amp activated protein kinase ampk protein  (Proteintech)

Journal: iScience

Article Title: Roseburia hominis enriched by baicalin reverses the non-response to metformin via upregulating linolenic acid metabolism

doi: 10.1016/j.isci.2025.113892

Figure Lengend Snippet: Baicalin combined with metformin improved the blood glucose levels in metformin non-responsive mice by activating the AMPK/ACC/CPT1 pathway (A–F) The serum concentrations of (A) TC, (B) LDL-C, (C) HDL-C, (D) IL-1β, (E) IL-6, and (F) IL-10. (G) Hematoxylin-eosin (H&E) staining of the liver, pancreas, epididymal adipose tissues, and oil red O staining of the liver (magnification, 30×; scale bars, 100 μm). (H) Western blot analysis of pAMPK (Thr172), ACC, and CPT1 proteins in the livers of mice after administration. Data are expressed as the mean ± SD (A‒F, n = 6–8; H, n = 3); ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001, as determined by one-way ANOVA with Holm-Sidak’s post hoc test (A, B, C, E, F, and H) and Kruskal-Wallis test (D). NCD, normal chow diet; NR, non-response; Met, metformin; BA, baicalin; TC, total cholesterol; LDL-C, low-density lipoprotein cholesterol; HDL-C, high-density lipoprotein cholesterol; IL-1β, interleukin-1β; IL-6, interleukin-6; IL-10, interleukin-10; AMPK, AMP-activated protein kinase; ACC, acetyl-CoA carboxylase; CPT1, carnitine palmitoyl transferase 1.

Article Snippet: These membranes were then incubated at 4°C overnight with primary antibodies against AMP-activated protein kinase (AMPK) (Proteintech, Cat# 10929-2-AP, RRID: AB_2169568 , 1:4,000), insulin receptor substrate 1 (IRS1) (Proteintech, Cat# 17509-1-AP, RRID: AB_10596914 , 1:1,000), phospho-AMPKα (Thr172) (40H9) (Cell Signaling Technology, Cat# 2535, RRID: AB_331250 , 1:1,000), phospho-IRS-1 (Ser636/639) (Cell Signaling Technology, Cat# 2388, RRID: AB_330339 , 1:1,000), phosphoenolpyruvate carboxykinase (PCK) (Proteintech, Cat# 16754-1-AP, RRID: AB_2160031 , 1:30,000), glucose-6-phosphatase (G6PC) (Proteintech, Cat# 66860-1-Ig, RRID: AB_2882199 , 1:5,000), mammalian target of rapamycin (mTOR) (Proteintech, Cat# 66888-1-Ig, RRID: AB_2882219 , 1:30,000), fatty acid desaturase 2 (FADS2) (Proteintech, Cat# 28034-1-AP, RRID: AB_2918142 , 1:4,000), phospholipase A2 group (PLA2G) (Proteintech, Cat# 18088-1-AP, RRID: AB_10859777 , 1:1,000), acetyl-CoA Carboxylase 1 (ACC1) (Proteintech, Cat# 21923-1-AP, RRID: AB_11042445 , 1:4,000), carnitine palmitoyl transferase 1 (CPT1) (Proteintech, Cat# 15184-1-AP, RRID: AB_2084676 , 1:50,000), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (Proteintech, Cat# 10494-1-AP, RRID: AB_2263076 , 1:20,000), β-tubulin (Proteintech, Cat# 10094-1-AP, RRID: AB_2210695 , 1:10,000).

Techniques: Staining, Western Blot

Journal: iScience

Article Title: Roseburia hominis enriched by baicalin reverses the non-response to metformin via upregulating linolenic acid metabolism

doi: 10.1016/j.isci.2025.113892

Figure Lengend Snippet: R. hominis treatment reversed the metformin NR phenotype in NR mice (A) Experimental protocol for administration of R. hominis in mice. (B) Oral glucose tolerance test (OGTT) curve and its area under the curve (AUC). (C) Insulin tolerance test (ITT) curve and its AUC. (D) Homeostasis model assessment of insulin resistance (HOMA-IR) after drug R. hominis . (E–J) The serum concentrations of (E) TC, (F) LDL-C, (G) HDL-C, (H) IL-1β, (I) IL-6, and (J) IL-10. (K) Hematoxylin-eosin (H&E) staining of the liver, pancreas, epididymal adipose tissues, and oil red O staining of the liver (magnification, 30×; scale bars, 100 μm). (L) Western blot analysis of AMPK/ACC/CPT1 proteins in the livers of mice after administration. Data are expressed as the mean ± SD ( n = 7–8); ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001, as determined by one-way ANOVA with Holm-Sidak’s post hoc test. ANOVA, analysis of variance; ABX, antibiotic mixed; FMT, fecal microbial transplantation; NCD, normal chow diet; R.h, Roseburia hominis ; TC, total cholesterol; LDL-C, low-density lipoprotein cholesterol; HDL-C, high-density lipoprotein cholesterol; IL-1β, interleukin-1β; IL-6, interleukin-6; IL-10, interleukin-10; AMPK, AMP-activated protein kinase; ACC, acetyl-CoA carboxylase; CPT1, carnitine palmitoyl transferase 1.

Article Snippet: These membranes were then incubated at 4°C overnight with primary antibodies against AMP-activated protein kinase (AMPK) (Proteintech, Cat# 10929-2-AP, RRID: AB_2169568 , 1:4,000), insulin receptor substrate 1 (IRS1) (Proteintech, Cat# 17509-1-AP, RRID: AB_10596914 , 1:1,000), phospho-AMPKα (Thr172) (40H9) (Cell Signaling Technology, Cat# 2535, RRID: AB_331250 , 1:1,000), phospho-IRS-1 (Ser636/639) (Cell Signaling Technology, Cat# 2388, RRID: AB_330339 , 1:1,000), phosphoenolpyruvate carboxykinase (PCK) (Proteintech, Cat# 16754-1-AP, RRID: AB_2160031 , 1:30,000), glucose-6-phosphatase (G6PC) (Proteintech, Cat# 66860-1-Ig, RRID: AB_2882199 , 1:5,000), mammalian target of rapamycin (mTOR) (Proteintech, Cat# 66888-1-Ig, RRID: AB_2882219 , 1:30,000), fatty acid desaturase 2 (FADS2) (Proteintech, Cat# 28034-1-AP, RRID: AB_2918142 , 1:4,000), phospholipase A2 group (PLA2G) (Proteintech, Cat# 18088-1-AP, RRID: AB_10859777 , 1:1,000), acetyl-CoA Carboxylase 1 (ACC1) (Proteintech, Cat# 21923-1-AP, RRID: AB_11042445 , 1:4,000), carnitine palmitoyl transferase 1 (CPT1) (Proteintech, Cat# 15184-1-AP, RRID: AB_2084676 , 1:50,000), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (Proteintech, Cat# 10494-1-AP, RRID: AB_2263076 , 1:20,000), β-tubulin (Proteintech, Cat# 10094-1-AP, RRID: AB_2210695 , 1:10,000).

Techniques: Staining, Western Blot, Transplantation Assay

Journal: Translational Neurodegeneration

Article Title: Deficient AMPK-SENP1-Sirt3 signaling impairs mitochondrial complex I function in Parkinson’s disease model

doi: 10.1186/s40035-025-00489-2

Figure Lengend Snippet: MPTP/MPP + attenuates AMPK activity and diminishes mitochondrial translocation of SENP1. a Relative intensities of AMPK (total lysates), P-AMPK/AMPK (total lysates), SENP1 (total lysates), SENP1 (mitochondrial lysates) and SUMO1-conjugated proteins (mitochondrial lysates) in the ventral midbrain of 3-month-old WT mice injected with MPTP or saline. Data were from 3 biological replicates, and are expressed as mean ± SD, n = 3 mice. b Left, western blotting for SENP1 (total lysates), SENP1 (mitochondrial lysates), AMPK (total lysates) and P-AMPK (total lysates) from SH-SY5Y cells treated with culture medium, MPP + , or MPP + and metformin (2 mmol/L) for 24 h, as well as blotting for SUMO1 in anti-Sirt3 immunoprecipitant from SH-SY5Y cell mitochondrial lysates. The total levels of Sirt3, SUMO1-conjugated proteins, and acetyl-conjugated proteins in mitochondrial lysates were also detected. Right, relative intensities of AMPK (total lysates), P-AMPK/AMPK (total lysates), SENP1 (total lysates), SENP1 (mitochondrial lysates), SUMOylated Sirt3 to total Sirt3 (mitochondrial lysates), SUMO1-conjugated proteins (mitochondrial lysates) and acetyl-conjugated proteins (mitochondrial lysates) from 3 biological replicates. Data expressed as mean ± SD, n = 3. c SH-SY5Y cells were treated with culture medium, MPP + , or MPP + + metformin for 24 h. CCK8 assay was performed to measure cell viability. Data expressed as mean ± SD, n = 3. One-way ANOVA with Tukey's post-hoc test; * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001

Article Snippet: For Western blotting assays, the following primary antibodies were used: anti-GAPDH (1:1000; Cell Signaling, 5174; Danvers, MA), anti-ATP5A (1:300; Abcam, 110273; Cambridge, MA), anti-Sirt3 (1:1000; Cell Signaling, 5490), anti-NDUFS2 (1:4000; Abcam, 110249), anti-NDUFA5 (1:1000; Proteintech, 16640; Rosemont, IL), anti-NDUFS3 (1:2000; Abcam, 14711), anti-NDUFA9 (1:1000; Abcam, 14713), anti-NDUFV1 (1:2000; Proteintech, 11238), anti-NDUFS7 (1:2000; Proteintech, 15728), anti-SENP1 (1:1000; Abcam, 108981), anti-AMP-activated protein kinase (AMPK) (1:1000; Cell Signaling, 2532), anti-SUMO1 (1:1000, custom-made by Cheng's team) [ ], anti-phosphorylated AMPK (P-AMPK) (1:1000; Cell Signaling, 2535), anti-Pan-AcK (Acetyl-conjugated proteins; 1:1000; Cell Signaling, 9441), and anti-tyrosine hydroxylase (TH)(1:200; Abcam, 112).

Techniques: Activity Assay, Translocation Assay, Injection, Saline, Western Blot, CCK-8 Assay