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Oxidative stress activated AMPK in macrophages. (A) The H 2 O 2 levels in renal tissues at day 0, 3, 7, and 28 post-IRI were detected (n = 4). (B) Representative flow cytometry histogram and (C) quantification of phospho-AMPK levels in renal macrophages (n = 5). (D) Immunoblots and (E) quantification of phospho-AMPK levels in WT BMDMs treated with PBS or 50 μM H 2 O 2 for 30 min. (F) Representative flow cytometry histogram and (G) quantification of phospho-AMPK levels in BMDMs exposed to TGF-β, IL-6, H 2 O 2 , and hypoxia (n = 5). (H) Intracellular calcium levels were detected by Fluo-4 stain using flow cytometry (n = 4). (I) Representative flow cytometry histogram and (J) quantification of phospho-AMPK levels in BMDMs treated with 50 μM H 2 O 2 or 50 μM H 2 O 2 + 10 <t>μM</t> <t>STO-609</t> (n = 5). The results represent mean ± SEM. ∗p < 0.05, ∗∗∗p < 0.001, NS no significance.
Camkkβ Inhibitor Sto 609, supplied by Selleck Chemicals, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Oxidative stress activated AMPK in macrophages. (A) The H 2 O 2 levels in renal tissues at day 0, 3, 7, and 28 post-IRI were detected (n = 4). (B) Representative flow cytometry histogram and (C) quantification of phospho-AMPK levels in renal macrophages (n = 5). (D) Immunoblots and (E) quantification of phospho-AMPK levels in WT BMDMs treated with PBS or 50 μM H 2 O 2 for 30 min. (F) Representative flow cytometry histogram and (G) quantification of phospho-AMPK levels in BMDMs exposed to TGF-β, IL-6, H 2 O 2 , and hypoxia (n = 5). (H) Intracellular calcium levels were detected by Fluo-4 stain using flow cytometry (n = 4). (I) Representative flow cytometry histogram and (J) quantification of phospho-AMPK levels in BMDMs treated with 50 μM H 2 O 2 or 50 μM H 2 O 2 + 10 <t>μM</t> <t>STO-609</t> (n = 5). The results represent mean ± SEM. ∗p < 0.05, ∗∗∗p < 0.001, NS no significance.
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Oxidative stress activated AMPK in macrophages. (A) The H 2 O 2 levels in renal tissues at day 0, 3, 7, and 28 post-IRI were detected (n = 4). (B) Representative flow cytometry histogram and (C) quantification of phospho-AMPK levels in renal macrophages (n = 5). (D) Immunoblots and (E) quantification of phospho-AMPK levels in WT BMDMs treated with PBS or 50 μM H 2 O 2 for 30 min. (F) Representative flow cytometry histogram and (G) quantification of phospho-AMPK levels in BMDMs exposed to TGF-β, IL-6, H 2 O 2 , and hypoxia (n = 5). (H) Intracellular calcium levels were detected by Fluo-4 stain using flow cytometry (n = 4). (I) Representative flow cytometry histogram and (J) quantification of phospho-AMPK levels in BMDMs treated with 50 μM H 2 O 2 or 50 μM H 2 O 2 + 10 <t>μM</t> <t>STO-609</t> (n = 5). The results represent mean ± SEM. ∗p < 0.05, ∗∗∗p < 0.001, NS no significance.
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Oxidative stress activated AMPK in macrophages. (A) The H 2 O 2 levels in renal tissues at day 0, 3, 7, and 28 post-IRI were detected (n = 4). (B) Representative flow cytometry histogram and (C) quantification of phospho-AMPK levels in renal macrophages (n = 5). (D) Immunoblots and (E) quantification of phospho-AMPK levels in WT BMDMs treated with PBS or 50 μM H 2 O 2 for 30 min. (F) Representative flow cytometry histogram and (G) quantification of phospho-AMPK levels in BMDMs exposed to TGF-β, IL-6, H 2 O 2 , and hypoxia (n = 5). (H) Intracellular calcium levels were detected by Fluo-4 stain using flow cytometry (n = 4). (I) Representative flow cytometry histogram and (J) quantification of phospho-AMPK levels in BMDMs treated with 50 μM H 2 O 2 or 50 μM H 2 O 2 + 10 <t>μM</t> <t>STO-609</t> (n = 5). The results represent mean ± SEM. ∗p < 0.05, ∗∗∗p < 0.001, NS no significance.
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Oxidative stress activated AMPK in macrophages. (A) The H 2 O 2 levels in renal tissues at day 0, 3, 7, and 28 post-IRI were detected (n = 4). (B) Representative flow cytometry histogram and (C) quantification of phospho-AMPK levels in renal macrophages (n = 5). (D) Immunoblots and (E) quantification of phospho-AMPK levels in WT BMDMs treated with PBS or 50 μM H 2 O 2 for 30 min. (F) Representative flow cytometry histogram and (G) quantification of phospho-AMPK levels in BMDMs exposed to TGF-β, IL-6, H 2 O 2 , and hypoxia (n = 5). (H) Intracellular calcium levels were detected by Fluo-4 stain using flow cytometry (n = 4). (I) Representative flow cytometry histogram and (J) quantification of phospho-AMPK levels in BMDMs treated with 50 μM H 2 O 2 or 50 μM H 2 O 2 + 10 μM STO-609 (n = 5). The results represent mean ± SEM. ∗p < 0.05, ∗∗∗p < 0.001, NS no significance.

Journal: Redox Biology

Article Title: Macrophage AMPK activated by oxidative stress drives profibrotic crosstalk with tubular cells to accelerate renal fibrosis after ischemic and reperfusion injury

doi: 10.1016/j.redox.2025.104002

Figure Lengend Snippet: Oxidative stress activated AMPK in macrophages. (A) The H 2 O 2 levels in renal tissues at day 0, 3, 7, and 28 post-IRI were detected (n = 4). (B) Representative flow cytometry histogram and (C) quantification of phospho-AMPK levels in renal macrophages (n = 5). (D) Immunoblots and (E) quantification of phospho-AMPK levels in WT BMDMs treated with PBS or 50 μM H 2 O 2 for 30 min. (F) Representative flow cytometry histogram and (G) quantification of phospho-AMPK levels in BMDMs exposed to TGF-β, IL-6, H 2 O 2 , and hypoxia (n = 5). (H) Intracellular calcium levels were detected by Fluo-4 stain using flow cytometry (n = 4). (I) Representative flow cytometry histogram and (J) quantification of phospho-AMPK levels in BMDMs treated with 50 μM H 2 O 2 or 50 μM H 2 O 2 + 10 μM STO-609 (n = 5). The results represent mean ± SEM. ∗p < 0.05, ∗∗∗p < 0.001, NS no significance.

Article Snippet: For immunoblot analysis of phospho-AMPKα, BMDMs were treated with 50 μM H 2 O 2 for 30 min. To investigate calcium-dependent signaling, BMDMs were pretreated with the CaMKKβ inhibitor STO-609 (Selleck, S8274, 10 μM) or vehicle for 1 h prior to H 2 O 2 stimulation, and AMPK phosphorylation was quantified by flow cytometry.

Techniques: Flow Cytometry, Western Blot, Staining