rabbit anti fkbp5  (Proteintech)

Journal: Scientific Reports

Article Title: FKBP51 disrupts the insulin signaling pathway and impairs mitochondrial bioenergetics in HepG2 cells

doi: 10.1038/s41598-026-40414-9

Figure Lengend Snippet: FKBP51 overexpression dampens insulin signaling in HepG2 cells but does not hinder its effects on glucose metabolism. HepG2 cells were transfected with either a plasmid coding Myc-DDK-tagged human FKBP51 (FKBP51) or an empty pCMV6 plasmid (Control) and were stimulated for 0.5 h with 100 nM insulin. ( A ) Representative blot ( n = 8), ( B ) FKBP51 protein levels, ( C ) Akt phosphorylation, ( D ) P70S6K phosphorylation, ( E ) FOXO1, ( F and G ) Glucose production assay ( n = 4), ( H ) mRNA levels (G6P, PCK1 y PDK4) ( n = 4), ( I ) Representative blot ( n = 3), GSK3β phosphorylation, ( J ) Representative imagens of glycogen synthesis assay and ( K ) Glycogen synthesis ( n = 4). One-way analysis of variance (ANOVA) followed by Tukey post hoc test. * p < 0.05 compared with control and square brackets with FKBP51 with insulin. Data are shown as means ± SEM.

Article Snippet: Proteins were resolved in SDS-polyacrylamide gels and subjected to immunoblotting overnight using antibodies specific for FKBP51 (1:1000, 12210 S, Cell Signaling, Massachusetts, USA), Akt (1:2,000, 2920 S, Cell Signaling), phospho-Ser 473 AKT (1:1000, 9271 S, Cell Signaling), phospho-p70S6K Thr389 (1:1000, 9206 S, Cell Signaling), p70S6K (1:2,000, 90205 S, Cell Signaling), phospho-FOXO1 Ser256 (1:1000, 9461 S, Cell Signaling), FOXO1 (1:1000, 14952 S, Cell Signaling), phospho-DRP1 Ser637 (1:1000, ab193216, Abcam, Cambridge, UK), DRP1 (1:1000, ab184247, Abcam), OPA1 (1:1000, ab157457, Abcam), Mfn1 (1:1000, 14739 S, Cell Signaling), Mfn2 (1:1000, 9482 S, Cell Signaling), mt-HSP70 (1:1000, MA3-D28, Invitrogen, Waltham, MA, USA), GAPDH (1:1000, 97166 S, Cell Signaling), OXPHOS (1:1000 ab110413, Abcam), phospo-GSK-3β Ser9 (1:1000, 5558, Cell Signaling) and GSK-3β (1:1,1000, 9832, Cell Signaling).

Techniques: Over Expression, Transfection, Plasmid Preparation, Control, Phospho-proteomics

Journal: Scientific Reports

Article Title: FKBP51 disrupts the insulin signaling pathway and impairs mitochondrial bioenergetics in HepG2 cells

doi: 10.1038/s41598-026-40414-9

Figure Lengend Snippet: FKBP51 is partially localized in mitochondria in HepG2 cells, but does not alter mitochondrial morphology. HepG2 cells were transfected with either a plasmid coding Myc-DDK-tagged human FKBP51 (FKBP51) or an empty pCMV6 plasmid (Control) and were stimulated for 3 h with 100 nM insulin and 200 nM CCCP for 2 h. ( A ) Representative image of immunofluorescence confocal microscopy, mtHSP70 (red) for mitochondria, FKBP51 (green). Segment lines represent the cellular contours. Scale bar: 24 μm, ( B ) Quantification of mitochondria (mtHSP70)-FKBP51 colocalization using Mander’s coefficient for cell imagens, ( C ) Quantification of FKBP51-mitochondria (mtHSP70) colocalization using Mander’s coefficient for cell imagens ( n = 6), ( D ) Representative blot mitochondria-cytosol subcellular fractionation ( n = 4), ( E ) Representative image of immunofluorescence confocal microscopy MitoTracker Green FM (200 nM for 0.2 h) in live-cells, ( F ) Quantification of average mitochondrial area, ( G ) Quantification of the of mitochondria number per cell in images cells and ( H ) Mean individual mitochondrial volume of HepG2 cells ( n = 5), ( I ) Representative image of Transmission Electron Microscopy, ( J ) Quantification of mitochondrial area, ( K ) Quantification of mitochondrial perimeter, ( L ) Quantification of mitochondrial aspect ratio. One-way analysis of variance (ANOVA) followed by Tukey post hoc test. * p < 0.05 compared with control. Data are shown as means ± SEM.

Article Snippet: Proteins were resolved in SDS-polyacrylamide gels and subjected to immunoblotting overnight using antibodies specific for FKBP51 (1:1000, 12210 S, Cell Signaling, Massachusetts, USA), Akt (1:2,000, 2920 S, Cell Signaling), phospho-Ser 473 AKT (1:1000, 9271 S, Cell Signaling), phospho-p70S6K Thr389 (1:1000, 9206 S, Cell Signaling), p70S6K (1:2,000, 90205 S, Cell Signaling), phospho-FOXO1 Ser256 (1:1000, 9461 S, Cell Signaling), FOXO1 (1:1000, 14952 S, Cell Signaling), phospho-DRP1 Ser637 (1:1000, ab193216, Abcam, Cambridge, UK), DRP1 (1:1000, ab184247, Abcam), OPA1 (1:1000, ab157457, Abcam), Mfn1 (1:1000, 14739 S, Cell Signaling), Mfn2 (1:1000, 9482 S, Cell Signaling), mt-HSP70 (1:1000, MA3-D28, Invitrogen, Waltham, MA, USA), GAPDH (1:1000, 97166 S, Cell Signaling), OXPHOS (1:1000 ab110413, Abcam), phospo-GSK-3β Ser9 (1:1000, 5558, Cell Signaling) and GSK-3β (1:1,1000, 9832, Cell Signaling).

Techniques: Transfection, Plasmid Preparation, Control, Immunofluorescence, Confocal Microscopy, Fractionation, Transmission Assay, Electron Microscopy

Journal: Scientific Reports

Article Title: FKBP51 disrupts the insulin signaling pathway and impairs mitochondrial bioenergetics in HepG2 cells

doi: 10.1038/s41598-026-40414-9

Figure Lengend Snippet: FKBP51 overexpression decreases Mitofusin 2 protein levels in HepG2 cells. Cells were transfected with either a plasmid coding Myc-DDK-tagged human FKBP51 (FKBP51) or an empty pCMV6 plasmid (Control) and were stimulated for 3 h with 100 nM insulin, mtHSP70 was used as loading control. ( A ) Representative blot of proteins of mitochondrial dynamics ( n = 10), ( B ) Mitofusin 1 protein levels (MFN1), ( C ) Mitofusin 2 protein levels (MFN2), ( E ) Ratio LOPA/SOPA1, ( D ) Dynamin-related protein 1 (Drp1) phosphorylation, ( F ) Mitochondrial fission protein 1 (Fis1 protein levels), ( G ) Dynamin-related protein 1 (Drp1), protein levels, ( H ) Representative blot of mitochondrial oxidative phosphorylation chain (OXPHOS), ( I ) NADH: ubiquinone oxidoreductase subunit B8 (NDUFB8, complex I) protein level, J Succinate dehydrogenase iron-sulfur subunit (SDHB, complex II), ( K ) Ubiquinol-cytochome c reductase core protein 2 (UQCRC2, complex III), ( L ) Cytochrome c oxidase subuni1 (MTCO1, complex IV) and ( M ) ATP synthase subunit alpha (ATP5A, complex V) ( n = 8). One-way analysis of variance (ANOVA) followed by Tukey post hoc test. * p < 0.05 compared with control. Data are shown as means ± SEM.

Article Snippet: Proteins were resolved in SDS-polyacrylamide gels and subjected to immunoblotting overnight using antibodies specific for FKBP51 (1:1000, 12210 S, Cell Signaling, Massachusetts, USA), Akt (1:2,000, 2920 S, Cell Signaling), phospho-Ser 473 AKT (1:1000, 9271 S, Cell Signaling), phospho-p70S6K Thr389 (1:1000, 9206 S, Cell Signaling), p70S6K (1:2,000, 90205 S, Cell Signaling), phospho-FOXO1 Ser256 (1:1000, 9461 S, Cell Signaling), FOXO1 (1:1000, 14952 S, Cell Signaling), phospho-DRP1 Ser637 (1:1000, ab193216, Abcam, Cambridge, UK), DRP1 (1:1000, ab184247, Abcam), OPA1 (1:1000, ab157457, Abcam), Mfn1 (1:1000, 14739 S, Cell Signaling), Mfn2 (1:1000, 9482 S, Cell Signaling), mt-HSP70 (1:1000, MA3-D28, Invitrogen, Waltham, MA, USA), GAPDH (1:1000, 97166 S, Cell Signaling), OXPHOS (1:1000 ab110413, Abcam), phospo-GSK-3β Ser9 (1:1000, 5558, Cell Signaling) and GSK-3β (1:1,1000, 9832, Cell Signaling).

Techniques: Over Expression, Transfection, Plasmid Preparation, Control, Phospho-proteomics

Journal: Scientific Reports

Article Title: FKBP51 disrupts the insulin signaling pathway and impairs mitochondrial bioenergetics in HepG2 cells

doi: 10.1038/s41598-026-40414-9

Figure Lengend Snippet: FKBP51 overexpression impairs mitochondrial bioenergetics. HepG2 cells were transfected with either a plasmid coding Myc-DDK-tagged human FKBP51 (FKBP51) or an empty pCMV6 plasmid (Control) and were stimulated for 3 h with 100 nM insulin. ( A ) Oxygen consumption rate (OCR) of cell, measured sequentially for 5 min, (CCCP 100 nM) ( n = 5), ( B ) Representative image of confocal microscopy of HepG2 cells loaded with TMRM then treated with CCCP 100 nM to dissipate the mitochondrial transmembrane potential. ΔB-C (ΔBasal-CCCP) was calculated as the mean fluorescence at 60 s before CCCP addition minus mean fluorescence during the last of the last 60 s of the recording, ( C ) Quantification of Δbasal-CCCP fluorescence ( n = 8), ( D ) Representative image of confocal microscopy MitoSOX 5 µM for 0.2 h with CCCP 200 nM for 1 h positive control, ( E ) Quantification of MitoSOX fluorescence ( n = 6), ( F ) Intracellular ATP levels of cells, ( G ) Graphical representation of the movement of calcium into the mitochondria by the Rhod-2 AM (4 µM for 0.2 h) probe in response to histamine 100 mM, ( H ) Area under the curve (AUC) of Ca 2+ movement to the mitochondria by the Rhod-2 AM probe in response to histamine and Slope the first 10 s in response to histamine by the Rhod-2 probe ( n = 4), ( I ) Graphical representation of the cytosolic Ca 2+ by the Fluo-4 AM (4.4 µM for 0.2 h) probe in response to histamine 100 mM, ( J ) Area under the curve (AUC) of Ca 2+ movement to the mitochondria by the Fluo-4 AM probe in response to histamine and Slope the first 10 s in response to histamine by the Fluo-4 AM probe ( n = 5). ( K ) proposed model. One-way analysis of variance (ANOVA) followed by Tukey post hoc test. * p < 0.05 compared with control and square brackets compared with FKBP51 with insulin. Data are shown as means + SEM.

Article Snippet: Proteins were resolved in SDS-polyacrylamide gels and subjected to immunoblotting overnight using antibodies specific for FKBP51 (1:1000, 12210 S, Cell Signaling, Massachusetts, USA), Akt (1:2,000, 2920 S, Cell Signaling), phospho-Ser 473 AKT (1:1000, 9271 S, Cell Signaling), phospho-p70S6K Thr389 (1:1000, 9206 S, Cell Signaling), p70S6K (1:2,000, 90205 S, Cell Signaling), phospho-FOXO1 Ser256 (1:1000, 9461 S, Cell Signaling), FOXO1 (1:1000, 14952 S, Cell Signaling), phospho-DRP1 Ser637 (1:1000, ab193216, Abcam, Cambridge, UK), DRP1 (1:1000, ab184247, Abcam), OPA1 (1:1000, ab157457, Abcam), Mfn1 (1:1000, 14739 S, Cell Signaling), Mfn2 (1:1000, 9482 S, Cell Signaling), mt-HSP70 (1:1000, MA3-D28, Invitrogen, Waltham, MA, USA), GAPDH (1:1000, 97166 S, Cell Signaling), OXPHOS (1:1000 ab110413, Abcam), phospo-GSK-3β Ser9 (1:1000, 5558, Cell Signaling) and GSK-3β (1:1,1000, 9832, Cell Signaling).

Techniques: Over Expression, Transfection, Plasmid Preparation, Control, Confocal Microscopy, Fluorescence, Positive Control

Journal: Journal of Neuroinflammation

Article Title: FKBP5 regulates interferon signaling leading to myeloid cell activation in multiple sclerosis

doi: 10.1186/s12974-026-03703-8

Figure Lengend Snippet: FKBP5 RNA expression is increased in myeloid cells from MS patients compared to healthy controls. A CSF myeloid cell clusters from published single-cell transcriptomic compendium . B Volcano plot of DEGs from CD14 + monocytes in MS compared to HCs. C Box plot of FKBP5 normalized counts in CD14 + monocytes from HCs ( n = 32) and MS ( n = 27). D Volcano plot of DEGs from CD36 + cDC2 in MS compared to HCs. E Box plot of FKBP5 normalized counts in CD36 + cDC2 from HCs ( n = 36) and MS ( n = 33). F IPA analysis of CD14 + monocytes significant genes. G IPA analysis of CD36 + cDC2 significant genes. H Box plot of FKBP5 expression from GEO234700 in microglia ( n = 5) and microglia nodules from MS NAWM ( n = 6). I Violin Plot of FKBP5 expression across different MS lesions from GSE180759 . J Violin Plot of FKBP5 from MIMS cells across different MS lesions. * P < 0.05 and ** P < 0.001 **** P < 0.00001 by False Discovery Rate (FDR) is calculated using the Benjamini–Hochberg method to control for multiple testing. Data are presented as mean ± SEM

Article Snippet: After blocking in 5% milk in TBST, primary antibodies FKBP5 (1:1000; Cell Signaling Technology (CST), 12210S, ULK (1:1000; CST, 8054), pULK (1:1000; CST, 5869), AKT (1:1000; CST, 9272), or LC3B (1:200; Santa Cruz Biotech, sc-376404) were incubated overnight (ON) in 5% milk in TBST at 4 °C.

Techniques: RNA Expression, Single Cell, Expressing, Control

Journal: Journal of Neuroinflammation

Article Title: FKBP5 regulates interferon signaling leading to myeloid cell activation in multiple sclerosis

doi: 10.1186/s12974-026-03703-8

Figure Lengend Snippet: Fkbp5 RNA and protein levels are increased during EAE in inflammatory phagocytes. A EAE clinical course of mice used for the FKBP5 immunoblot. Orange dots represent days in which SC and brain lysates were taken. B FKBP5 immunoblot and quantification of SC lysates from naïve and EAE mice. Microglia and infiltrating cells were isolated from naïve and EAE immunized mice after 18 and 28 days post immunization. C Histograms of Fkbp5-AF488 fluorescence in different cell types isolated from the CNS of EAE (orange) and naive (black) mice. D Immunofluorescent images showing EAE lower SC with lesion and infiltrating cells stained for Iba-1 (upper) or CD3 (lower) masked in red, DAPI in blue and FKBP5 signal is shown in green. E Volcano plot showing DEGs for inflammatory microglia/macrophage cells from SC of EAE mice with clinical score 4 versus 2. Genes labeled in orange have a log2FC > ± 2 and FDR < 0.01. Sequencing data analyzed by DESeq2, a list of DEGs is in Table S1. F GSEA of RNA-seq data, the columns represent normalized enrichment scores (NES) for upregulated gene sets in the EAE sicker mice. NES cutoff is −1.2 with FDR of less than 0.2. G Fkbp5 expression from EAE mice. H Fkbp5 expression from GSE99300 , reanalyzed by DESeq2 from the CNS of EAE immunized or naïve A.SW and SJL/J mice. I Fkbp5 expression (log scale) from GSE194071 from sorted microglia cells of Biozzi ABH immunized versus naïve mice. Scale bar = 100 μm. Experiments were repeated three times. FDR was calculated using the Benjamini–Hochberg method to control for multiple testing. Each dot represents one mouse. * P < 0.05, ** P < 0.01. Data presented as mean ± SEM. Scale bar = 100 μm

Article Snippet: After blocking in 5% milk in TBST, primary antibodies FKBP5 (1:1000; Cell Signaling Technology (CST), 12210S, ULK (1:1000; CST, 8054), pULK (1:1000; CST, 5869), AKT (1:1000; CST, 9272), or LC3B (1:200; Santa Cruz Biotech, sc-376404) were incubated overnight (ON) in 5% milk in TBST at 4 °C.

Techniques: Western Blot, Isolation, Fluorescence, Staining, Labeling, Sequencing, RNA Sequencing, Expressing, Control

Journal: Journal of Neuroinflammation

Article Title: FKBP5 regulates interferon signaling leading to myeloid cell activation in multiple sclerosis

doi: 10.1186/s12974-026-03703-8

Figure Lengend Snippet: Fkbp5-deficient microglia exhibited decreased proinflammatory phenotype and increased phagocytosis. A Volcano plot showing DEGs from sh96 compared to shCTR BV2 cell lines. Genes labeled in red are upregulated and have a log2FC > 1 and FDR < 0.05. Genes labeled in blue are downregulated with log2FC >—1 and FDR < 0.05. Sequencing data was analyzed by DESeq2, and a complete list of DEGs is included in Table S2. B IPA analysis of sh96 BV2 cell significant gene. C Heat map from the most downregulated gene from sh96 BV2 cell. D IPA analysis of disease related pathways from sh96 BV2 cell. E Representative myelin uptake graph from sh96 and shCTR cells after 5 h of incubation. F Quantification of CFSE uptake myelin in CD11b + CD45 + sh96 and shCTR BV2 cells. G Quantification of CFSE degraded myelin in CD11b + CD45 +. Degraded myelin was calculated by subtract CFSE + myelin frequency after the washes from 2 h CFSE + myelin uptake. H Quantification of the frequency of MHC-II + CD11b + CD45 + cells in sh96 and shCTR cells after LPS or cortisol stimulation. I Frequency of CFSE + CD4 + T cells co-cultured with shCTR or sh96 cells. Experiments were repeated three times. FDR was calculated using the Benjamini–Hochberg method to control for multiple testing. Each dot represents a single cell line replicate. * P < 0.05, ** P < 0.01 by unpaired t-tests. Data are presented as mean ± SEM

Article Snippet: After blocking in 5% milk in TBST, primary antibodies FKBP5 (1:1000; Cell Signaling Technology (CST), 12210S, ULK (1:1000; CST, 8054), pULK (1:1000; CST, 5869), AKT (1:1000; CST, 9272), or LC3B (1:200; Santa Cruz Biotech, sc-376404) were incubated overnight (ON) in 5% milk in TBST at 4 °C.

Techniques: Labeling, Sequencing, Incubation, Cell Culture, Control, Single Cell

Journal: Journal of Neuroinflammation

Article Title: FKBP5 regulates interferon signaling leading to myeloid cell activation in multiple sclerosis

doi: 10.1186/s12974-026-03703-8

Figure Lengend Snippet: Blocking Fkbp5 with SAFit2 in macrophages demonstrates resistance to IFN gamma response and improved phagocytosis. A Volcano plot showing DEGs from BMDM treated with SAFit2 compared to untreated BMDM. Genes labeled in red are upregulated and have a log2FC > 1 and FDR < 0.05. Genes labeled in blue are downregulated with log2FC >—1 and FDR < 0.05. Sequencing data was analyzed by DESeq2, and a complete list of DEGs is included in Table S3. B IPA analysis of SAFit2-treated BMDM significant genes. C Heat map from the most dysregulated genes from SAFit2-treated BMDM. D Quantification of CFSE uptake myelin in CD11b + CD45 + SAFit2 BMDM and control. E Quantification of CFSE degraded myelin in CD11b + CD45 +. Degraded myelin was calculated by subtract CFSE + myelin after the washes from CFSE + myelin uptake after 2 h of co-colture with BMDM. F Frequency of MHC-II + CD11b + CD45 + cells in untreated or SAFit2-treated cells after LPS or cortisol stimulation. G Frequency of CFSE + CD4 + T cells co-cultured with BMDM or SAFit2-treated BMDM, and representative graph of T cells proliferation. H-I Quantification of CFSE uptake and degraded myelin in CD11b + CD45 + SAFit2 BV2 and control. J Frequency of MHC-II + CD11b + CD45 + cells in untreated and SaFit2-BV2 cells after LPS or cortisol stimulation. K Frequency of CFSE + CD4 + T cells co-cultured with BV2 or SAFit2-treated BV2. L-M Quantification of MBP uptake and degradation in Iba-1 + SaFit2 treated primary microglia and control. Each dots represent fields/condition. N Representative image of MBP staining in primary microglia from myelin uptake or degradation. Experiments were repeated three times. FDR was calculated using the Benjamini–Hochberg method to control for multiple testing. Each dot represents a single BMDM or BV2 replicate. * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001 by unpaired t-tests. Data are presented as mean ± SEM. Scale bar = 50 μm

Article Snippet: After blocking in 5% milk in TBST, primary antibodies FKBP5 (1:1000; Cell Signaling Technology (CST), 12210S, ULK (1:1000; CST, 8054), pULK (1:1000; CST, 5869), AKT (1:1000; CST, 9272), or LC3B (1:200; Santa Cruz Biotech, sc-376404) were incubated overnight (ON) in 5% milk in TBST at 4 °C.

Techniques: Blocking Assay, Labeling, Sequencing, Control, Cell Culture, Staining

Journal: Journal of Neuroinflammation

Article Title: FKBP5 regulates interferon signaling leading to myeloid cell activation in multiple sclerosis

doi: 10.1186/s12974-026-03703-8

Figure Lengend Snippet: FKBP5 modulates Cxcl10 expression via IFNg response. A Venn diagram of significant downregulated genes from sh96 and SAFit2-treated BMDM. B GSEA analysis of Hallmark pathways from the 22 shared genes. C Predicted IFNg upstream regulators (IPA analysis) from shared significant genes in sh96 and SaFit2-treated BMDM. Blue represents downregulated genes and orange represents upregulated genes. D Cxcl10 expression analysis from BMDM treated with IFNg or with IFNg + SAFit2. E Cxcl10 expression analysis from shCTR and sh96 BV2 cells, treated with or without different concentration of IFNg. F Cxcl10 expression analysis from BV2 treated with IFNg or with IFNg + SAFit2. Experiments were repeated three times. Each dot represents a single BMDM or BV2 replicate. * P < 0.05, ** P < 0.01, and *** P < 0.001 by One-way ANOVA. Data are presented as mean ± SEM

Article Snippet: After blocking in 5% milk in TBST, primary antibodies FKBP5 (1:1000; Cell Signaling Technology (CST), 12210S, ULK (1:1000; CST, 8054), pULK (1:1000; CST, 5869), AKT (1:1000; CST, 9272), or LC3B (1:200; Santa Cruz Biotech, sc-376404) were incubated overnight (ON) in 5% milk in TBST at 4 °C.

Techniques: Expressing, Concentration Assay

Journal: Medical Gas Research

Article Title: Mechanisms by which sevoflurane affects cognitive function in aged marmosets and mice: up-regulation of FKBP5 expression in brain microglia

doi: 10.4103/mgr.MEDGASRES-D-24-00155

Figure Lengend Snippet: Identification of microglia in the hippocampus of marmosets after sevoflurane exposure. (A) Visualization of nine major classes of cells via UMAP from snRNA-seq. Each point represents a single cell (droplet barcode) colored by cluster. (B) UMAP visualization of the hippocampus in control and anesthetized marmosets. Green: control group; red: sevoflurane group. (C) Visualization of FKBP5 in the hippocampus of marmosets via UMAP. After anesthesia, FKBP5 was significantly increased in the microglial cluster. (D) Violin plot showing the expression of FKBP5 in nine major clusters of the hippocampus in control and anesthetized marmosets. Ctrl: Control; FKBP5: FK-506 binding protein; OL: oligodendrocytes; OPC: oligodendrocyte precursor cells; Sev: sevoflurane; snRNA-seq: single-cell nuclear RNA sequencing; UMAP: uniform manifold approximation and projection.

Article Snippet: The sections were incubated with rabbit anti-FKBP5 antibody (1:100) and mouse anti-ionized calcium binding adapter molecule 1 antibody (1:500, Servicebio, Wuhan, China, Cat# GB15105, RRID: AB_3663047) at 4°C overnight.

Techniques: Control, Expressing, Binding Assay, RNA Sequencing

Journal: Medical Gas Research

Article Title: Mechanisms by which sevoflurane affects cognitive function in aged marmosets and mice: up-regulation of FKBP5 expression in brain microglia

doi: 10.4103/mgr.MEDGASRES-D-24-00155

Figure Lengend Snippet: Identification of microglia in the hippocampus of mice after sevoflurane exposure. (A) Visualization of nine major classes of cells using UMAP from snRNA-seq. Each point represents a single cell (droplet barcode) colored by cluster. (B) UMAP visualization of the hippocampus in control and anesthetized mice. Green: Control group; red: sevoflurane group. (C) Visualization of FKBP5 in the hippocampus of mice via UMAP. After anesthesia, FKBP5 was significantly increased in the microglial cluster. (D) Violin plot showing the expression of FKBP5 in nine major clusters of the hippocampus between control and anesthetized mice. Ctrl: Control; FKBP5: FK-506 binding protein; Sev: sevoflurane; snRNA-seq: single-cell nuclear RNA sequencing; UMAP: uniform manifold approximation and projection.

Article Snippet: The sections were incubated with rabbit anti-FKBP5 antibody (1:100) and mouse anti-ionized calcium binding adapter molecule 1 antibody (1:500, Servicebio, Wuhan, China, Cat# GB15105, RRID: AB_3663047) at 4°C overnight.

Techniques: Control, Expressing, Binding Assay, RNA Sequencing

Journal: Medical Gas Research

Article Title: Mechanisms by which sevoflurane affects cognitive function in aged marmosets and mice: up-regulation of FKBP5 expression in brain microglia

doi: 10.4103/mgr.MEDGASRES-D-24-00155

Figure Lengend Snippet: The expression of FKBP5 in the hippocampus of mice after sevoflurane exposure. (A) Western blotting was used to validate the expression of FKBP5 in the brains of sevoflurane-treated mice. (B) Quantification of FKBP5 between the control group and sevoflurane group in A ( n = 5, P = 0.0102, t = 3.340). (C) Western blotting validated the expression of FKBP5 in the brains of sevoflurane/surgery-treated mice. (D) Quantification of FKBP5 between the control group and sevoflurane/surgery group in C ( n = 5, P = 0.0002, t = 6.470). The data are presented as mean ± SD. * P < 0.05, *** P < 0.001 (Student’s t -test). Ctrl: Control; Sev 6 h: sevoflurane anesthesia for 6 hours; Sev + Sur 6 h: sevoflurane anesthesia and surgery for 6 hours.

Article Snippet: The sections were incubated with rabbit anti-FKBP5 antibody (1:100) and mouse anti-ionized calcium binding adapter molecule 1 antibody (1:500, Servicebio, Wuhan, China, Cat# GB15105, RRID: AB_3663047) at 4°C overnight.

Techniques: Expressing, Western Blot, Control

Journal: Medical Gas Research

Article Title: Mechanisms by which sevoflurane affects cognitive function in aged marmosets and mice: up-regulation of FKBP5 expression in brain microglia

doi: 10.4103/mgr.MEDGASRES-D-24-00155

Figure Lengend Snippet: Sevoflurane/surgery induces cognitive decline through FKBP5 in microglia. (A) Immunofluorescence staining of the cortex in FKBP5 CKO mice and control mice. FKBP5 was knocked down in microglia of the FKBP5 CKO mouse brain after sevoflurane/surgery. Green: FKBP5, labeled with Alexa Fluor 488; red: IBA-1, a marker of microglia, labeled with CY3; blue: DAPI, labeled with cell nuclei; purple: merged. (B) The latency for male control mice and male FKBP5 CKO mice to identify the target hole in the training phase after exposure to sevoflurane/surgery ( n = 12, P = 0.0006, F (1, 88) = 12.53). (C) The percentage of total time that male control mice and male FKBP5 CKO mice spent in the target hole area during the test phase after exposure to sevoflurane/surgery ( P = 0.0002, t = 4.367). (D) The latency for female control mice and female FKBP5 CKO mice to identify the target hole in the training test after exposure to sevoflurane/surgery ( n = 14, P < 0.0001, F (1, 104) = 18.77). (E) The percentage of total time that female control mice and female FKBP5 CKO mice spent in the target hole area during the test phase after exposure to sevoflurane/surgery ( P = 0.0045, t = 3.110). Heterozygous mice were used as a control group. The data are presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.001 (Student’s t -test (C, E), two-way analysis of variance followed by Sidak’s multiple comparisons test (B, D)). Ctrl: Heterozygous mice; DAPI: 4′,6-diamidino-2′-phenylindole; FKBP5: FK-506 binding protein; FKBP5 CKO: microglia-specific FKBP5 conditional knockout; IBA-1: ionized calcium binding adapter molecule 1.

Article Snippet: The sections were incubated with rabbit anti-FKBP5 antibody (1:100) and mouse anti-ionized calcium binding adapter molecule 1 antibody (1:500, Servicebio, Wuhan, China, Cat# GB15105, RRID: AB_3663047) at 4°C overnight.

Techniques: Immunofluorescence, Staining, Control, Labeling, Marker, Binding Assay, Knock-Out

Journal: Medical Gas Research

Article Title: Mechanisms by which sevoflurane affects cognitive function in aged marmosets and mice: up-regulation of FKBP5 expression in brain microglia

doi: 10.4103/mgr.MEDGASRES-D-24-00155

Figure Lengend Snippet: Transcriptome sequencing analysis of the brains of FKBP5 CKO mice after sevoflurane/surgery. (A) Heatmapmap comparing FKBP5 CKO mice ( n = 10) and control mice ( n = 7) after sevoflurane/surgery. (B) Volcano plot comparing FKBP5 CKO mice ( n = 10) and control mice ( n = 7) after sevoflurane/surgery. The red dots represent RNAs significantly increased in FKBP5 CKO mice compared with control mice (fold change > 1.5, FDR < 0.05); the blue dots represent RNAs significantly decreased in FKBP5 CKO mice compared with control mice (fold change < 0.67, FDR < 0.05). (C–E) GO functional enrichment analysis in terms of biological process (BP) (C), molecular function (MF) (D), and cell composition (CC) (E). (F) KEGG pathway enrichment analysis. Heterozygous mice were used as a control group. Ctrl: Heterozygous mice; FDR: false discovery rate; FKBP5 CKO: microglia-specific FKBP5 conditional knockout.

Article Snippet: The sections were incubated with rabbit anti-FKBP5 antibody (1:100) and mouse anti-ionized calcium binding adapter molecule 1 antibody (1:500, Servicebio, Wuhan, China, Cat# GB15105, RRID: AB_3663047) at 4°C overnight.

Techniques: Sequencing, Control, Functional Assay, Knock-Out

Journal: Medical Gas Research

Article Title: Mechanisms by which sevoflurane affects cognitive function in aged marmosets and mice: up-regulation of FKBP5 expression in brain microglia

doi: 10.4103/mgr.MEDGASRES-D-24-00155

Figure Lengend Snippet: Identification of microglia in the hippocampus of marmosets after sevoflurane exposure. (A) Visualization of nine major classes of cells via UMAP from snRNA-seq. Each point represents a single cell (droplet barcode) colored by cluster. (B) UMAP visualization of the hippocampus in control and anesthetized marmosets. Green: control group; red: sevoflurane group. (C) Visualization of FKBP5 in the hippocampus of marmosets via UMAP. After anesthesia, FKBP5 was significantly increased in the microglial cluster. (D) Violin plot showing the expression of FKBP5 in nine major clusters of the hippocampus in control and anesthetized marmosets. Ctrl: Control; FKBP5: FK-506 binding protein; OL: oligodendrocytes; OPC: oligodendrocyte precursor cells; Sev: sevoflurane; snRNA-seq: single-cell nuclear RNA sequencing; UMAP: uniform manifold approximation and projection.

Article Snippet: The membranes were subsequently incubated with a rabbit anti-FKBP5 antibody (1:2000, Proteintech, Rosemont, IL, USA, Cat# 14155-1-AP, RRID: AB_2231625) and a mouse anti-glyceraldehyde-3-phosphate dehydrogenase antibody (1:5000, Abcam, Cambridge, UK, Cat#ab8245, RRID: AB_2107448) at 4°C overnight.

Techniques: Control, Expressing, Binding Assay, RNA Sequencing

Journal: Medical Gas Research

Article Title: Mechanisms by which sevoflurane affects cognitive function in aged marmosets and mice: up-regulation of FKBP5 expression in brain microglia

doi: 10.4103/mgr.MEDGASRES-D-24-00155

Figure Lengend Snippet: Identification of microglia in the hippocampus of mice after sevoflurane exposure. (A) Visualization of nine major classes of cells using UMAP from snRNA-seq. Each point represents a single cell (droplet barcode) colored by cluster. (B) UMAP visualization of the hippocampus in control and anesthetized mice. Green: Control group; red: sevoflurane group. (C) Visualization of FKBP5 in the hippocampus of mice via UMAP. After anesthesia, FKBP5 was significantly increased in the microglial cluster. (D) Violin plot showing the expression of FKBP5 in nine major clusters of the hippocampus between control and anesthetized mice. Ctrl: Control; FKBP5: FK-506 binding protein; Sev: sevoflurane; snRNA-seq: single-cell nuclear RNA sequencing; UMAP: uniform manifold approximation and projection.

Article Snippet: The membranes were subsequently incubated with a rabbit anti-FKBP5 antibody (1:2000, Proteintech, Rosemont, IL, USA, Cat# 14155-1-AP, RRID: AB_2231625) and a mouse anti-glyceraldehyde-3-phosphate dehydrogenase antibody (1:5000, Abcam, Cambridge, UK, Cat#ab8245, RRID: AB_2107448) at 4°C overnight.

Techniques: Control, Expressing, Binding Assay, RNA Sequencing

Journal: Medical Gas Research

Article Title: Mechanisms by which sevoflurane affects cognitive function in aged marmosets and mice: up-regulation of FKBP5 expression in brain microglia

doi: 10.4103/mgr.MEDGASRES-D-24-00155

Figure Lengend Snippet: The expression of FKBP5 in the hippocampus of mice after sevoflurane exposure. (A) Western blotting was used to validate the expression of FKBP5 in the brains of sevoflurane-treated mice. (B) Quantification of FKBP5 between the control group and sevoflurane group in A ( n = 5, P = 0.0102, t = 3.340). (C) Western blotting validated the expression of FKBP5 in the brains of sevoflurane/surgery-treated mice. (D) Quantification of FKBP5 between the control group and sevoflurane/surgery group in C ( n = 5, P = 0.0002, t = 6.470). The data are presented as mean ± SD. * P < 0.05, *** P < 0.001 (Student’s t -test). Ctrl: Control; Sev 6 h: sevoflurane anesthesia for 6 hours; Sev + Sur 6 h: sevoflurane anesthesia and surgery for 6 hours.

Article Snippet: The membranes were subsequently incubated with a rabbit anti-FKBP5 antibody (1:2000, Proteintech, Rosemont, IL, USA, Cat# 14155-1-AP, RRID: AB_2231625) and a mouse anti-glyceraldehyde-3-phosphate dehydrogenase antibody (1:5000, Abcam, Cambridge, UK, Cat#ab8245, RRID: AB_2107448) at 4°C overnight.

Techniques: Expressing, Western Blot, Control

Journal: Medical Gas Research

Article Title: Mechanisms by which sevoflurane affects cognitive function in aged marmosets and mice: up-regulation of FKBP5 expression in brain microglia

doi: 10.4103/mgr.MEDGASRES-D-24-00155

Figure Lengend Snippet: Sevoflurane/surgery induces cognitive decline through FKBP5 in microglia. (A) Immunofluorescence staining of the cortex in FKBP5 CKO mice and control mice. FKBP5 was knocked down in microglia of the FKBP5 CKO mouse brain after sevoflurane/surgery. Green: FKBP5, labeled with Alexa Fluor 488; red: IBA-1, a marker of microglia, labeled with CY3; blue: DAPI, labeled with cell nuclei; purple: merged. (B) The latency for male control mice and male FKBP5 CKO mice to identify the target hole in the training phase after exposure to sevoflurane/surgery ( n = 12, P = 0.0006, F (1, 88) = 12.53). (C) The percentage of total time that male control mice and male FKBP5 CKO mice spent in the target hole area during the test phase after exposure to sevoflurane/surgery ( P = 0.0002, t = 4.367). (D) The latency for female control mice and female FKBP5 CKO mice to identify the target hole in the training test after exposure to sevoflurane/surgery ( n = 14, P < 0.0001, F (1, 104) = 18.77). (E) The percentage of total time that female control mice and female FKBP5 CKO mice spent in the target hole area during the test phase after exposure to sevoflurane/surgery ( P = 0.0045, t = 3.110). Heterozygous mice were used as a control group. The data are presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.001 (Student’s t -test (C, E), two-way analysis of variance followed by Sidak’s multiple comparisons test (B, D)). Ctrl: Heterozygous mice; DAPI: 4′,6-diamidino-2′-phenylindole; FKBP5: FK-506 binding protein; FKBP5 CKO: microglia-specific FKBP5 conditional knockout; IBA-1: ionized calcium binding adapter molecule 1.

Article Snippet: The membranes were subsequently incubated with a rabbit anti-FKBP5 antibody (1:2000, Proteintech, Rosemont, IL, USA, Cat# 14155-1-AP, RRID: AB_2231625) and a mouse anti-glyceraldehyde-3-phosphate dehydrogenase antibody (1:5000, Abcam, Cambridge, UK, Cat#ab8245, RRID: AB_2107448) at 4°C overnight.

Techniques: Immunofluorescence, Staining, Control, Labeling, Marker, Binding Assay, Knock-Out

Journal: Medical Gas Research

Article Title: Mechanisms by which sevoflurane affects cognitive function in aged marmosets and mice: up-regulation of FKBP5 expression in brain microglia

doi: 10.4103/mgr.MEDGASRES-D-24-00155

Figure Lengend Snippet: Transcriptome sequencing analysis of the brains of FKBP5 CKO mice after sevoflurane/surgery. (A) Heatmapmap comparing FKBP5 CKO mice ( n = 10) and control mice ( n = 7) after sevoflurane/surgery. (B) Volcano plot comparing FKBP5 CKO mice ( n = 10) and control mice ( n = 7) after sevoflurane/surgery. The red dots represent RNAs significantly increased in FKBP5 CKO mice compared with control mice (fold change > 1.5, FDR < 0.05); the blue dots represent RNAs significantly decreased in FKBP5 CKO mice compared with control mice (fold change < 0.67, FDR < 0.05). (C–E) GO functional enrichment analysis in terms of biological process (BP) (C), molecular function (MF) (D), and cell composition (CC) (E). (F) KEGG pathway enrichment analysis. Heterozygous mice were used as a control group. Ctrl: Heterozygous mice; FDR: false discovery rate; FKBP5 CKO: microglia-specific FKBP5 conditional knockout.

Article Snippet: The membranes were subsequently incubated with a rabbit anti-FKBP5 antibody (1:2000, Proteintech, Rosemont, IL, USA, Cat# 14155-1-AP, RRID: AB_2231625) and a mouse anti-glyceraldehyde-3-phosphate dehydrogenase antibody (1:5000, Abcam, Cambridge, UK, Cat#ab8245, RRID: AB_2107448) at 4°C overnight.

Techniques: Sequencing, Control, Functional Assay, Knock-Out