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akt substrate 160  (Cell Signaling Technology Inc)


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    Structured Review

    Cell Signaling Technology Inc akt substrate 160
    Resistance exercise exceeds the benefits of endurance exercise in ameliorating metabolic dysfunction. Following 8 weeks of diet and exercise interventions, all mice were assessed for their metabolic function by GTT, ITT, and skeletal muscle response of Akt and <t>AS160</t> phosphorylation to injection of insulin measured by Western blot. (A–C) HOMA-IR taken after an overnight fast for baseline glucose and insulin. (D and E) GTT from 0–120 min and calculated AUC; colored * indicates significant difference from NC-SED. (F and G) ITT from 0–60 min and calculated AUC; colored * indicates significant difference from NC-SED. (H–N) pAkt stimulation, AS160 <t>S318,</t> and AS160 T642 in hindlimb muscles before and after insulin injection and the pre–post ∆ in phosphorylation. (O and P) Total and phosphorylated 4E-BP1. (Q–T) Western results for Raptor, COX4, LC3 II/I, and ubiquitin staining. Representative western blot images inset right. Data presented as mean ± standard error of the mean. Statistical analysis performed by analysis of variance between groups: * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001. NC-SED n : 8–16 (white); HFD-SED n : 8–18 (red); HFD-R EX n : 8–16 (blue); HFD-E EX n : 8–15 (green). 4E-BP1 = Eukaryotic translation initiation factor 4E binding protein; Akt = protein kinase B; AS160 = Akt substrate 160; COX4 = cytochrome c oxidase 4; CS = citrate sythase; E EX = endurance exercise; GAPDH = glyceraldehyde 3 phosphate dehydrogenase; GTT = glucose tolerance test; HFD = high fat diet; HOMA-IR = homeostatic model assessment for insulin resistance; iAUC = integrated area under the curve; ITT = insulin tolerance test; LC3 II/I = microtubule-associated protein light chain 3; NC = normal chow; pAkt = phospho-Akt; R EX = resistance exercise; SED = sedentary; Ub = ubiquitin.
    Akt Substrate 160, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 31 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/rabbit+antibody/Phospho-AS160+(Ser318)+Rabbit+mAb/pmc12816905-99-63-69
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    Images

    1) Product Images from "Weightlifting outperforms voluntary wheel running for improving adiposity and insulin sensitivity in obese mice"

    Article Title: Weightlifting outperforms voluntary wheel running for improving adiposity and insulin sensitivity in obese mice

    Journal: Journal of Sport and Health Science

    doi: 10.1016/j.jshs.2025.101100

    Resistance exercise exceeds the benefits of endurance exercise in ameliorating metabolic dysfunction. Following 8 weeks of diet and exercise interventions, all mice were assessed for their metabolic function by GTT, ITT, and skeletal muscle response of Akt and AS160 phosphorylation to injection of insulin measured by Western blot. (A–C) HOMA-IR taken after an overnight fast for baseline glucose and insulin. (D and E) GTT from 0–120 min and calculated AUC; colored * indicates significant difference from NC-SED. (F and G) ITT from 0–60 min and calculated AUC; colored * indicates significant difference from NC-SED. (H–N) pAkt stimulation, AS160 S318, and AS160 T642 in hindlimb muscles before and after insulin injection and the pre–post ∆ in phosphorylation. (O and P) Total and phosphorylated 4E-BP1. (Q–T) Western results for Raptor, COX4, LC3 II/I, and ubiquitin staining. Representative western blot images inset right. Data presented as mean ± standard error of the mean. Statistical analysis performed by analysis of variance between groups: * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001. NC-SED n : 8–16 (white); HFD-SED n : 8–18 (red); HFD-R EX n : 8–16 (blue); HFD-E EX n : 8–15 (green). 4E-BP1 = Eukaryotic translation initiation factor 4E binding protein; Akt = protein kinase B; AS160 = Akt substrate 160; COX4 = cytochrome c oxidase 4; CS = citrate sythase; E EX = endurance exercise; GAPDH = glyceraldehyde 3 phosphate dehydrogenase; GTT = glucose tolerance test; HFD = high fat diet; HOMA-IR = homeostatic model assessment for insulin resistance; iAUC = integrated area under the curve; ITT = insulin tolerance test; LC3 II/I = microtubule-associated protein light chain 3; NC = normal chow; pAkt = phospho-Akt; R EX = resistance exercise; SED = sedentary; Ub = ubiquitin.
    Figure Legend Snippet: Resistance exercise exceeds the benefits of endurance exercise in ameliorating metabolic dysfunction. Following 8 weeks of diet and exercise interventions, all mice were assessed for their metabolic function by GTT, ITT, and skeletal muscle response of Akt and AS160 phosphorylation to injection of insulin measured by Western blot. (A–C) HOMA-IR taken after an overnight fast for baseline glucose and insulin. (D and E) GTT from 0–120 min and calculated AUC; colored * indicates significant difference from NC-SED. (F and G) ITT from 0–60 min and calculated AUC; colored * indicates significant difference from NC-SED. (H–N) pAkt stimulation, AS160 S318, and AS160 T642 in hindlimb muscles before and after insulin injection and the pre–post ∆ in phosphorylation. (O and P) Total and phosphorylated 4E-BP1. (Q–T) Western results for Raptor, COX4, LC3 II/I, and ubiquitin staining. Representative western blot images inset right. Data presented as mean ± standard error of the mean. Statistical analysis performed by analysis of variance between groups: * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001. NC-SED n : 8–16 (white); HFD-SED n : 8–18 (red); HFD-R EX n : 8–16 (blue); HFD-E EX n : 8–15 (green). 4E-BP1 = Eukaryotic translation initiation factor 4E binding protein; Akt = protein kinase B; AS160 = Akt substrate 160; COX4 = cytochrome c oxidase 4; CS = citrate sythase; E EX = endurance exercise; GAPDH = glyceraldehyde 3 phosphate dehydrogenase; GTT = glucose tolerance test; HFD = high fat diet; HOMA-IR = homeostatic model assessment for insulin resistance; iAUC = integrated area under the curve; ITT = insulin tolerance test; LC3 II/I = microtubule-associated protein light chain 3; NC = normal chow; pAkt = phospho-Akt; R EX = resistance exercise; SED = sedentary; Ub = ubiquitin.

    Techniques Used: Phospho-proteomics, Injection, Western Blot, Muscles, Ubiquitin Proteomics, Staining, Binding Assay

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    Control:

    Article Title: GSTK1 suppresses HCC aggravation via L-carnitine metabolism by PGAM5/DRP1 complex-mediated mitochondrial quality control.
    Article Snippet: After 794 denaturation with 5x loading buffer (CWBIO, CW0028) under high 795 temperature, the protein samples were separated by SDS-PAGE and 796 transferred to PVDF membrane, followed by immunoblotting with 797 antibodies listed below. .. Rabbit antibody to Phospho-DRP1 (Ser616) (3455 798 [N/A] and 4494 [RRID: AB_11178659]), Mitofusin-1 (14739 [RRID: 799 AB_2744531]), Mitofusion-2 (11925 [RRID:AB_2750893]), SQSTM1/P62 800 (39749 [AB_2799160]), LC3A/B (12741 [RRID:AB_2617131]), Bcl-xL (2764 801 [RRID: AB_2228008]), IgG Isotype Control (3900 [RRID:AB_1550038]), 802 DYKDDDDK Tag (14793 [RRID: AB_2572291]) and mouse antibody to Bcl-2 803 (15071 [RRID: AB_2744528]) were obtained from Cell Signaling Technology. .. AR TIC LE IN PR ES S 40 804 Rabbit antibody to GSTK1 (ab134173 [RRID: AB_1580730]), MLKL 805 (ab184718 [RRID: AB_2755030]), DRP1 (ab184247 [RRID: AB_2895215]) 806 and mouse antibody to CPT1A (ab128568 [RRID: AB_11141632]) were 807 obtained from Abcam.

    Staining:

    Article Title: Composition for prevention or treatment of intractable epilepsy comprising mTOR inhibitor
    Article Snippet: .. Surgical tissue blocks were fixed overnight in freshly prepared phosphate-buffered (PB) 4% paraformaldehyde, ciyoprotected overnight in 20% buffered sucrose, and prepared into gelatin-embedded tissues blocks (7.5% gelatin in 10% sucrose/PB) before storage at −80° C. Cryostat-cut sections (10 um thick) were collected, placed on glass slides, and blocked in PBS-GT (0.2% gelatin and 0.2% Triton X-100 in PBS) at room temperature for 1 hr before staining with the following antibodies: rabbit antibody to phosphorylated S6 ribosomal protein (Ser240/Ser244) (1:100 dilution; 5364, Cell signaling Technology), and mouse antibody to NeuN (1:100 dilution; MAB377, Millipore). .. Subsequently, samples were washed in PBS and stained with the following secondary antibodies: Alexa Fluor 555-conjugated goat antibody to mouse (1:200 dilution; A21422, Invitrogen) and Alexa Fluor 488-conjugated goat antibody to rabbit (1:200 dilution; A11008, Invitrogen).

    Article Title: Composition for prevention or treatment of intractable epilepsy comprising mTOR inhibitor
    Article Snippet: .. The slides were then blocked in PBS-GT (0.2% gelatin and 0.2% Triton X-100 in PBS) at room temperature for 1 hr before staining with the following antibodies: rabbit antibody to phosphorylated S6 ribosomal protein (Ser240/Ser244) (1:100 dilution; 5364, Cell signaling Technology) and mouse antibody to NeuN (1:100 dilution; MAB377, Millipore). ..

    Membrane:

    Article Title: Maternal causation of early-onset pre-eclampsia: excessive endometrial gland-derived apolipoprotein D induces placental ferroptosis and developmental abnormalities.
    Article Snippet: .. The membrane was then blocked with 5% non-fat milk and incubated with antiPI3K/ anti-Phospho-AKT antibody/anti-AKT antibody (1:1000 dilution; Cell Signaling Technology #4249/Cell Signaling Technology #4060/Cell Signaling Technology #4691) (Table S7) and the reference protein anti-β-actin antibody followed by horseradish peroxidase-conjugated against rabbit antibody (1:1000 dilution; 7074 s, Cell Signaling Technology, USA). .. Blotting signals were revealed with ECL Plus (Beyotime, Shanghai, China).

    Article Title: Maternal causation of early-onset pre-eclampsia: excessive endometrial gland-derived apolipoprotein D induces placental ferroptosis and developmental abnormalities
    Article Snippet: .. The membrane was then blocked with 5% non-fat milk and incubated with anti-PI3K/ anti-Phospho-AKT antibody/anti-AKT antibody (1:1000 dilution; Cell Signaling Technology #4249/Cell Signaling Technology #4060/Cell Signaling Technology #4691) (Table S7) and the reference protein anti-β-actin antibody followed by horseradish peroxidase-conjugated against rabbit antibody (1:1000 dilution; 7074 s, Cell Signaling Technology, USA). .. Blotting signals were revealed with ECL Plus (Beyotime, Shanghai, China).

    Incubation:

    Article Title: Maternal causation of early-onset pre-eclampsia: excessive endometrial gland-derived apolipoprotein D induces placental ferroptosis and developmental abnormalities.
    Article Snippet: .. The membrane was then blocked with 5% non-fat milk and incubated with antiPI3K/ anti-Phospho-AKT antibody/anti-AKT antibody (1:1000 dilution; Cell Signaling Technology #4249/Cell Signaling Technology #4060/Cell Signaling Technology #4691) (Table S7) and the reference protein anti-β-actin antibody followed by horseradish peroxidase-conjugated against rabbit antibody (1:1000 dilution; 7074 s, Cell Signaling Technology, USA). .. Blotting signals were revealed with ECL Plus (Beyotime, Shanghai, China).

    Article Title: Maternal causation of early-onset pre-eclampsia: excessive endometrial gland-derived apolipoprotein D induces placental ferroptosis and developmental abnormalities
    Article Snippet: .. The membrane was then blocked with 5% non-fat milk and incubated with anti-PI3K/ anti-Phospho-AKT antibody/anti-AKT antibody (1:1000 dilution; Cell Signaling Technology #4249/Cell Signaling Technology #4060/Cell Signaling Technology #4691) (Table S7) and the reference protein anti-β-actin antibody followed by horseradish peroxidase-conjugated against rabbit antibody (1:1000 dilution; 7074 s, Cell Signaling Technology, USA). .. Blotting signals were revealed with ECL Plus (Beyotime, Shanghai, China).

    Article Title: The histologic and molecular alterations during the cold storage of the ascending human colon.
    Article Snippet: .. In brief, following deparaffinization, rehydration, and antigen retrieval using citrate buffer (10 mM, pH 6.0) sections were blocked and then incubated with primary rabbit antibody against cleaved Caspase-3 (1:100, #D175; Cell Signaling Technology, Danvers, MA) for 1 h at room temperature followed by incubation with an anti-rabbit probe, a rabbit alkaline phosphatase polymer and warp red chromogen. ..

    Article Title: Alpha-1 antitrypsin-glucocorticoid receptor axis: a new pathway in immune modulation.
    Article Snippet: .. Protein detection was performed by incubation with a primary anti-AAT antibody (Dako, Agilent, Santa Clara, CA, USA) or an antitetramethylrhodamine (TRITC) antibody (Invitrogen, Rockford, IL, USA) for 1.5 h, followed by incubation with a HRP-conjugated secondary rabbit antibody (Cell Signaling, Danvers, MA, USA) for 1 h. Signal development was achieved by addition of the HRP substrate 3,3′,5,5′-tetramethylbenzidine (TMB; Merck). .. The reaction was stopped with 0.16 M H2SO4, AR TIC LE IN PR ES S and the absorbance was measured at 450 nm using a Flexstation 3 multimode microplate reader (Molecular Devices, San Jose, CA, USA).

    Polymer:

    Article Title: The histologic and molecular alterations during the cold storage of the ascending human colon.
    Article Snippet: .. In brief, following deparaffinization, rehydration, and antigen retrieval using citrate buffer (10 mM, pH 6.0) sections were blocked and then incubated with primary rabbit antibody against cleaved Caspase-3 (1:100, #D175; Cell Signaling Technology, Danvers, MA) for 1 h at room temperature followed by incubation with an anti-rabbit probe, a rabbit alkaline phosphatase polymer and warp red chromogen. ..



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    Image Search Results


    Resistance exercise exceeds the benefits of endurance exercise in ameliorating metabolic dysfunction. Following 8 weeks of diet and exercise interventions, all mice were assessed for their metabolic function by GTT, ITT, and skeletal muscle response of Akt and AS160 phosphorylation to injection of insulin measured by Western blot. (A–C) HOMA-IR taken after an overnight fast for baseline glucose and insulin. (D and E) GTT from 0–120 min and calculated AUC; colored * indicates significant difference from NC-SED. (F and G) ITT from 0–60 min and calculated AUC; colored * indicates significant difference from NC-SED. (H–N) pAkt stimulation, AS160 S318, and AS160 T642 in hindlimb muscles before and after insulin injection and the pre–post ∆ in phosphorylation. (O and P) Total and phosphorylated 4E-BP1. (Q–T) Western results for Raptor, COX4, LC3 II/I, and ubiquitin staining. Representative western blot images inset right. Data presented as mean ± standard error of the mean. Statistical analysis performed by analysis of variance between groups: * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001. NC-SED n : 8–16 (white); HFD-SED n : 8–18 (red); HFD-R EX n : 8–16 (blue); HFD-E EX n : 8–15 (green). 4E-BP1 = Eukaryotic translation initiation factor 4E binding protein; Akt = protein kinase B; AS160 = Akt substrate 160; COX4 = cytochrome c oxidase 4; CS = citrate sythase; E EX = endurance exercise; GAPDH = glyceraldehyde 3 phosphate dehydrogenase; GTT = glucose tolerance test; HFD = high fat diet; HOMA-IR = homeostatic model assessment for insulin resistance; iAUC = integrated area under the curve; ITT = insulin tolerance test; LC3 II/I = microtubule-associated protein light chain 3; NC = normal chow; pAkt = phospho-Akt; R EX = resistance exercise; SED = sedentary; Ub = ubiquitin.

    Journal: Journal of Sport and Health Science

    Article Title: Weightlifting outperforms voluntary wheel running for improving adiposity and insulin sensitivity in obese mice

    doi: 10.1016/j.jshs.2025.101100

    Figure Lengend Snippet: Resistance exercise exceeds the benefits of endurance exercise in ameliorating metabolic dysfunction. Following 8 weeks of diet and exercise interventions, all mice were assessed for their metabolic function by GTT, ITT, and skeletal muscle response of Akt and AS160 phosphorylation to injection of insulin measured by Western blot. (A–C) HOMA-IR taken after an overnight fast for baseline glucose and insulin. (D and E) GTT from 0–120 min and calculated AUC; colored * indicates significant difference from NC-SED. (F and G) ITT from 0–60 min and calculated AUC; colored * indicates significant difference from NC-SED. (H–N) pAkt stimulation, AS160 S318, and AS160 T642 in hindlimb muscles before and after insulin injection and the pre–post ∆ in phosphorylation. (O and P) Total and phosphorylated 4E-BP1. (Q–T) Western results for Raptor, COX4, LC3 II/I, and ubiquitin staining. Representative western blot images inset right. Data presented as mean ± standard error of the mean. Statistical analysis performed by analysis of variance between groups: * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001. NC-SED n : 8–16 (white); HFD-SED n : 8–18 (red); HFD-R EX n : 8–16 (blue); HFD-E EX n : 8–15 (green). 4E-BP1 = Eukaryotic translation initiation factor 4E binding protein; Akt = protein kinase B; AS160 = Akt substrate 160; COX4 = cytochrome c oxidase 4; CS = citrate sythase; E EX = endurance exercise; GAPDH = glyceraldehyde 3 phosphate dehydrogenase; GTT = glucose tolerance test; HFD = high fat diet; HOMA-IR = homeostatic model assessment for insulin resistance; iAUC = integrated area under the curve; ITT = insulin tolerance test; LC3 II/I = microtubule-associated protein light chain 3; NC = normal chow; pAkt = phospho-Akt; R EX = resistance exercise; SED = sedentary; Ub = ubiquitin.

    Article Snippet: Primary antibodies used for analysis were from Cell Signaling Technologies (Danvers, MA, USA) and diluted 1:1000 unless otherwise stated as follows: protein kinase B (Akt; 1:500; #4691; Cell Signaling Technologies), phospho-Akt (pAkt) S473 (1:500; #9271; Cell Signaling Technologies), Ubiquitin (#3933; Cell Signaling Technologies), microtubule-associated protein light chain 3 (LC3 II/I; #4018; Cell Signaling Technologies), cytochrome c oxidase subunit 4(COX4; #11967; Cell Signaling Technologies), Akt substrate 160 (AS160 S318; #8619; Cell Signaling Technologies), AS160 T642 (#8881; Cell Signaling Technologies), eukaryotic translation initiation factor 4E binding protein (4E-BP1; #9452; Cell Signaling Technologies), and glyceraldehyde 3 phosphate dehydrogenase (GAPDH; #2118; Cell Signaling Technologies).

    Techniques: Phospho-proteomics, Injection, Western Blot, Muscles, Ubiquitin Proteomics, Staining, Binding Assay

    Resistance exercise exceeds the benefits of endurance exercise in ameliorating metabolic dysfunction. Following 8 weeks of diet and exercise interventions, all mice were assessed for their metabolic function by GTT, ITT, and skeletal muscle response of Akt and AS160 phosphorylation to injection of insulin measured by Western blot. (A–C) HOMA-IR taken after an overnight fast for baseline glucose and insulin. (D and E) GTT from 0–120 min and calculated AUC; colored * indicates significant difference from NC-SED. (F and G) ITT from 0–60 min and calculated AUC; colored * indicates significant difference from NC-SED. (H–N) pAkt stimulation, AS160 S318, and AS160 T642 in hindlimb muscles before and after insulin injection and the pre–post ∆ in phosphorylation. (O and P) Total and phosphorylated 4E-BP1. (Q–T) Western results for Raptor, COX4, LC3 II/I, and ubiquitin staining. Representative western blot images inset right. Data presented as mean ± standard error of the mean. Statistical analysis performed by analysis of variance between groups: * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001. NC-SED n : 8–16 (white); HFD-SED n : 8–18 (red); HFD-R EX n : 8–16 (blue); HFD-E EX n : 8–15 (green). 4E-BP1 = Eukaryotic translation initiation factor 4E binding protein; Akt = protein kinase B; AS160 = Akt substrate 160; COX4 = cytochrome c oxidase 4; CS = citrate sythase; E EX = endurance exercise; GAPDH = glyceraldehyde 3 phosphate dehydrogenase; GTT = glucose tolerance test; HFD = high fat diet; HOMA-IR = homeostatic model assessment for insulin resistance; iAUC = integrated area under the curve; ITT = insulin tolerance test; LC3 II/I = microtubule-associated protein light chain 3; NC = normal chow; pAkt = phospho-Akt; R EX = resistance exercise; SED = sedentary; Ub = ubiquitin.

    Journal: Journal of Sport and Health Science

    Article Title: Weightlifting outperforms voluntary wheel running for improving adiposity and insulin sensitivity in obese mice

    doi: 10.1016/j.jshs.2025.101100

    Figure Lengend Snippet: Resistance exercise exceeds the benefits of endurance exercise in ameliorating metabolic dysfunction. Following 8 weeks of diet and exercise interventions, all mice were assessed for their metabolic function by GTT, ITT, and skeletal muscle response of Akt and AS160 phosphorylation to injection of insulin measured by Western blot. (A–C) HOMA-IR taken after an overnight fast for baseline glucose and insulin. (D and E) GTT from 0–120 min and calculated AUC; colored * indicates significant difference from NC-SED. (F and G) ITT from 0–60 min and calculated AUC; colored * indicates significant difference from NC-SED. (H–N) pAkt stimulation, AS160 S318, and AS160 T642 in hindlimb muscles before and after insulin injection and the pre–post ∆ in phosphorylation. (O and P) Total and phosphorylated 4E-BP1. (Q–T) Western results for Raptor, COX4, LC3 II/I, and ubiquitin staining. Representative western blot images inset right. Data presented as mean ± standard error of the mean. Statistical analysis performed by analysis of variance between groups: * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001. NC-SED n : 8–16 (white); HFD-SED n : 8–18 (red); HFD-R EX n : 8–16 (blue); HFD-E EX n : 8–15 (green). 4E-BP1 = Eukaryotic translation initiation factor 4E binding protein; Akt = protein kinase B; AS160 = Akt substrate 160; COX4 = cytochrome c oxidase 4; CS = citrate sythase; E EX = endurance exercise; GAPDH = glyceraldehyde 3 phosphate dehydrogenase; GTT = glucose tolerance test; HFD = high fat diet; HOMA-IR = homeostatic model assessment for insulin resistance; iAUC = integrated area under the curve; ITT = insulin tolerance test; LC3 II/I = microtubule-associated protein light chain 3; NC = normal chow; pAkt = phospho-Akt; R EX = resistance exercise; SED = sedentary; Ub = ubiquitin.

    Article Snippet: Primary antibodies used for analysis were from Cell Signaling Technologies (Danvers, MA, USA) and diluted 1:1000 unless otherwise stated as follows: protein kinase B (Akt; 1:500; #4691; Cell Signaling Technologies), phospho-Akt (pAkt) S473 (1:500; #9271; Cell Signaling Technologies), Ubiquitin (#3933; Cell Signaling Technologies), microtubule-associated protein light chain 3 (LC3 II/I; #4018; Cell Signaling Technologies), cytochrome c oxidase subunit 4(COX4; #11967; Cell Signaling Technologies), Akt substrate 160 (AS160 S318; #8619; Cell Signaling Technologies), AS160 T642 (#8881; Cell Signaling Technologies), eukaryotic translation initiation factor 4E binding protein (4E-BP1; #9452; Cell Signaling Technologies), and glyceraldehyde 3 phosphate dehydrogenase (GAPDH; #2118; Cell Signaling Technologies).

    Techniques: Phospho-proteomics, Injection, Western Blot, Muscles, Ubiquitin Proteomics, Staining, Binding Assay

    RBC-MBs promotes the polarization of macrophages towards the M2 phenotype by inhibiting the TLR9-MyD88 and cGAS-STING pathways (A-F) RBC-MB treatment significantly attenuated CpG DNA-induced upregulation of pro-inflammatory cytokines (A-B), key proteins in the TLR9-MyD88 (C-D) and cGAS-STING (E-F) pathways. RAW264.7 cells were treated with 1 μg mL -1 CpG DNA for 24 h, followed by incubation with 0.5 mL RBC-MBs (concentration: 1 × 10 8 particles mL -1 ) for 24 h, and then subjected to western blot analysis (A, C, E). IL-1α, TNF-α, IL-6, TLR9, MyD88, STING, and cGAS levels were normalized to the β-actin level in each sample. p-NF-κB, p-TBK1, and p-IRF3 levels were normalized to their respective total protein levels (NF-κB, TBK1, and IRF3) in each sample. These normalized values were then used for statistical analysis (B, D, F), n = 6. (G) RBC-MB treatment significantly attenuated CpG DNA-induced upregulation of CD86 in macrophages. RAW264.7 cells were treated with 1 μg mL -1 CpG DNA for 24 h, followed by incubation with 0.5 mL RBC-MBs (concentration: 1 × 10 8 particles mL -1 ) for 24 h, then immunostained with an anti-CD86 antibody (green) and DAPI (blue, nuclei), and imaged by fluorescence microscopy. Representative images (left panel) and quantitative fluorescence graph (right panel) are shown. Scale bar: 50 μm, n = 6. (H-J) RBC-MB treatment significantly inhibited hepatic IRI-induced polarization of macrophages toward the M1 phenotype (H) and activation of the TLR9-MyD88 and cGAS-STING pathways (I-J). MASLD rats underwent 45 min of hepatic ischemia followed by reperfusion. At 0, 6, 12, 18, and 24 h post-reperfusion, the rats were treated with either Con-MBs or RBC-MBs. Primary macrophages were isolated from rat livers, then co-stained with CD68 and CD163 followed by flow cytometric analysis (H), n = 3/4, or subjected to western blotting (I-J). CD206, iNOS, TLR9, MyD88, STING, and cGAS levels were normalized to the β-actin level in each sample, and these normalized values were used for statistical analysis (J), n = 3. For (B, D, F, G right panel , H right panel , J), error bars represent mean ± standard deviation. P values were calculated using the unpaired Student's t-test (B, D, F, G right panel ) or one-way analysis of variance (ANOVA) (H right panel , J). *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; ns, not significant.(For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

    Journal: Materials Today Bio

    Article Title: Toll-like receptor 9-overexpressing red blood cell biomimetic microbubbles adsorb cell-free DNA to relieve steatotic liver ischemia-reperfusion injury

    doi: 10.1016/j.mtbio.2026.103558

    Figure Lengend Snippet: RBC-MBs promotes the polarization of macrophages towards the M2 phenotype by inhibiting the TLR9-MyD88 and cGAS-STING pathways (A-F) RBC-MB treatment significantly attenuated CpG DNA-induced upregulation of pro-inflammatory cytokines (A-B), key proteins in the TLR9-MyD88 (C-D) and cGAS-STING (E-F) pathways. RAW264.7 cells were treated with 1 μg mL -1 CpG DNA for 24 h, followed by incubation with 0.5 mL RBC-MBs (concentration: 1 × 10 8 particles mL -1 ) for 24 h, and then subjected to western blot analysis (A, C, E). IL-1α, TNF-α, IL-6, TLR9, MyD88, STING, and cGAS levels were normalized to the β-actin level in each sample. p-NF-κB, p-TBK1, and p-IRF3 levels were normalized to their respective total protein levels (NF-κB, TBK1, and IRF3) in each sample. These normalized values were then used for statistical analysis (B, D, F), n = 6. (G) RBC-MB treatment significantly attenuated CpG DNA-induced upregulation of CD86 in macrophages. RAW264.7 cells were treated with 1 μg mL -1 CpG DNA for 24 h, followed by incubation with 0.5 mL RBC-MBs (concentration: 1 × 10 8 particles mL -1 ) for 24 h, then immunostained with an anti-CD86 antibody (green) and DAPI (blue, nuclei), and imaged by fluorescence microscopy. Representative images (left panel) and quantitative fluorescence graph (right panel) are shown. Scale bar: 50 μm, n = 6. (H-J) RBC-MB treatment significantly inhibited hepatic IRI-induced polarization of macrophages toward the M1 phenotype (H) and activation of the TLR9-MyD88 and cGAS-STING pathways (I-J). MASLD rats underwent 45 min of hepatic ischemia followed by reperfusion. At 0, 6, 12, 18, and 24 h post-reperfusion, the rats were treated with either Con-MBs or RBC-MBs. Primary macrophages were isolated from rat livers, then co-stained with CD68 and CD163 followed by flow cytometric analysis (H), n = 3/4, or subjected to western blotting (I-J). CD206, iNOS, TLR9, MyD88, STING, and cGAS levels were normalized to the β-actin level in each sample, and these normalized values were used for statistical analysis (J), n = 3. For (B, D, F, G right panel , H right panel , J), error bars represent mean ± standard deviation. P values were calculated using the unpaired Student's t-test (B, D, F, G right panel ) or one-way analysis of variance (ANOVA) (H right panel , J). *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; ns, not significant.(For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

    Article Snippet: Primary antibodies used included: rabbit polyclonal antibodies against TLR9 (A14642, Abclonal), NF-κB (A2547, Abclonal), MyD88 (A0980, Abclonal), cGAS (A8335, Abclonal), IL-1α (A2170, Abclonal), TNF-α (A0277, Abclonal), IL-6 (A0286, Abclonal), CD206 (A8301, Abclonal), CD86 (A16805, Abclonal), CD47 (A1838, Abclonal), and ATP1A (A0643, Abclonal); rabbit monoclonal antibodies against STING (13647, CST, Beverly, MA, USA), p-NF-κB (3033, CST), p-TBK1 (5483, CST), TBK1 (ab40676, Abcam), p-IκB (2859, CST), IκB (4812, CST), p-IRF3 (4947, CST), and β-actin (AC026, Abclonal).

    Techniques: Incubation, Concentration Assay, Western Blot, Fluorescence, Microscopy, Activation Assay, Isolation, Staining, Standard Deviation