NRF2 levels after AAV9-Cx3cr1-Nrf2 injection. n = 3 mice. (C-D) Immobility time in TST (C) and FST (D). n = 8 mice. (E) TAC, MDA, SOD, and GSH/GSSG levels at 5 days after LPS injection. n = 4 mice. (F-L) Immunostaining of GFAP and IBA1 in the hippocampus area at 5 days after LPS injection (F); Quantification of (G) IBA1 + and (H) GFAP + cell counts, along with microglial morphology parameters: (I) number of branches, (J) average branch length, (K) total branch length, and (L) soma area. n = 4 mice. (M) qPCR analysis of TNF-α,IL-6, and IL-1β mRNA levels in the microglial sorted from the hippocampus brain tissue. n = 4 mice. (N) Western blot analysis of NRF2 downstream signaling component protein levels in microglia sorted from the hippocampus. n = 4 mice. Data are mean ± SEM. Statistical significance between two groups was determined by an unpaired two-tailed Student's t-test.

Article Snippet: Membranes were blocked with 5% non-fat milk in TBST for 1 h and incubated overnight at 4 °C with primary antibodies: SIRT6 (12486, Cell Signaling Technology), NRF2 (ab62352, Abcam), HO-1 (ab68477, Abcam), NQO1 (ab80588, Abcam), KEAP1 (8047, Cell Signaling Technology), NLRP3 (AG-20B-0014, AdipoGen), cleaved caspase-1 (4199, Cell Signaling Technology), cleaved IL-1β (83186, Cell Signaling Technology), and β-actin (A1978, Sigma-Aldrich).

Techniques: Activation Assay, In Vivo, Injection, Western Blot, Immunostaining, Two Tailed Test

Journal: Neurobiology of Stress

Article Title: Microglial SIRT6 confers protection against neuroinflammation-associated depression through NRF2-HO1 signaling

doi: 10.1016/j.ynstr.2026.100804

Figure Lengend Snippet: Overexpression of Sirt6 in microglial improved the depression behavior and impeded the microglial activation (A) Experimental timeline depicting tamoxifen administration, followed by tail vein injection of AAV9-Cx3cr1-Sirt6, subsequent LPS challenge, and finally, a series of behavioral tests. (B) Western blot analysis of SIRT6 expression after AAV9-Cx3cr1-Sirt6 injection. n = 3 mice. (C-D) The immobility time was quantified in TST (C) and FST (D). n = 8 mice. (E) TAC, MDA, SOD, and GSH/GSSG levels at 5 days after LPS injection. (F-L) Immunostaining of GFAP and IBA1 in the hippocampus area at 5 days after LPS injection (F); Quantification of (G) IBA1 + and (H) GFAP + cell counts, along with microglial morphology parameters: (I) number of branches, (J) average branch length, (K) total branch length, and (L) soma area. n = 4 mice.(M) qPCR analysis of TNF-α,IL-6, and IL-1β mRNA levels in the microglial sorted from the hippocampus brain tissue. n = 4 mice. Data are mean ± SEM. Statistical significance between two groups was determined by an unpaired two-tailed Student's t-test.

Article Snippet: Membranes were blocked with 5% non-fat milk in TBST for 1 h and incubated overnight at 4 °C with primary antibodies: SIRT6 (12486, Cell Signaling Technology), NRF2 (ab62352, Abcam), HO-1 (ab68477, Abcam), NQO1 (ab80588, Abcam), KEAP1 (8047, Cell Signaling Technology), NLRP3 (AG-20B-0014, AdipoGen), cleaved caspase-1 (4199, Cell Signaling Technology), cleaved IL-1β (83186, Cell Signaling Technology), and β-actin (A1978, Sigma-Aldrich).

Techniques: Over Expression, Activation Assay, Injection, Western Blot, Expressing, Immunostaining, Two Tailed Test

Journal: Neurobiology of Stress

Article Title: Microglial SIRT6 confers protection against neuroinflammation-associated depression through NRF2-HO1 signaling

doi: 10.1016/j.ynstr.2026.100804

Figure Lengend Snippet: UBCS039-mediated SIRT6 activation exerts antidepressant and anti-inflammatory effects (A) Schematic of the experimental timeline for UBCS039 administration, LPS injection, and behavioral testing. (B-C) The immobility time was measured in TST (B) and FST (C). n = 4 mice. (D-G) TAC (D), MDA (E), SOD (F), and GSH/GSSG (G) levels at 5 days after LPS injection. n = 4 mice. (H-I) Immunostaining of GFAP and IBA1 in the hippocampus area at 5 days after LPS injection (H); Quantification of IBA1 + and GFAP + cell counts, along with microglial morphology parameters: number of branches, average branch length, total branch length, and soma area (I). n = 4 mice. (J) qPCR analysis of TNF-α,IL-6, and IL-1β mRNA levels in the microglial sorted from the hippocampus brain tissue. n = 4 mice. Data are mean ± SEM. Statistical significance between two groups was determined by an unpaired two-tailed Student's t-test.

Article Snippet: Membranes were blocked with 5% non-fat milk in TBST for 1 h and incubated overnight at 4 °C with primary antibodies: SIRT6 (12486, Cell Signaling Technology), NRF2 (ab62352, Abcam), HO-1 (ab68477, Abcam), NQO1 (ab80588, Abcam), KEAP1 (8047, Cell Signaling Technology), NLRP3 (AG-20B-0014, AdipoGen), cleaved caspase-1 (4199, Cell Signaling Technology), cleaved IL-1β (83186, Cell Signaling Technology), and β-actin (A1978, Sigma-Aldrich).

Techniques: Activation Assay, Injection, Immunostaining, Two Tailed Test